Last data update: Apr 22, 2024. (Total: 46599 publications since 2009)
Records 1-30 (of 32 Records) |
Query Trace: Burkhalter KL [original query] |
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Field-collected ticks from Benton County, Arkansas, and prevalence of associated pathogens
Panella NA , Nicholson WL , Komar N , Burkhalter KL , Hughes HR , Theuret DP , Blocher BH , Sexton C , Connelly R , Rothfeldt L , Kenney JL . J Med Entomol 2024 The recovery of a Haemaphysalis longicornis Neumann (Acari: Ixodidae) tick from a dog in Benton County, Arkansas, in 2018 triggered a significant environmental sampling effort in Hobbs State Park Conservation Area. The objective of the investigation was to assess the tick population density and diversity, as well as identify potential tick-borne pathogens that could pose a risk to public health. During a week-long sampling period in August of 2018, a total of 6,154 ticks were collected, with the majority identified as Amblyomma americanum (L), (Acari: Ixodidae) commonly known as the lone star tick. No H. longicornis ticks were found despite the initial detection of this species in the area. This discrepancy highlights the importance of continued monitoring efforts to understand the dynamics of tick populations and their movements. The investigation also focused on pathogen detection, with ticks being pooled by species, age, and sex before being processed with various bioassays. The results revealed the presence of several tick-borne pathogens, including agents associated with ehrlichiosis (n = 12), tularemia (n = 2), and Bourbon virus (BRBV) disease (n = 1), as well as nonpathogenic rickettsial and anaplasmosis organisms. These findings emphasize the importance of public health messaging to raise awareness of the risks associated with exposure to tick-borne pathogens. Prevention measures, such as wearing protective clothing, using insect repellent, and conducting regular tick checks, should be emphasized to reduce the risk of tick-borne diseases. Continued surveillance efforts and research are also essential to improve our understanding of tick-borne disease epidemiology and develop effective control strategies. |
Fatal case of heartland virus disease acquired in the Mid-Atlantic Region, United States
Liu S , Kannan S , Meeks M , Sanchez S , Girone KW , Broyhill JC , Martines RB , Bernick J , Flammia L , Murphy J , Hills SL , Burkhalter KL , Laven JJ , Gaines D , Hoffmann CJ . Emerg Infect Dis 2023 29 (5) 992-996 Heartland virus (HRTV) disease is an emerging tickborne illness in the midwestern and southern United States. We describe a reported fatal case of HRTV infection in the Maryland and Virginia region, states not widely recognized to have human HRTV disease cases. The range of HRTV could be expanding in the United States. |
Laboratory and field evaluations of a commercially available real-time loop-mediated isothermal amplification assay for the detection of West Nile virus in mosquito pools
Burkhalter KL , O'Keefe M , Holbert-Watson Z , Green T , Savage HM , Markowski DM . J Am Mosq Control Assoc 2021 37 (4) 256-262 Although the specific cDNA amplification mechanisms of reverse-transcriptase polymerase chain reaction (RT-PCR) and RT loop-mediated isothermal amplification (RT-LAMP) are very different, both molecular assays serve as options to detect arboviral RNA in mosquito pools. Like RT-PCR, RT-LAMP uses a reverse transcription step to synthesize complementary DNA (cDNA) from an RNA template and then uses target-specific primers to amplify cDNA to detectable levels in a single-tube reaction. Using laboratory-generated West Nile virus (WNV) samples and field-collected mosquito pools, we evaluated the sensitivity and specificity of a commercially available WNV real-time RT-LAMP assay (Pro-AmpRT™ WNV; Pro-Lab Diagnostics, Inc., Round Rock, Texas) and compared the results to a validated real-time RT-PCR assay. Laboratory generated virus stock samples containing ≥ 2.3 log10 plaque-forming units (PFU)/ml and intrathoracically inoculated mosquitoes containing ≥ 2.4 log10 PFU/ml produced positive results in the Pro-AmpRT WNV assay. Of field-collected pools that were WNV positive by real-time RT-PCR, 74.5% (70 of 94) were also positive by the Pro-AmpRT WNV assay, resulting in an overall Cohen's kappa agreement of 79.4% between the 2 tests. The Pro-AmpRT WNV assay shows promise as a suitable virus screening tool for vector surveillance programs provided agencies are aware of its characteristics and limitations. |
