@article{mosichuk_wimbish_poplawski_birkenheuer_harrell_pierce_2024, title={Case report: Severe hepatopathy following rivaroxaban administration in a dog}, volume={11}, ISSN={["2297-1769"]}, DOI={10.3389/fvets.2024.1364677}, abstractNote={Rivaroxaban, a specific factor Xa inhibitor and commonly utilized anticoagulant, has been known to cause hepatotoxicity and liver failure in humans. Although rivaroxaban is frequently used in veterinary medicine, hepatotoxicity has not been previously reported in dogs. The current case report describes a dog that developed severe hepatopathy following rivaroxaban administration for a large right pulmonary artery thrombus. An estimated 6-year-old spayed female mixed-breed dog developed anorexia and lethargy 9 days after rivaroxaban administration began. Subsequent labwork revealed severe hepatocellular hepatopathy, and rivaroxaban was discontinued. Additional diagnostics did not reveal an underlying etiology, although hepatic cytology could be consistent with a toxic injury. The hepatopathy and clinical signs improved after rivaroxaban was discontinued. The time to onset, type of hepatopathy, and time to resolution were all similar to those reported for human cases. This case provides precedence to advocate for improved and closer monitoring of dogs receiving factor Xa inhibitors. In cases of suspected hepatotoxicity with no other identifiable cause, a risk–benefit analysis should be performed, and discontinuation of rivaroxaban administration or alternative anticoagulant medications should be considered.}, journal={FRONTIERS IN VETERINARY SCIENCE}, author={Mosichuk, Allison P. and Wimbish, Candace and Poplawski, Kristen and Birkenheuer, Adam and Harrell, Karyn and Pierce, Kursten V.}, year={2024}, month={Apr} }
 @article{weerarathne_maker_huang_taylor_cowan_hyatt_selvan_shatnawi_thomas_meinkoth_et al._2023, title={A Novel Vaccine Strategy to Prevent Cytauxzoonosis in Domestic Cats}, volume={11}, ISSN={["2076-393X"]}, DOI={10.3390/vaccines11030573}, abstractNote={Cytauxzoonosis is caused by Cytauxzoon felis (C. felis), a tick-borne parasite that causes severe disease in domestic cats in the United States. Currently, there is no vaccine to prevent this fatal disease, as traditional vaccine development strategies have been limited by the inability to culture this parasite in vitro. Here, we used a replication-defective human adenoviral vector (AdHu5) to deliver C. felis-specific immunogenic antigens and induce a cell-mediated and humoral immune response in cats. Cats (n = 6 per group) received either the vaccine or placebo in two doses, 4 weeks apart, followed by experimental challenge with C. felis at 5 weeks post-second dose. While the vaccine induced significant cell-mediated and humoral immune responses in immunized cats, it did not ultimately prevent infection with C. felis. However, immunization significantly delayed the onset of clinical signs and reduced febrility during C. felis infection. This AdHu5 vaccine platform shows promising results as a vaccination strategy against cytauxzoonosis.}, number={3}, journal={VACCINES}, author={Weerarathne, Pabasara and Maker, Rebekah and Huang, Chaoqun and Taylor, Brianne and Cowan, Shannon R. and Hyatt, Julia and Selvan, Miruthula Tamil and Shatnawi, Shoroq and Thomas, Jennifer E. and Meinkoth, James H. and et al.}, year={2023}, month={Mar} }
 @article{yang_ladouceur_baumgartner_marr_karounos_robertson_whitehurst_miller_birkenheuer_2023, title={A practical protocol to prepare paraffin-embedded whole tick histology sections}, volume={14}, ISSN={["1877-9603"]}, url={https://doi.org/10.1016/j.ttbdis.2023.102162}, DOI={10.1016/j.ttbdis.2023.102162}, abstractNote={Ticks are important ectoparasites that are capable of transmitting multiple classes of pathogens and are currently linked with many emerging tick-borne diseases worldwide. With increasing occurrences of tick-borne diseases in both humans and veterinary species, there is a continuous need to further our understanding of ticks and the pathogens they transmit. Whole tick histology provides a full scope of the tick internal anatomy, allowing researchers to examine multiple organs of interest in a single section. This is in contrast to other techniques that are more commonly utilized in tick-borne disease research, such as electron microscopy and light microscopy of individual organs. There is a lack of literature describing a practical technique to process whole tick histologic sections. Therefore, the current study aims to provide researchers with a workable protocol to prepare high quality paraffin-embedded whole tick histology sections. Amblyomma americanum adults were used as an example species for this study. After a series of pilot experiments using a combination of various fixatives, softening agents and processing techniques, we elected to compare two common fixatives, 10% neutral-buffered formalin (NBF) and Bouin's solution for whole ticks. Equal numbers of A. americanum unfed adults (n = 10/fixative) were processed identically and their whole tick histology coronal sections were individually scored. Higher scores were assigned to whole tick sections that contained more internal organs that are crucial for tick-borne disease research (e.g. salivary glands and midgut), high integrity of tissues and exoskeleton on the section, and good fixation and staining quality of the tissues. The mean total scores for Bouin's-fixed ticks were significantly higher compared to NBF-fixed ticks (p = 0.001). To further assess our preferred technique, we also demonstrated the feasibility of producing high quality whole tick sections for three other common tick species of medical importance (Rhipicephalus sanguineus, Ixodes scapularis, and Dermacentor variabilis) using Bouin's solution. While this technique may require further optimization for other tick species, we described a feasible protocol that uses commonly available tools, reagents and standard histologic equipment. This should allow any investigator to easily make adjustments to this protocol as needed based on their experimental goals.}, number={4}, journal={TICKS AND TICK-BORNE DISEASES}, author={Yang, Tzushan S. and LaDouceur, Elise E. B. and Baumgartner, Wes A. and Marr, Henry S. and Karounos, Michael and Robertson, James and Whitehurst, Nathan and Miller, Laura S. and Birkenheuer, Adam J.}, year={2023}, month={Jul} }
 @article{anderson_birkenheuer_moore_kendall_2023, title={A retrospective study of vector borne disease prevalence among anemic dogs in North Carolina}, volume={18}, ISSN={["1932-6203"]}, DOI={10.1371/journal.pone.0293901}, abstractNote={
Background
Anemia is an important cause of morbidity and mortality in dogs. Further understanding of the prevalence of vector borne diseases (VBD) in anemic dogs is needed.
}, number={11}, journal={PLOS ONE}, author={Anderson, Katie L. and Birkenheuer, Adam and Moore, George E. and Kendall, Allison}, year={2023}, month={Nov} }
 @article{yang_reichard_thomas_miller_marr_karounos_bell_birkenheuer_2023, title={Cytauxzoon felis in salivary glands of Amblyomma americanum}, volume={14}, ISSN={["1877-9603"]}, DOI={10.1016/j.ttbdis.2022.102056}, abstractNote={Cytauxzoon felis is a tick-borne piroplasmid hemoparasite that causes life-threatening disease in cats. Despite the critical role that ticks play in pathogen transmission, our knowledge regarding the C. felis life cycle remains limited to the feline hosts. Specific life stages of C. felis within the tick host have never been visualized microscopically and previous investigations have been limited to molecular detection by polymerase chain reaction (PCR). Sporozoites are the infectious stage of piroplasmids that are transmitted by ticks. In other tick-borne piroplasmids, sporozoite-based vaccines play a key role in disease prevention and management. We believe sporozoites have similar potential for cytauxzoonosis. Therefore, the objective of this study was to use different molecular and microscopic techniques to detect and evaluate C. felis sporozoites in tick salivary glands (SG). A total of 140 Amblyomma americanum adults that were fed on C. felis-infected cats as nymphs were included for this study. Specifically, dissected SGs were quartered and subjected to C. felis RT-PCR, RNAscope® in situ hybridization (ISH), histology, direct azure staining, and transmission electron microscopy (TEM). Cytauxzoon felis RT-PCR was also performed on half tick (HT) carcasses after SG dissection. Cytauxzoon felis RNA was detected in SGs of 17/140 ticks. Of these, 7/17 ticks had microscopic visualization via ISH and/or TEM. The remaining 10/17 ticks had only molecular detection of C. felis in SGs via RT-PCR without visualization. Cytauxzoon felis RNA was detected solely in HT carcasses via RT-PCR in 9/140 ticks. In ISH-positive tick SGs, hybridization signals were present in cytoplasms of SG acinar cells. TEM captured rare C. felis organisms with characteristic ultrastructural features of sporozoites. This study describes the first direct visualization of any developing stage of C. felis in ticks. Forthcoming studies should employ a combination of molecular and microscopic techniques to investigate the C. felis life cycle in A. americanum.}, number={1}, journal={TICKS AND TICK-BORNE DISEASES}, author={Yang, Tzushan S. and Reichard, Mason V and Thomas, Jennifer E. and Miller, Laura S. and Marr, Henry S. and Karounos, Michael and Bell, Aaron J. and Birkenheuer, Adam J.}, year={2023}, month={Jan} }
 @article{grady_ernst_secoura_price_birkenheuer_vaden_lidbury_gould_steiner_tolbert_2023, title={Gastric pH and serum gastrin concentration in age-matched healthy dogs and dogs with chronic kidney disease}, volume={10}, ISSN={["1939-1676"]}, DOI={10.1111/jvim.16907}, abstractNote={Abstract}, journal={JOURNAL OF VETERINARY INTERNAL MEDICINE}, author={Grady, Kylie and Ernst, Eli and Secoura, Patricia L. and Price, Josh and Birkenheuer, Adam and Vaden, Shelly L. and Lidbury, Jonathan and Gould, Emily and Steiner, Jeorg M. and Tolbert, M. Katherine}, year={2023}, month={Oct} }
 @article{yang_reichard_thomas_marr_karounos_hyatt_miller_birkenheuer_2023, title={Transmission of Cytauxzoon felis by injection of Amblyomma americanum salivary glands}, volume={95}, ISSN={["1873-0329"]}, DOI={10.1016/j.parint.2023.102753}, abstractNote={Cytauxzoonosis is a life-threatening disease of cats, caused by the tick-borne piroplasmid hemoparasite, Cytauxzoon felis. Current experimental models for cytauxzoonosis rely on either tick transmission or direct injection of infected cat tissues. These models require researchers to directly work with infected ticks or use cats with acute cytauxzoonosis. To improve the feasibility and accessibility, there is a need to establish sharable resources among researchers. In related piroplasmid parasites, sporozoite-based inoculums are routinely produced from tick salivary glands, cryopreserved and distributed to other investigators and facilities. For these parasites, sporozoites have been the basis for vaccine development and in vitro cultivation, both of which remain lacking for C. felis research. If infectious sporozoites can be similarly isolated for C. felis, it would significantly broaden our capabilities to study this parasite. Aims of this study was to determine if C. felis sporozoites inoculums collected from the salivary glands of Amblyomma americanum ticks were capable of inducing cytauxzoonosis in naïve cats. A. americanum nymphs were acquisition-fed on a donor cat chronically infected with C. felis and allowed to molt to adults. Four groups of adult ticks (n = 50/group) were either stimulation-fed for 4 days on naïve cats or were heated at 37 °C for 4 days. After these treatments, salivary glands (SG) of each group of ticks were collected to create inoculums. Infectivity of these inoculums was then tested by subcutaneous injection into naïve cats. The two naïve cats used for stimulation feeding and as controls both developed cytauxzoonosis, indicating these groups of ticks were capable of producing infectious sporozoites. Of the 2 cats that were injected with SGs from the stimulation-fed ticks, one cat developed cytauxzoonosis and C. felis infection was confirmed by both light microscopy and PCR. The other cat did not develop cytauxzoonosis and only had equivocal evidence of infection. Neither cat injected with SGs from the heated ticks developed cytauxzoonosis. One of these cats had equivocal evidence of infection and one had no evidence of infection. This study validates the feasibility of collecting infectious sporozoites from C. felis-infected ticks that can be used to infect naïve cats. While this model requires further optimization, it has the potential to expand resources to study C. felis and further advance research in this field.}, journal={PARASITOLOGY INTERNATIONAL}, author={Yang, Tzushan S. and Reichard, Mason V and Thomas, Jennifer E. and Marr, Henry S. and Karounos, Michael and Hyatt, Julia and Miller, Craig and Birkenheuer, Adam J.}, year={2023}, month={Aug} }
 @article{kao_spainhour_cowan_nafe_birkenheuer_reichard_miller_2022, title={A Serodiagnostic IgM ELISA to Detect Acute Cytauxzoonosis}, volume={11}, ISSN={["2076-0817"]}, DOI={10.3390/pathogens11101183}, abstractNote={Cytauxzoonosis is a tick-borne infectious disease affecting domestic cats with high mortality and limited treatment modalities. Because early diagnosis and therapeutic intervention are crucial to survival of infected cats, the objective of this study was to develop an ELISA capable of detecting cytauxzoonosis and differentiating acute vs. chronic infection in clinical feline blood samples. A microsphere immunoassay (MIA) was developed to evaluate the production of Cytauxzoon felis-specific IgM and IgG antibodies in serial plasma samples from cats with experimental C. felis infection by targeting a C. felis-specific transmembrane protein (c88). Recombinant c88 protein was utilized to develop indirect ELISAs to detect IgM and IgG antibodies in clinical plasma samples from: PCR-positive cats with acute C. felis infection (n = 36), C. felis-negative cats with pyrexia (n = 10), healthy C. felis-negative cats (n = 22), and chronic C. felis carriers (n = 4). Anti-c88 IgM antibodies were detectable at day 12 post-tick infestation in cats with experimental C. felis infection (within 24 hours of developing clinical signs), while anti-c88 IgG was detectable at day 15 post-tick infestation – indicating IgM could be used to detect early infection. Using a cut-off value of 19.85 percent positive, the C. felis IgM ELISA detected acute cytauxzoonosis in 94.44% (34/36) of cats presented with clinical signs of acute cytauxzoonosis with 100% specificity (indicating a “Strong Positive” result). When a lower cutoff of 8.60 percent positive was used, cytauxzoonosis was detected in the 2 remaining PCR-positive cats with 87.88% specificity (indicating of a “Weak Positive” result). One C. felis-negative, febrile cat had high IgG, and chronic carriers had variable IgM and IgG results. Combined interpretation of IgM and IgG ELISAs did not reliably differentiate acute vs. chronic infection. While further validation on assay performance is needed, the C. felis IgM ELISA is a promising test to detect acute cytauxzoonosis and can be utilized to develop a point-of-care test for clinical use.}, number={10}, journal={PATHOGENS}, author={Kao, Yun-Fan and Spainhour, Rebecca and Cowan, Shannon R. and Nafe, Laura and Birkenheuer, Adam and Reichard, Mason V and Miller, Craig A.}, year={2022}, month={Oct} }
 @article{dear_birkenheuer_2022, title={Babesia in North America An Update}, volume={52}, ISSN={["1878-1306"]}, DOI={10.1016/j.cvsm.2022.07.016}, abstractNote={Canine babesiosis results from infection of 1 of 5 identified protozoal species in the United States ( Babesia conradae , Babesia sp. “coco,” Babesia gibsoni , Babesia vogeli , and Babesia vulpes ). They are part of the Apicomplexa family of protozoa and are obligate intraerythrocytic parasites. Domestic and wild canids are suspected of being intermediate hosts. This updated article aims to provide practical guidance about the clinical manifestations of disease, treatment options, and outcomes. In addition, the authors hope to provide some clarity about the taxonomy and nomenclature of these organisms, as they have undergone multiple changes since their initial discovery.}, number={6}, journal={VETERINARY CLINICS OF NORTH AMERICA-SMALL ANIMAL PRACTICE}, author={Dear, Jonathan D. and Birkenheuer, Adam}, year={2022}, month={Nov}, pages={1193–1209} }
 @article{cerreta_yang_ramsay_birkenheuer_rahoi_qurollo_wilson_cushing_2022, title={DETECTION OF VECTOR-BORNE INFECTIONS IN LIONS AND TIGERS AT TWO ZOOS IN TENNESSEE AND OKLAHOMA, USA}, volume={53}, ISSN={["1937-2825"]}, DOI={10.1638/2020-0199}, abstractNote={Abstract: Protozoal and bacterial vector-borne infections are frequently diagnosed in domestic felids. However, with the exception of Mycoplasma haemofelis and Cytauxzoon felis, their occurrence in managed nondomestic felids housed in the United States is largely unknown. Following a case in February 2020 of fulminant cytauxzoonosis in an African lion (Panthera leo), EDTA–whole blood samples were collected opportunistically from February 2020 through June 2020 from 34 adult tigers (Panthera tigris) and eight adult African lions from the same sanctuary in eastern Tennessee as well as 14 adult tigers from a zoo in southern Oklahoma. Samples were analyzed for Cytauxzoon felis, Bartonella spp., hemotropic Mycoplasma, Rickettsia spp., Anaplasma spp., Ehrlichia spp., Babesia spp., and Hepatozoon spp. DNA by PCR amplification. All animals were asymptomatic at the time of collection. None of the Oklahoma animals were positive for vector-borne organisms, but these pathogens were detected in tigers at the Tennessee facility, including Cytauxzoon felis (11.8%), “Candidatus Mycoplasma haemominutum” (5.9%), and Ehrlichia ewingii (2.9%). During the study period, two animals developed clinical signs of cytauxzoonosis and were assessed for vector-borne infections as part of their diagnostic evaluation. This study documents the presence of tick-borne diseases in managed nondomestic felids in the southeastern United States and underscores that ectoparasite control measures should be practiced to minimize exposure of carnivores in managed care.}, number={1}, journal={JOURNAL OF ZOO AND WILDLIFE MEDICINE}, author={Cerreta, Anthony J. and Yang, Tzushan S. and Ramsay, Edward C. and Birkenheuer, Adam J. and Rahoi, Dane and Qurollo, Barbara and Wilson, James and Cushing, Andrew C.}, year={2022}, month={Mar}, pages={50–59} }
 @article{yang_reichard_marr_cohn_nafe_whitehurst_birkenheuer_2022, title={Direct injection of Amblyomma americanum ticks with Cytauxzoon felis}, volume={13}, ISSN={["1877-9603"]}, DOI={10.1016/j.ttbdis.2021.101847}, abstractNote={Cytauxzoon felis is a tick-borne hemoprotozoan parasite that causes life-threatening disease in domestic cats in the United States. Currently, the platforms for C. felis research are limited to natural or experimental infection of domestic cats. This study aims to develop an alternative model by infecting Amblyomma americanum ticks with C. felis via direct injection. Amblyomma americanum adults were injected with C. felis-infected feline erythrocytes through two routes: directly into the digestive tract through the anal pore (IA injection), or percutaneously into the tick hemocoel (IH injection). RNAscope® in situ hybridization (ISH) was used to visualize the parasites within the ticks at different time points after injection. Four months after injection, ticks were divided into 3 infestation groups based on injection methods and inoculum type and fed on 3 naïve cats to assess the ticks’ ability to transmit C. felis. Prior to the transmission challenge, selected ticks from each infestation group were tested for C. felis RNA via reverse transcription-PCR (RT-PCR). In both IA- and IH-injected ticks, ISH signals were observed in ticks up to 3 weeks after injection. The number of hybridization signals notably decreased over time, and no signals were detected by 4 months after injection. Prior to the transmission challenge, 37–57% of the sampled ticks were positive for C. felis RNA via RT-PCR. While the majority of injected ticks successfully attached and fed to repletion on all 3 cats during the transmission challenge, none of the cats became infected with C. felis. These results suggest that injected C. felis remained alive in ticks but was unable to progress to infective sporozoites after injection. It is unclear why this infection technique had been successful for other closely related tick-borne hemoprotozoa and not for C. felis. This outcome may be associated with uncharacterized differences in the C. felis life cycle, the lack of the feeding or molting in our model or absence of gametocytes in the inoculum. Nonetheless, our study demonstrated the potential of using ticks as an alternative model to study C. felis. Future improvement of a tick model for C. felis should consider other tick species for the injection model or utilize infection methods that more closely emulate the natural infection process.}, number={1}, journal={TICKS AND TICK-BORNE DISEASES}, author={Yang, Tzushan S. and Reichard, Mason V and Marr, Henry S. and Cohn, Leah A. and Nafe, Laura and Whitehurst, Nathan and Birkenheuer, Adam J.}, year={2022}, month={Jan} }
 @article{garrett_halseth_ruder_beasley_shock_birkenheuer_gabriel_fiorello_haire_olfenbuttel_et al._2022, title={Prevalence and genetic characterization of a Babesia microti-like species in the North American river otter (Lontra canadensis)}, volume={29}, ISSN={["2405-9390"]}, DOI={10.1016/j.vprsr.2022.100696}, abstractNote={A 4.5-month-old, male, North American river otter (Lontra canadensis) from Athens-Clarke County, Georgia, USA being temporarily housed at a rehabilitation facility, presented with a three-day history of lethargy, anorexia, and severe anemia. Antemortem blood smears revealed intraerythrocytic piroplasms. Supportive care and antiparasitic treatments were initiated, but the animal died three days following presentation. Gross necropsy revealed yellow discoloration of all adipose tissue throughout the carcass and a mildly enlarged, diffusely yellow to pale orange liver. Microscopically, moderate, centrilobular hepatocellular degeneration and necrosis were observed, consistent with hypoxia secondary to apparent hemolytic anemia. Piroplasms were frequently observed in red blood cells in histologic sections. The nearly full-length 18S rRNA gene sequence (1588 bp) was identical to a previously described piroplasm from North American river otters from North Carolina. Phylogenetically, based on the 18S rRNA gene sequence, the otter Babesia sp. was in a sister group with a clade that included several strains of Babesia microti-like species including Babesia sp. from badgers (Meles meles), Babesia vulpes, and Babesia sp. from raccoons (Procyon lotor). To better understand the distribution and genetic variability of this Babesia species, otters from four states in the eastern U.S. and California were tested. Overall, 30 of 57 (53%) otters were positive for Babesia sp. None of four otters from California were positive, but prevalences in eastern states were generally high, 5/9 (55%) in Georgia, 7/14 (50%) in South Carolina, 10/17 (59%) in North Carolina, and 8/13 (62%) in Pennsylvania). Partial 18S rRNA gene sequences from all populations were identical to the clinical case sequence. No Babesia sensu stricto infections were detected. There were six unique COI sequences (937 bp) detected in 18 positive otters. The most common lineage (A) was detected in 12 of 18 (67%) samples from Georgia, North Carolina, South Carolina, and Pennsylvania. Lineage B was found in two otters and the remaining lineage types were found in single otters. These six lineages were 99-99.8% similar to each other and were < 88% similar to related parasites such as B. vulpes, B. microti-like species of raccoons, B. microti, and B. rodhaini. Phylogenetically, the Babesia sp. of otters grouped together in a well-supported clade separate from a sister group including B. vulpes from fox (Vulpes vulpes) and domestic dogs. In conclusion, this report demonstrates that this piroplasm is a potential pathogen of North American river otters and the parasite is widespread in otter populations in the eastern United States.}, journal={VETERINARY PARASITOLOGY- REGIONAL STUDIES AND REPORTS}, author={Garrett, Kayla and Halseth, Ashlyn and Ruder, Mark G. and Beasley, James and Shock, Barbara and Birkenheuer, Adam J. and Gabriel, Mourad and Fiorello, Christine and Haire, M. Melanie and Olfenbuttel, Colleen and et al.}, year={2022}, month={Apr} }
 @article{hedgespeth_birkenheuer_friedenberg_olby_meurs_2021, title={A novel missense mutation of the NAT10 gene in a juvenile Schnauzer dog with chronic respiratory tract infections}, volume={35}, ISSN={["1939-1676"]}, url={https://doi.org/10.1111/jvim.16100}, DOI={10.1111/jvim.16100}, abstractNote={Abstract}, number={3}, journal={JOURNAL OF VETERINARY INTERNAL MEDICINE}, author={Hedgespeth, Barry A. and Birkenheuer, Adam J. and Friedenberg, Steven G. and Olby, Natasha J. and Meurs, Kathryn M.}, year={2021}, month={May}, pages={1542–1546} }
 @article{baneth_nachum-biala_birkenheuer_schreeg_prince_florin-christensen_schnittger_aroch_2020, title={A new piroplasmid species infecting dogs: morphological and molecular characterization and pathogeny of Babesia negevi n. sp.}, volume={13}, ISSN={["1756-3305"]}, DOI={10.1186/s13071-020-3995-5}, abstractNote={Abstract}, number={1}, journal={PARASITES & VECTORS}, author={Baneth, Gad and Nachum-Biala, Yaarit and Birkenheuer, Adam Joseph and Schreeg, Megan Elizabeth and Prince, Hagar and Florin-Christensen, Monica and Schnittger, Leonhard and Aroch, Itamar}, year={2020}, month={Apr} }
 @article{hartley_marr_birkenheuer_2020, title={Cytauxzoon felis cytochrome b gene mutation associated with atovaquone and azithromycin treatment}, volume={34}, ISSN={["1939-1676"]}, DOI={10.1111/jvim.15935}, abstractNote={Abstract}, number={6}, journal={JOURNAL OF VETERINARY INTERNAL MEDICINE}, author={Hartley, Ashley N. and Marr, Henry S. and Birkenheuer, Adam J.}, year={2020}, month={Nov}, pages={2432–2437} }
 @article{birkenheuer_buch_beall_braff_chandrashekar_2020, title={Global distribution of canine Babesia species identified by a commercial diagnostic laboratory}, volume={22}, ISSN={["2405-9390"]}, DOI={10.1016/j.vprsr.2020.100471}, abstractNote={Babesia species are important canine pathogens with a nearly worldwide distribution. Our understanding of the distribution of these parasites is continually improving. This is in large part, due to improved molecular diagnostic capabilities. However, it can be difficult to assimilate and compare previous reports from various regions due to differences in molecular methods. In this report, we characterize the results of over 100,000 canine samples from 52 different countries and territories spanning 4 continents that were submitted to a commercial diagnostic laboratory for Babesia testing by polymerase chain reaction. The same diagnostic algorithm was used for all samples and is designed to identify and differentiate B. gibsoni, B. canis, B. vogeli, B. rossi and B. conradae. Overall 3.4% of the samples submitted tested positive for the presence of Babesia sp. DNA and were differentiated to the species level. Babesia gibsoni was the most commonly identified species (48.8% of the positive results) followed by B. canis (35.2%) then B. vogeli (15.3%). Babesia gibsoni and B. vogeli were more widely distributed than B. canis, which was primarily found in Europe. This is the largest study of its type and these data provide a global overview of which Babesia species veterinarians could expect to find in their practice area.}, journal={VETERINARY PARASITOLOGY- REGIONAL STUDIES AND REPORTS}, author={Birkenheuer, Adam J. and Buch, Jesse and Beall, Melissa J. and Braff, Jennifer and Chandrashekar, Ramaswamy}, year={2020}, month={Dec} }
 @article{birkenheuer_royal_cerreta_hemstreet_lunn_gookin_mcgarvey_2020, title={Perceptions and attitudes of Small Animal Internal Medicine specialists toward the publication requirement for board certification}, volume={34}, ISSN={["1939-1676"]}, DOI={10.1111/jvim.15717}, abstractNote={Abstract}, number={2}, journal={JOURNAL OF VETERINARY INTERNAL MEDICINE}, author={Birkenheuer, Adam J. and Royal, Kenneth D. and Cerreta, Anthony and Hemstreet, Daniel and Lunn, Katharine F. and Gookin, Jody L. and McGarvey, Stephanie}, year={2020}, month={Mar}, pages={574–580} }
 @article{cohn_shaw_shoemake_birkenheuer_2020, title={Second illness due to subsequent Cytauxzoon felis infection in a domestic cat}, DOI={10.1177/2055116920908963}, abstractNote={Case summary A castrated male domestic shorthair cat from a wooded area in Missouri had recovered from typical severe cytauxzoonosis at 4 years of age, after intensive in-hospital supportive care and administration of atovaquone and azithromycin. At 11 years of age, the same cat again experienced an acute febrile illness compatible with cytauxzoonosis. Intraerythrocytic piroplasms typical of Cytauxzoon felis were identified by cytology. The owners opted for euthanasia but allowed collection of splenic and hepatic tissue for histopathologic examination. Schizont-laden macrophages were identified in both tissue specimens, confirming active cytauxzoonosis at the time of the cat’s death. }, journal={Journal of Feline Medicine and Surgery Open Reports}, author={Cohn, Leah A. and Shaw, Dan and Shoemake, Catherine and Birkenheuer, Adam J.}, year={2020} }
 @article{easley_holowaychuk_lashnits_nordone_marr_birkenheuer_2020, title={Serum procalcitonin concentrations in dogs with induced endotoxemia}, volume={1}, url={https://doi.org/10.1111/jvim.15711}, DOI={10.1111/jvim.15711}, abstractNote={Abstract}, journal={Journal of Veterinary Internal Medicine}, publisher={Wiley}, author={Easley, Frankie and Holowaychuk, Marie K. and Lashnits, Erin W. and Nordone, Shila K. and Marr, Henry and Birkenheuer, Adam J.}, year={2020} }
 @article{kendall_keenihan_kern_lindaberry_birkenheuer_moore_vaden_2020, title={Three-dimensional bladder ultrasound for estimation of urine volume in dogs compared with traditional 2-dimensional ultrasound methods}, volume={34}, ISSN={["1939-1676"]}, DOI={10.1111/jvim.15959}, abstractNote={Abstract}, number={6}, journal={JOURNAL OF VETERINARY INTERNAL MEDICINE}, author={Kendall, Allison and Keenihan, Erin and Kern, Zachary T. and Lindaberry, Crystal and Birkenheuer, Adam and Moore, George E. and Vaden, Shelly L.}, year={2020}, month={Nov}, pages={2460–2467} }
 @article{neupane_sevala_balakrishnan_marr_wilson_maggi_birkenheuer_lappin_chomel_breitschwerdt_2020, title={Validation of Bartonella henselae Western Immunoblotting for Serodiagnosis of Bartonelloses in Dogs}, volume={58}, ISSN={["1098-660X"]}, DOI={10.1128/JCM.01335-19}, abstractNote={
            Bartonella
            spp. are etiological agents of life-threatening zoonotic diseases in dogs worldwide. Due to the poor sensitivity of immunofluorescent-antibody assays (IFAs), a reliable serodiagnostic test for canine bartonelloses is of clinical importance. The utility of Western blotting (WB) for the serodiagnosis of canine bartonelloses has not been critically investigated. The objective of this study was to characterize WB immunodominant proteins that could be used to confirm a serodiagnosis of bartonelloses.
