@article{ponnusamy_sutton_mitchell_sonenshine_apperson_roe_2021, title={Tick Ecdysteroid Hormone, Global Microbiota/Rickettsia Signaling in the Ovary versus Carcass during Vitellogenesis in Part-Fed (Virgin) American Dog Ticks, Dermacentor variabilis}, volume={9}, ISSN={["2076-2607"]}, url={https://www.mdpi.com/2076-2607/9/6/1242}, DOI={10.3390/microorganisms9061242}, abstractNote={The transovarial transmission of tick-borne bacterial pathogens is an important mechanism for their maintenance in natural populations and transmission, causing disease in humans and animals. The mechanism for this transmission and the possible role of tick hormones facilitating this process have never been studied. Injections of physiological levels of the tick hormone, 20-hydroxyecdysone (20E), into part-fed (virgin) adult females of the American dog tick, Dermacentor variabilis, attached to the host caused a reduction in density of Rickettsia montanensis in the carcass and an increase in the ovaries compared to buffer-injected controls. This injection initiates yolk protein synthesis and uptake by the eggs but has no effect on blood feeding. Francisella sp. and R. montanensis were the predominant bacteria based on the proportionality in the carcass and ovary. The total bacteria load increased in the carcass and ovaries, and bacteria in the genus Pseudomonas increased in the carcass after the 20E injection. The mechanism of how the Rickettsia species respond to changes in tick hormonal regulation needs further investigation. Multiple possible mechanisms for the proliferation of R. montanensis in the ovaries are proposed.}, number={6}, journal={MICROORGANISMS}, author={Ponnusamy, Loganathan and Sutton, Haley and Mitchell, Robert D. and Sonenshine, Daniel E. and Apperson, Charles S. and Roe, Richard Michael}, year={2021}, month={Jun} }
 @article{kakumanu_ponnusamy_sutton_meshnick_nicholson_apperson_2018, title={Prevalence of Rickettsia Species (Rickettsiales: Rickettsiaceae) in Dermacentor variabilis Ticks (Acari: Ixodidae) in North Carolina}, volume={55}, ISSN={["1938-2928"]}, url={http://dx.doi.org/10.1093/jme/tjy074}, DOI={10.1093/jme/tjy074}, abstractNote={Abstract The American dog tick, Dermacentor variabilis (Say), is a vector of spotted fever group (SFG) rickettsiae, including Rickettsia rickettsii the causative organism of Rocky Mountain spotted fever (RMSF). In North Carolina, SFG rickettsioses (including RMSF) are a leading cause of tick-borne illness. Knowledge of the infection rate and geographic distribution of D. variabilis ticks infected with Rickettsia spp. provides information on the spatial distribution of public health risk. Accordingly, we extracted genomic DNA from adult D. variabilis collected from field habitats in 32 North Carolina counties from 2009 to 2013. A nested PCR assay of the 23S-5S intergenic spacer (IGS) region of Rickettsia coupled with reverse line blot hybridization (RLBH) with species-specific probes was used to detect and identify rickettsiae to species. Approximately half of the 532 tick DNA samples exhibited a band of the expected size on agarose gels, indicating infection with Rickettsia spp. RLBH analyses showed R. amblyommatis (formerly ‘Candidatus R. amblyommii’), R. parkeri, and R. montanensis were predominant, while other Rickettsia species detected included R. conorii-like, R. massiliae, R. rhipicephali, R. canadensis, R. bellii, and some unknown Rickettsia spp. Some ticks were infected with more than one Rickettsia species. Notably, several Rickettsia-positive ticks harbored R. rickettsii. DNA sequencing was performed on a portion of the 23S-5S IGS amplicons and the results were concordant with RLB assay results. We conclude that Rickettsia spp. are common in D. variabilis in North Carolina. Geographic patterns in the occurrence of Rickettsia-infected D. variabilis ticks across the counties sampled are discussed.}, number={5}, journal={JOURNAL OF MEDICAL ENTOMOLOGY}, author={Kakumanu, Madhavi L. and Ponnusamy, Loganathan and Sutton, Haley and Meshnick, Steven R. and Nicholson, William L. and Apperson, Charles S.}, year={2018}, month={Sep}, pages={1284–1291} }