Use of mosquitoes to indirectly assess West Nile virus activity among colonial waterbirds
Felix TA , Young G , Panella NA , Burkhalter KL , Komar N . Waterbirds 2021 43 314-320 West Nile virus activity was evaluated within an island waterbird nesting colony with > 1,250 birds at Riverside Reservoir, Weld County, Colorado, USA. To avoid disturbance of nesting birds, blood-engorged mosquitoes (Culex tarsalis) were used to sample blood indirectly from birds rather than capturing, sampling, and releasing live birds. Local virus activity was confirmed by West Nile virus-positive feather samples from 26% of 46 carcasses collected during monthly visits to the colony from June to September 2009, including American White Pelican (Pelecanus erythrorhynchos; n = 7), California Gull (Larus californicus; n = 1), Snowy Egret (Egretta thula; n = 2), and Cattle Egret (Bubulcus ibis; n = 2). Of 22 blood-engorged mosquitoes collected and the blood meal host identified to species, one West Nile virus infection was detected (putatively from a Snowy Egret), and West Nile virus-specific antibodies were detected in eight samples: Snowy Egret (n = 5), Great Blue Heron (Ardea herodias; n = 2), and American White Pelican (n = 1). The engorgement rate of female Culex tarsalis at the nesting colony was 34%, sixfold higher than that at a nearby mainland site of 5.3%. The utilization of mosquitoes for sampling blood from wild animals may have broader application, and potentially reduce human disturbance of sensitive nesting bird species. © 2021 The Waterbird Society. All rights reserved. |
Laboratory evaluation of the rapid analyte measurement platform assay to detect dengue virus in mosquito pools
Burkhalter KL , Savage HM . J Am Mosq Control Assoc 2021 37 (3) 152-156 We report the results of a laboratory sensitivity and specificity evaluation of the Rapid Analyte Measurement Platform (RAMP®) Dengue Virus (DENV) antigen detection assay, which is designed to detect all serotypes of DENV in mosquito pools. The RAMP DENV assay was able to detect geographically distinct strains of all 4 DENV serotypes in virus-spiked mosquito pools that contained at least 4.3 log10 plaque forming units/ml, although discrete sensitivity limits varied slightly for each serotype. The RAMP DENV assay also detected DENV 1-4 in mosquito pools containing a single infected mosquito and 24 laboratory-reared uninfected mosquitoes. No false positives were detected in negative control mosquito pools or in samples containing high titers of nontarget arboviruses. We found that while the kit-supplied RAMP buffer reduced the infectious titer of DENV, it did not completely inactivate all serotypes. We recommend adding a detergent, Triton X-100, to the buffer to ensure complete inactivation of DENV if the assay is to be conducted at a lower biosafety level than required for DENV handling. |
Experimental Infection of Amblyomma americanum (Acari: Ixodidae) With Bourbon Virus (Orthomyxoviridae: Thogotovirus)
Godsey MS , Rose D , Burkhalter KL , Breuner N , Bosco-Lauth AM , Kosoy OI , Savage HM . J Med Entomol 2021 58 (2) 873-879 Following the recent discovery of Bourbon virus (BRBV) as a human pathogen, and the isolation of the virus from Amblyomma americanum (L.) collected near the location of a fatal human case, we undertook a series of experiments to assess the laboratory vector competence of this tick species for BRBV. Larval ticks were infected using an immersion technique, and transstadial transmission of virus to the nymphal and then to the adult stages was demonstrated. Transstadially infected nymphs transmitted virus to adult ticks at very high rates during cofeeding, indicating the presence of infectious virus in the saliva of engorging ticks. Vertical transmission by transstadially infected females to their progeny occurred, but at a low rate. Rabbits fed on by infected ticks of all active life stages developed high titers of antibody to the virus, demonstrating host exposure to BRBV antigens/live virus during tick blood feeding. These results demonstrate that A. americanum is a competent vector of BRBV and indicate that cofeeding could be critical for enzootic maintenance. |
Genomic characterization of 99 viruses from the bunyavirus families Nairoviridae, Peribunyaviridae, and Phenuiviridae, including 35 previously unsequenced viruses.