          }, number={4}, journal={JOURNAL OF CLINICAL MICROBIOLOGY}, author={Neupane, Pradeep and Sevala, Sindhura and Balakrishnan, Nandhakumar and Marr, Henry and Wilson, James and Maggi, Ricardo and Birkenheuer, Adam and Lappin, Michael and Chomel, Bruno and Breitschwerdt, Edward B.}, year={2020}, month={Apr} }
 @article{garden_kidd_mexas_chang_jeffery_blois_fogle_macneill_lubas_birkenheuer_et al._2019, title={ACVIM consensus statement on the diagnosis of immune-mediated hemolytic anemia in dogs and cats}, volume={33}, ISSN={["1939-1676"]}, DOI={10.1111/jvim.15441}, abstractNote={Immune‐mediated hemolytic anemia (IMHA) is an important cause of morbidity and mortality in dogs. IMHA also occurs in cats, although less commonly. IMHA is considered secondary when it can be attributed to an underlying disease, and as primary (idiopathic) if no cause is found. Eliminating diseases that cause IMHA may attenuate or stop immune‐mediated erythrocyte destruction, and adverse consequences of long‐term immunosuppressive treatment can be avoided. Infections, cancer, drugs, vaccines, and inflammatory processes may be underlying causes of IMHA. Evidence for these comorbidities has not been systematically evaluated, rendering evidence‐based decisions difficult. We identified and extracted data from studies published in the veterinary literature and developed a novel tool for evaluation of evidence quality, using it to assess study design, diagnostic criteria for IMHA, comorbidities, and causality. Succinct evidence summary statements were written, along with screening recommendations. Statements were refined by conducting 3 iterations of Delphi review with panel and task force members. Commentary was solicited from several professional bodies to maximize clinical applicability before the recommendations were submitted. The resulting document is intended to provide clinical guidelines for diagnosis of, and underlying disease screening for, IMHA in dogs and cats. These should be implemented with consideration of animal, owner, and geographical factors.}, number={2}, journal={JOURNAL OF VETERINARY INTERNAL MEDICINE}, author={Garden, Oliver A. and Kidd, Linda and Mexas, Angela M. and Chang, Yu-Mei and Jeffery, Unity and Blois, Shauna L. and Fogle, Jonathan E. and MacNeill, Amy L. and Lubas, George and Birkenheuer, Adam and et al.}, year={2019}, pages={313–334} }
 @article{divincenti_garner_thomas_birkenheuer_2019, title={Babesia sp. infection in a zoo-housed polar bear (Ursus maritimus)}, DOI={10.1016/j.vprsr.2019.100350}, abstractNote={A 28-year-old female polar bear (Ursus maritimus) housed in a zoo in Upstate New York presented with acute inappetence and lethargy. The bear's condition rapidly deteriorated, and because laboratory testing indicated severe hepatic and renal disease, the bear was humanely euthanized. Examination of a blood smear from a sample collected just prior to euthanasia revealed the presence of intra-erythrocytic inclusions, which were identified as Babesia sp. by PCR. Although it is unclear if babesiosis contributed to this bear's clinical signs, this is the first report of Babesia sp. infection in this species. Zoological institutions exhibiting polar bears and located in tick-endemic areas, as well as managers of wild populations, should be aware of this species' susceptibility to babesiosis.}, journal={Veterinary Parasitology: Regional Studies and Reports}, author={DiVincenti, Louis and Garner, Michael and Thomas, Brittany and Birkenheuer, Adam}, year={2019} }
 @article{naor_lindemann_schreeg_marr_birkenheuer_carpenter_ryseff_2019, title={Clinical, morphological, and molecular characterization of an undetermined Babesia species in a maned wolf (Chrysocyon brachyurus)}, volume={10}, ISSN={["1877-9603"]}, DOI={10.1016/j.ttbdis.2018.09.005}, abstractNote={A possible novel Babesia species infection of a maned wolf (Chrysocyon brachyurus) was first reported in 2012. The current case details a confirmed report of a maned wolf with infection by an undetermined species of Babesia. As the mortality and morbidity of babesiosis is high, this may become a significant concern to captive maned wolves, which are considered a near-threatened species by the World Association of Zoos and Aquariums. The aim of this study is to report the clinical, morphological and molecular characterization of this Babesia species. A 2.5-year-old, intact female maned wolf was found laterally recumbent with pale mucous membranes and jaundice the morning of presentation. Hematological and serum biochemical data were consistent with babesiosis and showed a regenerative severe anemia, leukocytosis, thrombocytopenia, hyperbilirubinemia, azotemia, increased creatine phosphokinase and increase alanine aminotransferase. On blood film review, inclusion bodies were seen in the red blood cells with cytomorphological features that were most consistent with a small form Babesia species. A blood sample was sent for polymerase chain reaction (PCR) testing and multi-locus sequence analyses. These findings suggested a unique Babesia species that is most closely related to a Babesia species (Babesia sp. AJB-2006) that has been found to infect raccoons (Procyon lotor) in North America. Although the cytomorphological features of the piroplasms and the clinical presentation were similar in both the current and 2012 case, when comparing the 18S melt curve temperature of the two Babesia isolates, the peak temperature was different. Unfortunately, genetic material from the 2012 case was not available so comparison of multi-locus gene sequences could not be performed, excluding the possibility to definitively state if the Babesia spp. from both cases were distinct from each other. The maned wolf was treated with a whole blood transfusion, dexamethazone (0.28 mg/kg IM), azithromycin (10 mg/kg in NaCl SC), atavaquone (1.5 cc PO), and 2 imidocarb (6.6 mg/kg IM) injections, and clinically improved. These findings demonstrate the need to further characterize the molecular and epidemiological differences of the Babesia species in this case report and the Babesia species known to infect raccoons.}, number={1}, journal={TICKS AND TICK-BORNE DISEASES}, author={Naor, Adi Wasserkrug and Lindemann, Dana M. and Schreeg, Megan E. and Marr, Henry S. and Birkenheuer, Adam J. and Carpenter, James W. and Ryseff, Julia K.}, year={2019}, month={Jan}, pages={124–126} }
 @article{barash_thomas_birkenheuer_breitschwerdt_lemler_qurollo_2019, title={Prevalence of
            Babesia
            spp. and clinical characteristics of
            Babesia vulpes
            infections in North American dogs}, volume={7}, ISSN={0891-6640 1939-1676}, url={http://dx.doi.org/10.1111/jvim.15560}, DOI={10.1111/jvim.15560}, abstractNote={Abstract}, journal={Journal of Veterinary Internal Medicine}, publisher={Wiley}, author={Barash, Nanelle R. and Thomas, Brittany and Birkenheuer, Adam J. and Breitschwerdt, Edward B. and Lemler, Erica and Qurollo, Barbara A.}, year={2019}, month={Jul} }
 @article{barash_birkenheuer_vaden_jacob_2018, title={Agreement between Parallel Canine Blood and Urine Cultures: Is Urine Culture the Poor Man's Blood Culture?}, volume={56}, ISSN={["1098-660X"]}, DOI={10.1128/JCM.00506-18}, abstractNote={Bloodstream infections are a significant cause of morbidity and mortality in critically ill dogs, but due to cost and difficulties in sample acquisition, blood cultures are infrequently obtained. In ill dogs, urine cultures may be recommended as surrogates for blood cultures.}, number={9}, journal={JOURNAL OF CLINICAL MICROBIOLOGY}, author={Barash, Nanelle R. and Birkenheuer, Adam J. and Vaden, Shelly L. and Jacob, Megan E.}, year={2018}, month={Sep} }
 @article{barash_birkenheuer_vaden_jacob_2018, title={Agreement between parallel canine blood and urine cultures: Is urine culture the poor man's blood culture?}, DOI={10.1128/JCM.00506-1810}, journal={Journal of Clinical Microbiology}, author={Barash, Nanelle R. and Birkenheuer, Adam J. and Vaden, Shelly L. and Jacob, Megan E.}, year={2018} }
 @article{ullal_birkenheuer_vaden_2018, title={Azotemia and Proteinuria in Dogs Infected with Babesia gibsoni}, volume={54}, ISSN={["1547-3317"]}, DOI={10.5326/jaaha-ms-6693}, abstractNote={ABSTRACT}, number={3}, journal={JOURNAL OF THE AMERICAN ANIMAL HOSPITAL ASSOCIATION}, author={Ullal, Tarini and Birkenheuer, Adam and Vaden, Shelly}, year={2018}, pages={156–160} }
 @article{birkenheuer_marr_wilson_breitschwerdt_qurollo_2018, title={Babesia gibsoni
 cytochrome b mutations in canine blood samples submitted to a US veterinary diagnostic laboratory}, volume={32}, ISSN={0891-6640}, url={http://dx.doi.org/10.1111/jvim.15300}, DOI={10.1111/jvim.15300}, abstractNote={
Background: Babesiosis caused by Babesia gibsoni is recognized throughout the world and can be difficult to treat. Resistance to atovaquone is associated with mutations in the B. gibsoni mitochondrial genome, specifically the M128 position of cytochrome b (cytb). The prevalence of cytb mutations in North America has not been reported.}, number={6}, journal={Journal of Veterinary Internal Medicine}, publisher={Wiley}, author={Birkenheuer, Adam J. and Marr, Henry S. and Wilson, James M. and Breitschwerdt, Edward B. and Qurollo, Barbara A.}, year={2018}, month={Oct}, pages={1965–1969} }
 @article{neupane_hegarty_marr_maggi_birkenheuer_breitschwerdt_2018, title={Evaluation of cell culture-grown Bartonella
 antigens in immunofluorescent antibody assays for the serological diagnosis of bartonellosis in dogs}, volume={32}, ISSN={0891-6640}, url={http://dx.doi.org/10.1111/jvim.15301}, DOI={10.1111/jvim.15301}, abstractNote={BackgroundBecause of poor sensitivity and questionable specificity of immunofluorescent antibody assays (IFAs), serological diagnosis of Bartonella species infections in dogs remains challenging. Despite limitations, IFA testing is the historical “gold standard” for Bartonella serodiagnosis in animals and humans. Because most diagnostic laboratories test against only 1 or 2 Bartonella spp., testing against a broader panel of Bartonella antigens may enhance diagnostic sensitivity and specificity.}, number={6}, journal={Journal of Veterinary Internal Medicine}, publisher={Wiley}, author={Neupane, Pradeep and Hegarty, Barbara C. and Marr, Henry S. and Maggi, Ricardo G. and Birkenheuer, Adam J. and Breitschwerdt, Edward B.}, year={2018}, month={Oct}, pages={1958–1964} }
 @article{khana_peterson_stanton_schreeg_birkenheuer_tarigo_2018, title={Genetic conservation of Cytauxzoon felis antigens and mRNA expression in the schizont life-stage}, volume={263}, ISSN={["1873-2550"]}, DOI={10.1016/j.vetpar.2018.10.007}, abstractNote={Cytauxzoonosis is a highly fatal disease of domestic cats caused by the apicomplexan protozoan Cytauxzoon felis, which is most closely related to Theileria spp. The growing prevalence, high morbidity and mortality, and treatment cost of cytauxzoonosis emphasize the need for vaccine development. Traditional approaches for vaccine development, however, have been hindered by the inability to culture C. felis in vitro. Recent availability of the annotated C. felis genome combined with genome-based vaccine design and protein microarray immunoscreening allowed for high-throughput identification of C. felis antigens that could serve as vaccine candidates. This study assessed the suitability of three of these vaccine candidates (cf30, cf63, cf58) in addition to a previously reported vaccine candidate (cf76) based on two criteria: genetic conservation among diverse C. felis geographic isolates and expression in tissues containing the C. felis schizont life stage, which has been previously associated with the development of a protective immune response. A comparison of seventeen C. felis isolates across seven states demonstrated high sequence identity (99-100%) for cf30, cf63, and cf58, similar to the degree of conservation previously reported for cf76. RNAscope® in situ hybridization using acutely infected feline splenic tissue revealed robust levels of all transcripts in the schizont life stage of the parasite. These data support the suitability of these three antigens for further investigation as vaccine candidates against cytauxzoonosis.}, journal={VETERINARY PARASITOLOGY}, author={Khana, Daven B. and Peterson, David S. and Stanton, James B. and Schreeg, Megan E. and Birkenheuer, Adam J. and Tarigo, Jaime L.}, year={2018}, month={Nov}, pages={49–53} }
 @article{schreeg_marr_tarigo_sherrill_outi_scholl_bird_vigil_hung_nakajima_et al._2018, title={Identification of Cytauxzoon felis antigens via protein microarray and assessment of expression library immunization against cytauxzoonosis}, volume={15}, ISSN={["1559-0275"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-85059281263&partnerID=MN8TOARS}, DOI={10.1186/s12014-018-9218-9}, abstractNote={Cytauxzoonosis is a disease of felids in North America caused by the tick-transmitted apicomplexan parasite Cytauxzoon felis. Cytauxzoonosis is particularly virulent for domestic cats, but no vaccine currently exists. The parasite cannot be cultivated in vitro, presenting a significant limitation for vaccine development.Recent sequencing of the C. felis genome has identified over 4300 putative protein-encoding genes. From this pool we constructed a protein microarray containing 673 putative C. felis proteins. This microarray was probed with sera from C. felis-infected and naïve cats to identify differentially reactive antigens which were incorporated into two expression library vaccines, one polyvalent and one monovalent. We assessed the efficacy of these vaccines to prevent of infection and/or disease in a tick-challenge model.Probing of the protein microarray resulted in identification of 30 differentially reactive C. felis antigens that were incorporated into the two expression library vaccines. However, expression library immunization failed to prevent infection or disease in cats challenged with C. felis.Protein microarray facilitated high-throughput identification of novel antigens, substantially increasing the pool of characterized C. felis antigens. These antigens should be considered for development of C. felis vaccines, diagnostics, and therapeutics.}, number={1}, journal={CLINICAL PROTEOMICS}, author={Schreeg, Megan E. and Marr, Henry S. and Tarigo, Jaime L. and Sherrill, Meredith K. and Outi, Hilton K. and Scholl, Elizabeth H. and Bird, David M. and Vigil, Adam and Hung, Chris and Nakajima, Rie and et al.}, year={2018}, month={Dec} }
 @article{kovačević filipović_beletić_ilić božović_milanović_tyrrell_buch_breitschwerdt_birkenheuer_chandrashekar_2018, title={Molecular and Serological Prevalence of Anaplasma phagocytophilum, A. platys, Ehrlichia canis, E. chaffeenses, E. ewingii, Borrelia burgdorferi, Babesia canis, B. gibsoni and B. vogeli among Clinically Healthy Outdoor Dogs in Serbia}, volume={14}, ISSN={2405-9390}, url={http://dx.doi.org/10.1016/J.VPRSR.2018.10.001}, DOI={10.1016/J.VPRSR.2018.10.001}, abstractNote={Data concerning combined molecular and serological prevalence of emerging canine tick-borne pathogens in Serbia are lacking. A large population of outdoor living dogs in Belgrade, Serbia's' capital, present an excellent population for epidemiology study. Blood samples were collected from 111 dogs, including 46 shelter, 31 free roaming, and 34 hunting dogs. Species-specific real-time polymerase chain reaction (PCR) (IDEXX Laboratories, Inc., Westbrook Maine, USA) was applied for the molecular detection of Anaplasma phagocytophilum, A. platys, Ehrlichia canis, Babesia canis, B. gibsoni and B. vogeli. A research based SNAP assay (SNAP® M-A, IDEXX Laboratories, Inc., Westbrook Maine, USA) that uses genus and species-specific peptides was used to asses Anaplasma spp., A. phagocytophilum, A. platys, Ehrlichia spp., E. canis, E. chaffeensis, E. ewingii and Borrelia burgdorferi antibody status. B. canis, B. gibsoni and B. vogeli antibody status was assessed with an indirect immunofluorescence test (MegaCor Diagnostic, Horbranz, Austria). Anaplasma spp. and Ehrlichia spp. DNA was not amplified. One quarter of the dogs were A. phagocytophilum, one dog was A. platys, one was E. ewingii and two dogs were B. burgdorferi seroreactive with the SNAP® M-A. Babesia canis or B. gibsoni DNA was amplified by PCR from 16.2% of dogs, whereas 67.6% were seroreactive to one or more Babesia spp. Babesia vogeli was not PCR amplified. We conclude that outdoor dogs in this territory are reservoirs for B. canis and B. gibsoni and are frequently co-exposed to combinations of Anaplasma and Babesia spp.}, journal={Veterinary Parasitology: Regional Studies and Reports}, publisher={Elsevier BV}, author={Kovačević Filipović, Milica M. and Beletić, Anđelo D. and Ilić Božović, Anja V. and Milanović, Zorana and Tyrrell, Phyllis and Buch, Jesse and Breitschwerdt, Edward B. and Birkenheuer, Adam J. and Chandrashekar, Ramaswamy}, year={2018}, month={Dec}, pages={117–122} }
 @article{mylonakis_schreeg_chatzis_pearce_marr_saridomichelakis_birkenheuer_2018, title={Molecular detection of vector-borne pathogens in Greek cats}, volume={9}, ISSN={["1877-9603"]}, DOI={10.1016/j.ttbdis.2017.08.013}, abstractNote={Infectious diseases have been increasingly recognized in cats worldwide. The objective of this study was the molecular investigation of the prevalence of selected pathogens in healthy and sick cats from Greece, a country highly endemic for several canine vector-borne pathogens. Blood and/or bone marrow samples from 50 clinically healthy and 50 sick adult (>1 year-old) cats were retrospectively examined for the amplification of Bartonella spp., haemoplasmas, Ehrlichia spp., Anaplasma spp., Babesia spp., and Cytauxzoon spp. DNA. Overall, 14.9% of the cats were found to be infected or co-infected by haemoplasmas, including Candidatus Mycoplasma haemominutum and M. haemofelis. In addition, 8.5% of the cats were infected by Bartonella henselae, Bartonella clarridgeiae or Bartonella koehlerae. In contrast, DNA of Ehrlichia spp., Anaplasma spp., Babesia spp. and Cytauxzoon spp. was not amplified from the blood or bone marrow of any cat. There was no significant difference in either haemoplasma or Bartonella infection rates when comparing healthy and sick cats. This study represents the first description of Bartonella koehlerae in Greek cats.}, number={2}, journal={TICKS AND TICK-BORNE DISEASES}, author={Mylonakis, Mathios E. and Schreeg, Megan and Chatzis, Manolis K. and Pearce, Julian and Marr, Henry S. and Saridomichelakis, Manolis N. and Birkenheuer, Adam J.}, year={2018}, month={Feb}, pages={171–175} }
 @article{lashnits_correa_hegarty_birkenheuer_breitschwerdt_2017, title={Bartonella
 Seroepidemiology in Dogs from North America, 2008-2014}, volume={32}, ISSN={0891-6640}, url={http://dx.doi.org/10.1111/jvim.14890}, DOI={10.1111/jvim.14890}, abstractNote={BackgroundImproved understanding of Bartonella species seroepidemiology in dogs may aid clinical decision making and enhance current understanding of naturally occurring arthropod vector transmission of this pathogen.}, number={1}, journal={Journal of Veterinary Internal Medicine}, publisher={Wiley}, author={Lashnits, E. and Correa, M. and Hegarty, B.C. and Birkenheuer, A. and Breitschwerdt, E.B.}, year={2017}, month={Dec}, pages={222–231} }
 @article{levine_cianciolo_linder_bizikova_birkenheuer_brooks_salous_nordone_bellinger_marr_et al._2017, title={Endothelial alterations in a canine model of immune thrombocytopenia}, volume={30}, ISSN={0953-7104 1369-1635}, url={http://dx.doi.org/10.1080/09537104.2017.1378807}, DOI={10.1080/09537104.2017.1378807}, abstractNote={Abstract Bleeding heterogeneity amongst patients with immune thrombocytopenia (ITP) is poorly understood. Platelets play a role in maintaining endothelial integrity, and variable thrombocytopenia-induced endothelial changes may influence bleeding severity. Platelet-derived endothelial stabilizers and markers of endothelial integrity in ITP are largely underexplored. We hypothesized that, in a canine ITP model, thrombocytopenia would lead to alterations in the endothelial ultrastructure and that the Von Willebrand factor (vWF) would serve as a marker of endothelial injury associated with thrombocytopenia. Thrombocytopenia was induced in healthy dogs with an antiplatelet antibody infusion; control dogs received an isotype control antibody. Cutaneous biopsies were obtained prior to thrombocytopenia induction, at platelet nadir, 24 hours after nadir, and on platelet recovery. Cutaneous capillaries were assessed by electron microscopy for vessel thickness, the number of pinocytotic vesicles, the number of large vacuoles, and the number of gaps between cells. Pinocytotic vesicles are thought to represent an endothelial membrane reserve that can be used for repair of damaged endothelial cells. Plasma samples were assessed for vWF. ITP dogs had significantly decreased pinocytotic vesicle numbers compared to control dogs (P = 0.0357) and the increase in plasma vWF from baseline to 24 hours correlated directly with the endothelial large vacuole score (R = 0.99103; P < 0.0001). This direct correlation between plasma vWF and the number of large vacuoles, representing the vesiculo-vacuolar organelle (VVO), a permeability structure, suggests that circulating vWF could serve as a biomarker for endothelial alterations and potentially a predictor of thrombocytopenic bleeding. Overall, our results indicate that endothelial damage occurs in the canine ITP model and variability in the degree of endothelial damage may account for differences in the bleeding phenotype among patients with ITP.}, number={1}, journal={Platelets}, publisher={Informa UK Limited}, author={LeVine, Dana N. and Cianciolo, Rachel E. and Linder, Keith E. and Bizikova, Petra and Birkenheuer, Adam J. and Brooks, Marjory B. and Salous, Abdelghaffar K. and Nordone, Shila K. and Bellinger, Dwight A. and Marr, Henry and et al.}, year={2017}, month={Nov}, pages={88–97} }
 @article{qurollo_archer_schreeg_marr_birkenheuer_haney_thomas_breitschwerdt_2017, title={Improved molecular detection of Babesia infections in animals using a novel quantitative real-time PCR diagnostic assay targeting mitochondrial DNA}, volume={10}, ISSN={1756-3305}, url={http://dx.doi.org/10.1186/s13071-017-2064-1}, DOI={10.1186/s13071-017-2064-1}, abstractNote={Babesiosis is a protozoal, tick transmitted disease found worldwide in humans, wildlife and domesticated animals. Commonly used approaches to diagnose babesiosis include microscopic examination of peripheral blood smears, detection of circulating antibodies and PCR. To screen and differentiate canine Babesia infections many PCR assays amplify the 18S rRNA gene. These sequences contain hypervariable regions flanked by highly conserved regions allowing for amplification of a broad-range of Babesia spp. However, differences in the 18S rRNA gene sequence of distantly related clades can make it difficult to design assays that will amplify all Babesia species while excluding the amplification of other eukaryotes. By targeting Babesia mitochondrial genome (mtDNA), we designed a novel three primer qPCR with greater sensitivity and broader screening capabilities to diagnose and differentiate Babesia spp. Using 13 Babesia mtDNA sequences, a region spanning two large subunit rRNA gene fragments (lsu5-lsu4) was aligned to design three primers for use in a qPCR assay (LSU qPCR) capable of amplifying a wide range of Babesia spp. Plasmid clones were generated and used as standards to determine efficiency, linear dynamic range and analytical sensitivity. Animals naturally infected with vector-borne pathogens were tested retrospectively and prospectively to determine relative clinical sensitivity and specificity by comparing the LSU qPCR to an established 18S rDNA qPCR. The LSU qPCR efficiencies ranged between 92 and 100% with the limit of detection at five copies/reaction. The assay did not amplify mammalian host or other vector-borne pathogen gDNA except Cytauxzoon felis (a feline protozoal pathogen). The LSU qPCR assay amplified 12 different Babesia. sp. and C. felis from 31/31 (100%) archived samples, whereas the 18S qPCR amplified only 26/31 (83.9%). By prospective analysis, 19/394 diagnostic accessions (4.8%) were LSU qPCR positive, compared to 11/394 (2.8%) 18S rDNA qPCR positive. We have developed a more sensitive qPCR assay with a more expansive range of Babesia spp. detection by targeting a highly conserved region of mtDNA, when compared to an established 18S qPCR.}, number={1}, journal={Parasites & Vectors}, publisher={Springer Nature}, author={Qurollo, Barbara A. and Archer, Nikole R. and Schreeg, Megan E. and Marr, Henry S. and Birkenheuer, Adam J. and Haney, Kaitlin N. and Thomas, Brittany S. and Breitschwerdt, Edward B.}, year={2017}, month={Mar} }
 @article{manning_birkenheuer_briley_montgomery_harris_vanone_gookin_2016, title={Intermittent At-Home Suctioning of Esophageal Content for Prevention of Recurrent Aspiration Pneumonia in 4 Dogs with Megaesophagus}, volume={30}, ISSN={["1939-1676"]}, DOI={10.1111/jvim.14527}, abstractNote={BackgroundMegaesophagus carries a poor to guarded prognosis due to death from aspiration pneumonia. Options for medical management of regurgitation are limited to strategic oral or gastrostomy tube feeding.}, number={5}, journal={JOURNAL OF VETERINARY INTERNAL MEDICINE}, author={Manning, K. and Birkenheuer, A. J. and Briley, J. and Montgomery, S. A. and Harris, J. and Vanone, S. L. and Gookin, J. L.}, year={2016}, pages={1715–1719} }
 @article{schreeg_marr_tarigo_cohn_bird_scholl_levy_wiegmann_birkenheuer_2016, title={Mitochondrial Genome Sequences and Structures Aid in the Resolution of Piroplasmida phylogeny}, volume={11}, ISSN={["1932-6203"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-84994744879&partnerID=MN8TOARS}, DOI={10.1371/journal.pone.0165702}, abstractNote={The taxonomy of the order Piroplasmida, which includes a number of clinically and economically relevant organisms, is a hotly debated topic amongst parasitologists. Three genera (Babesia, Theileria, and Cytauxzoon) are recognized based on parasite life cycle characteristics, but molecular phylogenetic analyses of 18S sequences have suggested the presence of five or more distinct Piroplasmida lineages. Despite these important advancements, a few studies have been unable to define the taxonomic relationships of some organisms (e.g. C. felis and T. equi) with respect to other Piroplasmida. Additional evidence from mitochondrial genome sequences and synteny should aid in the inference of Piroplasmida phylogeny and resolution of taxonomic uncertainties. In this study, we have amplified, sequenced, and annotated seven previously uncharacterized mitochondrial genomes (Babesia canis, Babesia vogeli, Babesia rossi, Babesia sp. Coco, Babesia conradae, Babesia microti-like sp., and Cytauxzoon felis) and identified additional ribosomal fragments in ten previously characterized mitochondrial genomes. Phylogenetic analysis of concatenated mitochondrial and 18S sequences as well as cox1 amino acid sequence identified five distinct Piroplasmida groups, each of which possesses a unique mitochondrial genome structure. Specifically, our results confirm the existence of four previously identified clades (B. microti group, Babesia sensu stricto, Theileria equi, and a Babesia sensu latu group that includes B. conradae) while supporting the integration of Theileria and Cytauxzoon species into a single fifth taxon. Although known biological characteristics of Piroplasmida corroborate the proposed phylogeny, more investigation into parasite life cycles is warranted to further understand the evolution of the Piroplasmida. Our results provide an evolutionary framework for comparative biology of these important animal and human pathogens and help focus renewed efforts toward understanding the phylogenetic relationships within the group.}, number={11}, journal={PLOS ONE}, author={Schreeg, Megan E. and Marr, Henry S. and Tarigo, Jaime L. and Cohn, Leah A. and Bird, David M. and Scholl, Elizabeth H. and Levy, Michael G. and Wiegmann, Brian M. and Birkenheuer, Adam J.}, year={2016}, month={Nov} }