 @article{levine_apperson_levin_kelly_kakumanu_ponnusamy_sutton_salger_caldwell_szempruch_2017, title={Stable Transmission of Borrelia burgdorferi Sensu Stricto on the Outer Banks of North Carolina}, volume={64}, ISSN={1863-1959}, url={http://dx.doi.org/10.1111/zph.12302}, DOI={10.1111/zph.12302}, abstractNote={The spirochaete (Borrelia burgdorferi) associated with Lyme disease was detected in questing ticks and rodents during a period of 18 years, 1991–2009, at five locations on the Outer Banks of North Carolina. The black‐legged tick (Ixodes scapularis) was collected at varied intervals between 1991 and 2009 and examined for B. burgdorferi. The white‐footed mouse (Peromyscus leucopus), house mouse (Mus musculus) marsh rice rat (Oryzomys palustris), marsh rabbit (Sylvilagus palustris), eastern cottontail (Sylvilagus floridanus) and six‐lined racerunner (Cnemidophorus sexlineatus) were live‐trapped, and their tissues cultured to isolate spirochaetes. Borrelia burgdorferi isolates were obtained from questing adult I. scapularis and engorged I. scapularis removed from P. leucopus, O. palustris and S. floridanus. The prevalence of B. burgdorferi infection was variable at different times and sites ranging from 7 to 14% of examined questing I. scapularis. Mitochondrial (16S) rRNA gene phylogenetic analysis from 65 adult I. scapularis identified 12 haplotypes in two major clades. Nine haplotypes were associated with northern/Midwestern I. scapularis populations and three with southern I. scapularis populations. Sixteen isolates obtained from tick hosts in 2005 were confirmed to be B. burgdorferi by amplifying and sequencing of 16S rRNA and 5S‐23S intergenic spacer fragments. The sequences had 98–99% identity to B. burgdorferi sensu stricto strains B31, JD1 and M11p. Taken together, these studies indicate that B. burgdorferi sensu stricto is endemic in questing I. scapularis and mammalian tick hosts on the Outer Banks of North Carolina.}, number={5}, journal={Zoonoses and Public Health}, publisher={Wiley}, author={Levine, J. F. and Apperson, C. S. and Levin, M. and Kelly, T. R. and Kakumanu, M. L. and Ponnusamy, L. and Sutton, H. and Salger, S. A. and Caldwell, J. M. and Szempruch, A. J.}, year={2017}, month={Aug}, pages={337–354} }
 @article{kakumanu_ponnusamy_sutton_meshnick_nicholson_apperson_2016, title={Development and Validation of an Improved PCR Method Using the 23S-5S Intergenic Spacer for Detection of Rickettsiae in Dermacentor variabilis Ticks and Tissue Samples from Humans and Laboratory Animals}, volume={54}, ISSN={["1098-660X"]}, url={http://dx.doi.org/10.1128/jcm.02605-15}, DOI={10.1128/jcm.02605-15}, abstractNote={ABSTRACT A novel nested PCR assay was developed to detect Rickettsia spp. in ticks and tissue samples from humans and laboratory animals. Primers were designed for the nested run to amplify a variable region of the 23S-5S intergenic spacer (IGS) of Rickettsia spp. The newly designed primers were evaluated using genomic DNA from 11 Rickettsia species belonging to the spotted fever, typhus, and ancestral groups and, in parallel, compared to other Rickettsia-specific PCR targets (ompA, gltA, and the 17-kDa protein gene). The new 23S-5S IGS nested PCR assay amplified all 11 Rickettsia spp., but the assays employing other PCR targets did not. The novel nested assay was sensitive enough to detect one copy of a cloned 23S-5S IGS fragment from “Candidatus Rickettsia amblyommii.” Subsequently, the detection efficiency of the 23S-5S IGS nested assay was compared to those of the other three assays using genomic DNA extracted from 40 adult Dermacentor variabilis ticks. The nested 23S-5S IGS assay detected Rickettsia DNA in 45% of the ticks, while the amplification rates of the other three assays ranged between 5 and 20%. The novel PCR assay was validated using clinical samples from humans and laboratory animals that were known to be infected with pathogenic species of Rickettsia. The nested 23S-5S IGS PCR assay was coupled with reverse line blot hybridization with species-specific probes for high-throughput detection and simultaneous identification of the species of Rickettsia in the ticks. “Candidatus Rickettsia amblyommii,” R. montanensis, R. felis, and R. bellii were frequently identified species, along with some potentially novel Rickettsia strains that were closely related to R. bellii and R. conorii.}, number={4}, journal={JOURNAL OF CLINICAL MICROBIOLOGY}, author={Kakumanu, Madhavi L. and Ponnusamy, Loganathan and Sutton, Haley T. and Meshnick, Steven R. and Nicholson, William L. and Apperson, Charles S.}, year={2016}, month={Apr}, pages={972–979} }
