@article{travanty_apperson_ponnusamy_2019, title={A Diverse Microbial Community Supports Larval Development and Survivorship of the Asian Tiger Mosquito (Diptera: Culicidae)}, volume={56}, ISSN={["1938-2928"]}, url={https://doi.org/10.1093/jme/tjz003}, DOI={10.1093/jme/tjz003}, abstractNote={Abstract Laboratory microcosm experiments were conducted to evaluate effects of bacteria isolated from senescent white oak leaves on the growth and survivorship of larval Aedes albopictus (Skuse). Larvae hatched from surface-sterilized eggs were reared in microcosms containing individual bacterial isolates, combined isolates (Porphyrobacter sp., Enterobacter asburiae, Acidiphilium rubrum, Pseudomonas syringae, and Azorhizobium caulinodans), a positive control containing a microbial community from an infusion of white oak leaves, and a negative control consisting of sterile culture media. Experiments were conducted for 21 d after which microcosms were deconstructed, larval survivorship was calculated, and bacteria contained in pupae, and adults that developed were quantified to determine rates of transstadial transmission. Positive control microcosms containing diverse microbial communities had an average (±SE) pupation rate of 89.3 (±5.8)% and average larval survivorship of 96.0 (± 2.3)%. Pupation in microcosms with bacterial isolates only occurred twice among all experimental replications; average larval survivorship ranged from 19 to 56%, depending on treatment. Larval growth was not found to be dependent on bacterial isolate density or isolate species, and larval survivorship was dependent on bacterial isolate density, not on isolate species. Potential mechanisms for failed development of larvae in microcosms with bacterial isolates are discussed. Bacterial isolates alone did not support larval development. High larval survivorship in positive control microcosms suggests that a diverse microbial community is required to complete larval development. Additional studies are needed to evaluate larval growth and survivorship on nonbacterial microbes, such as fungi and protozoa.}, number={3}, journal={JOURNAL OF MEDICAL ENTOMOLOGY}, publisher={Oxford University Press (OUP)}, author={Travanty, Nicholas V. and Apperson, Charles S. and Ponnusamy, Loganathan}, year={2019}, month={May}, pages={632–640} }
 @article{shymanovich_faw_hajhashemi_teague_schal_ponnusamy_apperson_hatano_wasserberg_2019, title={Diel periodicity and visual cues guide oviposition behavior in Phlebotomus papatasi, vector of old-world cutaneous leishmaniasis}, volume={13}, ISSN={1935-2735}, url={http://dx.doi.org/10.1371/journal.pntd.0007165}, DOI={10.1371/journal.pntd.0007165}, abstractNote={Background Phlebotomine sand flies are vectors of human leishmaniases, important neglected tropical diseases. In this study, we investigated diel patterns of oviposition behavior, effects of visual cues on oviposition-site selection, and whether these affect the attraction of gravid Phlebotomus papatasi (Scopoli), the vector of old-world cutaneous leishmaniasis, to olfactory cues from oviposition sites. Methodology/principal findings To evaluate these questions, we conducted a series of experiments using attraction and oviposition assays within free-flight test chambers containing gravid females entrained under a 14:10 hrs light:dark photoperiod. By replacing sticky-screens or moist filter papers every three hours, we showed that oviposition site search occurs mainly in the latest part of the night whereas peak oviposition occurs during the early part of the night. Behavioral responses to olfactory oviposition cues are regulated by time-of-day and can be disrupted by transient exposure to a constant darkness photoperiod. Gravid females, but not any other stage, age, or sex, were attracted to dark, round oviposition jars, possibly resembling rodent burrow openings. This visual attraction disappeared in the absence of an illumination source. Egg deposition rate was not affected by jar color. Olfactory cues had the strongest effect when the visual cues were minimal. Conclusion and significance Our study showed, for the first time, that visual cues in the form of oviposition-site color, lighting level, and photoperiod are important in guiding the oviposition behavior of phlebotomine sand flies. Furthermore, such visual cues could modify the flies’ sensitivity to olfactory oviposition cues. Our results suggest that chemosensory and visual cues are complementary, with visual cues used to orient gravid females towards oviposition sites, possibly at long- to medium-ranges during crepuscular periods, while olfactory cues are used to approach the burrow in darkness and assess its suitability at close-range. Implications to sand fly control are discussed.}, number={3}, journal={PLOS Neglected Tropical Diseases}, publisher={Public Library of Science (PLoS)}, author={Shymanovich, Tatsiana and Faw, Lindsey and Hajhashemi, Nima and Teague, Jimmie and Schal, Coby and Ponnusamy, Loganathan and Apperson, Charles S. and Hatano, Eduardo and Wasserberg, Gideon}, editor={Oliveira, FabianoEditor}, year={2019}, month={Mar}, pages={e0007165} }