 @article{schreeg_marr_griffith_tarigo_bird_reichard_cohn_levy_birkenheuer_2016, title={PCR amplification of a multi-copy mitochondrial gene (cox3) improves detection of Cytauxzoon felis infection as compared to a ribosomal gene (18S)}, volume={225}, ISSN={["1873-2550"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-84975504702&partnerID=MN8TOARS}, DOI={10.1016/j.vetpar.2016.06.013}, abstractNote={Cytauxzoon felis is a tick-transmitted protozoan parasite that infects felids. Clinical disease caused by acute C. felis infection rapidly progresses in domestic cats, leading to high morbidity and mortality. Accurately diagnosing cytauxzoonosis as soon as possible during acute infection would allow for earlier initiation of antiprotozoal therapy which could lead to higher survival rates. Molecular detection of parasite rRNA genes (18S) by PCR has previously been shown to be a sensitive method of diagnosing C. felis infections. Based on evidence from related apicomplexan species, we hypothesized that C. felis mitochondrial genes would exist at higher copy numbers than 18S and would be a more sensitive diagnostic target. In this study we have designed a PCR assay targeting the C. felis mitochondrial gene cytochrome c oxidase subunit III (cox3). Herein we demonstrate that (1) the cox3 PCR can detect as low as 1 copy of DNA target and can detect C. felis in samples with known mitochondrial sequence heterogeneity, (2) cox3 copy number is increased relative to 18S in blood and tissue samples from acutely infected cats, and (3) the cox3 PCR is more sensitive than 18S PCR for detection of C. felis during early infections.}, journal={VETERINARY PARASITOLOGY}, author={Schreeg, Megan E. and Marr, Henry S. and Griffith, Emily H. and Tarigo, Jaime L. and Bird, David M. and Reichard, Mason V. and Cohn, Leah A. and Levy, Michael G. and Birkenheuer, Adam J.}, year={2016}, month={Jul}, pages={123–130} }
 @article{wardrop_birkenheuer_blais_callan_kohn_lappin_sykes_2016, title={Update on Canine and Feline Blood Donor Screening for Blood-Borne Pathogens}, volume={30}, ISSN={["1939-1676"]}, DOI={10.1111/jvim.13823}, abstractNote={An update on the 2005 American College of Veterinary Internal Medicine (ACVIM) Consensus Statement on blood donor infectious disease screening was presented at the 2015 ACVIM Forum in Indianapolis, Indiana, followed by panel and audience discussion. The updated consensus statement is presented below. The consensus statement aims to provide guidance on appropriate blood‐borne pathogen testing for canine and feline blood donors in North America.}, number={1}, journal={JOURNAL OF VETERINARY INTERNAL MEDICINE}, author={Wardrop, K. J. and Birkenheuer, A. and Blais, M. C. and Callan, M. B. and Kohn, B. and Lappin, M. R. and Sykes, J.}, year={2016}, pages={15–35} }
 @article{conner_hanel_brooks_cohn_birkenheuer_2015, title={Coagulation abnormalities in 5 cats with naturally occurring cytauxzoonosis}, volume={25}, ISSN={["1476-4431"]}, DOI={10.1111/vec.12326}, abstractNote={Abstract}, number={4}, journal={JOURNAL OF VETERINARY EMERGENCY AND CRITICAL CARE}, author={Conner, Bobbi J. and Hanel, Rita M. and Brooks, Marjory B. and Cohn, Leah A. and Birkenheuer, Adam J.}, year={2015}, pages={538–545} }
 @article{vandersea_birkenheuer_litaker_vaden_renschler_gookin_2015, title={Identification of Parabodo caudatus (class Kinetoplastea) in urine voided from a dog with hematuria}, volume={27}, ISSN={["1943-4936"]}, DOI={10.1177/1040638714562827}, abstractNote={A voided urine sample, obtained from a 13-year-old intact male dog residing in a laboratory animal research facility, was observed to contain biflagellate protozoa 5 days following an episode of gross hematuria. The protozoa were identified as belonging to the class Kinetoplastea on the basis of light microscopic observation of Wright–Giemsa-stained urine sediment in which the kinetoplast was observed basal to 2 anterior flagella. A polymerase chain reaction (PCR) assay using primers corresponding with conserved regions within the 18S ribosomal RNA gene of representative kinetoplastid species identified nucleotide sequences with 100% identity to Parabodo caudatus. Parabodo caudatus organisms were unable to be demonstrated cytologically or by means of PCR in samples collected from the dog’s environment. The dog had a history of 50 complete urinalyses performed over the 12-year period preceding detection of P. caudatus, and none of these were noted to contain protozoa. Moreover, the gross hematuria that was documented 5 days prior to detection of P. caudatus had never before been observed in this dog. Over the ensuing 2.5 years of the dog’s life, 16 additional complete urinalyses were performed, none of which revealed the presence of protozoa. Bodonids are commonly found in soil as well as in freshwater and marine environments. However, P. caudatus, in particular, has a 150-year-long, interesting, and largely unresolved history in people as either an inhabitant or contaminant of urine. This historical conundrum is revisited in the current description of P. caudatus as recovered from the urine of a dog.}, number={1}, journal={JOURNAL OF VETERINARY DIAGNOSTIC INVESTIGATION}, author={Vandersea, Mark W. and Birkenheuer, Adam J. and Litaker, R. Wayne and Vaden, Shelly L. and Renschler, Janelle S. and Gookin, Jody L.}, year={2015}, month={Jan}, pages={117–120} }
 @article{rizzi_reichard_cohn_birkenheuer_taylor_meinkoth_2015, title={Prevalence of Cytauxzoon felis infection in healthy cats from enzootic areas in Arkansas, Missouri, and Oklahoma}, volume={8}, ISSN={["1756-3305"]}, DOI={10.1186/s13071-014-0618-z}, abstractNote={Infection with Cytauxzoon felis in domestic cats can cause fever, lethargy, depression, inappetence, icterus, and often death. With a high mortality rate, cytauxzoonosis was historically considered a fatal disease. Within the last 15 years, cats with or without treatment have been recognized as chronically infected survivors of C. felis infection. Our objective was to determine the prevalence of C. felis in healthy domestic cats from Arkansas, Missouri, and Oklahoma.Infection with C. felis was determined using DNA extracted from anticoagulated whole blood and PCR amplification using C. felis-specific primers. Chi-square, Fisher's exact tests, and odds ratios were used to compare proportions of cats infected with C. felis.Blood samples were collected from 902 healthy domestic cats between October 2008 and April 2012. DNA from Cytauxzoon felis was detected in 56 of 902 (6.2%; 95% confidence interval, 4.7-7.9) samples. The highest prevalence of C. felis infection (15.5%; 10.3-21.7) was observed in cats from Arkansas, followed by cats from Missouri (12.9%; 6.1-24.0), and cats from Oklahoma (3.4%; 2.2-5.1). Cats sampled in Arkansas and Missouri were 5.1 and 4.2, respectively, times more likely to be chronically infected with C. felis than cats from Oklahoma.Infection with C. felis is common in domestic cats through Arkansas, Missouri, and Oklahoma. The high prevalence of C. felis reported herein suggests that infected domestic cats are likely reservoirs of infection for naive felines. The high prevalence of C. felis substantiates the importance for the use of approved acaricides on cats to prevent cytauxzoonosis.}, journal={PARASITES & VECTORS}, author={Rizzi, Theresa E. and Reichard, Mason V. and Cohn, Leah A. and Birkenheuer, Adam J. and Taylor, Jared D. and Meinkoth, James H.}, year={2015}, month={Jan} }
 @article{schreeg_marr_tarigo_cohn_levy_birkenheuer_2015, title={Rapid High-Resolution Melt Analysis of Cytauxzoon felis Cytochrome b To Aid in the Prognosis of Cytauxzoonosis}, volume={53}, ISSN={["1098-660X"]}, DOI={10.1128/jcm.00635-15}, abstractNote={ABSTRACT}, number={8}, journal={JOURNAL OF CLINICAL MICROBIOLOGY}, author={Schreeg, Megan E. and Marr, Henry S. and Tarigo, Jaime L. and Cohn, Leah A. and Levy, Michael G. and Birkenheuer, Adam J.}, year={2015}, month={Aug}, pages={2517–2524} }
 @article{levine_birkenheuer_brooks_nordone_bellinger_jones_fischer_oglesbee_frey_brinson_et al._2014, title={A novel canine model of immune thrombocytopenia: has immune thrombocytopenia (ITP) gone to the dogs?}, volume={167}, ISSN={0007-1048}, url={http://dx.doi.org/10.1111/bjh.13005}, DOI={10.1111/bjh.13005}, abstractNote={Summary}, number={1}, journal={British Journal of Haematology}, publisher={Wiley}, author={LeVine, Dana N. and Birkenheuer, Adam J. and Brooks, Marjory B. and Nordone, Shila K. and Bellinger, Dwight A. and Jones, Sam L. and Fischer, Thomas H. and Oglesbee, Stephen E. and Frey, Kahlina and Brinson, Nicole S. and et al.}, year={2014}, month={Jul}, pages={110–120} }
 @article{maggi_birkenheuer_hegarty_bradley_levy_breitschwerdt_2014, title={Comparison of serological and molecular panels for diagnosis of vector-borne diseases in dogs}, volume={7}, ISSN={1756-3305}, url={http://dx.doi.org/10.1186/1756-3305-7-127}, DOI={10.1186/1756-3305-7-127}, abstractNote={Canine vector-borne diseases (CVBD) are caused by a diverse array of pathogens with varying biological behaviors that result in a wide spectrum of clinical presentations and laboratory abnormalities. For many reasons, the diagnosis of canine vector-borne infectious diseases can be challenging for clinicians. The aim of the present study was to compare CVBD serological and molecular testing as the two most common methodologies used for screening healthy dogs or diagnosing sick dogs in which a vector-borne disease is suspected.We used serological (Anaplasma species, Babesia canis, Bartonella henselae, Bartonella vinsonii subspecies berkhoffii, Borrelia burgdorferi, Ehrlichia canis, and SFG Rickettsia) and molecular assays to assess for exposure to, or infection with, 10 genera of organisms that cause CVBDs (Anaplasma, Babesia, Bartonella, Borrelia, Ehrlichia, Francisella, hemotropic Mycoplasma, Neorickettsia, Rickettsia, and Dirofilaria). Paired serum and EDTA blood samples from 30 clinically healthy dogs (Group I) and from 69 sick dogs suspected of having one or more canine vector-borne diseases (Groups II-IV), were tested in parallel to establish exposure to or infection with the specific CVBDs targeted in this study.Among all dogs tested (Groups I-IV), the molecular prevalences for individual CVBD pathogens ranged between 23.3 and 39.1%. Similarly, pathogen-specific seroprevalences ranged from 43.3% to 59.4% among healthy and sick dogs (Groups I-IV). Among these representative sample groupings, a panel combining serological and molecular assays run in parallel resulted in a 4-58% increase in the recognition of exposure to or infection with CVBD.We conclude that serological and PCR assays should be used in parallel to maximize CVBD diagnosis.}, number={1}, journal={Parasites & Vectors}, publisher={Springer Nature}, author={Maggi, Ricardo G and Birkenheuer, Adam J and Hegarty, Barbara C and Bradley, Julie M and Levy, Michael G and Breitschwerdt, Edward B}, year={2014}, pages={127} }
 @article{pritchard_birkenheuer_hanel_wood_2014, title={Copperhead (Agkistrodon contortrix) envenomation of dogs: 52 cases (2004-2011)}, DOI={10.5326/JAAHA-MS-6131}, abstractNote={Copperhead envenomation is common within the US, and no studies exist describing the clinical course of copperhead envenomation in dogs. Almost all treatment decisions regarding those bites are extrapolated from retrospective studies evaluating the clinical course of rattlesnake bites. Because copperheads and rattlesnakes produce venom with different potency, assumptions that treatment of the different envenomations should be similar may be incorrect. The purpose of this retrospective study was to evaluate the clinical course of copperhead envenomation in dogs and administered treatments. Medical records of 52 dogs treated for copperhead envenomation were reviewed, and owners were contacted regarding outcome. The most common clinical signs associated with copperhead envenomation included swelling, pain, and ecchymosis. Clinicopathological abnormalities (e.g., thrombocytopenia, elevated clotting times, leukocytosis) were mild, and red blood cell morphology changes and coagulopathies were rare. Most dogs were treated with antimicrobials, analgesics, and fluid therapy. No dogs in this study required the use of antivenin and all survived to discharge. This study found that the clinical course after copperhead envenomation is generally limited to local rather than systemic illness. Copperhead envenomation in dogs is largely self-limiting and responsive to supportive care with hospitalization for monitoring.}, journal={Journal of the American Animal Hospital Association}, author={Pritchard, Jessica C. and Birkenheuer, Adam J. and Hanel, Rita M. and Wood, Michael W.}, year={2014} }
 @article{lewis_cohn_marr_birkenheuer_2014, title={Failure of efficacy and adverse events associated with dose-intense diminazene diaceturate treatment of chronic Cytauxzoon felis infection in five cats}, volume={16}, ISSN={["1532-2750"]}, DOI={10.1177/1098612x13502974}, abstractNote={ Cytauxzoon felis is a hemoprotozoan parasite of cats. While many infected cats die of acute illness, some enter a chronic carrier state. To date, no treatment has been documented to clear the chronic carrier state, leaving recovered cats to act as a potential indirect source of infection via a tick vector. Diminazene diaceturate is an anti-protozoal therapy that has been suggested for use in the treatment of acute cytauxzoonosis, but which failed to clear the carrier state at the dose used in acute illness. We hypothesized that a dose-intensified regimen of diminazene could reduce or eliminate parasitemia from five domestic cats naturally infected with C felis. Cats were administered 4 mg/kg of diminazene diaceturate intramuscularly for 5 consecutive days. Clearance of the organism was assessed via semi-quantitative polymerase chain reaction and light microscopy 1, 3, 6 and 10 weeks after starting treatment. Additionally, cats were monitored for adverse drug reactions by daily observation and examination. Complete blood count, biochemical profile and urinalysis were performed at 1, 3 and 10 weeks. Adverse events were common and included profuse salivation and nausea at the time of injection, monoparesis in the injected leg, proteinuria and potential hepatotoxicity. Severity of parasitemia was not reduced. Diminazene diaceturate cannot be recommended for elimination of the carrier state of C felis infection. }, number={2}, journal={JOURNAL OF FELINE MEDICINE AND SURGERY}, author={Lewis, Kristin M. and Cohn, Leah A. and Marr, Henry S. and Birkenheuer, Adam J.}, year={2014}, month={Feb}, pages={157–163} }
 @article{floras_holowaychuk_hodgins_marr_birkenheuer_sharif_bersenas_bienzle_2014, title={Investigation of a Commercial ELISA for the Detection of Canine Procalcitonin}, volume={28}, ISSN={["1939-1676"]}, DOI={10.1111/jvim.12309}, abstractNote={BackgroundRapid identification of sepsis enables prompt administration of antibiotics and is essential to improve patient survival. Procalcitonin (PCT) is a biomarker used to diagnose sepsis in people. Commercial assays to measure canine PCT peptide have not been validated.}, number={2}, journal={JOURNAL OF VETERINARY INTERNAL MEDICINE}, author={Floras, A. N. K. and Holowaychuk, M. K. and Hodgins, D. C. and Marr, H. S. and Birkenheuer, A. and Sharif, S. and Bersenas, A. M. E. and Bienzle, D.}, year={2014}, month={Mar}, pages={599–602} }
 @article{balakrishnan_pritchard_ericson_grindem_phillips_jennings_mathews_tran_birkenheuer_breitschwerdt_et al._2014, title={Prostatitis, Steatitis, and Diarrhea in a Dog following Presumptive Flea-Borne Transmission of Bartonella henselae}, volume={52}, ISSN={0095-1137 1098-660X}, url={http://dx.doi.org/10.1128/JCM.00942-14}, DOI={10.1128/jcm.00942-14}, abstractNote={ABSTRACT}, number={9}, journal={Journal of Clinical Microbiology}, publisher={American Society for Microbiology}, author={Balakrishnan, N. and Pritchard, J. and Ericson, M. and Grindem, C. and Phillips, K. and Jennings, S. and Mathews, K. and Tran, H. and Birkenheuer, A. J. and Breitschwerdt, Edward and et al.}, editor={Munson, E.Editor}, year={2014}, month={Jun}, pages={3447–3452} }
 @article{yancey_hegarty_qurollo_levy_birkenheuer_weber_diniz_breitschwerdt_2014, title={Regional Seroreactivity and Vector-Borne Disease Co-Exposures in Dogs in the United States from 2004–2010: Utility of Canine Surveillance}, volume={14}, ISSN={1530-3667 1557-7759}, url={http://dx.doi.org/10.1089/vbz.2014.1592}, DOI={10.1089/vbz.2014.1592}, abstractNote={Vector-borne disease (VBD) pathogens remain an emerging health concern for animals and humans throughout the world. Surveillance studies of ticks and humans have made substantial contributions to our knowledge of VBD epidemiology trends, but long-term VBD surveillance data of dogs in the United States is limited. This seroreactivity study assessed US temporal and regional trends and co-exposures to Anaplasma, Babesia, Bartonella, Borrelia burgdorferi, Dirofilaria immitis, Ehrlichia spp., and spotted fever group Rickettsia in dogs from 2004-2010. Dog serum samples (N=14,496) were submitted to the North Carolina State University, College of Veterinary Medicine, Vector Borne Disease Diagnostic Laboratory for vector-borne pathogens diagnostic testing using immunofluorescent antibody (IFA) and enzyme-linked immunosorbent assay (ELISA) assays. These convenience samples were retrospectively reviewed and analyzed. The largest proportion of samples originated from the South (47.6%), with the highest percent of seroreactive samples observed in the Midatlantic (43.4%), compared to other US regions. The overall seroreactivity of evaluated VBD antigens were Rickettsia rickettsia (10.4%), B. burgdorferi (5.2%), Ehrlichia spp. (4.3%), Bartonella henselae (3.8%), Anaplasma spp. (1.9%), Bartonella vinsonii subsp. berkhoffii (1.5%), Babesia canis (1.1%), and D. immitis (0.8%). Significant regional and annual seroreactivity variation was observed with B. burgdorferi, Ehrlichia, and Rickettsia exposures. Seasonal seroreactivity variation was evident with Rickettsia. Seroreactivity to more than one antigen was present in 16.5% of exposed dogs. Nationally, the most prevalent co-exposure was Rickettsia with Ehrlichia spp. (5.3%), and the highest odds of co-exposure was associated with Anaplasma spp. and B. burgdorferi (odds ratio=6.6; 95% confidence interval 5.0, 8.8). Notable annual and regional seroreactivity variation was observed with certain pathogens over 7 years of study, suggesting canine surveillance studies may have value in contributing to future VBD knowledge.}, number={10}, journal={Vector-Borne and Zoonotic Diseases}, publisher={Mary Ann Liebert Inc}, author={Yancey, Caroline B. and Hegarty, Barbara C. and Qurollo, Barbara A. and Levy, Michael G. and Birkenheuer, Adam J. and Weber, David J. and Diniz, Pedro P.V.P. and Breitschwerdt, Edward B.}, year={2014}, month={Oct}, pages={724–732} }
 @article{li_d’annibale-tolhurst_adler_fang_yin_birkenheuer_levy_jones_sung_hawkins_et al._2013, title={A Myristoylated Alanine-Rich C Kinase Substrate–Related Peptide Suppresses Cytokine mRNA and Protein Expression in LPS-Activated Canine Neutrophils}, volume={48}, ISSN={1044-1549 1535-4989}, url={http://dx.doi.org/10.1165/rcmb.2012-0278OC}, DOI={10.1165/rcmb.2012-0278oc}, abstractNote={Myristoylated alanine-rich C kinase substrate (MARCKS) is a ubiquitously expressed protein kinase C substrate that has emerged as a potential therapeutic target for the amelioration of mucin secretion and inflammation in patients with chronic obstructive pulmonary disease. MARCKS also plays a key role in regulating the adhesion, migration, and degranulation of neutrophils. Moreover, given its biological role in epithelial and immune cells, we hypothesized that MARCKS may play an integral role in cytokine secretion by neutrophils. Because the amino terminus of MARCKS is highly conserved across vertebrate species, we successfully applied the well-characterized human MARCKS inhibitory peptide, myristoylated N-terminal sequence (MANS), to attenuate the function of MARCKS in isolated canine neutrophils. Pretreatment of canine neutrophils with MANS peptide significantly reduced both mRNA and protein expression in a broad range of LPS-induced cytokines, including IL-8, a chemokine (C-X-C motif) ligand-1 orthologue, and TNF-α, in comparison with untreated cells or those treated with a control peptide. This reduction in cytokine expression was observed even when neutrophils were treated with MANS 2 hours after LPS exposure. The observed reduction in cytokine secretion was not attributable to protein retention or cell death, but was associated with reduced cytokine transcript synthesis. These observations identify MARCKS protein as a promising therapeutic target in the treatment of inflammatory diseases or syndromes attributed to neutrophil influx and inflammatory cytokine production, such as sepsis, acute lung injury, and acute respiratory distress syndrome.}, number={3}, journal={American Journal of Respiratory Cell and Molecular Biology}, publisher={American Thoracic Society}, author={Li, Jingjing and D’Annibale-Tolhurst, Melissa A. and Adler, Kenneth B. and Fang, Shijing and Yin, Qui and Birkenheuer, Adam J. and Levy, Michael G. and Jones, Samuel L. and Sung, Eui Jae and Hawkins, Eleanor C. and et al.}, year={2013}, month={Mar}, pages={314–321} }
 @article{tarigo_scholl_bird_brown_cohn_dean_levy_doolan_trieu_nordone_et al._2013, title={A Novel Candidate Vaccine for Cytauxzoonosis Inferred from Comparative Apicomplexan Genomics}, volume={8}, ISSN={["1932-6203"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-84882652524&partnerID=MN8TOARS}, DOI={10.1371/journal.pone.0071233}, abstractNote={Cytauxzoonosis is an emerging infectious disease of domestic cats (Felis catus) caused by the apicomplexan protozoan parasite Cytauxzoon felis. The growing epidemic, with its high morbidity and mortality points to the need for a protective vaccine against cytauxzoonosis. Unfortunately, the causative agent has yet to be cultured continuously in vitro, rendering traditional vaccine development approaches beyond reach. Here we report the use of comparative genomics to computationally and experimentally interpret the C. felis genome to identify a novel candidate vaccine antigen for cytauxzoonosis. As a starting point we sequenced, assembled, and annotated the C. felis genome and the proteins it encodes. Whole genome alignment revealed considerable conserved synteny with other apicomplexans. In particular, alignments with the bovine parasite Theileria parva revealed that a C. felis gene, cf76, is syntenic to p67 (the leading vaccine candidate for bovine theileriosis), despite a lack of significant sequence similarity. Recombinant subdomains of cf76 were challenged with survivor-cat antiserum and found to be highly seroreactive. Comparison of eleven geographically diverse samples from the south-central and southeastern USA demonstrated 91–100% amino acid sequence identity across cf76, including a high level of conservation in an immunogenic 226 amino acid (24 kDa) carboxyl terminal domain. Using in situ hybridization, transcription of cf76 was documented in the schizogenous stage of parasite replication, the life stage that is believed to be the most important for development of a protective immune response. Collectively, these data point to identification of the first potential vaccine candidate antigen for cytauxzoonosis. Further, our bioinformatic approach emphasizes the use of comparative genomics as an accelerated path to developing vaccines against experimentally intractable pathogens.}, number={8}, journal={PLOS ONE}, author={Tarigo, Jaime L. and Scholl, Elizabeth H. and Bird, David McK and Brown, Corrie C. and Cohn, Leah A. and Dean, Gregg A. and Levy, Michael G. and Doolan, Denise L. and Trieu, Angela and Nordone, Shila K. and et al.}, year={2013}, month={Aug} }
 @article{qurollo_davenport_sherbert_grindem_birkenheuer_breitschwerdt_2013, title={Infection with Panola MountainEhrlichiasp. in a Dog with Atypical Lymphocytes and Clonal T-Cell Expansion}, volume={27}, ISSN={0891-6640}, url={http://dx.doi.org/10.1111/jvim.12148}, DOI={10.1111/jvim.12148}, abstractNote={An 11-year-old, castrated male Scottish Terrier from Raleigh, NC that lived predominantly indoors and had no travel history was referred to the North Carolina State University Veterinary Health Complex (NCSU-VHC) in October 2012 for routine reevaluation of hepatobiliary disease. The dog's previous 3-year medical history included biliary mucocele (2010); neutrophilic hepatitis (2011); recurrent Escherichia coli urinary tract infections, esophageal dysmotility, aspiration pneumonia, and transient thrombocytopenia (2012); and food responsive enteropathy (2009–2012). All of these medical problems were well controlled at the time of examination and no clinical abnormalities were reported by the dog's owners. In the months before the present examination, ticks occasionally were noted and fleas were commonly found on the dog despite reported use of preventive therapies. On physical examination, the dog was obese (body condition score, 7 out of 9) and had mild hepatomegaly, both of which had been present for more than a year. Notable CBC findings included mild thrombocytopenia (platelet count, 143,000/μL; reference interval [RI], 190,000–468,000/μL) and an increased number of atypical lymphocytes (1,773/μL), with normal appearing lymphocytes within the laboratory reference range (1,854/μL; RI, 594–3,305/μL) and an otherwise normal differential cell count. A review of the blood smear by a pathologist identified a population of immature lymphocytes with angular nuclei and cell shape, multiple nucleoli, and deeply basophilic, vacuolated cytoplasm that had a tendency to mold into surrounding cells (Fig 1). These morphologic abnormalities raised the suspicion of possible lymphoid neoplasia, although these changes can be seen secondary to reactive processes such as chronic inflammatory or infectious diseases. Serum biochemical abnormalities included an increase in alkaline phosphatase activity (ALP; 817 IU/L; RI, 16–140 IU/L) and alanine aminotransferase activity (ALT; 60 IU/L; RI, 12–54). During the previous 12-month period, ALT activity varied between normal and 80 IU/L and ALP activity varied between 359 and 534 IU/L. Because these hematological and serum biochemical abnormalities were present 2 weeks later, abdominal ultrasound examination was repeated and identified mottled splenic parenchyma and several previously identified changes including hyperechoic nodular hepatomegaly; mild dystrophic mineralization of the spleen, liver, kidneys, and prostate; and, a thickened urinary bladder apex. Splenic cytology identified a mixed lymphoid population and mild extramedullary hematopoiesis. Liver cytology identified normal hepatocytes and an expansion of intermediate lymphocytes, raising concern for lymphoma. Cytology of a palpably normal popliteal lymph node identified expansion of intermediate to large lymphocytes, most consistent with lymphoma (Fig 2). T-cell clonality was documented in a liver aspirate after submission to NCSU-CVM Clinical Immunology Laboratory for polymerase chain reaction (PCR) to identify antigen receptor rearrangements (PARR). Aseptically collected ethylenediamine tetraacetic acid (EDTA)-anticoagulated whole blood and serum were submitted to the NCSU-CVM Vector Borne Diseases Diagnostic Laboratory (VBDDL) for a vector-borne pathogen serology panel and Bartonella alpha-Proteobacteria Growth Medium (BAPGM) enrichment blood culture PCR platform. The dog was seronegative for antibodies to Anaplasma phagocytophilum, Anaplasma platys, and Borrelia burgdorferi and for Dirofilaria immitis antigen using a commercial enzyme-linked immunosorbent assay based kit (SNAP® 4Dx® Plusa); the dog was seronegative for antibodies to Babesia canis, Babesia gibsoni, Bartonella henselae, and Bartonella vinsonii subsp. berkhoffii, using indirect immunofluorescent antibody (IFA) testing. By IFA, the dog was seroreactive to Rickettsia rickettsii (1:64; laboratory cut-off value, 1:64) and Ehrlichia canis (1:1028; laboratory cut-off value, 1:64) antigens, but seronegative to Ehrlichia spp. peptides in the SNAP 4Dx Plusa kit, which is known to detect antibodies against E. canis, E. chaffeensis, and E. ewingii. Bartonella enrichment blood culture and PCR were negative. Nine months before this presentation, the dog was seronegative in the NCSU-CVM-VBDDL to the vector-borne pathogens tested above. Because of the discrepancy between the E. canis IFA and SNAP 4Dx Plusa test results, Ehrlichia spp. PCR was performed on DNA extracted from the dog's EDTA-anticoagulated whole blood using a previously described 16S ribosomal RNA PCR assay.1 A 351 base pair portion of the 16S rRNA gene was amplified, sequenced, and