Kapuscinski ML , Bergren NA , Russell BJ , Lee JS , Borland EM , Hartman DA , King DC , Hughes HR , Burkhalter KL , Kading RC , Stenglein MD . PLoS Pathog 2021 17 (3) e1009315 Bunyaviruses (Negarnaviricota: Bunyavirales) are a large and diverse group of viruses that include important human, veterinary, and plant pathogens. The rapid characterization of known and new emerging pathogens depends on the availability of comprehensive reference sequence databases that can be used to match unknowns, infer evolutionary and pathogenic potential, and make response decisions in an evidence-based manner. In this study, we determined the coding-complete genome sequences of 99 bunyaviruses in the Centers for Disease Control and Prevention's Arbovirus Reference Collection, focusing on orthonairoviruses (family Nairoviridae), orthobunyaviruses (Peribunyaviridae), and phleboviruses (Phenuiviridae) that either completely or partially lacked genome sequences. These viruses had been collected over 66 years from 27 countries from vertebrates and arthropods representing 37 genera. Many of the viruses had been characterized serologically and through experimental infection of animals but were isolated in the pre-sequencing era. We took advantage of our unusually large sample size to systematically evaluate genomic characteristics of these viruses, including reassortment, and co-infection. We corroborated our findings using several independent molecular and virologic approaches, including Sanger sequencing of 197 genome segments, and plaque isolation of viruses from putative co-infected virus stocks. This study contributes to the described genetic diversity of bunyaviruses and will enhance the capacity to characterize emerging human pathogenic bunyaviruses. |
Powassan virus infection likely acquired through blood transfusion presenting as encephalitis in a kidney transplant recipient
Taylor L , Stevens T , Destrampe EM , Brown JA , McGavic J , Gould CV , Chambers TV , Kosoy OI , Burkhalter KL , Annambhotla P , Basavaraju SV , Groves J , Osborn RA , Weiss J , Stramer SL , Misch EA . Clin Infect Dis 2020 72 (6) 1051-1054 A kidney transplant patient without known tick exposure developed encephalitis three weeks after transplantation. During the transplant hospitalization, the patient had received a blood transfusion from an asymptomatic donor later discovered to have been infected with Powassan virus. This report describes a probable instance of transfusion-transmitted Powassan virus infection. |
Focal amplification and suppression of West Nile virus transmission associated with communal bird roosts in northern Colorado
Komar N , Panella NA , Burkhalter KL . J Vector Ecol 2018 43 (2) 220-234 To explain the patchy distribution of West Nile virus (WNV), we propose that avian immunity encountered by Culex vectors regulates WNV transmission, particularly at communal bird roosts. To test this hypothesis, we selected two test sites with communally roosting American robins (Turdus migratorius) and two control sites that lacked communal roosts. The density of vector-vertebrate contacts, represented by engorged Culex pipiens, was 23-fold greater at test sites compared to control sites, and the density of blood-engorged Cx. pipiens measured in resting mosquito traps correlated positively with the presence of robins and negatively with the presence of other birds, confirming an attraction to robins for blood feeding. WNV transmission was alternately up-regulated (amplification) and down-regulated (suppression) at both test sites. At one test site, infection in resting Cx. pipiens surged from zero to 37.2 per thousand within four weeks, and robin immunity rose from 8.4% to 64% before reducing to 33%. At this site, ten potentially infectious contacts between vector and vertebrates (including nine robins and a mourning dove [Zenaida macroura]) were documented. Infectious vector-vertebrate contacts were absent from control sites. The use of infectious vector-vertebrate contacts, rather than infected mosquitoes, to evaluate a transmission focus is novel. |
Notes from the field: Spatially associated coincident and noncoincident cases of La Crosse encephalitis - North Carolina, 2002-2017