 @article{travanty_ponnusamy_kakumanu_nicholson_apperson_2019, title={Diversity and structure of the bacterial microbiome of the American dog tick, Dermacentor variabilis, is dominated by the endosymbiont Francisella}, volume={79}, ISSN={["1878-7665"]}, url={https://doi.org/10.1007/s13199-019-00642-2}, DOI={10.1007/s13199-019-00642-2}, number={3}, journal={SYMBIOSIS}, publisher={Springer Science and Business Media LLC}, author={Travanty, Nicholas V. and Ponnusamy, Loganathan and Kakumanu, Madhavi L. and Nicholson, William L. and Apperson, Charles S.}, year={2019}, month={Nov}, pages={239–250} }
 @article{deguenon_travanty_zhu_carr_denning_reiskind_watson_roe_ponnusamy_2019, title={Exogenous and endogenous microbiomes of wild-caught Phormia regina (Diptera: Calliphoridae) flies from a suburban farm by 16S rRNA gene sequencing}, volume={9}, ISSN={["2045-2322"]}, url={http://dx.doi.org/10.1038/s41598-019-56733-z}, DOI={10.1038/s41598-019-56733-z}, abstractNote={Abstract The black blow fly, Phormia regina (Meigen) (Diptera: Calliphoridae) is one of the most abundant carrion flies in North America. Calliphorids are important in agriculture and animal production, veterinary sciences, forensics and medical entomology. While the role of flies in the epidemiology of human and animal diseases is an active area of research, little is known about the microorganisms associated with these insects. We examined the diversity of wild-caught black blow fly endogenous (internal body) and exogenous (external body) microbial communities using 16S rRNA gene sequencing. Overall, 27 phyla, 171 families and 533 genera were detected, and diversity was significantly higher ( P < 0.05) on external body surfaces. At the genus level, Dysgonomonas , Ignatzschineria , Acinetobacter , Vagococcus , Myroides , and Wohlfahrtiimonas were predominant. Cloning and sequencing of nearly full-length fragments of the 16S rRNA gene showed that some of the species identified are known to be pathogenic to humans, animals, and plants. Myroides odoratimimus and Acinetobacter radioresistens are well-known, multi-drug resistant bacteria. These results provide a snapshot of the microbial communities harbored by adult black blow flies and call for more comprehensive studies to better characterize the role these flies may play in the transmission of pathogenic microorganisms.}, journal={SCIENTIFIC REPORTS}, author={Deguenon, Jean M. and Travanty, Nicholas and Zhu, Jiwei and Carr, Ann and Denning, Steven and Reiskind, Michael H. and Watson, David W. and Roe, R. Michael and Ponnusamy, Loganathan}, year={2019}, month={Dec} }
 @article{ponnusamy_willcox_roe_davidson_linsuwanon_schuster_richards_meshnick_apperson_2018, title={Bacterial microbiome of the chigger mite Leptotrombidium imphalum varies by life stage and infection with the scrub typhus pathogen Orientia tsutsugamushi}, volume={13}, ISSN={["1932-6203"]}, url={https://doi.org/10.1371/journal.pone.0208327}, DOI={10.1371/journal.pone.0208327}, abstractNote={Scrub typhus is a mites-borne rickettsiosis caused by the obligate intracellular Gram-negative bacterium Orientia tsutsugamushi. The disease is potentially life threatening and is prevalent in tropical Asia, islands of the western Pacific Ocean and northern Australia where an estimated one million cases occur annually. Orientia tsutsugamushi is transmitted by the bite of larval mites in the genus Leptotrombidium. In the present study, the composition of the microbiome in larvae, deutonymphs and adult males and females from laboratory colonies of L. imphalum that were infected as well as uninfected with O. tsutsugamushi were investigated by high-throughput sequencing of the bacterial 16S rRNA gene. Notably, the bacterial microbiomes of infected adult females were dominated by sequences of O. tsutsugamushi and an unidentified species of Amoebophilaceae, which together comprised 98.2% of bacterial sequences. To improve the taxonomic resolution of the Amoebophilaceae OTU a nearly full length sequence of the 16S rRNA gene was amplified, cloned, and Sanger sequenced. Infected female mites had 89 to 92% nucleotide identity with the Amoebophilaceae family, indicating that the bacterium was likely to be a species of a novel genus. The species composition of bacterial communities varied between mite life stages regardless of their infection status. Uninfected adults exhibited greater species diversity than adults infected with O. tsutsugamushi. In the infected colony, the rate of filial infection with Orientia was less than 100%. Larval and male mites that were PCR-negative for Orientia contained low numbers of sequences of Amoebophilaceae (0.01 and 0.06%, respectively) in their taxonomic profiles, suggesting that a mutualistic relationship exists between the novel species of Amoebophilaceae and O. tsutsugamushi. Our study findings provide the basis for further research to determine the influence of the novel Amoebophilaceae species on the bacterial microbiome and on vector susceptibility to and transovarial transmission of O. tsutsugamushi.}, number={12}, journal={PLOS ONE}, publisher={Public Library of Science (PLoS)}, author={Ponnusamy, Loganathan and Willcox, Alexandra C. and Roe, R. Michael and Davidson, Silas A. and Linsuwanon, Piyada and Schuster, Anthony L. and Richards, Allen L. and Meshnick, Steven R. and Apperson, Charles S.}, editor={Perotti, M. AlejandraEditor}, year={2018}, month={Dec} }