investigated in the NCBI GenBank nucleotide database. The partial 16S rRNA gene identified infection with the Panola Mountain Ehrlichia sp. (PME) with 100% coverage (331 bp) and 100% identity to PME 16S ribosomal RNA gene (DQ324367.1). To confirm these results, alternative PME genes, including gltA and map1 were targeted. Furthermore, sodb, a gene not previously sequenced for PME, was amplified using newly developed primers designed to amplify Ehrlichia spp. sodb genes. Primers designed to amplify a 311 bp portion of PME gltA (EPM-gltA-Forward, 5′-CGTGTTTTTCTGCCTTAGCTGCAC and EPM-gltA-Reverse, 5′-CGGCCCAGAAGAACC TGTCA) based on a published PME gltA sequence (DQ363995.1) and a second set of primers designed to target a 480 bp portion of PME map1 (EPM-map1-Forward-3, 5′-CGAGAGCCAACGTTTACAT and EPM-map1-Reverse-2, 5′-GTACCAATACCTGCACATAC) based on published PME map1 sequences (DQ324368.1, EU272373.1 and EU272355.1) were used. Primers designed to amplify a 300 bp portion of sodb (sodb-Forward, 5′-TTTAATAATGCTGGTCAAGTATGGAATCAT and sodb-Reverse, 5′-AAGCGTGTTCCCATACATCCATAG) based on published Ehrlichia spp. sodb sequences (AF392615.1, CP000107.1) were used. Reactions contained 5 μL of DNA extract, 12.5 μL of MyTaqHS-2Xb, 0.125 μM (or 0.25 μM for sodb primers) of each primerc and RNAse-free, molecular-grade water to a final volume of 25 μL. All reactions were performed in a thermocyclerd with an aluminum block under the following conditions, with respective primer annealing temperatures specified: initial temperature at 94°C for 3 minutes, 55 cycles consisting of denaturation at 94°C for 10 seconds, annealing at 68°C (gltA), 62°C (map1), 58°C (sodb) for 10 seconds, extension at 72°C for 15 seconds, and a final extension at 72°C for 30 seconds. Negative controls (RNAse-free, molecular-grade water and uninfected canine genomic DNA) were included in all assays. Amplified PCR products were sequenced directlye and alignments were made with GenBank sequences using AlignX software.f Sequence identities for the partial gltA and map1 genes are as follows, all with 100% coverage: gltA, 100% similar (269 bp) to PME partial gltA gene from an infected goat (DQ363995.1) and Amblyomma americanum tick (EU272374.1); map1, 100% similar (460 bp) to PME partial map1 gene from an infected goat (DQ324368.1) and A. americanum tick (EU272373.1). Before this study, there was no sequence for PME sodb deposited in GenBank. The highest sequence identities assigned by BLASTg for the partial gene amplified using Ehrlichia spp. sodb primers are as follows, all with 100% coverage: sodb, 89% (264/295 bp) similar to multiple strains of E. ruminantium sodb gene, with the highest Max scores reported for Senegal (DQ647026.1), Pokoase (DQ647024.1), and Kumm1 (DQ647023.1) strains, 83% (245/295 bp) similar to E. canis strain Jake sodb (CP000107.1) and 80% (236/295 bp) similar to E. chaffeensis strain Arkansas sodb (AF392615.1). The PME partial sodb gene sequence was submitted to Genbank (Accession number KC702804). Primer sets specific for Rickettsia ompA, E. canis p30, E. ewingii p28, and E. chaffeensis p28 did not amplify DNA from the PME-infected blood sample, suggesting this dog was not likely coinfected with Rickettsia or other Ehrlichia species. Retrospective PCRs using DNA extracted from predoxycycline splenic cytologic smears were negative (16S rRNA, gltA, map1, sodb, E. canis p30, E. ewingii p28, and E. chaffeensis p28) for pathogen DNA. Treatment for suspected lymphoma was not initiated because of the possibility that lymphocytosis and other lymphocytic abnormalities were due to a reactive process secondary to ehrlichiosis.2-4 Treatment with doxycycline (approximately 5 mg/kg PO q12h) was commenced for 30 days. After starting doxycycline treatment, occasional vomiting was noted. Resolution of the thrombocytopenia and lymphocytosis occurred 1 week after starting the treatment. ALP and ALT activities remained increased at 1,989 IU/L and 119 IU/L, respectively. One week after completion of doxycycline, repeat aspiration cytology identified a mixed lymphoid population with expansion of intermediate lymphocytes and mild extramedullary hematopoiesis within the spleen, a mixed lymphoid population within the popliteal lymph node, and an expansion of intermediate lymphocytes and mild vacuolar change within the liver. Flow cytometry of the liver showed a population of large, granular cells with positive intracellular staining for CD3, consistent with T-cell lymphoma. When cytology was repeated again 4 weeks after completion of doxycycline, there was resolution of the abnormal lymphoid population in liver and lymph nodes, but there were increased numbers of lymphoblasts and mild persistent extramedullary hematopoiesis within the spleen (Fig 3). ALP and ALT activities had decreased to 1,352 IU/L and 66 IU/L, respectively. Throughout the 3-month time period described in this case report, the owners reported no clinical abnormalities other than occasional gastrointestinal signs, but felt in retrospect that the dog may have been slightly lethargic as they reported it was more energetic after completion of the doxycycline treatment regimen. Furthermore, 5 subsequent CBCs performed over the next 6 months remained normal. EDTA-anticoagulated whole blood and convalescent serum samples collected from the dog approximately 2 weeks after starting doxycycline treatment were PCR negative for PME (16S rRNA, gltA, map1 and sodb) and the SNAP 4Dx Plus was positive for anti-Ehrlichia spp. antibodies, respectively. Convalescent serum, collected approximately 5 weeks after completion of antibiotic treatment, was minimally reactive (1:32) to R. rickettsii antigens, whereas the dog remained E. canis seroreactive by both IFA (1:512) and SNAP 4Dx Plusa (weak Ehrlichia spp. positive). EDTA whole blood collected at this time remained PCR negative (16S rRNA, gltA, map1 and sodb) for PME. In this report, we provide molecular evidence of PME in a thrombocytopenic dog with abnormal lymphocytosis and clonal T-cell expansion. Treatment with doxycycline resulted in resolution of thrombocytopenia, abnormal lymphocytosis, and abnormal lymphoid cells in liver and lymph nodes, supporting a potential role for PME as a cause of host immune dysregulation. These findings also support a potential pathogenic role for PME as a cause of thrombocytopenia in dogs from the United States. Cytopathology and flow cytometry in this case identified a large number of atypical lymphocytes in the peripheral blood and in hepatic and splenic tissue aspirates with a high number of intermediate lymphocytes, granular CD3+ cells, and clonal T-cell expansion. Intracellular infections that induce cell-mediated immunity can cause cytological changes similar to malignancy.5, 6 A study characterizing peripheral blood smears in human ehrlichiosis patients infected with either E. chaffeensis or E. ewingii documented prominent large granular lymphocytes with atypical, folded, hyperchromatic nuclei that might be confused with neoplastic NK or NK-like T-cells.6 Immunophenotypes compared between dogs with naturally acquired canine monocytic ehrlichiosis (CME) and healthy dogs found that dogs with CME had higher relative numbers of CD3+ T-cells in peripheral blood than did healthy dogs.7 Additional studies found that dogs experimentally infected with E. canis had transitory CD8+ lymphocytosis in both peripheral blood and lymph nodes.8, 9 Additional studies describe clinically ill, E. canis-seropositive dogs with an increasing percentage of peripheral blood CD8+ lymphocytes.3, 10 PARR results from the dog in this case demonstrated clonal T-cell expansion, typically associated with malignancy. At least 2 reports, however, describe this phenomenon in dogs with E. canis infections.2, 4 In a previous report, an E. canis-seropositive dog with pancytopenia and clonal T-cell expansion was treated with doxycycline, which resolved the pancytopenia and the dog remained healthy 2 years later.2 Further investigation is needed to determine if expanded T-cells in E. canis infections are transitory or potentially could develop into lymphoid malignancy, particularly in association with chronic undiagnosed infections. As current evidence suggests, a role for Ehrlichia spp. in immune dysregulation is likely, but the extent may be subject to host factors, species and strain virulence, as well as phase of the disease. CME has acute, subclinical, and chronic phases representing different infection durations and variations in clinical disease manifestations. One recent study, however, found no statistically significant differences among CD3+, CD8+, and, CD4+ cells in peripheral blood samples from dogs with clinical or subclinical CME.11 Signs of lymphoid malignancy continue to be monitored in the dog of this report. Resolution of the immunologic abnormalities noted in liver, lymph node, and blood after doxycycline treatment, however, strongly supports an infectious etiology. To the authors’ knowledge, this is the first report of PME infection in a dog. Genetically and antigenically similar to E. ruminantium, PME was first identified by PCR in a goat experimentally infested with A. americanum ticks collected from Panola Mountain State Park, Georgia.12 Goats infected with PME developed serous nasal discharge and febrile illness with hematologic changes consisting of decreased ALP activities and neutropenia; rare morulae in mononuclear cells were identified in 1 goat.12, 13 PME also was detected by PCR in whole blood from a man in Atlanta, GA who developed myalgia after being bitten by a nymphal A. americanum tick.14 No hematologic abnormalities were reported, and clinical signs resolved after doxycycline treatment. The role, if any, of PME in the overall pathogenesis of the various disease manifestations described in the dog of this report over its 3 year history of illness is impossible to assess. At the time PME was identified, the dog was asymptomatic and hematological abnormalities were limited to thrombocytopenia, increased numbers of atypical lymphocytes, and progressively increasing ALP activity (after initiation of doxycycline). The most notable characteristics of illness that potentially were related to PME infection in this dog included thrombocytopenia, cytological changes in the liver, lymph node and spleen, and clonal T-cell expansion. After treatment with doxycycline, sequential resolution of most of these abnormalities occurred. It is not clear when the dog became infected with PME, but serum screened for exposure to vector-borne pathogens 8 months earlier was negative, and the dog's owners reported flea and tick exposure over the previous 6-month time period. PME exposure presumably occurred in central NC because the dog had no travel history outside of the state. This is not a surprising finding, given documentation of PME in A. americanum from the eastern United States, with PME-positive ticks detected in FL, GA, KY, NJ, and NY.15 In addition, deer from AR, NC, and VA were PCR positive for PME and were shown to be competent reservoirs for the pathogen.16 Documentation of PME in this dog supports the possibility that this tick-borne organism may represent an unrecognized human or ruminant pathogen in NC and surrounding states. Currently, serological diagnostics specific for PME are not available. Serum from goats infected with PME was weakly IFA seropositive to E. chaffeensis and seropositive by ELISA to the MAP1 protein from E. ruminantium.12, 13 Serum from the dog of this report reacted strongly with E. canis antigen by IFA, but less strongly with the synthetic antigens used in a commercial ELISA (SNAP 4Dx Plus). Antibodies to other Ehrlichia spp. such as E. chaffeensis have been shown to cross-react with E. canis antigens, and it is likely that the reactivity from this PME-infected dog also represented a cross-reaction.17 Whereas E. canis, E. ewingii, and E. chaffeensis were not detected by PCR, we cannot rule out the possibility that seroreactivity was because of previous exposure with one of these pathogens. In addition to highlighting the emergence of vector-borne pathogens in a novel host species, the findings in this case report underscore the challenges faced in diagnosing canine lymphocytic malignancies and reinforce the need to better understand the immunopathology of canine ehrlichiosis, which can be caused by E. canis, E. chaffeensis, E. ewingii, E. muris, and PME in dogs in North America. Documentation of abnormal or expanded lymphocyte populations in the blood or tissues of dogs should prompt diagnostic consideration of infection with an intracellular pathogen, including Ehrlichia spp. Conflict of Interest: Authors disclose no conflict of interest.}, number={5}, journal={Journal of Veterinary Internal Medicine}, publisher={Wiley}, author={Qurollo, B.A. and Davenport, A.C. and Sherbert, B.M. and Grindem, C.B. and Birkenheuer, A.J. and Breitschwerdt, E.B.}, year={2013}, month={Jul}, pages={1251–1255} }
 @article{schreeg_marr_tarigo_cohn_levy_birkenheuer_2013, title={Pharmacogenomics of Cytauxzoon felis Cytochrome b: Implications for Atovaquone and Azithromycin Therapy in Domestic Cats with Cytauxzoonosis}, volume={51}, ISSN={["0095-1137"]}, DOI={10.1128/jcm.01407-13}, abstractNote={ABSTRACT}, number={9}, journal={JOURNAL OF CLINICAL MICROBIOLOGY}, author={Schreeg, Megan E. and Marr, Henry S. and Tarigo, Jaime and Cohn, Leah A. and Levy, Michael G. and Birkenheuer, Adam J.}, year={2013}, month={Sep}, pages={3066–3069} }
 @article{palerme_brown_marks_birkenheuer_2013, title={Splenosystemic Shunts in Cats: A Retrospective of 33 Cases (2004-2011)}, volume={27}, ISSN={["1939-1676"]}, DOI={10.1111/jvim.12188}, abstractNote={BackgroundPortosystemic shunts are uncommonly reported in cats. The majority of reports describe congenital shunts in young cats originating from the left gastric vein. Although they are only rarely reported, acquired portosystemic shunts in cats appear to be more variable in their anatomic location.}, number={6}, journal={JOURNAL OF VETERINARY INTERNAL MEDICINE}, author={Palerme, J-S. and Brown, J. C. and Marks, S. L. and Birkenheuer, A. J.}, year={2013}, month={Nov}, pages={1347–1353} }
 @article{owens_downey_pressler_birkenheuer_chandler_scott-moncrieff_2012, title={Congenital Adrenal Hyperplasia Associated with Mutation in an 11 ss-Hydroxylase-Like Gene in a Cat}, volume={26}, ISSN={["0891-6640"]}, DOI={10.1111/j.1939-1676.2012.00971.x}, abstractNote={A 3-year-old indeterminate sex domestic medium-hair cat was presented to the Purdue University Veterinary Teaching Hospital for further evaluation of chronic polyuria and polydipsia, foul-smelling urine, and increased serum androgen concentrations. The cat had been surrendered to a shelter 4 months previously for undetermined reasons. Diagnostic tests performed before referral included a serum biochemistry panel, CBC, urinalysis, urine culture, and ACTH stimulation testing (cortisol, aldosterone, and sex hormone assay). Abnormalities included minimally concentrated urine (specific gravity. 1.018), decreased baseline and stimulated serum cortisol and aldosterone concentrations, increased baseline progesterone, and increased baseline and stimulated 17-OH progesterone and androstenedione concentrations. The cat had a small body frame, thickened skin, gynecomastia, a fully formed penis with barbs, and an intact but empty scrotum. Initial indirect systolic blood pressure was 180 mm Hg, and remained persistently elevated during hospitalization. Clinicopathologic abnormalities included increased serum urea nitrogen concentration (41 mg/dL, reference range 15–35 mg/dL), hypernatremia (158 mmol/L, reference range 148–157 mmol/L), and hyperglobulinemia (5.2 g/dL, reference range 2.3–3.8 g/dL). Feline leukemia and feline immunodeficiency virus status were both negative, and a CBC did not reveal abnormalities. Urine specific gravity was 1.007, with no abnormalities on urine dipstick or sediment examinations. A repeated ACTH stimulation test including sex hormone assay confirmed the previously noted abnormalities (Table 1). Endogenous ACTH concentration was increased (>1,250 pg/mL, reference range 25–50 pg/mL).1 Abdominal radiographs did not reveal abnormalities and abdominal sonographic examination revealed anatomically normal adrenal glands and no identifiable internal genitalia. Baseline serum deoxycorticosterone (DOC), 11-desoxycortisol (11-DES), dehydroepiandrosterone (DHEA), pregnenolone, and corticosterone concentrations were measured by high-performance liquid chromatography with tandem mass spectrometry by specific assays validated to bioanalytical standards, and compared with 4 age-matched healthy cats (2 neutered males, 2 spayed females).1 Serum concentrations of DOC (1,177 ng/dL [healthy control cats, 4.6–18.0]), 11-DES (3,647 ng/dL [healthy control cats, 9–29]), DHEA (155 ng/dL [healthy control cats, 9–47]), and pregnenolone (552 ng/dL [healthy control cats, 115–311]) were markedly greater than results for any of the 4 healthy cats, whereas serum corticosterone concentration (9 ng/dL [healthy control cats, 164–550]) was lower. Exploratory laparotomy was performed with the aim to identify internal testes that would explain the cat's foul-smelling urine and physical examination abnormalities. The vas deferens and spermatic cords were bilaterally identified traversing through the inguinal rings and into the scrotum, consistent with prior castration; the identity of this tissue was confirmed histopathologically. The adrenal glands were free of gross lesions. The cat was confirmed as genetically male by DNA extraction of whole blood with identification of X and Y chromosome linked markers.2 The cat was complacent, easy to work with, and good natured during initial hospitalization. After evaluation the cat was adopted into a home with 2 other cats, and immediately demonstrated intercat and owner-directed aggression during play and handling. A diagnosis of intermale aggression was made, presumed secondary to excess androgen concentration or other hormonal imbalance. Behavior modification and fluoxetine (0.6 mg/kg PO q24h) failed to modify the undesired behavior. A presumptive diagnosis of 11β-hydroxylase deficiency resulting in congenital adrenal hyperplasia (CAH) was made based on results of the physical examination abnormalities and adrenal steroid testing, and prednisone therapy (0.2 mg/kg, PO, q24h) was prescribed. Two weeks after initiation of prednisone, the cat's mammary glands had decreased in size and systolic blood pressure had decreased to 150 mm Hg. Because of the partial but incomplete response, after 8 weeks the dose of prednisone was increased (0.5 mg/kg PO q24h). Complete regression of mammary tissue and gradual regression of the penile barbs were noted after 3 weeks at the increased prednisone dose: systolic blood pressure (165 mm Hg) and endogenous ACTH concentration (235 pg/mL), however, were still increased. The prednisone dose was increased further (0.8 mg/kg/d), which resulted in normalization of systolic blood pressure (120 mm Hg) and resolution of polyuria, polydipsia, and malodorous urine within 4 weeks. Secondary sex characteristics continued to resolve, with subjective reduction in skin thickening and regression of the penile barbs. However, despite resolution of clinical signs, repeat endocrine testing revealed persistently increased endogenous ACTH (617 pg/mL) and androstenedione (>10 ng/mL) concentrations. Therapy was changed to prednisolone (0.62 mg/kg PO q24h) because of increased bioavailability.2 The cat's systolic blood pressure (110 mm Hg) and endogenous ACTH (26 pg/mL) decreased to within reference range within 2 weeks of the change in therapy. Sex hormone assay revealed baseline concentrations of androstenedione (0.39 ng/mL), progesterone (<0.03 ng/mL), 17 OH progesterone (0.09 ng/mL), and testosterone (0.02 ng/mL) within or below reference ranges. Serum aldosterone concentration remained below reference range (<11 pg/mL). Eight weeks after therapy was changed to prednisolone, the cat was diagnosed with Klebsiella pyelonephritis based on acute lethargy and inappetance, abdominal pain, an inflammatory leukogram, azotemia, and a positive urine culture, and successfully treated with 6 weeks of amoxicillin/clavulanic acid3 (15.5 mg/kg/d PO divided q12h). After antibiotic treatment the prednisolone dose was decreased (0.5 mg/kg PO q24h). Nine months later the cat remained normotensive (systolic blood pressure 140 mm Hg), endogenous ACTH was within reference range (11.6 pg/mL), DOC was 102 ng/dL, and sex hormone concentrations were markedly reduced pre- and post-ACTH stimulation (Table 1). Mild inte-male aggression persisted; however, owner-directed aggression was almost nonexistent at this time and the cat is more relaxed in his home environment. Genetic mutations associated with 11β-hydroxylase deficiency in people have thus far all been identified within the coding regions of CYP11B1, the 11β-hydroxylase gene.3 Alignment4 of the 9 Homo sapiens CYP11B1 exons (GenBank accession no. NG_007954) to the 2X Felis catus whole genome shotgun (GenBank accession no. PRJNA10762) identified a CYP11B1-like gene within 3 nonoverlapping feline genome contigs (GenBank accession nos. ACBE01524914.1, ACBE01524915.1, ACBE01524916.1). To determine whether 11β-hydroxylase deficiency in the cat of this report was likewise because of mutation of the CYP11B1-like coding region, genomic sequence analysis of this cat's and clinically healthy control cats' CYP11B1-like gene was performed. The complete feline CYP11B1-like gene was amplified in 13 overlapping amplicons for the cat reported here and 1 healthy control cat with total DNA isolated5 from 200 μL whole blood (Table 2). After PCR, appropriately sized amplicons were isolated, purified as needed,6 and directly sequenced.7 Amplicons with unresolvable chromatograms were presumed to be secondary to intronic variation; sequencing was repeated7 after cloning into a plasmid vector,8 , 9 Chromatograms were manually inspected for heterogeneity, and contigs assembled by commercially available software.10 Intron-exon boundaries were annotated by alignment with human 11-β hydroxylase exons,10 with putative exons translated in tandem in silico. The initial CYP11B1-like gene sequence from the cat of this report suggested that 1 or more mutations were present in the region homologous to exon 7 of human CYP11B1 (Fig 1). The PCR amplification of the putative exon 7 region from the cat of this report and 5 healthy cats was performed using a proofreading polymerase (Table 2). Two exonic mutations within the putative CYP11B1 coding sequence were found exclusively in the cat of this report, including a silent (ie, not associated with a change in amino acid) guanosine-to-adenosine mutation in exon 1, and a guanosine-to-adenosine mutation in exon 7 that results in an arginine to glutamine amino acid substitution; this latter mutation results in 11β-hydroxylase deficiency-associated CAH in people.4 CAH syndrome is because of 1 or more adrenal enzyme deficiencies resulting in inadequate cortisol biosynthesis and altered production of androgens.5 CAH is the most common genetic endocrine disorder in humans.6 While mutations associated with dysfunction of any of the 5 adrenal enzymes required for cortisol biosynthesis could result in CAH, mutations of CYP21A (90–95% of cases) and CYP11B1 (5–8% of cases), the genes encoding 21-hydroxylase and 11β-hydroxylase, respectively, are most commonly implicated in people. Inheritance of all forms of CAH is autosomal recessive, with an overall worldwide prevalence of 1 in 15,000 live births.7 CAH is rare in domestic animals, reported thus far in 1 female cat and 1 Great Dane (sex not reported), suspected to be secondary to 11β-hydroxylase deficiency in the cat and 17-hydroxylase deficiency in the dog.8 , 11 11β-hydroxylase is a mitochondrial cytochrome P450 adrenal enzyme that converts 11-deoxycortisol to cortisol, and deoxycorticosterone to corticosterone.5 11β-hydroxylase deficiency results in insufficient cortisol production and secondary inadequate negative feedback to the hypothalamus and cranial pituitary gland, leading to overproduction of ACTH.5 Uninhibited ACTH production continuously stimulates the adrenal cortex to produce steroid precursors, which are inappropriately shunted toward production of sex hormones such as androstenedione. Clinical signs therefore result from increased circulating concentrations of androgens and mineralocorticoids. The hallmark of common forms of CAH in people is virilism of females.9 Human girls can be misclassified as males at birth because of ambiguous genitalia as excess androgen concentrations in utero can result in an enlarged penis-like clitoris, a common urogenital sinus replacing a divided urethra and vagina, and partial fusion of the labia majora, which may be mistaken for a scrotum.9 Despite the presence of external male genitalia in the cat of this report, medical records before initial evaluation were unavailable, and it was unclear if he had been neutered or if the cat was a genotypic female with ambiguous genitalia; without this information definitive sexual classification as a male required histopathology and genotyping. Additional phenotypic changes observed in people with 11β-hydroxylase deficiency include precocious puberty, gynecomastia, and benign testicular nodules in males,5 hirsutism in females,9 and premature physeal closure, stunted growth, acne, and infertility in both sexes.5 Although these findings may be difficult to identify in domestic animals, the cat reported here did have gynecomastia, a greasy haircoat, and a small body frame. Whether or not precocious puberty, testicular nodules, or infertility would have been encountered is unknown because of the cat's age at presentation and prior castration. 21-hydroxylase deficiency, the most common cause of CAH in people, results in defective DOC production, absence of aldosterone, massive salt-wasting, and life-threatening systemic hypotension, whereas 11β-hydroxylase deficiency-associated CAH results in excess DOC, salt retention, volume expansion, and hypertension. Despite the weak mineralocorticoid effects (one-thirtieth the potency of aldosterone) of DOC, 10–100-fold increases in production of this hormone in 11β-hydroxylase deficient CAH patients often results in hypertension-associated adverse effects.10 Treatment with corticosteroids reduces the production of DOC and may resolve hypertension. The 1 previously reported 11β-hydroxylase deficiency cat was too aggressive for accurate blood pressure measuring; therefore this is the first report in which hypertension, and successful therapy, is documented in a cat with CAH.8 Consistent with reports of CAH in people, the systolic blood pressure of the cat in this report normalized synchronously with reduction of endogenous ACTH and serum androgen concentrations.5 Treatment of choice for human patients with 11β-hydroxylase deficiency is supplementation with corticosteroids, usually hydrocortisone because of its similarity to cortisol.11, 12 Differences in 24-hour cortisol demand, and at different life stages, frequently complicates therapy in people: for example, infants and adolescents routinely require higher doses of glucocorticoids than do adults.13 A more aggressive alternative to medical therapy, particularly when adverse effects secondary to increased exogenous glucocorticoid administration become intolerable, is bilateral adrenalectomy and treatment for hypoadrenocorticism.14 Serum concentrations of renin, 17-OH progesterone, androstenedione, testosterone (in females), and DOC can be used in people as markers of adequate hormonal control.5, 15 We used multiple criteria to evaluate adequate adrenal suppression for the cat of this report, including endogenous ACTH concentration, serum androgen concentrations, systemic blood pressure, physical examination findings, and serum DOC concentration. Psychological disorders are strongly associated with CAH in people, including higher aggression scores in females presumptively secondary to increased in utero androgen concentrations,16 increased anxiety and depression, and overall impaired general quality of life.6 The behavioral abnormalities noted in the cat of this report might be consistent with these findings, as standard behavior modification both with and without psychopharmacologic therapy was not effective until after the cat's hormonal status was equivalent to that of a castrated male cat, and therefore more consistent with intermale aggression rather than territorial aggression. 11β-hydroxylase is capable of catalyzing the terminal biosynthesis reactions of both aldosterone and cortisol.17 CYP11B1 encodes 11β-hydroxylase, which is responsible for cortisol biosynthesis in the zona fasciculata and zona reticularis, whereas CYP11B2 encodes aldosterone synthase, the enzyme necessary for aldosterone biosynthesis in the zona glomerulosa.18 These 2 unique genes have been identified in people and several rodent species,17-21 whereas other animals, including frogs,22 pigs,8 and cows22 have a single isozyme that catalyzes both reactions. It is unclear whether or not cats have 2 distinct enzymes or a single isozyme. In people, serum aldosterone concentration is decreased with 11β-hydroxylase deficiency because increased DOC concentration provides adequate and even excessive amounts of mineralocorticoid activity, and aldosterone production (which is stimulated by hyperkalemia and the renin-angiotensin system) is down-regulated.5, 23, 24 When ACTH production decreases after corticosteroid supplementation, DOC production declines as well, eliminating the low-renin hypertension and returning the sodium-potassium balance to normal: serum aldosterone concentrations should consequently return to normal, precluding the need for mineralocorticoid supplementation.5 It is assumed that reduction in DOC concentrations in species with a single isozyme would result in a state of mineralocorticoid deficiency, with life-threatening hyperkalemia and hyponatremia. Hyponatremia or hyperkalemia were never recorded in the cat of this report despite persistent hypoaldosteronism, likely because DOC was maintained at high enough concentrations. Although there is not sufficient clinicopathologic evidence to support 2 functional isoenzymes in this cat, it is still possible that 2 distinct isoenzymes are present but DOC concentration did not decrease low enough to allow stimulation of aldosterone biosynthesis. Congenital adrenal hyperplasia is the most common genetic endocrine disorder in humans, yet this is just the 2nd report of the disease in a cat. Although a genetic disorder, clinical signs might not be clinically apparent until later in life and could vary considerably from neonatal hypovolemic shock to male precocious puberty and mild hypertension. While many patients with this disorder might not survive the neonatal period, many others may go unnoticed with subtle clinical signs. CAH should be a differential diagnosis for cats with unexplained hypertension, polyuria, polydipsia, presence of secondary sex characteristics postneutering, and behavioral abnormalities, including intercat aggression. The information presented in this manuscript was presented in part at the Society of Comparative Endocrinology in Fort Collins, CO in May 2011. We acknowledge Dr Alison Muehrcke, for her referral of this case to the Purdue University Veterinary Teaching Hospital. Work described here was not supported by any grant or funding agency. Conflict of Interest: Dr Pressler is an Associate Editor for the Journal of Veterinary Internal Medicine.}, number={5}, journal={JOURNAL OF VETERINARY INTERNAL MEDICINE}, author={Owens, S. L. and Downey, M. E. and Pressler, B. M. and Birkenheuer, A. J. and Chandler, D. W. and Scott-Moncrieff, J. C.}, year={2012}, pages={1221–1226} }