Byrd BD , Williams CJ , Staples JE , Burkhalter KL , Savage HM , Doyle MS . MMWR Morb Mortal Wkly Rep 2018 67 (39) 1104-1105 La Crosse virus (LACV) is the most common cause of pediatric arthropod-borne viral (arboviral) encephalitis in the United States (1). It is a California serogroup bunyavirus primarily transmitted by the eastern tree-hole mosquito (Aedes triseriatus) (2). LACV encephalitis is a reportable condition in North Carolina and is a nationally notifiable disease. In North Carolina, LACV encephalitis is the most common endemic arboviral disease reported in humans, with seven western counties accounting for approximately 80% of confirmed cases since 2003 (3). The fatality rate for LACV encephalitis is <1%, with most patients recovering without overt clinical sequelae; however, long-term neurologic sequelae reported in some patients include recurrent seizures, hemiparesis, and cognitive and neurobehavioral abnormalities (4). |
Susceptibility and vectorial capacity of American Aedes albopictus and Aedes aegypti (Diptera: Culicidae) to American Zika virus strains
Lozano-Fuentes S , Kenney JL , Varnado W , Byrd BD , Burkhalter KL , Savage HM . J Med Entomol 2018 56 (1) 233-240 The rapid expansion of Zika virus (ZIKV), following the recent outbreaks of Chikungunya virus, overwhelmed the public health infrastructure in many countries and alarmed many in the scientific community. Aedes aegypti (L.) (Diptera: Culicidae) and Aedes albopictus (Skuse) (Diptera: Culicidae) have previously been incriminated as the vectors of these pathogens in addition to dengue virus. In our study, we challenged low generation Ae. aegypti (Chiapas, Mexico) and Ae. albopictus (North Carolina, Mississippi), with three strains of ZIKV, Puerto Rico (GenBank: KU501215), Honduras (GenBank: KX694534), and Miami (GenBank: MF988743). Following an oral challenge with 107.5 PFU/ml of the Puerto Rico strain, we observed high infection and dissemination rates in both species (95%). We report estimated transmission rates for both species (74 and 33%, for Ae. aegypti (L.) (Diptera: Culicidae) and Ae. albopictus (Skuse) (Diptera: Culicidae), respectively), and the presence of a probable salivary gland barrier in Ae. albopictus to Zika virus. Finally, we calculated vectorial capacity for both species and found that Ae. albopictus had a slightly lower vectorial capacity when compared with Ae. aegypti.Second Language La rapida expansion del virus Zika, poco despues de las epidemias de chikungunya, rebaso la infraestructura de salud publica en muchos paises y sorprendio a muchos en la comunidad cientifica. Notablemente, Aedes aegypti y Aedes albopictus transmiten estos patogenos ademas del virus del dengue. En este estudio se expusieron con tres cepas americanas de virus Zika a grupos de Aedes aegypti y Aedes albopictus de generacion reciente. Encontramos altos porcentajes de infeccion y diseminacion en ambas especies (95%). Se reporta, la transmision viral en ambas especies (74 y 33%, para Aedes aegypti and Aedes albopictus, respectivamente) y una probable barrera a nivel de glandulas salivarias. Finalmente, calculamos la capacidad vectorial para ambas especies. |
Surveillance for Heartland and Bourbon viruses in Eastern Kansas, June 2016
Savage HM , Godsey MS Jr , Tatman J , Burkhalter KL , Hamm A , Panella NA , Ghosh A , Raghavan RK . J Med Entomol 2018 55 (6) 1613-1616 In June 2016, we continued surveillance for tick-borne viruses in eastern Kansas following upon a larger surveillance program initiated in 2015 in response to a fatal human case of Bourbon virus (BRBV) (Family Orthomyxoviridae: Genus Thogotovirus). In 4 d, we collected 14,193 ticks representing four species from four sites. Amblyomma americanum (L.) (Acari: Ixodidae) accounted for nearly all ticks collected (n = 14,116, 99.5%), and the only other species identified were Amblyomma maculatum Koch (Acari: Ixodidae), Dermacentor variabilis (Say) (Acari: Ixodidae) and Ixodes scapularis Say (Acari: Ixodidae). All ticks were tested for both BRBV and Heartland virus (Family Bunyaviridae: Genus Phlebovirus) in 964 pools. Five Heartland virus positive tick pools were detected and confirmed by real-time reverse transcription PCR (rRT-PCR), while all pools tested negative for BRBV. Each Heartland positive pool was composed of 25 A. americanum nymphs with positive pools collected at three different sites in Bourbon County. A. americanum is believed to be the primary vector of both Heartland and BRBVs to humans based upon multiple detections of virus in field-collected ticks, its abundance, and its aggressive feeding behavior on mammals including humans. However, it is possible that A. americanum encounters viremic vertebrate hosts of BRBV less frequently than viremic hosts of Heartland virus, or that BRBV is less efficiently passed among ticks by co-feeding, or less efficiently passed vertically from infected female ticks to their offspring resulting in lower field infection rates. |