 @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{dhammi_krestchmar_ponnusamy_bacheler_reisig_herbert_del pozo-valdivia_roe_2016, title={Biology, pest status, microbiome and control of kudzu bug (Hemiptera: Heteroptera: Plataspidae): a new invasive pest in the U.S.}, volume={17}, ISSN={["1422-0067"]}, url={http://dx.doi.org/10.3390/ijms17091570}, DOI={10.3390/ijms17091570}, abstractNote={Soybean is an important food crop, and insect integrated pest management (IPM) is critical to the sustainability of this production system. In recent years, the introduction into the United States of the kudzu bug currently identified as Megacopta cribraria (F.), poses a threat to soybean production. The kudzu bug was first discovered in the state of Georgia, U.S. in 2009 and since then has spread to most of the southeastern states. Because it was not found in the North American subcontinent before this time, much of our knowledge of this insect comes from research done in its native habitat. However, since the U.S. introduction, studies have been undertaken to improve our understanding of the kudzu bug basic biology, microbiome, migration patterns, host selection and management in its expanding new range. Researchers are not only looking at developing IPM strategies for the kudzu bug in soybean, but also at its unique relationship with symbiotic bacteria. Adult females deposit bacterial packets with their eggs, and the neonates feed on these packets to acquire the bacteria, Candidatus Ishikawaella capsulata. The kudzu bug should be an informative model to study the co-evolution of insect function and behavior with that of a single bacteria species. We review kudzu bug trapping and survey methods, the development of bioassays for insecticide susceptibility, insecticide efficacy, host preferences, impact of the pest on urban environments, population expansion, and the occurrence of natural enemies. The identity of the kudzu bug in the U.S. is not clear. We propose that the kudzu bug currently accepted as M. cribraria in the U.S. is actually Megacopta punctatissima, with more work needed to confirm this hypothesis.}, number={9}, journal={Int. J. Mol. Sci}, publisher={MDPI AG}, author={Dhammi, Anirudh and Krestchmar, Jaap B. and Ponnusamy, Loganathan and Bacheler, Jack S. and Reisig, Dominic D. and Herbert, Ames and Del Pozo-Valdivia, Alejandro I. and Roe, R. Michael}, year={2016} }
 @article{paz-soldan_yukich_soonthorndhada_giron_apperson_ponnusamy_schal_morrison_keating_wesson_2016, title={Design and Testing of Novel Lethal Ovitrap to Reduce Populations of Aedes Mosquitoes: Community-Based Participatory Research between Industry, Academia and Communities in Peru and Thailand}, volume={11}, ISSN={1932-6203}, url={http://dx.doi.org/10.1371/journal.pone.0160386}, DOI={10.1371/journal.pone.0160386}, abstractNote={Background Dengue virus (and Chikungunya and Zika viruses) is transmitted by Aedes aegypti and Aedes albopictus mosquitoes and causes considerable human morbidity and mortality. As there is currently no vaccine or chemoprophylaxis to protect people from dengue virus infection, vector control is the only viable option for disease prevention. The purpose of this paper is to illustrate the design and placement process for an attractive lethal ovitrap to reduce vector populations and to describe lessons learned in the development of the trap. Methods This study was conducted in 2010 in Iquitos, Peru and Lopburi Province, Thailand and used an iterative community-based participatory approach to adjust design specifications of the trap, based on community members’ perceptions and feedback, entomological findings in the lab, and design and research team observations. Multiple focus group discussions (FGD) were held over a 6 month period, stratified by age, sex and motherhood status, to inform the design process. Trap testing transitioned from the lab to within households. Results Through an iterative process of working with specifications from the research team, findings from the laboratory testing, and feedback from FGD, the design team narrowed trap design options from 22 to 6. Comments from the FGD centered on safety for children and pets interacting with traps, durability, maintenance issues, and aesthetics. Testing in the laboratory involved releasing groups of 50 gravid Ae. aegypti in walk-in rooms and assessing what percentage were caught in traps of different colors, with different trap cover sizes, and placed under lighter or darker locations. Two final trap models were mocked up and tested in homes for a week; one model was the top choice in both Iquitos and Lopburi. Discussion The community-based participatory process was essential for the development of novel traps that provided effective vector control, but also met the needs and concerns of community members.}, number={8}, journal={PLOS ONE}, publisher={Public Library of Science (PLoS)}, author={Paz-Soldan, Valerie A. and Yukich, Josh and Soonthorndhada, Amara and Giron, Maziel and Apperson, Charles S. and Ponnusamy, Loganathan and Schal, Coby and Morrison, Amy C. and Keating, Joseph and Wesson, Dawn M.}, editor={Paul, RichardEditor}, year={2016}, month={Aug}, pages={e0160386} }