 @article{lewis_cohn_marr_birkenheuer_2012, title={Diminazene Diaceturate for Treatment of Chronic Cytauxzoon felis Parasitemia in Naturally Infected Cats}, DOI={10.1111/j.1939-1676.2012.01003.x}, abstractNote={BackgroundCytauxzoon felis is a hemoprotozoal parasite that causes substantial morbidity and mortality during the acute phase of infection in cats. However, cats that survive the acute illness remain persistently infected and may serve as a reservoir for the tick‐transmitted pathogen.}, journal={Journal of Veterinary Internal Medicine}, author={Lewis, K. M. and Cohn, L. A. and Marr, H. S. and Birkenheuer, A. J.}, year={2012} }
 @article{lewis_cohn_downey_whitney_birkenheuer_2012, title={Evaluation of Cytauxzoon felis infection status in captive-born wild felids housed in an area endemic for the pathogen}, volume={241}, ISSN={["0003-1488"]}, DOI={10.2460/javma.241.8.1088}, abstractNote={Abstract}, number={8}, journal={JAVMA-JOURNAL OF THE AMERICAN VETERINARY MEDICAL ASSOCIATION}, author={Lewis, Kristin M. and Cohn, Leah A. and Downey, Megan E. and Whitney, Marlyn S. and Birkenheuer, Adam J.}, year={2012}, month={Oct}, pages={1088–1092} }
 @article{holowaychuk_birkenheuer_li_marr_boll_nordone_2012, title={Hypocalcemia and Hypovitaminosis D in Dogs with Induced Endotoxemia}, volume={26}, ISSN={["0891-6640"]}, DOI={10.1111/j.1939-1676.2012.00886.x}, abstractNote={BackgroundHypocalcemia is a documented electrolyte disturbance in people and animals with sepsis, but its mechanism is poorly understood.}, number={2}, journal={JOURNAL OF VETERINARY INTERNAL MEDICINE}, author={Holowaychuk, M. K. and Birkenheuer, A. J. and Li, J. and Marr, H. and Boll, A. and Nordone, S. K.}, year={2012}, pages={244–251} }
 @article{lewis_cohn_birkenheuer_2012, title={Lack of evidence for perinatal transmission of Cytauxzoon felis in domestic cats}, volume={188}, ISSN={["0304-4017"]}, DOI={10.1016/j.vetpar.2012.02.019}, abstractNote={Cytauxzoon felis is a hemoprotozoan parasite of cats capable of causing severe, often fatal disease during acute infection, but cats that survive the acute stage of disease become chronic carriers. These otherwise healthy carriers are capable of transmitting the infection to other cats via the bite of a vector tick. A variety of other hematoprotozoan parasites are capable of vertical transmission from mother to offspring. If this were possible for C. felis, it could be an important part of the explanation for the apparent emergence of this disease with an increased incidence in an expanding geographic area. We investigated the possibility of perinatal transmission of C. felis from chronically infected cats to their offspring. Two queens produced a total of 14 healthy kittens in three litters. All kittens tested negative for C. felis by microscopic slide review and PCR until they were adopted to private homes at approximately 12 weeks of age. While this does not rule out the possibility of perinatal transmission, it is unlikely to be a common phenomenon.}, number={1-2}, journal={VETERINARY PARASITOLOGY}, author={Lewis, Kristin M. and Cohn, Leah A. and Birkenheuer, Adam J.}, year={2012}, month={Aug}, pages={172–174} }
 @article{stowe_birkenheuer_grindem_2012, title={Pathology in practice. Intraerythrocytic infection with organisms consistent with a  large Babesia sp.}, volume={241}, DOI={10.2460/javma.241.8.1029}, number={8}, journal={Journal of the American Veterinary Medical Association}, author={Stowe, Devorah Marks and Birkenheuer, Adam and Grindem, Carol B}, year={2012}, month={Oct}, pages={1029–1031} }
 @article{lewis_cohn_birkenheuer_papich_2012, title={Pharmacokinetics of diminazene diaceturate in healthy cats}, volume={35}, ISSN={["0140-7783"]}, DOI={10.1111/j.1365-2885.2011.01359.x}, abstractNote={Survival of five of six cats treated for acute cytauxzoonosis with diminazene aceturate has been reported (Greene et al., 1999). Diminazene, an aromatic diamidine, has been used to treat babesiosis and trypanosomiasis in dogs, leishmaniasis in humans, and a variety of other protozoal diseases in domestic livestock. It was administered at a dose of 2 mg ⁄ kg IM (two doses, 1 week apart) in the few cats with C. felis infection. Diminazene was administered to seven cats experimentally infected with Trypanosoma evansi at a dose of 3.5 mg ⁄ kg via intramuscular injection on five consecutive days (DaSilva et al., 2009). The treatment was 85.7% efficacious in eliminating the parasite, and no adverse effects were observed. Although commercially available in both aceturate (MW 515.5) and diaceturate (MW 587.6) salts, the drug is not Food and Drug Administration approved for use in any species in the United States of America. Feline cytauxzoonosis is a disease with high morbidity and mortality. Left untreated, 97% of cats with clinical cytauxzoonosis will die of the disease (Birkenheuer et al., 2006). Recently, the combination of the anti-malarial drug atovaquone with the antibiotic imidocarb dipropionate produced improved survival, but 40% of treated cats still died of the disease (Cohn et al., 2011). Additionally, this combination therapy requires thrice-daily oral dosing and can be prohibitively expensive. Diminazene, if found to be effective for the treatment of C. felis, would offer advantages both in practicality of dosing and in cost. Before additional clinical studies can be performed, the disposition of diminazene needs investigation. Therefore, our study objective was to characterize the pharmacokinetic profile of diminazene diaceturate in cats. Four adult cats (two male and two female) weighing 3.1– 5.3 kg were used. The cats were healthy based on physical examination, CBC, biochemical panel, and urinalysis. The day prior to study, cats were sedated with propofol (3–5 mg ⁄ kg to effect), and indwelling jugular catheters were placed to facilitate sample collection for the first 48 h. After that, samples were collected via jugular or medial saphenous venipuncture. Heparinized blood samples (2.0 mL) were collected just before (hour 0) and 0.5, 1, 2, 4, 8, 12, 18, 24, 36, 48, 72, 120, and 168 hours after intramuscular administration of diminazene diaceturate. Plasma was separated by centrifugation within 30 min of collection and frozen ()80 C) until analysis. Cats were observed several times daily to monitor for adverse events such as anorexia, lethargy, injection site pain, abnormal mentation, seizures, vomiting, ptylism, or diarrhea. Temperature, heart rate, respiratory rate, and effort were recorded 0, 12, 24, 48, 72, 120, and 168 h after drug administration. At 168 h after injection, CBC, biochemical panel, and urinalysis were repeated. All procedures were approved by the University of Missouri Animal Care and Use Committee. A powdered commercial drug formulation of diminazene diaceturate (Veriben , CevaSante Animale, Libourne, France) was freshly reconstituted with sterile water to a concentration of 7 mg ⁄ mL and the solution was filtered (0.2 micron disk filter; B Braun, Bethlehem, PA, USA) prior to administration. The dose was calculated at 3 mg ⁄ kg diminazene diaceturate (equivalent to 1.68 mg ⁄ kg base or 2.6 mg ⁄ kg of diminazene aceturate). The total volume of reconstituted drug varied from 1.3 to 2.3 mL per cat; all volumes exceeding 2.0 mL were administered in two separate intramuscular (IM) locations. Concentrations of diminazene were measured by highpressure liquid chromatography (HPLC) analysis using UV absorption and ion-pairing conditions. Because the assay detects the amount of diminazene base, it can be used for detection of either diminazene aceturate or diminazene diaceturate. The reference standard of diminazene (Sigma Aldrich, St Louis, MO, USA) was dissolved in distilled water to make up a 1 mg ⁄ mL stock solution. From this stock solution, further dilutions were prepared in distilled water to generate calibration curves in plasma and to generate quality control samples. The calibration curve included eight standards, including zero (blank). The calibration curve linear range was 0.01–10 lg ⁄ mL. The mobile phase consisted of 73% distilled water, 27% acetonitrile, and an ion-pairing reagent added as a mobile phase modifier. As J. vet. Pharmacol. Therap. doi: 10.1111/j.1365-2885.2011.01359.x SHORT COMMUNICATION}, number={6}, journal={JOURNAL OF VETERINARY PHARMACOLOGY AND THERAPEUTICS}, author={Lewis, K. M. and Cohn, L. A. and Birkenheuer, A. J. and Papich, M. G.}, year={2012}, month={Dec}, pages={608–610} }
 @article{di cicco_downey_beeler_marr_cyrog_kidd_diniz_cohn_birkenheuer_2012, title={Re-emergence of Babesia conradae and effective treatment of infected dogs with atovaquone and azithromycin}, volume={187}, ISSN={["0304-4017"]}, DOI={10.1016/j.vetpar.2012.01.006}, abstractNote={Babesia conradae (B. conradae) causes hemolytic anemia in dogs. This organism has not been reported clinically since it was originally described in southern California in 1991. To date, no anti-protozoal therapies have been associated with clearance of B. conradae. This report describes the use of atovaquone and azithromycin for the treatment of dogs naturally infected with B. conradae and report the re-emergence of B. conradae in southern California. Twelve dogs naturally infected with B. conradae were identified by practicing veterinarians and public health officials in southern California. Treatments consisted of a 10 day course of atovaquone (13.3mg/kg PO q 8h) and azithromycin (10-12.5mg/kg PO q 24h). Four dogs were treated in a randomized blinded placebo-controlled fashion, four additional cases were treated in a non-random, non-blinded fashion and one dog received no treatment. All dogs were tested for B. conradae DNA by polymerase chain reaction (PCR) initially and then once or 3 times post treatment (60-210 days). B. conradae infected dogs that received treatment did not have any detectable Babesia DNA by PCR after treatment. In contrast, dogs receiving placebo had detectable Babesia DNA by PCR throughout the study period. Combination therapy with atovaquone and azithromycin appears to be effective for acute and chronic babesiosis caused by B. conradae.}, number={1-2}, journal={VETERINARY PARASITOLOGY}, author={Di Cicco, Michael F. and Downey, Megan E. and Beeler, Emily and Marr, Henry and Cyrog, Peter and Kidd, Linda and Diniz, Pedro Paulo V. P. and Cohn, Leah A. and Birkenheuer, Adam J.}, year={2012}, month={Jun}, pages={23–27} }
 @article{shock_birkenheuer_patton_olfenbuttel_beringer_grove_peek_butfiloski_hughes_lockhart_et al._2012, title={Variation in the ITS-1 and ITS-2 rRNA genomic regions of Cytauxzoon felis from bobcats and pumas in the eastern United States and comparison with sequences from domestic cats}, volume={190}, ISSN={0304-4017}, url={http://dx.doi.org/10.1016/j.vetpar.2012.06.010}, DOI={10.1016/j.vetpar.2012.06.010}, abstractNote={Cytauxzoon felis, a tick-borne protozoan parasite, is the causative agent of cytauxzoonosis in domestic cats in the United States. The natural reservoir for this parasite is the bobcat (Lynx rufus), which typically does not develop clinical signs. Although not likely important reservoirs, C. felis has also been detected in pumas (Puma concolor) in Florida and Louisiana. Recent studies suggest that specific genotypes of C. felis that circulate in domestic cats may be associated with variable clinical outcomes and specific spatial locations. In the current study, we investigated the intraspecific variation of the C. felis internal transcribed spacer (ITS)-1 and ITS-2 rRNA regions from 145 wild felids (139 bobcats and six pumas) from 11 states (Florida, Georgia, Kansas, Kentucky, Louisiana, Missouri, North Carolina, North Dakota, South Carolina, Oklahoma, and Pennsylvania). Unambiguous ITS-1 and ITS-2 data were obtained for 144 and 112 samples, respectively, and both ITS-1 and ITS-2 sequences were obtained for 111 (77%) samples. For the ITS-1 region, sequences from 65 samples collected from wild felids were identical to those previously reported in domestic cats, while the other 79 sequences were unique. C. felis from 45 bobcats and one puma had ITS-1 sequences identical to the most common sequence reported from domestic cats. Within the ITS-2 region, sequences from 49 bobcats were identical to those previously reported in domestic cats and 63 sequences were unique (with some occurring in more than one bobcat). The most common ITS-2 sequence from domestic cats was also common in wild felids (31 bobcats and a puma). Samples from three pumas from Florida and two bobcats from Missouri had a 40- or 41-bp insert in the ITS-2 similar to one described previously in a domestic cat from Arkansas. Additionally, a previously undescribed 198- or 199-bp insert was detected in the ITS-2 sequence from four bobcats. Collectively, based on combined ITS-1 and ITS-2 sequences, five different genotypes were detected in the wild felids. Genotype ITSa was the most common genotype (11 bobcats and one puma) and fewer numbers of ITSb, ITSe, ITSg, and ITSi were detected in bobcats. These data indicate that, based on ITS-1 and ITS-2 sequences, numerous C. felis strains may circulate in wild felids.}, number={1-2}, journal={Veterinary Parasitology}, publisher={Elsevier BV}, author={Shock, Barbara C. and Birkenheuer, Adam J. and Patton, Laura L. and Olfenbuttel, Colleen and Beringer, Jeff and Grove, Daniel M. and Peek, Matt and Butfiloski, Joseph W. and Hughes, Daymond W. and Lockhart, J. Mitchell and et al.}, year={2012}, month={Nov}, pages={29–35} }
 @article{cohn_birkenheuer_brunker_ratcliff_craig_2011, title={Efficacy of Atovaquone and Azithromycin or Imidocarb Dipropionate in Cats with Acute Cytauxzoonosis}, volume={25}, ISSN={["0891-6640"]}, DOI={10.1111/j.1939-1676.2010.0646.x}, abstractNote={Background:  Imidocarb or a combination of atovaquone and azithromycin (A&A) has been suggested for treatment of cats with cytauxzoonosis, but neither has been prospectively evaluated for efficacy.}, number={1}, journal={JOURNAL OF VETERINARY INTERNAL MEDICINE}, author={Cohn, L. A. and Birkenheuer, A. J. and Brunker, J. D. and Ratcliff, E. R. and Craig, A. W.}, year={2011}, pages={55–60} }
 @article{li_birkenheuer_marr_levy_yoder_nordone_2011, title={Expression and function of triggering receptor expressed on myeloid cells-1 (TREM-1) on canine neutrophils}, volume={35}, ISSN={0145-305X}, url={http://dx.doi.org/10.1016/j.dci.2011.03.021}, DOI={10.1016/j.dci.2011.03.021}, abstractNote={The dog is both a valued veterinary species and a widely used translational model for sepsis research. However, relatively little work has been performed evaluating potential biomarkers present during canine infection. Triggering receptor expressed on myeloid cells-1 (TREM-1) has shown promise as a biomarker for infection and pneumonia in humans. Here we describe, for the first time, the expression and function of the canine orthologue of TREM-1. Expression of TREM-1 on canine neutrophils is significantly up-regulated by stimulation with microbial agonists of TLR2/6, TLR1/2, and TLR4/MD2. Kinetics of TREM-1 protein up-regulation are rapid, with significant increases observed within 2 hr of neutrophil activation. Functionally, canine TREM-1 synergistically enhances LPS-induced production of IL-8, TNF-α and a canine orthologue of CXCL1. Collectively, these data suggest that TREM-1 expression in dogs, as it is in humans, is an amplifier of pro-inflammatory responses to microbial products. These results have direct application to veterinary diagnostics as well as the potential to enhance the utility of canine disease models in the assessment of potential therapeutics in the treatment of human sepsis.}, number={8}, journal={Developmental & Comparative Immunology}, publisher={Elsevier BV}, author={Li, Jingjing and Birkenheuer, Adam J. and Marr, Henry S. and Levy, Michael G. and Yoder, Jeffrey A. and Nordone, Shila K.}, year={2011}, month={Aug}, pages={872–880} }
 @article{levy_lappin_glaser_birkenheuer_anderson_edinboro_2011, title={Prevalence of infectious diseases in cats and dogs rescued following Hurricane Katrina}, volume={238}, ISSN={["1943-569X"]}, DOI={10.2460/javma.238.3.311}, abstractNote={Abstract}, number={3}, journal={JAVMA-JOURNAL OF THE AMERICAN VETERINARY MEDICAL ASSOCIATION}, author={Levy, Julie K. and Lappin, Michael R. and Glaser, Amy L. and Birkenheuer, Adam J. and Anderson, Tara C. and Edinboro, Charlotte H.}, year={2011}, month={Feb}, pages={311–317} }
 @article{hummel_grooters_davidson_jennings_nicklas_birkenheuer_2011, title={Successful management of gastrointestinal pythiosis in a dog using itraconazole, terbinafine, and mefenoxam}, volume={49}, ISSN={["1369-3786"]}, DOI={10.3109/13693786.2010.543705}, abstractNote={Medical therapy for pythiosis is hampered by a lack of efficacious drugs. The present report describes a case of canine gastrointestinal pythiosis in which lesions were resolved through the administration of itraconazole, terbinafine, and the agricultural fungicide mefenoxam. No substantial adverse effects occurred in association with administration of the latter compound. Additional studies are needed to evaluate the pharmacokinetics of mefenoxam and to further assess its tolerability and potential efficacy for the treatment of pythiosis in dogs.}, number={5}, journal={MEDICAL MYCOLOGY}, author={Hummel, James and Grooters, Amy and Davidson, Gigi and Jennings, Samuel and Nicklas, Jodi and Birkenheuer, Adam}, year={2011}, month={Jul}, pages={539–542} }
 @article{clancey_horney_burton_birkenheuer_mcburney_tefft_2010, title={Babesia (Theileria) annae in a Red Fox (Vulpes vulpes) from Prince Edward Island, Canada}, volume={46}, ISSN={0090-3558}, url={http://dx.doi.org/10.7589/0090-3558-46.2.615}, DOI={10.7589/0090-3558-46.2.615}, abstractNote={A 4–6-mo-old female red fox (Vulpes vulpes) was presented to the Atlantic Veterinary College (AVC) Teaching Hospital, Prince Edward Island, Canada. On presentation, the fox was weak and had pale mucous membranes. A complete blood count and a serum biochemistry profile were performed. Blood smear examination revealed low numbers of erythrocytes containing centrally to paracentrally located, single, rarely multiple, approximately 1×2 μm, oval to round organisms with morphology similar to Babesia microti. Polymerase chain reaction testing and DNA sequencing of the Babesia species 18S rRNA gene were performed on DNA extracted from whole blood. Results were positive for a Babesia microti–like parasite genetically identical to Babesia (Theileria) annae. The fox was euthanized due to poor prognosis for recovery. Necropsy examination revealed multifocal to locally extensive subacute nonsuppurative meningoencephalitis, an eosinophilic bronchopneumonia, a moderate diffuse vacuolar hepatopathy, and lesions associated with blunt trauma to the left abdominal region. This is the first reported case of a red fox in Canada infected with a piroplasm. It remains uncertain whether the presence of this hemoparasite in this fox was pathogenic or an incidental finding. The potential for competent vectors of Babesia species on Prince Edward Island, the potential for this Babesia microti–like parasite to infect other wild and domestic canids, and the significance of this parasite to the health of infected individuals are yet to be determined.}, number={2}, journal={Journal of Wildlife Diseases}, publisher={Wildlife Disease Association}, author={Clancey, Noel and Horney, Barbara and Burton, Shelley and Birkenheuer, Adam and McBurney, Scott and Tefft, Karen}, year={2010}, month={Apr}, pages={615–621} }
 @article{kamani_sannusi_dogo_tanko_egwu_tafarki_ogo_kemza_onovoh_shamaki_et al._2010, title={Babesia canis and Babesia rossi co-infection in an untraveled Nigerian dog}, volume={173}, ISSN={["1873-2550"]}, DOI={10.1016/j.vetpar.2010.06.040}, abstractNote={A sexually intact 6-month-old female Alsatian dog was presented to the Veterinary Clinic of the National Veterinary Research Institute, Vom, Plateau State, Nigeria, for the following complaints: anorexia, hemoglobinuria, fever, tick infestation and general malaise. Microscopy revealed piroplasms with a wide range of sizes (1-5 μm in length) in red blood cells, raising a suspicion of a co-infection with two or more Babesia species. Specific PCR assays for canine Babesia spp. and DNA sequencing revealed the presence of Babesia canis and Babesia rossi co-infection. This study constitutes the first report of co-infection with B. canis and B. rossi in the West African sub-region and the first report of autochthonous B. canis on the African continent. Practitioners should be aware of potential changes in the species/sub-species of Babesia causing canine babesiosis in this region.}, number={3-4}, journal={VETERINARY PARASITOLOGY}, author={Kamani, Joshua and Sannusi, Abdulrahim and Dogo, A. Goni and Tanko, James T. and Egwu, Kinsley O. and Tafarki, Agbadu E. and Ogo, Isaac N. and Kemza, Sarah and Onovoh, Emmanuel and Shamaki, David and et al.}, year={2010}, month={Oct}, pages={334–335} }
 @article{birkenheuer_horney_bailey_scott_sherbert_catto_marr_camacho_ballman_2010, title={Babesia microti-like infections are prevalent in North American foxes}, volume={172}, ISSN={["1873-2550"]}, DOI={10.1016/j.vetpar.2010.05.020}, abstractNote={Babesia microti-like organisms have recently been identified as a cause of hemolytic anemia and azotemia in European dogs. A genetically and morphologically similar B. microti-like parasite has been identified in two foxes from North America. In order to assess the prevalence of this parasite in North American wild canids we screened blood samples from coyotes (Canis latrans) and red foxes (Vulpes vulpes) from eastern Canada and red foxes and gray foxes (Urocyon cinereoargenteus) from North Carolina, USA for the presence B. microti-like DNA by polymerase chain reaction. Thirty-nine percent (50/127) of the red fox samples, 26% (8/31) of the gray fox samples and none (0/12) from the coyote samples tested positive for the presence of B. microti-like DNA. Partial 18S ribosomal ribonucleic acid and beta-tubulin genes from the North American B. microti-like parasites of foxes were sequenced and samples from six domestic dogs from Spain that were infected with a B. microti-like parasite were analyzed for comparison. Partial 18S ribosomal ribonucleic acid and beta-tubulin gene sequences from the North American B. microti-like parasites of foxes were nearly identical to those previously reported from foxes as well as those from domestic dogs from Spain characterized in this study. Interestingly, partial beta-tubulin gene sequences characterized from the B. microti-like parasites of domestic dogs from Spain in this study were different from those previously reported from a Spanish domestic dog sample which is believed to be a pseudogene. The ability of the North American B. microti-like parasite to infect and induce disease in domestic dogs remains unknown. Further studies investigating the pathogenic potential of the North American B. microti-like parasite in domestic dogs are indicated.}, number={3-4}, journal={VETERINARY PARASITOLOGY}, author={Birkenheuer, Adam J. and Horney, Barbara and Bailey, Matthew and Scott, McBurney and Sherbert, Brittany and Catto, Victoria and Marr, Henry S. and Camacho, Angel-Tomas and Ballman, Anne E.}, year={2010}, month={Sep}, pages={179–182} }
 @article{sikorski_birkenheuer_holowaychuk_mccleary-wheeler_davis_littman_2010, title={Babesiosis Caused by a Large Babesia Species in 7 Immunocompromised Dogs}, volume={24}, ISSN={["1939-1676"]}, DOI={10.1111/j.1939-1676.2009.0440.x}, abstractNote={Background:  A large unnamed Babesia species was detected in a dog with lymphoma. It was unknown if this was an underrecognized pathogen.}, number={1}, journal={JOURNAL OF VETERINARY INTERNAL MEDICINE}, author={Sikorski, L. E. and Birkenheuer, A. J. and Holowaychuk, M. K. and McCleary-Wheeler, A. L. and Davis, J. M. and Littman, M. P.}, year={2010}, pages={127–131} }
 @article{barber_li_diniz_porter_breitschwerdt_claiborne_birkenheuer_levine_levine_chandler_et al._2010, title={Evaluation of Brain Tissue or Cerebrospinal Fluid with Broadly Reactive Polymerase Chain Reaction for Ehrlichia, Anaplasma, Spotted Fever Group Rickettsia, Bartonella, and Borrelia Species in Canine Neurological Diseases (109 Cases)}, volume={24}, ISSN={0891-6640 1939-1676}, url={http://dx.doi.org/10.1111/j.1939-1676.2009.0466.x}, DOI={10.1111/j.1939-1676.2009.0466.x}, abstractNote={BACKGROUND
Vector-transmitted microorganisms in the genera Ehrlichia, Anaplasma, Rickettsia, Bartonella, and Borrelia are commonly suspected in dogs with meningoencephalomyelitis (MEM), but the prevalence of these pathogens in brain tissue and cerebrospinal fluid (CSF) of dogs with MEM is unknown.


HYPOTHESIS/OBJECTIVES
To determine if DNA from these genera is present in brain tissue and CSF of dogs with MEM, including those with meningoencephalitis of unknown etiology (MUE) and histopathologically confirmed cases of granulomatous (GME) and necrotizing meningoencephalomyelitis (NME).


ANIMALS
Hundred and nine dogs examined for neurological signs at 3 university referral hospitals.