 @article{lee_kakumanu_ponnusamy_vaughn_funkhouser_thornton_meshnick_apperson_2014, title={Prevalence of Rickettsiales in ticks removed from the skin of outdoor workers in North Carolina}, volume={7}, ISSN={["1756-3305"]}, url={http://dx.doi.org/10.1186/s13071-014-0607-2}, DOI={10.1186/s13071-014-0607-2}, abstractNote={Tick-transmitted rickettsial diseases, such as ehrlichiosis and spotted fever rickettsiosis, are significant sources of morbidity and mortality in the southern United States. Because of their exposure in tick-infested woodlands, outdoor workers experience an increased risk of infection with tick-borne pathogens. As part of a double blind randomized-controlled field trial of the effectiveness of permethrin-treated clothing in preventing tick bites, we identified tick species removed from the skin of outdoor workers in North Carolina and tested the ticks for Rickettsiales pathogens.Ticks submitted by study participants from April-September 2011 and 2012 were identified to species and life stage, and preliminarily screened for the genus Rickettsia by nested PCR targeting the 17-kDa protein gene. Rickettsia were further identified to species by PCR amplification of 23S-5S intergenic spacer (IGS) fragments combined with reverse line blot hybridization with species-specific probes and through cloning and nucleotide sequence analysis of 23S-5S amplicons. Ticks were examined for Ehrlichia and Anaplasma by nested PCR directed at the gltA, antigen-expressing gene containing a variable number of tandem repeats, 16S rRNA, and groESL genes.The lone star tick (Amblyomma americanum) accounted for 95.0 and 92.9% of ticks submitted in 2011 (n = 423) and 2012 (n = 451), respectively. Specimens of American dog tick (Dermacentor variabilis), Gulf Coast tick (Amblyomma maculatum) and black-legged tick (Ixodes scapularis) were also identified. In both years of our study, 60.9% of ticks tested positive for 17-kDa. "Candidatus Rickettsia amblyommii", identified in all four tick species, accounted for 90.2% (416/461) of the 23S-5S-positive samples and 52.9% (416/787) of all samples tested. Nucleotide sequence analysis of Rickettsia-specific 23S-5S IGS, ompA and gltA gene fragments indicated that ticks, principally A. americanum, contained novel species of Rickettsia. Other Rickettsiales, including Ehrlichia ewingii, E. chaffeensis, Ehrlichia sp. (Panola Mountain), and Anaplasma phagocytophilum, were infrequently identified, principally in A. americanum.We conclude that in North Carolina, the most common rickettsial exposure is to R. amblyommii carried by A. americanum. Other Rickettsiales bacteria, including novel species of Rickettsia, were less frequently detected in A. americanum but are relevant to public health nevertheless.}, journal={PARASITES & VECTORS}, author={Lee, Sangmi and Kakumanu, Madhavi L. and Ponnusamy, Loganathan and Vaughn, Meagan and Funkhouser, Sheana and Thornton, Haley and Meshnick, Steven R. and Apperson, Charles S.}, year={2014}, month={Dec} }
 @article{stell_roe_arellano_kennedy_thornton_saavedra-rodriguez_wesson_black_apperson_2013, title={Proof of concept for a novel insecticide bioassay based on sugar feeding by adult Aedes aegypti (Stegomyia aegypti)}, volume={27}, ISSN={["1365-2915"]}, DOI={10.1111/j.1365-2915.2012.01048.x}, abstractNote={Aedes aegypti L. (Stegomyia aegypti) (Diptera: Culicidae) is the principal vector of dengue and yellow fever viruses in tropical and subtropical regions of the world. Disease management is largely based on mosquito control achieved by insecticides applied to interior resting surfaces and through space sprays. Population monitoring to detect insecticide resistance is a significant component of integrated disease management programmes. We developed a bioassay method for assessing insecticide susceptibility based on the feeding activity of mosquitoes on plant sugars. Our prototype sugar‐insecticide feeding bioassay system was composed of inexpensive, disposable components, contained minimal volumes of insecticide, and was compact and highly transportable. Individual mosquitoes were assayed in a plastic cup that contained a sucrose‐permethrin solution. Trypan blue dye was added to create a visual marker in the mosquito's abdomen for ingested sucrose‐permethrin solution. Blue faecal spots provided further evidence of solution ingestion. With the sugar‐insecticide feeding bioassay, the permethrin susceptibility of Ae. aegypti females from two field‐collected strains was characterized by probit analysis of dosage‐response data. The field strains were also tested by forced contact of females with permethrin residues on filter paper. Dosage‐response patterns were similar, indicating that the sugar‐insecticide feeding bioassay had appropriately characterized the permethrin susceptibility of the two strains.}, number={3}, journal={MEDICAL AND VETERINARY ENTOMOLOGY}, author={Stell, F. M. and Roe, R. M. and Arellano, C. and Kennedy, L. and Thornton, H. and Saavedra-Rodriguez, K. and Wesson, D. M. and Black, W. C. and Apperson, C. S.}, year={2013}, month={Sep}, pages={284–297} }