 @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{choi_grieco_apperson_schal_ponnusamy_wesson_achee_2016, title={Effect of Spatial Repellent Exposure on Dengue Vector Attraction to Oviposition Sites}, volume={10}, ISSN={1935-2735}, url={http://dx.doi.org/10.1371/journal.pntd.0004850}, DOI={10.1371/journal.pntd.0004850}, abstractNote={Background Aedes aegypti is a primary vector of dengue virus (DENV), the causative agent of dengue fever, an arthropod-borne disease of global importance. Although a vaccine has been recommended for prevention, current dengue prevention strategies rely on vector control. Recently, volatile pyrethroids—spatial repellents—have received interest as a novel delivery system for adult Ae. aegypti control. Understanding the full range of behavioral effects spatial repellents elicit in mosquito species will be critical to understanding the overall impact these products have on vector populations and will guide expectations of efficacy against DENV transmission. Methodology/Principal Findings The current study quantified changes in attraction of gravid Ae. aegypti to experimental oviposition sites following exposure to the spatial repellent transfluthrin. Responses were measured with two-choice olfaction bioassays using ‘sticky-screens’ covering cups to prevent contact with the oviposition substrate. Two cups contained a bacterial attractant composed of four species of bacteria in calcium alginate beads in water and two cups contained only deionized water. Results from 40 replicates (n = 780 females total per treatment) indicated an estimated difference in attraction of 9.35% ± 0.18 (p ≤ 0.003), implying that the transfluthrin-exposed mosquitoes were more attracted to the experimental oviposition sites than the non-exposed mosquitoes. Conclusions/Significance Findings from this study will further characterize the role of spatial repellents to modify Ae. aegypti behavior related to dengue prevention specifically, and encourage innovation in vector control product development more broadly.}, number={7}, journal={PLOS Neglected Tropical Diseases}, publisher={Public Library of Science (PLoS)}, author={Choi, Diane B. and Grieco, John P. and Apperson, Charles S. and Schal, Coby and Ponnusamy, Loganathan and Wesson, Dawn M. and Achee, Nicole L.}, editor={Kittayapong, PattamapornEditor}, year={2016}, month={Jul}, pages={e0004850} }
 @article{wallace_nicholson_perniciaro_vaughn_funkhouser_juliano_lee_kakumanu_ponnusamy_apperson_et al._2016, title={Incident Tick-Borne Infections in a Cohort of North Carolina Outdoor Workers}, volume={16}, ISSN={["1557-7759"]}, url={http://dx.doi.org/10.1089/vbz.2015.1887}, DOI={10.1089/vbz.2015.1887}, abstractNote={Tick-borne diseases cause substantial morbidity throughout the United States, and North Carolina has a high incidence of spotted fever rickettsioses and ehrlichiosis, with sporadic cases of Lyme disease. The occupational risk of tick-borne infections among outdoor workers is high, particularly those working on publicly managed lands. This study identified incident tick-borne infections and examined seroconversion risk factors among a cohort of North Carolina outdoor workers. Workers from the North Carolina State Divisions of Forestry, Parks and Recreation, and Wildlife (n = 159) were followed for 2 years in a randomized controlled trial evaluating the effectiveness of long-lasting permethrin-impregnated clothing. Antibody titers against Rickettsia parkeri, Rickettsia rickettsii, "Rickettsia amblyommii," and Ehrlichia chaffeensis were measured at baseline (n = 130), after 1 year (n = 82), and after 2 years (n = 73). Titers against Borrelia burgdorferi were measured at baseline and after 2 years (n = 90). Baseline seroprevalence, defined as indirect immunofluorescence antibody titers of 1/128 or greater, was R. parkeri (24%), R. rickettsii (19%), "R. amblyommii" (12%), and E. chaffeensis (4%). Incident infection was defined as a fourfold increase in titer over a 1-year period. There were 40 total seroconversions to at least one pathogen, including R. parkeri (n = 19), "R. amblyommii" (n = 14), R. rickettsii (n = 9), and E. chaffeensis (n = 8). There were no subjects whose sera were reactive to B. burgdorferi C6 antigen. Thirty-eight of the 40 incident infections were subclinical. The overall risk of infection by any pathogen during the study period was 0.26, and the risk among the NC Division of Forest Resources workers was 1.73 times that of workers in other divisions (95% confidence interval [CI]: 1.02, 2.92). The risk of infection was lower in subjects wearing permethrin-impregnated clothing, but not significantly (risk ratio = 0.81; 95% CI: 0.47, 1.39). In summary, outdoor workers in North Carolina are at high risk of incident tick-borne infections, most of which appear to be asymptomatic.}, number={5}, journal={VECTOR-BORNE AND ZOONOTIC DISEASES}, author={Wallace, John W. and Nicholson, William L. and Perniciaro, Jamie L. and Vaughn, Meagan F. and Funkhouser, Sheana and Juliano, Jonathan J. and Lee, Sangmi and Kakumanu, Madhavi L. and Ponnusamy, Loganathan and Apperson, Charles S. and et al.}, year={2016}, month={May}, pages={302–308} }