METHODS
Brain tissue and CSF were collected prospectively from dogs with neurological disease and evaluated by broadly reactive polymerase chain reaction (PCR) for Ehrlichia, Anaplasma, Spotted Fever Group Rickettsia, Bartonella, and Borrelia species. Medical records were evaluated retrospectively to identify MEM and control cases.


RESULTS
Seventy-five cases of MUE, GME, or NME, including brain tissue from 31 and CSF from 44 cases, were evaluated. Brain tissue from 4 cases and inflammatory CSF from 30 cases with infectious, neoplastic, compressive, vascular, or malformative disease were evaluated as controls. Pathogen nucleic acids were detected in 1 of 109 cases evaluated. Specifically, Bartonella vinsonii subsp. berkhoffii DNA was amplified from 1/6 dogs with histopathologically confirmed GME.


CONCLUSION AND CLINICAL IMPORTANCE
The results of this investigation suggest that microorganisms in the genera Ehrlichia, Anaplasma, Rickettsia, and Borrelia are unlikely to be directly associated with canine MEM in the geographic regions evaluated. The role of Bartonella in the pathogenesis of GME warrants further investigation.}, number={2}, journal={Journal of Veterinary Internal Medicine}, publisher={Wiley}, author={Barber, R.M. and Li, Q. and Diniz, P.P.V.P. and Porter, B.F. and Breitschwerdt, E.B. and Claiborne, M.K. and Birkenheuer, A.J. and Levine, J.M. and Levine, G.J. and Chandler, K. and et al.}, year={2010}, month={Mar}, pages={372–378} }
 @article{chinnadurai_birkenheuer_blanton_maggi_belfiore_marr_breitschwerdt_stoskopf_2010, title={Prevalence of Selected Vector-borne Organisms and Identification of Bartonella Species DNA in North American River Otters (Lontra canadensis)}, volume={46}, ISSN={0090-3558}, url={http://dx.doi.org/10.7589/0090-3558-46.3.947}, DOI={10.7589/0090-3558-46.3.947}, abstractNote={Trapper-killed North American river otters (Lontra canadensis) in North Carolina, USA, were screened for multiple vector-borne bacteria known to be pathogenic to mammals. Blood was collected from 30 carcasses in 2006, from 35 in 2007, and from one live otter in 2008. Samples were screened using conventional polymerase chain reaction (PCR) tests for DNA from Bartonella spp., Ehrlichia spp., and spotted fever group Rickettsia spp. All samples were negative for Rickettsia spp. Twelve of 30 samples from 2006 produced amplicons using the assay designed to detect Ehrlichia spp., but sequencing revealed that the amplified DNA fragment was from a novel Wolbachia sp., thought to be an endosymbiote of a Dirofilaria sp. Between 2006 and 2007, DNA from a novel Bartonella sp. was detected in 19 of 65 animals (29%). Blood from one live otter captured in 2008 was found positive for this Bartonella sp. by both PCR and culture. The pathogenicity of this Bartonella species in river otters or other mammals is unknown.}, number={3}, journal={Journal of Wildlife Diseases}, publisher={Wildlife Disease Association}, author={Chinnadurai, Sathya K. and Birkenheuer, Adam J. and Blanton, Hunter L. and Maggi, Ricardo G. and Belfiore, Natalia and Marr, Henry S. and Breitschwerdt, Edward B. and Stoskopf, Michael K.}, year={2010}, month={Jul}, pages={947–950} }
 @article{bartlett_abou-madi_messick_birkenheuer_kollias_2009, title={DIAGNOSIS AND TREATMENT OF BABESIA ODOCOILEI IN CAPTIVE REINDEER (RANGIFER TARANDUS TARANDUS) AND RECOGNITION OF THREE NOVEL HOST SPECIES}, volume={40}, ISSN={["1937-2825"]}, DOI={10.1638/2008-0011.1}, abstractNote={Abstract Two captive reindeer (Rangifer tarandus tarandus) at a New York zoological institution were diagnosed with Babesia odocoilei. Clinical signs consistent with acute babesiosis included fever, hemoglobinuria, and hemolytic anemia. Both episodes were precipitated by stressful events that may have compromised their immunocompetence. The diagnosis was confirmed by visualization of intraerythrocytic parasites on stained blood smears, polymerase chain reaction, and speciation of the Babesia by sequencing a hypervariable region of the 18S rRNA gene. One reindeer died with gross and histopathologic lesions, including pigmentary nephrosis with severe acute tubular degeneration and necrosis secondary to intravascular hemolysis. A second reindeer was successfully treated with supportive care and an antiprotozoal, imidocarb dipropionate (Imizol, 12%, Schering-Plough Animal Health, Union, New Jersey 07083, USA) at 3 mg/kg s.c. or i.m. s.i.d. on days 1, 2, 6, 9, and 21. Two other reindeer in the exhibit tested negative for Babesia by polymerase chain reaction but were treated with imidocarb dipropionate as prophylaxis while final testing results were pending. Additionally, B. odocoilei was identified in three novel asymptomatic host species within the collection: yak (Bos grunniens), muntjac (Muntiacus reevesi), and markhor goat (Capra falconeri). Due to the high morbidity and mortality associated with acute babesiosis, captive reindeer should receive tick prevention, be tested for subclinical infections in endemic areas, and receive aggressive treatment for acute infections when clinical babesiosis is suspected.}, number={1}, journal={JOURNAL OF ZOO AND WILDLIFE MEDICINE}, author={Bartlett, Susan L. and Abou-Madi, Noha and Messick, Joanne B. and Birkenheuer, Adam and Kollias, George V.}, year={2009}, month={Mar}, pages={152–159} }
 @article{levine_zhou_ghiloni_fields_birkenheuer_gookin_roberston_malloy_feldman_2009, title={Hereditary 1,25-Dihydroxyvitamin D-Resistant Rickets in a Pomeranian Dog Caused by a Novel Mutation in the Vitamin D Receptor Gene}, volume={23}, ISSN={["1939-1676"]}, DOI={10.1111/j.1939-1676.2009.0405.x}, abstractNote={Hypocalcemic rickets encompasses a group of disorders in which intestinal absorption of calcium is insufficient to meet the calcium demands of a growing skeleton. Causes include dietary calcium deficiency and insufficient intestinal absorption of calcium caused by vitamin D deficiency or decreased vitamin D activity. Hereditary disorders of vitamin D documented in humans include vitamin D-dependent rickets type I (VDDR-I), in which there is a deficiency of the renal enzyme (1α-hydroxylase) that converts 25-hydroxyvitamin D3 to the active hormone 1α,25-dihydroxyvitamin D3 (1,25(OH)2D3 or calcitriol) and vitamin D-dependent rickets type II (VDDR-II) in which there is end-organ resistance to the active hormone. The latter is usually secondary to defects in the vitamin D receptor (VDR). VDDR-II is currently more commonly termed hereditary vitamin D-resistant rickets (HVDRR). We describe a case of rickets in a juvenile dog that led us to investigate potential hereditary vitamin D disorders and resulted in the documentation of HVDRR in a dog. The disease is characterized by hypocalcemia, secondary hyperparathyroidism, hypomineralization of bones, rickets, and in some cases, alopecia. An intact female Pomeranian dog was initially evaluated by one of the authors at 2 months of age for nonpruritic alopecia of the head and rear limbs. The remainder of the dog's physical examination at that time was unremarkable. A skin scraping was negative. The dog and her half-sibling (same sire, different dam) had been obtained from a breeder a few days before examination. The half-sibling was the same age as the affected dog, and was normal in appearance. Both dogs were eating a commercial puppy food. At 4 months of age, the affected dog appeared to tire easily when playing. On physical examination, there was lateral bowing of the antebrachium of both forelimbs and palpable thickening of the distal radii. The alopecia had become more generalized and remained nonpruritic (Fig 1). A serum biochemistry profile revealed marked hypocalcemia (total calcium 5.5 mg/dL; reference range 8.9–11.4 mg/dL) with a normal albumin concentration, hyperphosphatemia (phosphate 7.3 mg/dL; reference range 2.5–6 mg/dL), and increased alkaline phosphatase (ALP) (829 U/L; reference range 5–131 U/L). Blood urea nitrogen (BUN) and creatinine, complete blood count, and serum thyroxine (total thyroxine 2.7 μg/dL; reference range 1.0–4.0 μg/dL) were all within normal limits. Photograph of the affected dog at 3 months of age demonstrating the remarkable alopecia. The photograph shows partial alopecia on the ventral chest, in the perineal area, and on the caudo-medial aspect of thighs. In the areas where hair is present, it is thin, likely because of a decreased amount of secondary hairs. The normal half-sibling is pictured to the right for comparison. The dog was reevaluated at 5 months of age for intermittent lameness while playing. Although her alopecia was resolving, her abnormal forelimb conformation persisted. Radiographs showed widening and abnormal lucency (decreased mineralization) of the distal radial and ulnar physes, with sclerosis and widening of the adjacent epiphyses and metaphyses (Fig 2A and B). Persistent hypocalcemia (total calcium 5.7 mg/dL) and low-ionized calcium (0.66 mmol/L; reference range 1.24–1.43 mmol/L) were documented, along with hyperparathyroidism (intact parathyroid hormone [PTH] 739 pg/mL; reference range 27–155 pg/mL).a Pre- and postprandial serum concentrations of bile acids were within reference range. On the basis of hypocalcemia, hyperparathyroidism, and the skeletal abnormalities, a provisional diagnosis was a vitamin D-dependent form of rickets. Treatment with oral calcium supplementation (calcium carbonateb 29 mg/kg PO q24h) and 1,25(OH)2D3c (43 ng/kg PO q24h) was initiated. Lateral (A) and dorsopalmar (B) radiographs of the right carpus acquired at 5 months of age before initiation of treatment. There is widening and increased lucency of the distal radial and ulnar physes with irregular (frayed) metaphyseal margins and flaring of the adjacent epiphyses and metaphyses. Lateral (C) and dorsopalmar (D) radiographs of the right carpus acquired at 8.5 months of age, 3.5 months after initiation of treatment. The distal antebrachial physes appear more normal in width, opacity, and margination. Evidence of distal metaphyseal remodeling is present. Total serum calcium concentrations were persistently low (ranging from 4.9 to 6.2 mg/dL) despite consistent increases in oral calcium and 1,25(OH)2D3 supplementation to doses well above standard recommendations (25–50 mg/kg/d of calcium carbonate and 20–30 ng/kg/d 1,25(OH)2D3)1 (Table 1). At 7 months of age, the dog had a generalized seizure that was controlled by administration of diazepam. At the time of the seizure, total serum calcium concentration was 5.7 mg/dL (reference range 7.9–12 mg/dL).d The dog was referred to the Internal Medicine Service of North Carolina State University, College of Veterinary Medicine (NCSU CVM) for further evaluation. On examination, the dog was bright and alert and interactive. Her conformation was unusual, with the forelimbs being shorter than the hind limbs, and slightly bowed laterally. Despite prior resolution of the alopecia, the affected dog's coat was of poorer quality than that of her half-sibling. The remainder of the physical examination was unremarkable. A serum biochemistry profile confirmed persistent hypocalcemia (6.3 mg/dL; reference range 9.2–11.6 mg/dL), with normal serum phosphorous concentration (5.7 mg/dL; 2–6.7 mg/dL) and renal values (BUN and creatinine). There was high PTH (91 pmol/L; 3–17 pmol/L),e increased ALP (486 U/L; 14–120 U/L), low 25-hydroxyvitamin D (43 nmol/L; 60–215 nmol/L),e and low-ionized calcium (0.94 mmol/L; 1.25–1.45 mmol/L)e concentration in serum. Rickets due to vitamin D resistance was suspected because of the lack of response to high-dose therapy with 1,25(OH)2D3. Since the dog did not respond to oral calcium supplementation, calcium (12.5 mg/kg SQ q24h)f was administered parenterally. 1,25(OH)2D3 therapy (75 ng/kg PO q24h) was continued. At 8 months of age, while still receiving 1,25(OH)2D3, serum 1,25(OH)2D3 concentrationg was measured to definitively differentiate between vitamin D deficiency and the two types of vitamin D-dependent rickets, type-I and type-II (HVDRR). The dog's serum 1,25(OH)2D3 concentration was over 20-fold greater than the upper end of the reference range (1,017 pb/mL; 25–50 pg/mL) despite persistent hypocalcemia (total calcium 5.4 mg/dL; 8.9–11.4 mg/dL). The increased concentration of 1,25 (OH)2D3 was consistent with a diagnosis of HVDRR. In this case, even though the dog was being supplemented with 1,25(OH)2D3, HVDRR was considered likely because high-circulating concentrations of 1,25(OH)2D3 were ineffective in raising serum calcium concentrations, confirming 1,25(OH)2D3 resistance. Oral calcium carbonate therapy was reinitiated at this time (75 mg/kg PO q24h) in addition to the 1,25(OH)2D3 supplementation and daily SQ calcium injections. At 8.5 months of age, the dog became acutely paraplegic in the hind limbs and depressed. The dog was evaluated by her local veterinarian, where total calcium concentration was determined to be 4.0 mg/dL (reference range 7.9–12 mg/dL). The veterinarian administered calcium IM (25 mg/kg IM once)f and referred the dog to the Emergency Service at NCSU. On examination, the dog was opisthotonic and tetanic, stuporous, weak, and believed to be having frequent focal seizures. Ionized calcium was 0.61 mmol/L (reference range 1.11–1.40 mmol/L). Calcium gluconate (100 mg/kg IV once)h was administered slowly, followed by a continuous rate infusion of calcium (10 mg/kg/h IV).h The dog continued to have seizure-like activity and muscle tetany, despite normalization of serum-ionized calcium (0.91 mmol/L) and a normal ECG. Tetany and seizure activity were controlled with midazolami (0.5 mg/kg IV once, then 0.3 mg/kg/h IV). After initial stabilization, a neurologic examination revealed signs attributable to diffuse forebrain disease (variably obtunded to stuporous, intermittent lateral strabismus in both eyes, bilateral miotic pupils, and reduced nasal sensation bilaterally) and hind limb paresis (exaggerated pelvic limb segmental spinal reflexes, absent deep pain in both pelvic limbs and the tail). The forebrain signs were attributed to hypocalcemia exacerbated by potential cerebral edema. The pelvic limb paresis suggested a T3–L3 spinal lesion. Pain management was instituted with fentanyl (2 mcg/kg IV PRN).j Spinal radiographs were suggestive of a compression fracture of the vertebral body of T11, although a congenital deformity could not be ruled out (Fig 3A). Computed tomography confirmed the fracture of T11 and showed accompanying spinal cord compression. Flaring of the ribs at the costochondral junctions (so-called rachitic rosary in people) was also readily apparent on radiographs (Fig 3B). Radiographs of the distal radial and ulnar growth plates, however, showed progressive improvement in mineralization compared with the images taken at 5 months of age (Fig 2C and D). Radiographs at 8.5 months, 3.5 months after initiation of treatment. (A) Lateral radiograph of the spine at the thoracolumbar junction. There is generalized osteopenia, the T11 vertebral body is wedged shaped, and there is concavity of the ventral margin of the vertebral body. These changes are highly suggestive of a pathologic compression fracture, although a preexisting vertebral body anomaly could not be ruled out. (B) Lateral radiograph of the costochondral junctions. The junctions are “flared” and moderately sclerotic. This radiographic finding is frequently present in human patients with rickets, and is commonly called a rachitic rosary, because the lesions palpate like beads on a rosary. Given the severity of the dog's spinal injury (functionally complete), the chance of recovery was very low and would have been compromised by the difficulty in surgical repair of osteopenic bones. As a result, the dog was euthanized. Gross postmortem evaluation revealed signs of rickets with diffusely enlarged costochrondral junctions (beading), a fracture callus on the left eight rib, and displacement of the T11 vertebra. Microscopically, failure of endochondral ossification with retained cartilage cores was noted in several locations (ribs, radius, nasal turbinates). A compression fracture of T11 with subluxation was confirmed. In the region of T11, locally extensive, severe myelomalacia, and fibrinoid vasculitis of the spinal cord were seen. The brain was normal, suggesting that severe hypocalcemia was the cause for the dog's forebrain signs. The parathyroid was diffusely hyperplastic and hypertrophied, consistent with the persistently increased PTH levels. HVDRR in humans is almost always due to mutations in the VDR.2 For this reason, before euthanasia, blood samples were drawn from both the affected dog and the normal half-sibling for genomic sequence analysis of the VDR gene. With owner consent, immediately before euthanasia, the affected dog was anesthetized with propofolk to obtain skin biopsies from her ventral abdomen from which fibroblast cultures were derived. These cultures were subsequently used for analysis of VDR expression and as a source of RNA for VDR sequence analysis. The fibroblasts were grown in minimal essential media containing 10% calf seruml and maintained in an atmosphere of 95% air, 5% CO2 at 37°C. Genomic DNA was isolated from blood using a commercial kit.m Primers were designed based on the DNA sequence of Canis lupus familiaris VDR obtained from GenBank® (accession number NC_006609, Gene ID: 486588) (Table 2). Exons 2–9 of the VDR gene from the affected dog and half-sibling were amplified by PCR and directly sequenced at the Stanford University protein and nucleic acid facility. A unique single base deletion (guanine) was identified at the exon 4-intron junction (Fig 4A) in the affected dog's genomic DNA. The sequence of the exon 4-intron junction of the half-sibling was homozygous for the wild-type sequence (Fig 4B). DNA sequence of exon 4 of the VDR gene in the dog. (A) Genomic DNA sequence of the affected dog's VDR gene. A unique 1 bp (G) deletion was identified in the exon 4-intron junction (B) DNA sequence of the half-sibling's VDR gene. (C) Sequence of the affected dog's VDR cDNA. Deletion of a single guanine nucleotide base was identified in the dog's VDR cDNA resulting in a frameshift that introduced a downstream premature termination codon. The premature stop codon is predicted to be at codon 216, which is 232 amino acids upstream of the normal stop codon. The translated amino acid sequence is shown above the nucleotide sequence. The normal amino acid sequence and that of the affected dog diverge after amino acid R175. From the sequence in GenBank®, the normal VDR exon4-intron sequence is AGG|gtacgtggcc (exon 4 sequence in capitals and intron sequence in lower case with bolded letters indicating splicing recognition sequence). The deletion of a single guanine base in the GG|g triplet would generate the sequence AG|gtacgtggcc and leave the 5′-splice site intact. On the other hand, deletion of guanine in the coding sequence would cause a frameshift in exon 5. In order to document this frameshift, RNA was isolated from the dog's cultured fibroblasts. The RNA was reverse transcribedn and amplifiedo by PCR using gene specific primers (forward primer 5′-ctgcggcccaagctgtcgga and reverse primer 5′-ttggatgctgtaactgaccag). The VDR cDNA products were purifiedp and directly sequenced. As shown in Fig 4C, the single guanine nucleotide deletion in the VDR cDNA led to a frameshift that caused the amino acid sequence to diverge after R175. (It should be noted that the canine R175 corresponds to R154 in humans, because the canine VDR sequence is 21 amino acids longer at the N-terminus compared with the human sequence.) An additional 41 amino acids were in frame before a downstream termination signal occurred. The mutation deleted 273 amino acids of the VDR ligand-binding domain (LBD) and was expected to abolish 1,25(OH)2D3 binding. Although we were able to amplify the VDR cDNA from the dog's cells, the mutant VDR mRNA transcript may be subjected to nonsense-mediated mRNA decay and the affected dog may not express any VDR. To determine whether the mutation abolished 1,25(OH)2D3 binding, we examined [3H]1,25(OH)2D3 binding in the dog's fibroblasts compared with normal human fibroblasts. Cell extracts were incubated with 1 nM [3H]1,25(OH)2D3 (specific activity 158 Ci/mmol) with or without 250-fold excess of radioinert 1,25(OH)2D3 as described previously.3 Hydroxylapatite was used to separate bound and free hormone. Protein concentrations were determined by the Bradford method.4 No detectable binding was observed in extracts from the dog's cells. This report describes a VDR gene mutation resulting in HVDRR in a dog. The molecular basis for HVDRR was attributed to a 1 bp deletion in exon 4 that caused a frameshift and introduced a premature stop in exon 5. We expect that this mutation would lead to the affected dog's failure to respond to 1,25(OH)2D3 in one of 2 ways: either the mutation truncated a major portion of the LBD, leading to the inability of the dog's VDR to bind 1,25(OH)2D3, or the mRNA was subjected to nonsense-mediated degradation and this dog had a complete failure of VDR expression. The absence of 1,25(OH)2D3 binding in the dog fibroblast binding assay confirms that the affected dog's cells lack a VDR capable of responding to 1,25(OH)2D3, although it does not differentiate between these two causes of defective VDR binding. Over 100 human cases of HVDRR have been reported.2 In children, HVDRR is characterized by hypocalcemia, secondary hyperparathyroidism, and severe rickets.5,6 The disease is almost always because of mutations in the VDR, with many different mutations identified, including mutations in the LBD and in the DNA-binding domain.5,6 The VDR mutations result in end-organ resistance to 1,25(OH)2D3.2 Resistance leads to defective intestinal calcium absorption, hypocalcemia, and secondary hyperparathyroidism, which in turn causes failure of bone mineral deposition, especially failure of calcium deposition.5,7 As a result, metaphyseal cartilage accumulates, but cannot be properly mineralized and rickets results.7 A syndrome that is similar to HVDRR has been described in cats,7–9 although a genetic mutation in the VDR was not established in any of these animals. These case reports suggest that cats fare relatively well once they reach the end of their growth period, with the exception of one cat, which died suddenly at 13 months of age, and another that was euthanized at 9 years of age for hip pain from osteoarthritis.7–9 Inadequate dietary intake of vitamin D resulting in nutritional secondary hyperparathyroidism has been reported previously in dogs,10 but could be readily ruled out in this case because the dog failed to respond to high concentrations of calcitriol. VDDR-I has been reported in a Saint Bernard dog and a domestic short hair cat.11,12 In the dog reported here, clinical signs of early onset rickets, laboratory findings of hypocalcemia, secondary hyperparathyroidism, and markedly increased serum 1,25(OH)2D3 levels, and radiographic findings of abnormally wide growth plates are all consistent with those reported in people with HVDRR. The dog's poor response to therapy appears to be closely aligned with the human condition in how challenging it is to maintain normal calcium levels and adequate bone mineralization without functional VDRs. Most mutations in people result in a defective VDR that can no longer respond to even high doses of 1,25(OH)2D3. Some children can be treated by high doses of 1,25(OH)2D3 that overcome the defect in binding affinity for 1,25(OH)2D3, however, treatment usually requires high doses of IV calcium administered daily over many months to eventually correct the hypocalcemia, suppress the secondary hyperparathyroidism, and heal the rachitic bone abnormalities.13,14 IV calcium infusions bypass the intestinal calcium absorption defect caused by the disease. After the total body calcium deficit is restored to normal, some children can be transitioned to high doses of oral calcium to maintain their improved metabolic state and prevent rickets from recurring. It is possible that aggressive initial management of this dog with oral calcium therapy and injectable calcium supplementation may have improved the outcome in this case. Based on the VDR LBD mutation in this dog, she could not have responded to even high concentrations of calcitriol. Evidence that the calcium treatment regime used in this case had some efficacy was shown by the increase in bone mineralization seen in her appendicular skeletal radiographs from 5 to 8.5 months of age (Fig 2). Alternatively, an even more aggressive initial approach with vascular access port placement15 and daily IV calcium supplementation (with concurrent ECG monitoring) could have been undertaken as it is in human patients.16 This approach would be technically challenging, would likely be associated with port infections, and would require significant owner commitment. It is noteworthy that the alopecia observed in this dog is also present in many children with HVDDR.5,6 It is proposed that the product of the hairless gene (HR) acts as a corepressor of the VDR, and that the HR and the unliganded VDR are involved in regulation of the hair cycle.17–22 The details of the mechanism of alopecia in HVDDR remain unknown. Large cystic follicles filled with keratin were seen in this dog on postmortem evaluation along with thin epithelium, and reduced number and size of adnexal structures such as sebaceous and apocrine glands. The dog's secondary hair follicles were smaller than normal. In people with HVDRR, the hair follicles may appear normal on microscopic examination, but hair shafts are absent.5 It is interesting that the alopecia had resolved at the time of the dog's death, yet obvious dermal changes were still microscopically present. In contrast to this dog, alopecia persists in human patients with HVDRR that undergo successful therapy or that show spontaneous improvement in calcium homeostasis.5 In humans, all cases VDR mutations causing premature termination codons have life-long alopecia, even if medical treatment improves the hypocalcemia and rickets. We have no explanation for the improvement of the alopecia in this dog. Dogs may potentially have a compensatory mechanism in the hair cycle lacked by humans that can replace the absent VDR function. In humans, patients with alopecia generally have more severe resistance to 1,25(OH)2D3 than those without alopecia, which would correlate with the severity of this dog's disease.23 HVDRR is an autosomal recessive genetic disorder in humans.5 Unfortunately, the breeder of this dog was not interested in progenitor testing to help establish the pattern of heredity in this family. aAntech Diagnostics, Lake Success, NY bRoxane Laboratories Inc, Columbus, OH cCalcitriol, 100 ng/mL Fixed Oil Solution, compounded, Health Park Pharmacy, Raleigh, NC dIdexx VetTest 8008 Chemistry Analyzer, Idexx Laboratories, Westbrook, ME eDiagnostic Center for Population and Animal Health, Michigan State University, Lansing, MI fCalcium glycerophosphate and calcium lactate, Calphosan, Glenwood, Englewood, NJ gHeartland Assays, Ames, IA hCalcium gluconate 10%, AAP Pharmaceuticals LLC, Schaumburg, IL iMidazolam, Bedford Laboratories, Bedford, OH jFentanyl, Hospira, Lake Forest, IL kPropofol, Teva Pharmaceuticals, North Wales, PA lHyclone, Logan, UT mQIAamp DNA blood mini kit, Qiagen, Valencia, CA nSuperscript first strand CDNA synthesis kit, Invitrogen, Carlsbad, CA oTAQ DNA polymerase, New England Biolabs, Ipswich, MA pQIAquick PCR purification kit This work was supported by NIH grant DK42482 (to DF) and NCSU CVM teaching initiative funds (to DNL).}, number={6}, journal={JOURNAL OF VETERINARY INTERNAL MEDICINE}, author={LeVine, D. N. and Zhou, Y. and Ghiloni, R. J. and Fields, E. L. and Birkenheuer, A. J. and Gookin, J. L. and Roberston, I. D. and Malloy, P. J. and Feldman, D.}, year={2009}, pages={1278–1283} }