Zika virus MB16-23 in mosquitoes, Miami-Dade County, Florida, USA, 2016
Mutebi JP , Hughes HR , Burkhalter KL , Kothera L , Vasquez C , Kenney JL . Emerg Infect Dis 2018 24 (4) 808-10 We isolated a strain of Zika virus, MB16-23, from Aedes aegypti mosquitoes collected in Miami Beach, Florida, USA, on September 2, 2016. Phylogenetic analysis suggests that MB16-23 most likely originated from the Caribbean region. |
Laboratory evaluation of commercially available platforms to detect West Nile and Zika viruses from honey cards
Burkhalter KL , Wiggins K , Burkett-Cadena N , Alto BW . J Med Entomol 2018 55 (3) 717-722 Commercially available assays utilizing antigen or nucleic acid detection chemistries provide options for mosquito control districts to screen their mosquito populations for arboviruses and make timely operational decisions regarding vector control. These assays may be utilized even more advantageously when combined with honey-soaked nucleic acid preservation substrate ('honey card') testing by reducing or replacing the time- and labor-intensive efforts of identifying and processing mosquito pools. We tested artificially inoculated honey cards and cards fed upon individually by West Nile virus (WNV) and Zika virus (ZIKV)-infected mosquitoes with three assays to compare detection rates and the limit of detection for each platform with respect to virus detection of a single infected mosquito and quantify the time interval of virus preservation on the cards. Assays evaluated included CDC protocols for real-time reverse transcriptase polymerase chain reaction (RT-PCR) for WNV and ZIKV, Pro-Lab Diagnostics ProAmpRT WNV loop-mediated amplification (LAMP) and ZIKV LAMP assays, and the Rapid Analyte Measurement Platform (RAMP) WNV assay. Real-time RT-PCR was the most sensitive assay and the most robust to viral RNA degradation over time. To maximize the detection of virus, honey cards should be left in the traps </=1 d if using LAMP assays and </=3 d if using real-time RT-PCR to detect viruses from field samples. The WNV RAMP assay, although effective for pool screening, lacks sensitivity required for honey card surveillance. Future studies may determine the minimum number of infectious mosquitoes required to feed on a honey card that would be reliably detected by the LAMP or RAMP assays. |
Surveillance for tick-borne viruses near the location of a fatal human case of Bourbon virus (Family Orthomyxoviridae: Genus Thogotovirus) in Eastern Kansas, 2015
Savage HM , Godsey MS Jr , Panella NA , Burkhalter KL , Manford J , Trevino-Garrison IC , Straily A , Wilson S , Bowen J , Raghavan RK . J Med Entomol 2018 55 (3) 701-705 Bourbon virus (Family Orthomyxoviridae: Genus Thogotovirus) was first isolated from a human case-patient residing in Bourbon County, Kansas, who subsequently died. Before becoming ill in late spring of 2014, the patient reported several tick bites. In response, we initiated tick surveillance in Bourbon County and adjacent southern Linn County during spring and summer of 2015. We collected 20,639 host-seeking ticks representing four species from 12 sites. Amblyomma americanum (L.) (Acari: Ixodidae) and Dermacentor variabilis (Say) (Acari: Ixodidae) accounted for nearly all ticks collected (99.99%). Three tick pools, all composed of adult A. americanum ticks collected in Bourbon County, were virus positive. Two pools were Heartland virus (Family Bunyaviridae: Genus Phlebovirus) positive, and one was Bourbon virus positive. The Bourbon virus positive tick pool was composed of five adult females collected on a private recreational property on June 5. Detection of Bourbon virus in the abundant and aggressive human-biting tick A. americanum in Bourbon County supports the contention that A. americanum is a vector of Bourbon virus to humans. The current data combined with virus detections in Missouri suggest that Bourbon virus is transmitted to humans by A. americanum ticks, including both the nymphal and adult stages, that ticks of this species become infected as either larvae, nymphs or both, perhaps by feeding on viremic vertebrate hosts, by cofeeding with infected ticks, or both, and that Bourbon virus is transstadially transmitted. Multiple detections of Heartland virus and Bourbon virus in A. americanum ticks suggest that these viruses share important components of their transmission cycles. |