 @article{marayati_schal_ponnusamy_apperson_rowland_wasserberg_2015, title={Attraction and oviposition preferences of Phlebotomus papatasi (Diptera: Psychodidae), vector of Old-World cutaneous leishmaniasis, to larval rearing media}, volume={8}, ISSN={1756-3305}, url={http://dx.doi.org/10.1186/s13071-015-1261-z}, DOI={10.1186/s13071-015-1261-z}, abstractNote={As part of a project aimed at developing oviposition attractants for the control and surveillance of Phlebotomus papatasi (a vector of Old-World cutaneous leishmaniasis), we tested the hypothesis that gravid sand flies are attracted to chemical cues emanating from the growth medium of conspecific larvae - predominantly larvae-conditioned host feces that represents a suitable oviposition site. We report the results of a systematic assessment of media from various developmental stages of the sand fly using oviposition and olfactometer behavioral assays.We conducted multiple-choice oviposition assays in 500 mL Nalgene jars. Six treatments were placed on separate filter paper discs at the bottom of the jar: 2(nd)/3(rd) larval instar medium, 4(th) larval instar/pupae medium, frass from expired colonies, larval food (aged rabbit chow and rabbit feces mix), rabbit feces, and a solvent (water) control. Fifty gravid females were introduced into each jar. Cumulative number of eggs laid on each filter paper per jar was counted at different time intervals from digital images. Attraction of gravid sand flies to these six treatments was assayed with a 3-chamber linear olfactometer. Twenty gravid females were transferred to the middle chamber of the olfactometer and their distribution in treatment and control chambers was recorded after 3 h.Almost no eggs were oviposited during the first 72 h following a blood-meal. Cumulative egg deposition increased drastically in the next 24 h (hours 73-96), with a slight non-significant increasing trend thereafter. Comparing mean cumulative egg deposition among the six treatments, we found that significantly more eggs were oviposited on 2(nd)/3(rd) larval rearing medium followed by 4(th) instar/pupae rearing medium. Oviposition preference did not vary over time. The olfactometer results were consistent with the oviposition assays, with 2(nd)/3(rd) larval rearing medium being the most attractive, followed by 4(th) instar/pupae rearing medium.The key finding of this study is that gravid, laboratory reared, Ph. papatasi sand flies are significantly more attracted to rearing medium of the most biologically active larval stages (2(nd)/3(rd) instar and 4(th) instar/pupae). This finding indicates that sand fly-digested host food and feces is attractive to gravid females and suggests that the larvae and larval gut microbiome may be involved in conditioning the oviposition substrate and possibly the production of oviposition attractants and stimulants.}, number={1}, journal={Parasites & Vectors}, publisher={Springer Science and Business Media LLC}, author={Marayati, Bahjat Fadi and Schal, Coby and Ponnusamy, Loganathan and Apperson, Charles S. and Rowland, Tobin E. and Wasserberg, Gideon}, year={2015}, month={Dec} }
 @article{guagliardo_morrison_barboza_wesson_ponnusamy_astete_vazquez-prokopec_kitron_2015, title={Evidence for Aedes aegypti (Diptera: Culicidae) oviposition on boats in the Peruvian Amazon}, volume={52}, number={4}, journal={Journal of Medical Entomology}, author={Guagliardo, S. A. and Morrison, A. C. and Barboza, J. L. and Wesson, D. M. and Ponnusamy, L. and Astete, H. and Vazquez-Prokopec, G. and Kitron, U.}, year={2015}, pages={726–729} }
 @article{ponnusamy_schal_wesson_arellano_apperson_2015, title={Oviposition responses of Aedes mosquitoes to bacterial isolates from attractive bamboo infusions}, volume={8}, journal={Parasites & Vectors}, author={Ponnusamy, L. and Schal, C. and Wesson, D. M. and Arellano, C. and Apperson, C. S.}, year={2015} }
 @article{van treuren_ponnusamy_brinkerhoff_gonzalez_parobek_juliano_andreadis_falco_ziegler_hathaway_et al._2015, title={Variation in the Microbiota of Ixodes Ticks with Regard to Geography, Species, and Sex}, volume={81}, ISSN={["1098-5336"]}, url={http://dx.doi.org/10.1128/aem.01562-15}, DOI={10.1128/aem.01562-15}, abstractNote={ABSTRACT Ixodes scapularis is the principal vector of Lyme disease on the East Coast and in the upper Midwest regions of the United States, yet the tick is also present in the Southeast, where Lyme disease is absent or rare. A closely related species, I. affinis, also carries the pathogen in the South but does not seem to transmit it to humans. In order to better understand the geographic diversity of the tick, we analyzed the microbiota of 104 adult I. scapularis and 13 adult I. affinis ticks captured in 19 locations in South Carolina, North Carolina, Virginia, Connecticut, and New York. Initially, ticks from 4 sites were analyzed by 454 pyrosequencing. Subsequently, ticks from these sites plus 15 others were analyzed by sequencing with an Illumina MiSeq machine. By both analyses, the microbiomes of female ticks were significantly less diverse than those of male ticks. The dissimilarity between tick microbiomes increased with distance between sites, and the state in which a tick was collected could be inferred from its microbiota. The genus Rickettsia was prominent in all locations. Borrelia was also present in most locations and was present at especially high levels in one site in western Virginia. In contrast, members of the family Enterobacteriaceae were very common in North Carolina I. scapularis ticks but uncommon in I. scapularis ticks from other sites and in North Carolina I. affinis ticks. These data suggest substantial variations in the Ixodes microbiota in association with geography, species, and sex.