 @article{duncan_marr_birkenheuer_maggi_williams_correa_breitschwerdt_2008, title={Bartonella DNA in the Blood and Lymph Nodes of Golden Retrievers with Lymphoma and in Healthy Controls}, volume={22}, ISSN={0891-6640 1939-1676}, url={http://dx.doi.org/10.1111/j.1939-1676.2007.0018.x}, DOI={10.1111/j.1939-1676.2007.0018.x}, abstractNote={Background:Although lymphoma is the most common neoplastic process reported in dogs, its precise etiology is unknown. Golden Retrievers are more likely to develop lymphoma, suggesting a breed predisposition; however, other factors, including environment, immunity, and infection, are likely contributors to oncogenesis.}, number={1}, journal={Journal of Veterinary Internal Medicine}, publisher={Wiley}, author={Duncan, A.W. and Marr, H.S. and Birkenheuer, A.J. and Maggi, R.G. and Williams, L.E. and Correa, M.T. and Breitschwerdt, E.B.}, year={2008}, month={Jan}, pages={89–95} }
 @article{birkenheuer_marr_warren_acton_mucker_humphreys_tucker_2008, title={Cytauxzoon felis infections are present in bobcats (Lynx rufus) in a region where cytauxzoonosis is not recognized in domestic cats}, volume={153}, ISSN={["1873-2550"]}, DOI={10.1016/j.vetpar.2008.01.020}, abstractNote={This study was performed to determine the prevalence of Cytauxzoon felis (C. felis) infections in bobcats (Lynx rufus) from a region where C. felis is recognized in domestic cats, North Carolina (NC), and a region where C. felis is not recognized in domestic cats, Pennsylvania (PA). Samples from NC (n=32) were obtained post-mortem via cardiac puncture from legally trapped bobcats. Samples from PA (n=70) were collected post-mortem onto Nobuto blood collecting strips by the PA Game Commission. Each sample was tested using a C. felis specific PCR assay as well as a PCR assay targeting host DNA to rule out the presence of PCR inhibitors. Three samples were excluded due to the presence of PCR inhibitors. Thirty-three percent (10/30) of the samples from NC and 7% (5/69) of the samples from PA tested positive for the presence of C. felis. The proportion of C. felis positive bobcats from NC was significantly different than that from PA (P<0.005). Despite the lower prevalence of C. felis infections in bobcats from PA this finding is unique and indicates the potential for C. felis infections in domestic cats in the northeastern USA if the appropriate tick vectors are present. Veterinary practitioners in PA should be on alert for cytauxzoonosis in domestic cats. Further studies about the epidemiology and transmission of C. felis infections among both domestic cats and bobcats are needed.}, number={1-2}, journal={VETERINARY PARASITOLOGY}, author={Birkenheuer, Adam J. and Marr, Henry S. and Warren, Camille and Acton, Anne E. and Mucker, Eric M. and Humphreys, Jan G. and Tucker, Melissa D.}, year={2008}, month={May}, pages={126–130} }
 @article{stauffer_birkenheuer_levy_marr_gookin_2008, title={Evaluation of four DNA extraction methods for the detection of Tritrichomonas foetus in feline stool specimens by polymerase chain reaction}, volume={20}, ISSN={["1943-4936"]}, DOI={10.1177/104063870802000518}, abstractNote={ Feces are increasingly valued as practical samples for molecular diagnosis of infectious disease. However, extraction of polymerase chain reaction (PCR) quality DNA from fecal samples can be challenging because of coextraction of PCR inhibitors. Because the type and quantity of PCR inhibitors is influenced by diet, endogenous flora, and concurrent disease, it is unlikely that extraction method performance with human feces can be directly extrapolated to that of domestic cats. In the present study, 4 commercially available DNA extraction methods were examined for their influence on the sensitivity of PCR for the detection of Tritrichomonas foetus in feline stool. DNA was extracted from serially diluted feline-origin T. foetus trophozoites in the absence or presence of feline feces. The ZR Fecal DNA kit was identified as affording the greatest analytical sensitivity and reproducibility and was able to detect ≥10 T. foetus organisms per 100 mg feces in 100% of PCR reactions. Further, the identified extraction method could be completed in the shortest time of all kits tested. }, number={5}, journal={JOURNAL OF VETERINARY DIAGNOSTIC INVESTIGATION}, author={Stauffer, Stephen H. and Birkenheuer, Adam J. and Levy, Michael G. and Marr, Henry and Gookin, Jody L.}, year={2008}, month={Sep}, pages={639–641} }
 @article{hawkins_johnson_guptill_marr_breitschwerdt_birkenheuer_2008, title={Failure to identify an association between serologic or molecular evidence ofBartonellainfection and idiopathic rhinitis in dogs}, volume={233}, ISSN={0003-1488}, url={http://dx.doi.org/10.2460/javma.233.4.597}, DOI={10.2460/javma.233.4.597}, abstractNote={Abstract}, number={4}, journal={Journal of the American Veterinary Medical Association}, publisher={American Veterinary Medical Association (AVMA)}, author={Hawkins, Eleanor C. and Johnson, Lynelle R. and Guptill, Lynn and Marr, Henry S. and Breitschwerdt, Edward B. and Birkenheuer, Adam J.}, year={2008}, month={Aug}, pages={597–599} }
 @article{lehtinen_birkenheuer_droleskey_holman_2008, title={In vitro cultivation of a newly recognized Babesia sp in dogs in North Carolina}, volume={151}, ISSN={["0304-4017"]}, DOI={10.1016/j.vetpar.2007.10.022}, abstractNote={A novel large Babesia sp. from an infected dog was cultivated in vitro by microaerophilous stationary phase culture methodology. A primary culture initiated in enriched RPMI-1640 medium supplemented with 40% canine serum and incubated in a 2% oxygen environment supported parasite growth in vitro. Subsequent subcultures into enriched HL-1 medium with 20% fetal bovine serum also supported parasite propagation. Cultures were successfully introduced to 5% carbon dioxide in air atmosphere at passage 4. To date, the parasites have been continuously cultured through 35 passages, although the parasitemias are low, ranging from 0.2 to 0.3%. Parasites cultured in RPMI with canine serum were cryopreserved and successfully recovered from liquid nitrogen storage. The small subunit ribosomal rRNA gene sequence was identical in blood-derived and culture-derived parasites, differing in a single base position from the previously reported sequence for this Babesia sp. The ultrastructure of the parasite was consistent with that of other large Babesia spp., except that the spherical body contained numerous round particles unlike the inclusions previously described in Babesia spp.}, number={2-4}, journal={VETERINARY PARASITOLOGY}, author={Lehtinen, Lauren E. and Birkenheuer, Adam J. and Droleskey, Robert E. and Holman, Patricia J.}, year={2008}, month={Feb}, pages={150–157} }
 @article{birkenheuer_marr_hladio_acton_2008, title={Molecular evidence of prevalent dual piroplasma infections in North American raccoons (Procyon lotor)}, volume={135}, ISSN={["1469-8161"]}, DOI={10.1017/S0031182007003538}, abstractNote={SUMMARY}, number={1}, journal={PARASITOLOGY}, author={Birkenheuer, A. J. and Marr, H. S. and Hladio, N. and Acton, A. E.}, year={2008}, month={Jan}, pages={33–37} }
 @article{small_atkins_gordon_birkenheuer_booth-sayer_keene_fujii_miller_2008, title={Use of a nitinol gooseneck snare catheter for removal of adult Dirofilaria immitis in two cats}, volume={233}, ISSN={["0003-1488"]}, DOI={10.2460/javma.233.9.1441}, abstractNote={Abstract}, number={9}, journal={JAVMA-JOURNAL OF THE AMERICAN VETERINARY MEDICAL ASSOCIATION}, author={Small, Merrilee T. and Atkins, Clarke E. and Gordon, Sonya G. and Birkenheuer, Adam J. and Booth-Sayer, Margaret A. and Keene, Bruce W. and Fujii, Yoko and Miller, Matthew W.}, year={2008}, month={Nov}, pages={1441–1445} }
 @article{puskar_lemons_papich_vaden_birkenheuer_2007, title={Antibiotic-resistant Corynebacterium jeikeium urinary tract infection in a cat}, volume={43}, ISSN={["0587-2871"]}, DOI={10.5326/0430061}, abstractNote={A 10-year-old, castrated male, domestic longhaired cat with a history of urinary tract disease and perineal urethrostomy was presented for evaluation of persistent urinary tract inflammation. Prior to referral, diphtheroid organisms had been cultured from a urine sample obtained by cystocentesis, and they were interpreted as sample contamination. Subsequent urine culture and gene sequencing identified Corynebacterium jeikeium, which was resistant to antibiotics and appeared to be the cause of the urinary tract infection.}, number={1}, journal={JOURNAL OF THE AMERICAN ANIMAL HOSPITAL ASSOCIATION}, author={Puskar, Michelle and Lemons, Carol and Papich, Mark G. and Vaden, Shelley L. and Birkenheuer, Adam}, year={2007}, pages={61–64} }
 @article{valentine_harms_cadenas_birkenheuer_marr_braun-mcneill_maggi_breitschwerdt_2007, title={Bartonella DNA in Loggerhead Sea Turtles}, volume={13}, ISSN={1080-6040 1080-6059}, url={http://dx.doi.org/10.3201/eid1306.061551}, DOI={10.3201/eid1306.061551}, abstractNote={To the Editor: Bartonella are fastidious, aerobic, gram-negative, facultative, intracellular bacteria that infect erythrocytes, erythroblasts, endothelial cells, monocytes, and dendritic cells, and are transmitted by arthropod vectors or by animal scratches or bites (1–6). Currently, 20 species or subspecies of Bartonella have been characterized, of which 8 are known zoonotic pathogens (7). B. henselae has been recently identified from canine blood (8) and from harbor porpoises (9). Pathogenic bacteria are an important threat in terrestrial and marine environments, and in the case of B. henselae, reservoir hosts may be more diverse than currently recognized.}, number={6}, journal={Emerging Infectious Diseases}, publisher={Centers for Disease Control and Prevention (CDC)}, author={Valentine, K. Hope and Harms, Craig A. and Cadenas, Maria B. and Birkenheuer, Adam J. and Marr, Henry S. and Braun-McNeill, Joanne and Maggi, Ricardo G. and Breitschwerdt, Edward B.}, year={2007}, month={Jun}, pages={949–950} }
 @article{hawkins_birkenheuer_marr_rogala_large_adler_2007, title={Quantification of mucin gene expression in tracheobronchial epithelium of healthy dogs and dogs with chronic bronchitis}, volume={68}, ISSN={["1943-5681"]}, DOI={10.2460/ajvr.68.4.435}, abstractNote={Abstract}, number={4}, journal={AMERICAN JOURNAL OF VETERINARY RESEARCH}, author={Hawkins, Eleanor C. and Birkenheuer, Adam J. and Marr, Henry S. and Rogala, Allison R. and Large, Edward E. and Adler, Kenneth B.}, year={2007}, month={Apr}, pages={435–440} }
 @article{haber_tucker_marr_levy_burgess_lappin_birkenheuer_2007, title={The detection of Cytauxzoon felis in apparently healthy free-roaming cats in the USA}, volume={146}, ISSN={["1873-2550"]}, DOI={10.1016/j.vetpar.2007.02.029}, abstractNote={Cytauxzoon felis typically causes fatal disease in domestic cats. Survival after infection and persistent parasitemia without clinical illness has been documented in a few cases. To our knowledge there are no prevalence studies of C. felis in domestic cats. The purpose of this study was to estimate the prevalence of C. felis infected cats that were presented to trap-neuter-return programs in Florida, North Carolina and Tennessee. Cats that were presented to trap-neuter-return programs were tested using a C. felis-specific PCR assay. A total of 961 domestic cats were tested (494 from Florida; 392 from North Carolina; 75 from Tennessee). Prevalence of C. felis infection in this population was 0.3%. Two cats from Florida and one cat from Tennessee tested positive for the presence of C. felis DNA. These amplicons were sequenced and confirmed to be C. felis. The cat from Tennessee was alive without evidence of illness 2 months post-surgery. The other two cats were alive 24 h post-surgery, but were then lost to follow-up. This is the first report documenting C. felis infections in free-roaming cats. Despite the low prevalence rate, the presence of apparently healthy infected free-roaming cats suggests that they may have the capacity to serve as an additional reservoir host for C. felis. Further investigations should evaluate the potential vector competence of domestic cats as well as the role of chronically infected cats in areas in which cytauxzoonosis appears hyperendemic.}, number={3-4}, journal={VETERINARY PARASITOLOGY}, author={Haber, Marion D. and Tucker, Melissa D. and Marr, Henry S. and Levy, Julie K. and Burgess, Jill and Lappin, Michael R. and Birkenheuer, Adam J.}, year={2007}, month={May}, pages={316–320} }
 @article{birkenheuer_harms_neel_marr_tucker_acton_tuttle_stoskopf_2007, title={The identification of a genetically unique piroplasma in North American river otters (Lontra canadensis)}, volume={134}, ISSN={["1469-8161"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-34249725062&partnerID=MN8TOARS}, DOI={10.1017/S0031182006002095}, abstractNote={SUMMARY}, number={5}, journal={PARASITOLOGY}, author={Birkenheuer, A. J. and Harms, C. A. and Neel, J. and Marr, H. S. and Tucker, M. D. and Acton, A. E. and Tuttle, A. D. and Stoskopf, M. K.}, year={2007}, month={May}, pages={631–635} }
 @article{lennon_boyle_hutchins_friedenthal_correa_bissett_moses_papich_birkenheuer_2007, title={Use of basal serum or plasma cortisol concentrations to rule out a diagnosis of hypoadrenocorticism in dogs: 123 cases (2000-2005)}, volume={231}, ISSN={["0003-1488"]}, DOI={10.2460/javma.231.3.413}, abstractNote={Abstract}, number={3}, journal={JAVMA-JOURNAL OF THE AMERICAN VETERINARY MEDICAL ASSOCIATION}, author={Lennon, Elizabeth M. and Boyle, Tonya E. and Hutchins, Rae Grace and Friedenthal, Arit and Correa, Maria T. and Bissett, Sally A. and Moses, Lorra S. and Papich, Mark G. and Birkenheuer, Adam J.}, year={2007}, month={Aug}, pages={413–416} }
 @article{birkenheuer_le_valenzisi_tucker_levy_breitschwerdt_2006, title={Cytauxzoon felisinfection in cats in the mid-Atlantic states: 34 cases (1998–2004)}, volume={228}, ISSN={0003-1488}, url={http://dx.doi.org/10.2460/javma.228.4.568}, DOI={10.2460/javma.228.4.568}, abstractNote={Abstract}, number={4}, journal={Journal of the American Veterinary Medical Association}, publisher={American Veterinary Medical Association (AVMA)}, author={Birkenheuer, Adam J. and Le, Jaime A. and Valenzisi, Amy M. and Tucker, Melissa D. and Levy, Michael G. and Breitschwerdt, Edward B.}, year={2006}, month={Feb}, pages={568–571} }
 @article{birkenheuer_marr_alleman_levy_breitschwerdt_2006, title={Development and evaluation of a PCR assay for the detection of Cytauxzoon felis DNA in feline blood samples}, volume={137}, ISSN={["1873-2550"]}, DOI={10.1016/j.vetpar.2005.12.007}, abstractNote={Cytauxzoonosis is an emerging tick borne infectious disease of domestic cats in the United States, caused by the organism Cytauxzoon felis (C. felis). In naturally infected domestic cats the disease is almost always fatal. Currently there are no commercially available molecular or serologic tests to facilitate the antemortem diagnosis of C. felis infection. Clinical and pathological diagnosis of cytauxzoonosis is based on microscopic identification of parasites in tissues or on blood smears. We have developed and evaluated the sensitivity and specificity of a polymerase chain reaction (PCR) based assay for the diagnosis of C. felis infections in feline blood samples. The assay is sensitive enough to detect one copy of a cloned fragment of the C. felis 18S rRNA gene. This PCR assay can be used for the rapid clinical diagnosis of cytauxzoonosis and for epidemiological studies that will better define the geographic distribution of C. felis infection in cats.}, number={1-2}, journal={VETERINARY PARASITOLOGY}, author={Birkenheuer, AJ and Marr, H and Alleman, AR and Levy, MG and Breitschwerdt, EB}, year={2006}, month={Apr}, pages={144–149} }
 @article{gookin_copple_papich_poore_stauffer_birkenheuer_twedt_levy_2006, title={Efficacy of ronidazole for treatment of feline Tritrichomonas foetus infection}, volume={20}, ISSN={["1939-1676"]}, DOI={10.1892/0891-6640(2006)20[536:EORFTO]2.0.CO;2}, abstractNote={Objectives: To determine the efficacy of ronidazole (RDZ), tinidazole (TDZ), and metronidazole (MDZ) against Tritrichomonas foetus in vitro and of RDZ for treatment of feline naturally occurring or experimentally induced T foetus infection. Animals: A cat naturally infected with T foetus infection and diarrhea. Ten specific-pathogen-free (SPF) kittens. Procedure: RDZ, TDZ, and MDZ were tested for activity against 3 different feline isolates of T foetus in vitro. RDZ then was administered to a naturally infected cat at 10 mg/kg PO q24h for 10 days. SPF kittens were infected orogastrically with feline T foetus and treated with either placebo or RDZ (10 mg/kg PO q12h for 14 days). Cats with relapsing infection or those receiving placebo were treated subsequently with RDZ (either 30 or 50 mg/kg PO q12h for 14 days). Feces were examined for T foetus by direct microscopy, culture, and polymerase chain reaction (PCR) testing weekly. Results: Both RDZ and TDZ killed T foetus at concentrations .0.1 mg/mL in vitro. In the naturally infected cat, RDZ abolished diarrhea and T foetus infection for 85 days after treatment, at which time infection and diarrhea relapsed. Retreatment with RDZ eradicated diarrhea and T foetus infection for over 407 days. In experimentally induced infection, RDZ at 10 mg/kg caused initial improvement, but infection relapsed in all 5 cats 2 to 20 weeks after treatment. At 30 or 50 mg/kg, 10/10 cats were negative for T foetus infection for follow-up durations of 21 to 30 weeks after treatment. Conclusions and Clinical Relevance: Oral administration of RDZ at 30 to 50 mg/kg q12h for 14 days resolved diarrhea and eradicated infection (on the basis of polymerase chain reaction [PCR] testing) in 1 naturally infected cat and 10 experimentally inoculated cats receiving a different isolate of T foetus.}, number={3}, journal={JOURNAL OF VETERINARY INTERNAL MEDICINE}, author={Gookin, Jody L. and Copple, Christina N. and Papich, Mark G. and Poore, Matthew F. and Stauffer, Stephen H. and Birkenheuer, Adam J. and Twedt, David C. and Levy, Michael G.}, year={2006}, pages={536–543} }
 @article{birkenheuer_whittington_neel_large_barger_levy_breitschwerdt_2006, title={Molecular characterization of a Babesia species identified in a North American raccoon}, volume={42}, ISSN={["1943-3700"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-33748280658&partnerID=MN8TOARS}, DOI={10.7589/0090-3558-42.2.375}, abstractNote={Piroplasmosis was first described in raccoons (Procyon lotor) in 1926, and the official description of a small piroplasm as Babesia lotori was done in 1981. Babesia microti-like gene sequences have been characterized in raccoons in both North American and Japan. It is well documented that the microscopic appearance of piroplasms does not always accurately predict the genotype and phylogenetic classification. Discrepancies using phenotype to predict genotype have been reported most frequently when evaluating small piroplasms. We amplified and sequenced the full-length 18S rRNA gene from a small piroplasm identified in a raccoon and used this sequence for phylogenetic analyses. Based on these analyses, the organism was placed in the Babesia sensu stricto clade, confirming that it is a true Babesia sp. This documents that at least two Babesia spp. can infect raccoons. The data generated in this study can be used to design molecular diagnostic tests for detection of this Babesia sp., which will be useful for epidemiologic and comparative phylogenetic studies. As piroplasmosis has been documented with increased frequency in humans in recent years, the results of this study will aid in the recognition of zoonotic babesiosis.}, number={2}, journal={JOURNAL OF WILDLIFE DISEASES}, author={Birkenheuer, Adam J. and Whittington, Julia and Neel, Jennifer and Large, Edward and Barger, Anne and Levy, Michael G. and Breitschwerdt, Edward B.}, year={2006}, month={Apr}, pages={375–380} }
 @article{wardrop_reine_birkenheuer_hale_hohenhaus_crawford_lappin_2005, title={Canine and feline blood donor screening for infectious disease}, volume={19}, ISSN={["1939-1676"]}, DOI={10.1892/0891-6640(2005)19<135:CAFBDS>2.0.CO;2}, abstractNote={Journal of Veterinary Internal MedicineVolume 19, Issue 1 p. 135-142 Open Access Canine and Feline Blood Donor Screening for Infectious Disease K. Jane Wardrop, Corresponding Author K. Jane Wardrop College of Veterinary Medicine, Washington State University, Pullman, WA (Wardrop) DVM, MS, DACVP, College of Veterinary Medicine, Washington State University, Pullman, WA; e-mail: [email protected].Search for more papers by this authorNyssa Reine, Nyssa Reine Department of Medicine, Animal Medical Center, New York, NY (Reine, Hohenhaus)Search for more papers by this authorAdam Birkenheuer, Adam Birkenheuer College of Veterinary Medicine, North Carolina State University, Raleigh, NC (Birkenheuer)Search for more papers by this authorAnne Hale, Anne Hale Midwest Animal Blood Services Inc, Stockbridge, MI (Hale)Search for more papers by this authorAnn Hohenhaus, Ann Hohenhaus Department of Medicine, Animal Medical Center, New York, NY (Reine, Hohenhaus)Search for more papers by this authorCynda Crawford, Cynda Crawford the Department of Small Animal Clinical Sciences, College of Veterinary Medicine, University of Florida, Gainesville, FL (Crawford)Search for more papers by this authorMichael R. Lappin, Michael R. Lappin the Department of Clinical Sciences, College of Veterinary Medicine and Biomedical Sciences, Colorado State University, Fort Collins, CO (Lappin).Search for more papers by this author K. Jane Wardrop, Corresponding Author K. Jane Wardrop College of Veterinary Medicine, Washington State University, Pullman, WA (Wardrop) DVM, MS, DACVP, College of Veterinary Medicine, Washington State University, Pullman, WA; e-mail: [email protected].Search for more papers by this authorNyssa Reine, Nyssa Reine Department of Medicine, Animal Medical Center, New York, NY (Reine, Hohenhaus)Search for more papers by this authorAdam Birkenheuer, Adam Birkenheuer College of Veterinary Medicine, North Carolina State University, Raleigh, NC (Birkenheuer)Search for more papers by this authorAnne Hale, Anne Hale Midwest Animal Blood Services Inc, Stockbridge, MI (Hale)Search for more papers by this authorAnn Hohenhaus, Ann Hohenhaus Department of Medicine, Animal Medical Center, New York, NY (Reine, Hohenhaus)Search for more papers by this authorCynda Crawford, Cynda Crawford the Department of Small Animal Clinical Sciences, College of Veterinary Medicine, University of Florida, Gainesville, FL (Crawford)Search for more papers by this authorMichael R. Lappin, Michael R. Lappin the Department of Clinical Sciences, College of Veterinary Medicine and Biomedical Sciences, Colorado State University, Fort Collins, CO (Lappin).Search for more papers by this author First published: 28 June 2008 https://doi.org/10.1111/j.1939-1676.2005.tb02672.xCitations: 80AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onEmailFacebookTwitterLinkedInRedditWechat Abstract Consensus Statements of the American College of Veterinary Internal Medicine (ACVIM) provide veterinarians with guidelines regarding the pathophysiology, diagnosis, or treatment of animal diseases. The foundation of the Consensus Statement is evidence-based medicine, but if such evidence is conflicting or lacking, the panel provides interpretive recommendations on the basis of their collective expertise. The Consensus Statement is intended to be a guide for veterinarians, but it is not a statement of standard of care or a substitute for clinical judgment. Topics of statements and panel members to draft the statements are selected by the Board of Regents with input from the general membership. A draft prepared and input from Diplomates is solicited at the Forum and via the ACVIM Web site and incorporated in a final version. 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Infectious Disease of the Dog and Cat. Philadelphia , PA : WB Saunders; 1998: 248–257. 16 Wanke MM. Canine brucellosis. Anim Reprod Sci 2004; 82–83: 195–207. 17 Barr SC, Gossett KA, Klei TR. Clinical, clinicopathologic, and parasitologic observations of trypanosomiasis in dogs infected with North American Trypanosoma cruzi isolates. Am J Vet Res 1991; 52: 954–960. 18 Bradley KK, Bergman DK, Woods JP. Prevalence of American trypanosomiasis (Chagas disease) among dogs in Oklahoma. J Am Vet Med Assoc 2000; 217: 1853–1857. 19 Breitschwerdt EB, Blann KR, Stebbins ME, et al. Clinicopathological abnormalities and treatment response in 24 dogs seroreactive to Bartonella vinsonii (berkhoffii) antigens. J Am Anim Hosp Assoc 2004; 40: 92–101. 20 Honadel TE, Chomel BB, Yamamoto K., et al. Seroepidemiology of Bartonella vinsonii subsp berkhoffii exposure among healthy dogs. J Am Vet Med Assoc 2001; 219: 480–484. 21 Pappalardo BL, Brown TT, Tompkins M., et al. Immunopathology of Bartonella vinsonii (berkhoffii) in experimentally infected dogs. Vet Immunol Immunopathol 2001; 83: 125–147. 22 Brinson J., Messick J. Use of a polymerase chain reaction assay for detection of Haemobartonella canis in a dog. J Am Vet Med Assoc 2001; 218: 1943–1945. 23 Fritz CL, Kjemtrup AM. Lyme borreliosis. J Am Vet Med Assoc 2003; 223: 1261–1270. 24 Magnarelli LA, Anderson JF, Schreier AB. Persistence of antibodies to Borrelia burgdorferi in dogs in New York and Connecticut. J Am Vet Med Assoc 1990; 196: 1064–1068. 25 Wormser GP, Bittker S., Cooper D., et al. Yield of large-volume blood cultures in patients with early Lyme disease. J Infect Dis 2001; 184: 1070–1072. 26 Gerber MA, Shapiro ED, Krause PJ, et al. The risk of acquiring Lyme disease or babesiosis from a blood transfusion. J Infect Dis 1994; 170: 231–234. 27 Straubinger RK. PCR-based quantification of Borrelia burgdorferi organisms in canine tissues over a 500-day postinfection period. J Clin Microbiol 2000; 38: 2191–2199. 28 Messick JB. Hemotrophic mycoplasmas (hemoplasmas): A review and new insights into pathogenic potential. Vet Clin Pathol 2004; 33: 2–13. 29 Foley JE, Harrus S., Poland A., et al. Molecular, clinical, and pathologic comparison of two distinct strains of Haemobartonella felis in domestic cats. Am J Vet Res 1998; 59: 1581–1588. 30 Jensen WA, Lappin MR, Kamkar S., et al. Use of a polymerase chain reaction assay to detect and differentiate two strains of Haemobartonella felis in naturally infected cats. Am J Vet Res 2001; 62: 604–608. 31 Westfall DS, Jensen WA, Reagan W., et al. Inoculation of two genotypes of Haemobartonella felis (California and Ohio variants) to induce infection in cats and response to treatment with azithromycin. Am J Vet Res 2001; 62: 687–691. 32 Hackett TB, Jensen WA, Lehman T., et al. Prevalence of Mycoplasma spp., Bartonella spp., Ehrlichia spp., and Anaplasma phagocytophilum DNA in blood of cats used as blood donors in the United States. J Am Vet Med Assoc In review, 2004. 33 Berent LM, Messick JB, Cooper SK. Detection of Haemobartonella felis in cats with experimentally induced acute and chronic infections, using a polymerase chain reaction assay. Am J Vet Res 1998; 59: 1215–1220. 34 Lappin MR, Black JC. Bartonella spp infection as a possible cause of uveitis in a cat. J Am Vet Med Assoc 1999; 214: 1205–1207. 35 Chomel BB, Wey AC, Kasten RW, et al. Fatal case of endocarditis associated with Bartonella henselae type I infection in a domestic cat. J Clin Microbiol 2003; 41: 5337–5339. 36 Finkelstein JL, Brown TP, O'Reilly KL, et al. Studies on the growth of Bartonella henselae in the cat flea (Siphonaptera: Pulicidae) J Med Entomol 2002; 39: 915–919. 37 Rolain JM, Locatelli C., Chabanne L., et al. Prevalence of Bartonella clarridgeiae and Bartonella henselae in domestic cats from France and detection of the organisms in erythrocytes by immunofluorescence. Clin Diagn Lab Immunol 2004; 11: 423–425. 38 Kordick DL, Breitschwerdt EB. Relapsing bacteremia after blood transmission of Bartonella henselae to cats. Am J Vet Res 1997; 58: 492–497. 39 Windsor JJ. Cat-scratch disease; epidemiology, aetiology and treatment. Br J Biomed Sci 2001; 58: 101–110. 40 Jameson P., Greene C., Regnery R., et al. Prevalence of Bartonella henselae antibodies in pet cats throughout regions of North America. J Infect Dis 1995; 172: 1145–1149. 41 Heller R., Artois M., Xemar V., et al. Prevalence of Bartonella henselae and Bartonella clarridgeiae in stray cats. J Clin Microbiol 1997; 35: 1327–1331. 42 Kordick DL, Papich MG, Breitschwerdt EB. Efficacy of enrofloxacin or doxcycline for treatment of Bartonella henselae or Bartonella clarridgeiae infection in cats. Antimicrob Agents Chemother 1997; 41: 2448–2455. 43 Hardy WD, Hess PW, Essex M., et al. Horizontal transmission of feline leukemia virus in cats. Bibl Haematol 1975; 40: 67–74. 44 Callanan J., Thompson H., Toth S., et al. Clinical and pathological findings in feline immunodeficiency virus experimental infection. Vet Immunol Immunopathol 1992; 35: 3–13. 45 Levy JK, Crawford PC, Slater MR. Effect of vaccination against feline immunodeficiency virus on serological testing. J Am Vet Med Assoc 2004. In press. 46 Crawford PC, Slater MR, Leutenegger CM, Levy JK. PCR for diagnosis of FIV infection. J Am Vet Med Assoc 2004. In press. 47 Wagner JE, Ferris DH, Kier AB, et al. Experimentally induced cytauxzoonosis-like disease in domestic cats. Vet Parasitol 1980; 6: 305–311. 48 Meinkoth J., Kocan AA, Whitworth L., et al. Cats surviving natural infection with Cytauxzoon felis: 18 cases (1997–1998). J Vet Intern Med 2000; 14: 521–525. 49 Breitschwert EB, Abrams-Ogg AC, Lappin MR, et al. Molecular evidence supporting Ehrlichia canis-like infection in cats. J Vet Intern Med 2002; 16: 642–649. 50 Lappin MR, Breitschwerdt EB, Jensen WA, et al. Molecular and serological evidence of Anaplasma phagocytophilum infection of cats in North America. J Am Vet Med Assoc 2004. In press. 51 Bjoersdorff A., Svendenius L., Owens JH, et al. Feline granulocytic ehrlichiosis—A report of a new clinical entity and characterization of the infectious agent. J Sm Anim Pract 1999; 40: 20–24. 52 Dawson JE, Abeygunawardena I., Holland CJ, et al. Susceptibility of cats to infection with Ehrlichia risticii, causative agent of equine monocytic ehrlichiosis. Am J Vet Res 1988; 49: 2096–2100. 53 Burney DP, Spilker M., McReynolds L., et al. Detection of Toxoplasma gondii parasitemia in experimentally inoculated cats. J Paras-itol 1999; 5: 947–951. Citing Literature Volume19, Issue1January 2005Pages 135-142 ReferencesRelatedInformation}, number={1}, journal={JOURNAL OF VETERINARY INTERNAL MEDICINE}, author={Wardrop, KJ and Reine, N and Birkenheuer, A and Hale, A and Hohenhaus, A and Crawford, C and Lappin, MR}, year={2005}, pages={135–142} }