Bourbon virus in field-collected ticks, Missouri, USA
Savage HM , Burkhalter KL , Godsey MS Jr , Panella NA , Ashley DC , Nicholson WL , Lambert AJ . Emerg Infect Dis 2017 23 (12) 2017-2022 Bourbon virus (BRBV) was first isolated in 2014 from a resident of Bourbon County, Kansas, USA, who died of the infection. In 2015, an ill Payne County, Oklahoma, resident tested positive for antibodies to BRBV, before fully recovering. We retrospectively tested for BRBV in 39,096 ticks from northwestern Missouri, located 240 km from Bourbon County, Kansas. We detected BRBV in 3 pools of Amblyomma americanum (L.) ticks: 1 pool of male adults and 2 pools of nymphs. Detection of BRBV in A. americanum, a species that is aggressive, feeds on humans, and is abundant in Kansas and Oklahoma, supports the premise that A. americanum is a vector of BRBV to humans. BRBV has not been detected in nonhuman vertebrates, and its natural history remains largely unknown. |
Entomological investigations during early stages of a chikungunya outbreak in the United States Virgin Islands, 2014
Kenney JL , Burkhalter KL , Scott ML , McAllister J , Lang FE , Webster S , Maduro DJ , Johannes J , Liburd A , Mutebi JP . J Am Mosq Control Assoc 2017 33 (1) 8-15 During the 2014 chikungunya (CHIK) outbreak in the Caribbean, we performed entomological surveys on 3 United States Virgin Islands (USVI): St. Croix, St. Thomas, and St. John. We aimed to evaluate the potential for chikungunya virus (CHIKV) transmission in the USVI. The surveys took place between June 19, 2014, and June 29, 2014, during the dry season in USVI. A total of 1,929 adult mosquitoes belonging to 4 species - Culex quinquefasciatus (68.4%), Aedes aegypti (29.7%), Ae. mediovittatus (1.3%), and Ae. sollicitans (<1%) - were detected. Environmental investigations showed that between 73% and 87% of the homes had containers that could serve as mosquito larval habitats. In addition, 47% of the homes did not have air conditioning and between 69% and 79% of homes showed evidence of frequent outdoor activity exhibited by residents. Taken together, these observations suggest a high potential for CHIKV transmission in USVI. The relative abundance of Ae. aegypti on St. John's, St. Thomas, and St. Croix was 21.0, 11.0, and 3.0 mosquitoes/trap per day, respectively, suggesting that the former 2 islands were at the highest risk of CHIKV outbreaks. Insecticide resistance testing detected high levels of resistance to malathion and permethrin in several local populations of Ae. aegypti on St. Croix Island, which suggested that these 2 insecticides should not be used during CHIK outbreaks. |
Transmission incompetence of Culex quinquefasciatus and Culex pipiens pipiens from North America for Zika virus
Kenney JL , Romo H , Duggal NK , Tzeng WP , Burkhalter KL , Brault AC , Savage HM . Am J Trop Med Hyg 2017 96 (5) 1235-1240 AbstractIn late 2014, Zika virus (ZIKV; Flaviviridae, Flavivirus) emerged as a significant arboviral disease threat in the Western hemisphere. Aedes aegypti and Aedes albopictus have been considered the principal vectors of ZIKV in the New World due to viral isolation frequency and vector competence assessments. Limited reports of Culex transmission potential have highlighted the need for additional vector competence assessments of North American Culex species. Accordingly, North American Culex pipiens and Culex quinquefasciatus were orally exposed and intrathoracically inoculated with the African prototype ZIKV strain and currently circulating Asian lineage ZIKV strains to assess infection, dissemination, and transmission potential. Results indicated that these two North American Culex mosquito species were highly refractory to oral infection with no dissemination or transmission observed with any ZIKV strains assessed. Furthermore, both Culex mosquito species intrathoracically inoculated with either Asian or African lineage ZIKVs failed to expectorate virus in saliva. These in vivo results were further supported by the observation that multiple mosquito cell lines of Culex species origin demonstrated significant growth restriction of ZIKV strains compared with Aedes-derived cell lines. In summation, no evidence for the potential of Cx. pipiens or Cx. quinquefasciatus to serve as a competent vector for ZIKV transmission in North America was observed. |
Detection of Zika Virus in Desiccated Mosquitoes by Real-Time Reverse Transcription PCR and Plaque Assay.
