@article{jarred_lewbart_stover_thomas_maggi_breitschwerdt_2018, title={Identification of Hemotropic Mycoplasmas in an Eastern Box Turtle (Terrapene carolina carolina) and a Yellow-bellied Slider (Trachemys scripta scripta) from North Carolina, USA}, volume={54}, ISSN={0090-3558}, url={http://dx.doi.org/10.7589/2017-07-153}, DOI={10.7589/2017-07-153}, abstractNote={Abstract:  Mycoplasma spp. are known from several chelonian and other reptilian species. We determined if turtles obtained by the Turtle Rescue Team at North Carolina State University are carriers of hemotropic Mycoplasma or Bartonella spp. Spleen samples were collected at necropsy during May through July, 2014 from 53 turtles of seven species. All turtles were dead or were euthanized upon arrival due to severe traumatic injuries, or they died shortly after beginning treatment. We used PCR amplification for both bacterial genera; Bartonella spp. DNA was not amplified. Based upon sequencing of the 16S rRNA subunit, one eastern box turtle (Terrapene carolina carolina) and one yellow-bellied slider (Trachemys scripta scripta) were infected with Mycoplasma spp. that have genetic similarities to strains that infect other animals.}, number={2}, journal={Journal of Wildlife Diseases}, publisher={Wildlife Disease Association}, author={Jarred, Jo and Lewbart, Gregory A. and Stover, Kelsey and Thomas, Brittany and Maggi, Ricardo and Breitschwerdt, Edward B.}, year={2018}, month={Apr}, pages={371–374} }
 @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{hegarty_qurollo_thomas_park_chandrashekar_beall_thatcher_breitschwerdt_2015, title={Serological and molecular analysis of feline vector-borne anaplasmosis and ehrlichiosis using species-specific peptides and PCR}, volume={8}, ISSN={1756-3305}, url={http://dx.doi.org/10.1186/s13071-015-0929-8}, DOI={10.1186/s13071-015-0929-8}, abstractNote={With the exception of Bartonella spp. or Cytauxzoon felis, feline vector-borne pathogens (FVBP) have been less frequently studied in North America and are generally under-appreciated as a clinical entity in cats, as compared to dogs or people. This study investigated selected FVBP seroreactivity and PCR prevalence in cats using archived samples. Feline blood samples submitted to the Vector Borne Diseases Diagnostic Laboratory (VBDDL) at North Carolina State University College of Veterinary Medicine (NCSU-CVM) between 2008 and 2013 were tested using serological assays and PCR. An experimental SNAP® Multi-Analyte Assay (SNAP® M-A) (IDEXX Laboratories, Inc. Westbrook, Maine, USA) was used to screen all sera for antibodies to Anaplasma and Ehrlichia genus peptides and A.phagocytophilum, A.platys, B.burgdorferi, E.canis, E.chaffeensis, and E.ewingii species-specific peptides. PCR assays were used to amplify Anaplasma or Ehrlichia DNA from extracted ethylenediaminetetraacetic acid (EDTA)-anti-coagulated blood samples. Amplicons were sequenced to identify species. Overall, 7.8 % (56/715) of cats were FVBP seroreactive and 3.2 % (13/406) contained Anaplasma or Ehrlichia DNA. Serologically, B.burgdorferi (5.5 %) was the most prevalent FVBP followed by A.phagocytophilum (1.8 %). Ehrlichia spp. antibodies were found in 0.14 % (12/715) of cats with species-specific seroreactivity to E.canis (n = 5), E.ewingii (n = 2) and E.chaffeensis (n = 1). Of seropositive cats, 16 % (9/56) were exposed to more than one FVBP, all of which were exposed to B.burgdorferi and either A.phagocytophilum (n = 7) or E.ewingii (n = 2). Based upon PCR and DNA sequencing, 4, 3, 3, 2, and 1 cat were infected with A.phagocytophilum, A.platys, E. ewingii, E. chaffeensis and E.canis, respectively. Cats are exposed to and can be infected with vector-borne pathogens that commonly infect dogs and humans. To our knowledge, this study provides the first evidence for E.chaffeensis and E.ewingii infection in naturally-exposed cats in North America. Results from this study support the need for regional, serological and molecular FVBP prevalence studies, the need to further optimize serodiagnostic and PCR testing for cats, and the need for prospective studies to better characterize clinicopathological disease manifestations in cats infected with FVBP.}, number={1}, journal={Parasites & Vectors}, publisher={Springer Science and Business Media LLC}, author={Hegarty, Barbara C. and Qurollo, Barbara A. and Thomas, Brittany and Park, Karen and Chandrashekar, Ramaswamy and Beall, Melissa J. and Thatcher, Brendon and Breitschwerdt, Edward B.}, year={2015}, month={Jun}, pages={320} }