 @article{birkenheuer_correa_levy_breitschwerdt_2005, title={Geographic distribution of babesiosis among dogs in the United States and association with dog bites: 150 cases (2000-2003)}, volume={227}, ISSN={0003-1488}, url={http://dx.doi.org/10.2460/javma.2005.227.942}, DOI={10.2460/javma.2005.227.942}, abstractNote={Abstract}, number={6}, journal={Journal of the American Veterinary Medical Association}, publisher={American Veterinary Medical Association (AVMA)}, author={Birkenheuer, Adam J. and Correa, Maria T. and Levy, Michael G. and Breitschwerdt, Edward B.}, year={2005}, month={Sep}, pages={942–947} }
 @article{haber_birkenheuer_2005, title={Icterus and pancytopenia in a cat}, volume={3}, ISBN={1542-4014}, number={7}, journal={NAVC Clinician's Brief}, author={Haber, M. and Birkenheuer, A.}, year={2005}, pages={21} }
 @article{gookin_birkenheuer_st john_spector_levy_2005, title={Molecular characterization of trichomonads from feces of dogs with diarrhea}, volume={91}, ISSN={["1937-2345"]}, DOI={10.1645/ge-474r.1}, abstractNote={Trichomonads are occasionally observed in the feces of dogs with diarrhea. On the basis of superficial morphological appearance, these infections have been attributed to opportunistic overgrowth of the commensal, Pentatrichomonas hominis. However, molecular characterization of canine trichomonads has never been reported. This study was performed to determine, by means of rRNA gene sequence analysis, the identity of trichomonads observed in feces from dogs with diarrhea. Total DNA was isolated from fecal samples obtained from a 3-mo-old mixed breed dog and litter of German Shepherd puppies having profuse liquid diarrhea containing numerous trichomonads. Total DNA was subject to PCR amplification of partial 18S rRNA gene or 5.8S, ITS1, ITS2, and partial 18S and 28S rRNA genes using species-specific and universal primers, respectively. Products of 642 and 1864 base-pair length were amplified and cloned. On the basis of rRNA gene sequence, the trichomonads observed in the single dog and the litter of puppies shared 100% identity with Tritrichomonas foetus and P. hominis, respectively. The present study is the first to establish the molecular identity of trichomonads infecting dogs with diarrhea. These studies validate the longstanding assumption that canine trichomoniasis may be attributed to P. hominis. Importantly, these studies additionally recognize that canine trichomoniasis may also be caused by infection with T. foetus.}, number={4}, journal={JOURNAL OF PARASITOLOGY}, author={Gookin, JL and Birkenheuer, AJ and St John, V and Spector, M and Levy, MG}, year={2005}, month={Aug}, pages={939–943} }
 @article{birkenheuer_neel_ruslander_levy_breitschwerdt_2004, title={Detection and molecular characterization of a novel large Babesia species in a dog}, volume={124}, ISSN={0304-4017}, url={http://dx.doi.org/10.1016/j.vetpar.2004.07.008}, DOI={10.1016/j.vetpar.2004.07.008}, abstractNote={Babesia canis has generally been considered the only large Babesia to infect dogs. Here we describe the molecular characterization of a large Babesia species that was detected in the blood and bone marrow of a dog with clinical and hematological abnormalities consistent with babesiosis. Analysis of the 18S rRNA genes revealed a unique sequence that shared 93.9% sequence identity with B. bigemina and 93.5% sequence identity with B. caballi, compared to 91.2-91.6% identity with B. canis canis, B. c. vogeli, and B. c. rossi. Cross-reactive antibodies against B. canis, B. gibsoni (Asian genotype), or B. gibsoni (California genotype) antigens were not detected in acute or convalescent serum samples. The dog was treated with imidocarb diproprionate, which resulted in the resolution of clinical signs, and subsequently Babesia DNA was not detectable by PCR in post-treatment samples. The organism described in this report represents a genetically unique large Babesia sp. and is the eighth genetically distinct piroplasm capable of infecting the domestic dog.}, number={3-4}, journal={Veterinary Parasitology}, publisher={Elsevier BV}, author={Birkenheuer, A.J. and Neel, J. and Ruslander, D. and Levy, M.G. and Breitschwerdt, E.B.}, year={2004}, month={Oct}, pages={151–160} }
 @article{birkenheuer_levy_breitschwerdt_2004, title={Efficacy of Combined Atovaquone and Azithromycin for Therapy of Chronic Babesia gibsoni (Asian Genotype) Infections in Dogs}, DOI={10.1111/j.1939-1676.2004.tb02573.x}, abstractNote={Babesiosis caused byBabesia gibsoni(Asian genotype) is an emerging disease in dogs in the United States. To date, no drugs have been shown to eliminateB gibsoni(Asian genotype) infections from dogs. Twenty‐two dogs that remained persistently infected withB gibsoni(Asian genotype) after either imidocarb diproprionate and or diminazine aceturate therapy were identified and randomly and evenly distributed into 2 groups. One group was treated with atovaquone and azithromycin combination therapy, and the other group received a placebo. Eight of 10 dogs in the treatment group had no detectableB gibsoni(Asian genotype) DNA, as determined by a sensitive and specific polymerase chain reaction (PCR) assay, in any of their posttreatment samples. In contrast,B gibsoni(Asian genotype) DNA was detectable by PCR in the posttreatment samples from 11 of 11 of the placebo‐treated dogs. One dog in the treatment group was excluded from the treatment outcome analysis. This dog had 2 consecutive negative PCR assay results and was euthanized because of ongoing degenerative joint disease prior to completion of the study. No adverse effects of treatment were reported in any dog during the study period. A combination of atovaquone and azithromycin is the 1st described treatment that will either eliminateB gibsoni(Asian genotype) infections or suppress the parasitemia below the limit of detection in the majority of treated dogs.}, journal={Journal of Veterinary Internal Medicine}, author={Birkenheuer, Adam J. and Levy, Michael G. and Breitschwerdt, Edward B.}, year={2004} }
 @article{birkenheuer_levy_breitschwerdt_2004, title={Efficacy of combined atovaquone and azithromycin for therapy of chronic Babesia gibsoni (Asian genotype) infections in dogs}, volume={18}, ISSN={["1939-1676"]}, DOI={10.1892/0891-6640(2004)18<494:EOCAAA>2.0.CO;2}, abstractNote={Babesiosis caused by Babesia gibsoni (Asian genotype) is an emerging disease in dogs in the United States. To date, no drugs have been shown to eliminate B gibsoni (Asian genotype) infections from dogs. Twenty-two dogs that remained persistently infected with B gibsoni (Asian genotype) after either imidocarb diproprionate and or diminazine aceturate therapy were identified and randomly and evenly distributed into 2 groups. One group was treated with atovaquone and azithromycin combination therapy, and the other group received a placebo. Eight of 10 dogs in the treatment group had no detectable B gibsoni (Asian genotype) DNA, as determined by a sensitive and specific polymerase chain reaction (PCR) assay, in any of their posttreatment samples. In contrast, B gibsoni (Asian genotype) DNA was detectable by PCR in the posttreatment samples from 11 of 11 of the placebo-treated dogs. One dog in the treatment group was excluded from the treatment outcome analysis. This dog had 2 consecutive negative PCR assay results and was euthanized because of ongoing degenerative joint disease prior to completion of the study. No adverse effects of treatment were reported in any dog during the study period. A combination of atovaquone and azithromycin is the 1st described treatment that will either eliminate B gibsoni (Asian genotype) infections or suppress the parasitemia below the limit of detection in the majority of treated dogs.}, number={4}, journal={JOURNAL OF VETERINARY INTERNAL MEDICINE}, author={Birkenheuer, AJ and Levy, MG and Breitschwerdt, EB}, year={2004}, pages={494–498} }
 @article{tuttle_birkenheuer_juopperi_levy_breitschwerdt_2003, title={Concurrent bartonellosis and babesiosis in a dog with persistent thrombocytopenia}, volume={223}, ISSN={0003-1488}, url={http://dx.doi.org/10.2460/javma.2003.223.1306}, DOI={10.2460/javma.2003.223.1306}, abstractNote={A 12-year-old castrated male West Highland White Terrier was referred because of recurrent episodes of collapsing. The dog was mildly anemic and severely thrombocytopenic and had high serum alanine aminotransferase activity. Infection with Bartonella vinsonii (berkhoffii) was initially diagnosed on the basis of serologic testing. Despite treatment with a series of antimicrobials and prolonged use of immunosuppressive drugs, thrombocytopenia persisted. After 5 months of treatment, Babesia canis organisms were seen during examination of a direct blood smear. The dog was treated with imidocarb dipropionate for babesiosis, after which thrombocytopenia resolved, and administration of immunosuppressive drugs was discontinued. Retrospective review of blood smears failed to identify organisms; however, polymerase chain reaction (PCR) analysis of multiple stored blood samples obtained during the 5-month period of persistent thrombocytopenia identified DNA of B. canis vogeli. Babesiosis may cause persistent, unexplained thrombocytopenia in dogs that are not anemic. A PCR assay can facilitate a diagnosis of babesiosis when organisms are not evident or when serologic testing fails to detect Babesia-specific antibodies.}, number={9}, journal={Journal of the American Veterinary Medical Association}, publisher={American Veterinary Medical Association (AVMA)}, author={Tuttle, Allison D. and Birkenheuer, Adam J. and Juopperi, Tarja and Levy, Michael G. and Breitschwerdt, Edward B.}, year={2003}, month={Nov}, pages={1306–1310} }
 @article{birkenheuer_levy_breitschwerdt_2003, title={Development and Evaluation of a Seminested PCR for Detection and Differentiation of Babesia gibsoni (Asian Genotype) and B. canis DNA in Canine Blood Samples}, volume={41}, ISSN={0095-1137}, url={http://dx.doi.org/10.1128/jcm.41.9.4172-4177.2003}, DOI={10.1128/JCM.41.9.4172-4177.2003}, abstractNote={ABSTRACT}, number={9}, journal={Journal of Clinical Microbiology}, publisher={American Society for Microbiology}, author={Birkenheuer, A. J. and Levy, M. G. and Breitschwerdt, E. B.}, year={2003}, month={Sep}, pages={4172–4177} }
 @article{birkenheuer_levy_stebbins_poore_breitschwerdt_2003, title={Serosurvey of antiBabesia antibodies in stray dogs and American pit bull terriers and American Staffordshire terriers from North Carolina}, volume={39}, ISSN={["0587-2871"]}, DOI={10.5326/0390551}, abstractNote={Stray dogs (n=359) and kennel dogs (n=149) from North Carolina were tested for evidence of antiBabesia antibodies. AntiBabesia antibodies were detected in 21/359 and 22/149 of the stray and kennel dogs, respectively. A total of 57 dogs from both groups were tested for babesiasis by light microscopy and polymerase chain reaction (PCR). Babesia deoxyribonucleic acid (DNA) was detected in 3/28 of the stray dogs and 14/29 of the kennel dogs. When Babesia DNA was detected by PCR, the species-specific PCR results differed from the Babesia species antibody titer results in 6/17 of the PCR-positive dogs. There was no association between antiBabesia antibodies and the presence of ticks. There are currently Babesia gibsoni epizootics affecting American pit bull terrier kennels.}, number={6}, journal={JOURNAL OF THE AMERICAN ANIMAL HOSPITAL ASSOCIATION}, author={Birkenheuer, AJ and Levy, MG and Stebbins, M and Poore, M and Breitschwerdt, E}, year={2003}, pages={551–557} }
 @article{stegeman_birkenheuer_kruger_breitschwerdt_2003, title={Transfusion-associated Babesia gibsoni infection in a dog}, volume={222}, ISSN={0003-1488}, url={http://dx.doi.org/10.2460/javma.2003.222.959}, DOI={10.2460/javma.2003.222.959}, abstractNote={A 2.5-year-old spayed female German Shepherd Dog was referred for evaluation of progressive anemia, lethargy, and weight loss. Seventeen days earlier, the dog had received a whole blood transfusion to manage hemorrhage after ovariohysterectomy. Mild fever, splenomegaly, and thrombocytopenia were also identified. Von Willebrand disease and Babesia gibsoni infection were diagnosed. Because of the serologic cross-reactivity of B gibsoni and B canis in the immunofluorescent antibody assay for IgG antibodies against these organisms, polymerase chain reaction amplification of parasite DNA was required to identify the infecting Babesia sp. The source of the B gibsoni infection was traced to an apparently healthy American Pit Bull Terrier blood donor. Despite resolution of clinical signs in the dog of this report, a series of antiparasitic treatments failed to eliminate the B gibsoni infection. Screening of potential blood donor dogs for Babesia spp is becoming increasingly important in the United States.}, number={7}, journal={Journal of the American Veterinary Medical Association}, publisher={American Veterinary Medical Association (AVMA)}, author={Stegeman, Julie R. and Birkenheuer, Adam J. and Kruger, John M. and Breitschwerdt, Edward B.}, year={2003}, month={Apr}, pages={959–963} }
 @article{levy_gookin_poore_birkenheuer_dykstra_litaker_2003, title={Tritrichomonas foetus and not Pentatrichomonas hominis is the etiologic agent of feline trichomonal diarrhea}, volume={89}, ISSN={["1937-2345"]}, DOI={10.1645/0022-3395(2003)089[0099:TFANPH]2.0.CO;2}, abstractNote={Recently, several investigators have reported large-bowel diarrhea in cats associated with intestinal trichomonad parasites. These reports have presumptively identified the flagellates as Pentatrichomonas hominis, an organism putatively capable of infecting the intestinal tracts of a number of mammalian hosts, including cats, dogs, and man. The purpose of the present study was to determine the identity of this recently recognized flagellate by means of rRNA gene sequence analysis; restriction enzyme digest mapping; and light, transmission, and scanning electron microscopy (SEM).}, number={1}, journal={JOURNAL OF PARASITOLOGY}, author={Levy, MG and Gookin, JL and Poore, M and Birkenheuer, AJ and Dykstra, MJ and Litaker, RW}, year={2003}, month={Feb}, pages={99–104} }
 @article{birkenheuer_breitschwerdt_alleman_pitulle_2002, title={Differentiation of Haemobartonella canis and Mycoplasma haemofelis on the basis of comparative analysis of gene sequences}, volume={63}, ISSN={0002-9645}, url={http://dx.doi.org/10.2460/ajvr.2002.63.1385}, DOI={10.2460/ajvr.2002.63.1385}, abstractNote={Abstract}, number={10}, journal={American Journal of Veterinary Research}, publisher={American Veterinary Medical Association (AVMA)}, author={Birkenheuer, Adam J. and Breitschwerdt, Edward B. and Alleman, A. Rick and Pitulle, Christian}, year={2002}, month={Oct}, pages={1385–1388} }
 @article{rotstein_taylor_birkenhauer_roelke-parker_homer_2002, title={Retrospective study of proliferative papillary vulvitis in Florida panthers}, volume={38}, ISSN={["0090-3558"]}, DOI={10.7589/0090-3558-38.1.115}, abstractNote={Proliferative, papillary vulvitis was identified in 16 of 34 (47%) free-ranging and captive female Florida panthers (Puma concolor coryi) monitored over a period from 1983–98. Gross lesions were characterized by extensive papilliferous proliferation in the mucosa of the vestibulum vaginae. Within lesions, the mean length and width of vestibular papillae were 1.07 ± 0.39 mm (CV = 36%) and 0.55 ± 0.11 mm (CV = 20%) respectively. Histologically, three to 12 layers of non-cornified stratified squamous epithelium with various degrees of basal cell spongiosis and rete ridge formation covered fibrous papillae. Mixed leukocytic mucosal inflammation also was observed. Infectious organisms were not observed, and immunohistochemical testing for the presence of papillomavirus antigens in specimens from seven panthers was negative. Lesions in nearly all of the panthers were first observed during a six-year period (1986–92), with one each in 1983, 1996 and 1998. There were no significant differences between the number of females having litters, the number of litters between age-matched and interval-matched females, and the interval between litters among lesion positive and lesion negative females over the 15 yr period. The severity of lesions did not appear to differ between parous and nulliparous free-ranging lesion-positive females. The cause of proliferative vulvitis remains unknown. However, the lesion did not appear to have a significant effect on reproduction.}, number={1}, journal={JOURNAL OF WILDLIFE DISEASES}, author={Rotstein, DS and Taylor, SK and Birkenhauer, A and Roelke-Parker, M and Homer, BL}, year={2002}, month={Jan}, pages={115–123} }
 @article{gookin_birkenheuer_breitschwerdt_levy_2002, title={Single-Tube Nested PCR for Detection of Tritrichomonasfoetus in Feline Feces}, volume={40}, ISSN={0095-1137}, url={http://dx.doi.org/10.1128/jcm.40.11.4126-4130.2002}, DOI={10.1128/JCM.40.11.4126-4130.2002}, abstractNote={ABSTRACT}, number={11}, journal={Journal of Clinical Microbiology}, publisher={American Society for Microbiology}, author={Gookin, J. L. and Birkenheuer, A. J. and Breitschwerdt, E. B. and Levy, M. G.}, year={2002}, month={Nov}, pages={4126–4130} }
 @article{gaskin_schantz_jackson_birkenheuer_tomlinson_gramiccia_levy_steurer_kollmar_hegarty_et al._2002, title={Visceral leishmaniasis in a New York foxhound kennel}, volume={16}, ISSN={["0891-6640"]}, DOI={10.1892/0891-6640(2002)016<0034:VLIANY>2.3.CO;2}, abstractNote={Although endemic throughout much of the world, autochthonous visceral leishmaniasis has been reported on only 3 previous occasions in North America. After diagnosis of visceral leishmaniasis in 4 foxhounds from a kennel in Dutchess County, New York (index kennel), serum and ethylenediamine-tetraacetic acid (EDTA)-anticoagulated blood were collected from the remaining 108 American or cross-bred foxhounds in the index kennel and from 30 Beagles and Basset Hounds that were periodically housed in the index kennel. Samples were analyzed for antibodies to or DNA of tickborne disease pathogens and Leishmania spp. Most dogs had antibodies to Rickettsia spp., Ehrlichia spp., Babesia spp., or some combination of these pathogens but not to Bartonella vinsonii (berkhoffi). However, DNA of rickettsial, ehrlichial, or babesial agents was detected in only 9 dogs. Visceral leishmaniasis was diagnosed in 46 of 112 (41%) foxhounds from the index kennel but was not diagnosed in any of the Beagles and Basset Hounds. A positive Leishmania status was defined by 1 or more of the following criteria: a Leishmania antibody titer > or = 1:64, positive Leishmania polymerase chain reaction (PCR), positive Leishmania culture, or identification of Leishmania amastigotes by cytology or histopathology. The species and zymodeme of Leishmania that infected the foxhounds was determined to be Leishmania infantum MON-1 by isoenzyme electrophoresis. Foxhounds that were > 18 months of age or that had traveled to the southeastern United States were more likely to be diagnosed with visceral leishmaniasis. Transmission of Leishmania spp. in kennel outbreaks may involve exposure to an insect vector, direct transmission, or vertical transmission.}, number={1}, journal={JOURNAL OF VETERINARY INTERNAL MEDICINE}, author={Gaskin, AA and Schantz, P and Jackson, J and Birkenheuer, A and Tomlinson, L and Gramiccia, M and Levy, M and Steurer, F and Kollmar, E and Hegarty, BC and et al.}, year={2002}, pages={34–44} }
 @article{gaskin_schantz_jackson_birkenheuer_tomlinson_gramiccia_levy_steurer_kollmar_hegarty_et al._2002, title={Visceral leishmaniasis in a New York foxhound kennel}, DOI={10.1111/j.1939-1676.2002.tb01604.x}, abstractNote={Although endemic throughout much of the world, autochthonous visceral leishmaniasis has been reported on only 3 previous occasions in North America. After diagnosis of visceral leishmaniasis in 4 foxhounds from a kennel in Dutchess County, New York (index kennel), serum and ethylenediamine‐tetraacetic acid (EDTA)‐anticoagulated blood were collected from the remaining 108 American or cross‐bred foxhounds in the index kennel and from 30 Beagles and Basset Hounds that were periodically housed in the index kennel. Samples were analyzed for antibodies to or DNA of tickborne disease pathogens andLeishmaniaspp. Most dogs had antibodies toRickettsiaspp.,Ehrlichiaspp.,Babesiaspp., or some combination of these pathogens but not toBartonella vinsonii(berkhoffi). However, DNA of rickettsial, ehrlichial, or babesial agents was detected in only 9 dogs. Visceral leishmaniasis was diagnosed in 46 of 112 (41%) foxhounds from the index kennel but was not diagnosed in any of the Beagles and Basset Hounds. A positiveLeishmaniastatus was defined by 1 or more of the following criteria: aLeishmaniaantibody titer ≥1:64, positiveLeishmaniapolymerase chain reaction (PCR), positiveLeishmaniaculture, or identification ofLeishmaniaamastigotes by cytology or histopathology. The species and zymodeme ofLeishmaniathat infected the foxhounds was determined to beLeishmania infantumMON‐1 by isoenzyme electrophoresis. Foxhounds that were >18 months of age or that had traveled to the southeastern United States were more likely to be diagnosed with visceral leishmaniasis. Transmission ofLeishmaniaspp. in kennel outbreaks may involve exposure to an insect vector, direct transmission, or vertical transmission.}, journal={Journal of Veterinary Internal Medicine}, author={Gaskin, Amanda A. and Schantz, Peter and Jackson, Joan and Birkenheuer, Adam and Tomlinson, Lindsay and Gramiccia, Marina and Levy, Michael and Steurer, Frank and Kollmar, Eleanor and Hegarty, Barbara C. and et al.}, year={2002} }
 @article{gasser_birkenheuer_breitschwerdt_2001, title={Canine Rocky Mountain spotted fever: A retrospective study of 30 cases}, volume={37}, ISSN={["0587-2871"]}, DOI={10.5326/15473317-37-1-41}, abstractNote={Rocky Mountain spotted fever (RMSF) was diagnosed in 30 dogs examined at North Carolina State University, Veterinary Teaching Hospital between 1984 and 1997. Historical, physical examination, and laboratory abnormalities were reviewed. Diagnostic criteria included a four-fold rise in antibody titer to Rickettsia rickettsii (R. rickettsii) (n=15) or a single R. rickettsii antibody titer of 1:1,024 or greater (n=15; when this initial titer was determined one week or more after the onset of clinical signs). Fifteen (50%) dogs were greater than seven years of age, and 13 (43%) dogs were between two and seven years of age. There was no sex predilection. Only five (17%) dogs had a history of known tick exposure. Presumably due to delayed diagnosis, dogs with antibody titers of 1:1,024 or greater at the time of presentation had a higher incidence of more severe neurological dysfunction (e.g., ataxia, hyperesthesia, vestibular disease, and seizures) and cutaneous lesions (e.g., hyperemia, edema, petechiae, ecchymoses, and necrosis). Laboratory findings included anemia, leukocytosis accompanied by toxic granulation of neutrophils, hypoalbuminemia, and coagulation abnormalities; signs were generally more severe in the 15 dogs with R. rickettsii antibody titers of 1:1,024 or greater at the time of presentation. Twelve (40%) dogs in this study were severely thrombocytopenic (less than 75 x10(3) platelets/microl; reference range, 200 to 450 x 10(3)/microl), without clinical evidence of fulminant disseminated intravascular coagulation. In this study, the survival rate following R. rickettsii infection was 100%.}, number={1}, journal={JOURNAL OF THE AMERICAN ANIMAL HOSPITAL ASSOCIATION}, author={Gasser, AM and Birkenheuer, AJ and Breitschwerdt, EB}, year={2001}, pages={41–48} }
 @article{kjemtrup_kocan_whitworth_meinkoth_birkenheuer_cummings_boudreaux_stockham_irizarry-rovira_conrad_2000, title={There are at least three genetically distinct small piroplasms from dogs}, volume={30}, ISSN={["0020-7519"]}, DOI={10.1016/S0020-7519(00)00120-X}, abstractNote={The 18S nuclear subunit ribosomal RNA (18S rRNA) gene of small piroplasms isolated from dogs from Okinawa (Japan), Oklahoma, North Carolina, Indiana, Missouri, and Alabama, was isolated and sequenced. Phylogenetic analysis of these sequences and comparisons with sequences from other Babesia, Cytauxzoon, and Theileria species revealed that all canine small babesial isolates, with the exception of isolates from California and Spain, were placed in a group containing the Babesia spp. sensu stricto. Within the Babesia spp. sensu stricto, there was support for separating the small canine piroplasms from the large canine piroplasm, Babesia canis. The isolate from California was in a distinct phylogenetic clade, closely related to babesial isolates from wildlife and humans from the Western US. The canine isolate from Spain was closely related to Babesia microti. These results suggest that there are multiple small piroplasm species in dogs. The isolates from the Midwestern and Eastern US and the one from Japan probably represent a single species with wide geographic distribution.}, number={14}, journal={INTERNATIONAL JOURNAL FOR PARASITOLOGY}, author={Kjemtrup, AM and Kocan, AA and Whitworth, L and Meinkoth, J and Birkenheuer, AJ and Cummings, J and Boudreaux, MK and Stockham, SL and Irizarry-Rovira, A and Conrad, PA}, year={2000}, month={Dec}, pages={1501–1505} }
 @article{birkenheuer_levy_savary_gager_breitschwerdt_1999, title={Babesia gibsoni infections in dogs from North Carolina}, volume={35}, ISSN={["0587-2871"]}, DOI={10.5326/15473317-35-2-125}, abstractNote={The recognition of canine babesiosis in North Carolina caused by Babesia gibsoni documents the expansion of the previously reported endemic area of this disease. Clinical signs ranged from severe hemolytic anemia and thrombocytopenia to subclinical infections. No infected dogs had traveled to endemic areas. Antibabesial treatment failed to eradicate the organism from infected dogs.}, number={2}, journal={JOURNAL OF THE AMERICAN ANIMAL HOSPITAL ASSOCIATION}, author={Birkenheuer, AJ and Levy, MG and Savary, KCM and Gager, RB and Breitschwerdt, EB}, year={1999}, pages={125–128} }