}, number={18}, journal={APPLIED AND ENVIRONMENTAL MICROBIOLOGY}, author={Van Treuren, Will and Ponnusamy, Loganathan and Brinkerhoff, R. Jory and Gonzalez, Antonio and Parobek, Christian M. and Juliano, Jonathan J. and Andreadis, Theodore G. and Falco, Richard C. and Ziegler, Lorenza Beati and Hathaway, Nicholas and et al.}, year={2015}, month={Sep}, pages={6200–6209} }
 @article{ponnusamy_gonzalez_van treuren_weiss_parobek_juliano_knight_roe_apperson_meshnickh_2014, title={Diversity of Rickettsiales in the Microbiome of the Lone Star Tick, Amblyomma americanum}, volume={80}, ISSN={["1098-5336"]}, url={http://dx.doi.org/10.1128/aem.02987-13}, DOI={10.1128/aem.02987-13}, abstractNote={ABSTRACT}, number={1}, journal={APPLIED AND ENVIRONMENTAL MICROBIOLOGY}, author={Ponnusamy, Loganathan and Gonzalez, Antonio and Van Treuren, Will and Weiss, Sophie and Parobek, Christian M. and Juliano, Jonathan J. and Knight, Rob and Roe, R. Michael and Apperson, Charles S. and Meshnickh, Steven R.}, year={2014}, month={Jan}, pages={354–359} }
 @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{ponnusamy_böröczky_wesson_schal_apperson_2011, title={Bacteria Stimulate Hatching of Yellow Fever Mosquito Eggs}, volume={6}, ISSN={1932-6203}, url={http://dx.doi.org/10.1371/journal.pone.0024409}, DOI={10.1371/journal.pone.0024409}, abstractNote={Background Aedes aegypti Linnaeus is a peridomestic mosquito that lays desiccation-resistant eggs in water-filled human-made containers. Previous investigations connected egg hatching with declining dissolved oxygen (DO) that is associated with bacterial growth. However, past studies failed to uncouple DO from other potential stimulatory factors and they contained little quantitative information about the microbial community; consequently, a direct role for bacteria or compounds associated with bacteria in stimulating egg hatching cannot be dismissed. Methodology/Principal Findings Environmental factors stimulating hatch of Ae. aegypti eggs were investigated using non-sterile and sterile white oak leaf (WOL) infusions and a bacterial culture composed of a mix of 14 species originally isolated from bamboo leaf infusion. In WOL infusion with active microbes, 92.4% of eggs hatched in 2-h at an average DO concentration of 2.4 ppm. A 24-h old bacterial culture with a DO concentration of 0.73 ppm also stimulated 95.2% of eggs hatch within 1-h. In contrast, only 4.0% of eggs hatched in sterile infusion, whose DO averaged 7.4 ppm. Effects of bacteria were uncoupled from DO by exposing eggs to bacterial cells suspended in NaCl solution. Over a 4-h exposure period, 93.8% of eggs hatched while DO concentration changed minimally from 7.62 to 7.50 ppm. Removal of bacteria by ultra-filtration and cell-free filtrate resulted in only 52.0% of eggs hatching after 4-h at an average DO concentration of 5.5 ppm. Conclusions/Significance Collectively, the results provide compelling evidence that bacteria or water-soluble compounds secreted by bacteria, not just low DO concentration, stimulate hatching of Ae. aegypti eggs. However, the specific cues involved remain to be identified. These research findings contribute new insight into an important aspect of the oviposition biology of Ae. aegypti, a virus vector of global importance, providing the basis for a new paradigm of environmental factors involved in egg hatching.}, number={9}, journal={PLoS ONE}, publisher={Public Library of Science (PLoS)}, author={Ponnusamy, Loganathan and Böröczky, Katalin and Wesson, Dawn M. and Schal, Coby and Apperson, Charles S.}, editor={Leulier, FrançoisEditor}, year={2011}, month={Sep}, pages={e24409} }
 @article{ponnusamy_xu_böröczky_wesson_abu ayyash_schal_apperson_2010, title={Oviposition Responses of the Mosquitoes Aedes aegypti and Aedes albopictus to Experimental Plant Infusions in Laboratory Bioassays}, volume={36}, ISSN={0098-0331 1573-1561}, url={http://dx.doi.org/10.1007/s10886-010-9806-2}, DOI={10.1007/s10886-010-9806-2}, abstractNote={Attraction of the mosquitoes Aedes aegypti and Ae. albopictus to plant infusions was evaluated by using a modified sticky-screen bioassay that improved the resolution of mosquito responses to odorants. Under bioassay conditions, solid-phase microextraction-gas chromatographic analyses of the volatile marker chemical indole showed that odorants diffused from bioassay cups, forming a concentration gradient. Infusions were prepared by separately fermenting senescent leaves of eight plant species in well water. Plant infusions were evaluated over an 8-fold range of leaf biomass and/or a 28 d fermentation period. The responses of gravid females of both mosquito species varied with the plant species and biomass of plant materials used to make infusions, and with the length of the fermentation period. Infusions made from senescent bamboo (Arundinaria gigantea) and white oak (Quercus alba) leaves were significantly attractive to both mosquitoes. In general, infusions prepared by using low biomass of plant material over a 7–14 d fermentation period were most attractive to Ae. aegypti. In contrast, Ae. albopictus was attracted to infusions made using a wider range of plant biomass and over a longer fermentation period. Both mosquito species were more attracted to a non-sterile white oak leaf infusion than to white oak leaf infusion that was prepared using sterilized plant material and water, thus suggesting a role for microbial activity in the production of odorants that mediate the oviposition response of gravid mosquitoes.}, number={7}, journal={Journal of Chemical Ecology}, publisher={Springer Science and Business Media LLC}, author={Ponnusamy, Loganathan and Xu, Ning and Böröczky, Katalin and Wesson, Dawn M. and Abu Ayyash, Luma and Schal, Coby and Apperson, Charles S.}, year={2010}, month={Jun}, pages={709–719} }
 @article{ponnusamy_wesson_arellano_schal_apperson_2010, title={Species Composition of Bacterial Communities Influences Attraction of Mosquitoes to Experimental Plant Infusions}, volume={59}, ISSN={0095-3628 1432-184X}, url={http://dx.doi.org/10.1007/s00248-009-9565-1}, DOI={10.1007/s00248-009-9565-1}, abstractNote={In the container habitats of immature mosquitoes, catabolism of plant matter and other organic detritus by microbial organisms produces metabolites that mediate the oviposition behavior of Aedes aegypti and Aedes albopictus. Public health agencies commonly use oviposition traps containing plant infusions for monitoring populations of these mosquito species, which are global vectors of dengue viruses. In laboratory experiments, gravid females exhibited significantly diminished responses to experimental infusions made with sterilized white oak leaves, showing that attractive odorants were produced through microbial metabolic activity. We evaluated effects of infusion concentration and fermentation time on attraction of gravid females to infusions made from senescent bamboo or white oak leaves. We used plate counts of heterotrophic bacteria, total counts of 4′,6-diamidino-2-phenylindole-stained bacterial cells, and 16S ribosomal DNA (rDNA) polymerase chain reaction–denaturing gradient gel electrophoresis (DGGE) to show that changes in the relative abundance of bacteria and the species composition of bacterial communities influenced attraction of gravid A. aegypti and A. albopictus mosquitoes to infusions. DGGE profiles showed that bacterial species composition in infusions changed over time. Principal components analysis indicated that oviposition responses to plant infusions were in general most affected by bacterial diversity and abundance. Analysis of bacterial 16S rDNA sequences derived from DGGE bands revealed that Proteobacteria (Alpha-, Beta-, Delta-, and Gamma-) were the predominant bacteria detected in both types of plant infusions. Gravid A. aegypti were significantly attracted to a mix of 14 bacterial species cultured from bamboo leaf infusion. The oviposition response of gravid mosquitoes to plant infusions is strongly influenced by abundance and diversity of bacterial species, which in turn is affected by plant species, leaf biomass, and fermentation time.}, number={1}, journal={Microbial Ecology}, publisher={Springer Science and Business Media LLC}, author={Ponnusamy, Loganathan and Wesson, Dawn M. and Arellano, Consuelo and Schal, Coby and Apperson, Charles S.}, year={2010}, month={Jan}, pages={158–173} }
 @article{ponnusamy_xu_stav_wesson_schal_apperson_2008, title={Diversity of Bacterial Communities in Container Habitats of Mosquitoes}, volume={56}, ISSN={["1432-184X"]}, url={http://dx.doi.org/10.1007/s00248-008-9379-6}, DOI={10.1007/s00248-008-9379-6}, abstractNote={We investigated the bacterial diversity of microbial communities in water-filled, human-made and natural container habitats of the mosquitoes Aedes aegypti and Aedes albopictus in suburban landscapes of New Orleans, Louisiana in 2003. We collected water samples from three classes of containers, including tires (n = 12), cemetery urns (n = 23), and miscellaneous containers that included two tree holes (n = 19). Total genomic DNA was extracted from water samples, and 16S ribosomal DNA fragments (operational taxonomic units, OTUs) were amplified by PCR and separated by denaturing gradient gel electrophoresis (DGGE). The bacterial communities in containers represented diverse DGGE-DNA banding patterns that were not related to the class of container or to the local spatial distribution of containers. Mean richness and evenness of OTUs were highest in water samples from tires. Bacterial phylotypes were identified by comparative sequence analysis of 90 16S rDNA DGGE band amplicons. The majority of sequences were placed in five major taxa: Alpha-, Beta- and Gammaproteobacteria, Actinobacteria, Bacteroidetes, Cyanobacteria, Firmicutes, and an unclassified group; Proteobacteria and Bacteroidetes were the predominant heterotrophic bacteria in containers. The bacterial communities in human-made containers consisted mainly of undescribed species, and a phylogenetic analysis based on 16S rRNA sequences suggested that species composition was independent of both container type and the spatial distribution of containers. Comparative PCR-based, cultivation-independent rRNA surveys of microbial communities associated with mosquito habitats can provide significant insight into community organization and dynamics of bacterial species.}, number={4}, journal={MICROBIAL ECOLOGY}, author={Ponnusamy, Loganathan and Xu, Ning and Stav, Gil and Wesson, Dawn M. and Schal, Coby and Apperson, Charles S.}, year={2008}, month={Nov}, pages={593–603} }
 @article{ponnusamy_xu_nojima_wesson_schal_apperson_2008, title={Identification of bacteria and bacteria-associated chemical cues that mediate oviposition site preferences by Aedes aegypti}, volume={105}, ISSN={0027-8424 1091-6490}, url={http://dx.doi.org/10.1073/pnas.0802505105}, DOI={10.1073/pnas.0802505105}, abstractNote={The yellow fever mosquito,Aedes aegypti, the global vector of dengue and yellow fever, is inexorably linked to water-filled human-made containers for egg laying and production of progeny. Oviposition is stimulated by cues from water containers, but the nature and origin of these cues have not been elucidated. We showed that mosquito females directed most of their eggs to bamboo and white-oak leaf infusions, and only a small fraction of the eggs were laid in plain water containers. In binary choice assays, we demonstrated that microorganisms in leaf infusions produced oviposition-stimulating kairomones, and using a combination of bacterial culturing approaches, bioassay-guided fractionation of bacterial extracts, and chemical analyses, we now demonstrate that specific bacteria-associated carboxylic acids and methyl esters serve as potent oviposition stimulants for gravidAe. aegypti. Elucidation of these compounds will improve understanding of the chemical basis of egg laying behavior ofAe. aegypti, and the kairomones will likely enhance the efficacy of surveillance and control programs for this disease vector of substantial global public health importance.}, number={27}, journal={Proceedings of the National Academy of Sciences}, publisher={Proceedings of the National Academy of Sciences}, author={Ponnusamy, L. and Xu, N. and Nojima, S. and Wesson, D. M. and Schal, C. and Apperson, C. S.}, year={2008}, month={Jul}, pages={9262–9267} }
 @article{richards_ponnusamy_unnasch_hassan_apperson_2006, title={Host-feeding patterns of Aedes albopictus (Diptera : Culicidae) in relation to availability of human and domestic animals in suburban landscapes of central north Carolina}, volume={43}, DOI={10.1603/0022-2585(2006)43[543:HPOAAD]2.0.CO;2}, abstractNote={Abstract Aedes albopictus (Skuse) (Diptera: Culicidae) is a major nuisance mosquito and a potential arbovirus vector. The host-feeding patterns of Ae. albopictus were investigated during the 2002 and 2003 mosquito seasons in suburban neighborhoods in Wake County, Raleigh, NC. Hosts of blood-fed Ae. albopictus (n = 1,094) were identified with an indirect enzyme-linked immunosorbent assay, by using antisera made in New Zealand White rabbits to the sera of animals that would commonly occur in peridomestic habitats. Ae. albopictus fed predominantly on mammalian hosts (83%). Common mammalian hosts included humans (24%), cats (21%), and dogs (14%). However, a notable proportion (7%) of bloodmeals also was taken from avian hosts. Some bloodmeals taken from birds were identified to species by a polymerase chain reaction-heteroduplex assay (PCR-HDA). Ae. albopictus fed predominantly on chickens and a northern cardinal. PCR-HDA failed to produce detectable products for 29 (58%) of 50 bloodmeals for which DNA had been amplified, indicating that these mosquitoes took mixed bloodmeals from avian and nonavian hosts. Ae. albopictus preference for humans, dogs, and cats was determined by calculating host-feeding indices for the three host pairs based on the proportion of host specific blood-fed mosquitoes collected in relation to the number of specific hosts per residence as established by a door-to-door survey conducted in 2003. Estimates of the average amount of time that residents and their pets (cats and dogs) spent out of doors were obtained. Host-feeding indices based only on host abundance indicated that Ae. albopictus was more likely to feed on domestic animals. However, when feeding indices were time-weighted, Ae. albopictus fed preferentially upon humans. Ae. albopictus blood feeding on humans was investigated using a STR/PCR-DNA profiling technique that involved amplification of three short tandem repeats loci. Of 40 human bloodmeals, 32 (80%) were from a single human, whereas eight (20%) were multiple bloodmeals taken from more than one human host. We conclude that the blood-feeding preference of Ae. albopictus for mammals will limit acquisition of arboviruses by this species from infected avian amplification hosts. This feeding preference likely limits the vector potential of Ae. albopictus for North American arboviruses.}, number={3}, journal={Journal of Medical Entomology}, author={Richards, S. L. and Ponnusamy, L. and Unnasch, T. R. and Hassan, H. K. and Apperson, C. S.}, year={2006}, pages={543–551} }