Burkhalter KL , Savage HM . Emerg Infect Dis 2017 23 (4) 680-681 We assayed Zika virus-infected mosquitoes stored at room temperature for <30 days for live virus by using plaque assay and virus RNA by using real-time reverse transcription PCR. Viable virus was detected in samples stored <10 days, and virus RNA was detected in samples held for 30 days. |
Transmission of Heartland virus (Bunyaviridae: Phlebovirus) by experimentally infected Amblyomma americanum (Acari: Ixodidae)
Godsey MS Jr , Savage HM , Burkhalter KL , Bosco-Lauth AM , Delorey MJ . J Med Entomol 2016 53 (5) 1226-1233 Heartland virus (HRTV; Bunyaviridae: Phlebovirus) is a recently described cause of human illness in the United States. After field studies conducted in 2012 implicated Amblyomma americanum (L.) as a vector of HRTV, we initiated experiments to assess the vector competence of A. americanum Larval and nymphal ticks were immersed in high-titered suspensions of HRTV, and then tested for virus at various intervals postimmersion. In a later trial larval ticks were immersed in HRTV, followed by engorgement on a rabbit. A subset of postmolt nymphs was tested for HRTV to document transstadial transmission. Putatively infected nymphs were cofed with uninfected colony larvae to assess nonviremic transmission. In another trial, nymphs were fed on a rabbit and allowed to molt to the adult stage. Male and female ticks fed and mated upon a rabbit, and females were allowed to oviposit. Male and spent female ticks were tested for HRTV, and offspring of infected females were tested to assess vertical transmission. Infection rates of ≤50% were observed in immersed larvae and nymphs tested at intervals following immersion. Transstadial transmission from larvae to nymphs and then to adults was documented. HRTV was detected in a pool of nymphs molted from uninfected larvae cofed with infected nymphs. Vertical transmission of HRTV was observed in progeny of infected females. Infected females took longer to oviposit and produced fewer offspring. Serologic conversions (without viremia) in rabbits fed upon by immersed larvae or transstadially infected ticks indicate horizontal transmission of HRTV. |
A simple modification to the mosquito homogenization protocol safely inactivates West Nile virus and allows virus detection by the Rapid Analyte Measurement Platform (RAMP) ASSAY
Burkhalter KL , Biggerstaff BJ , Horiuchi K , Savage HM . J Am Mosq Control Assoc 2016 32 (2) 77-82 We evaluated the ability of the Rapid Analyte Measurement Platform (RAMP(R)) mosquito-grinding buffer to inactivate West Nile virus (WNV) by subjecting WNV-positive samples ground in RAMP buffer to incubation intervals ranging from 5 min to 60 min. At each time point an aliquot was removed and serially diluted in bovine albumin (BA)-1 cell culture media to stop the inactivation process by RAMP buffer. Each BA-1 sample was tested for viable virus using Vero 6-well cell culture plaque assay and observed for plaques. We observed very limited inactivation of WNV (1-2 log10 plaque-forming units/ml) by RAMP buffer. Concerned for RAMP operators who may be using this assay in low-level biocontainment facilities, we developed an alternate sample homogenization protocol using Triton X-100 detergent that ensures complete WNV inactivation without compromising the performance of the RAMP assay. |
Surveillance for Heartland virus (Bunyaviridae: Phlebovirus) in Missouri during 2013: First detection of virus in adults of Amblyomma americanum (Acari: Ixodidae)
Savage HM , Godsey MS Jr , Panella NA , Burkhalter KL , Ashley DC , Lash RR , Ramsay B , Patterson T , Nicholson WL . J Med Entomol 2016 53 (3) 607-612 During 2013, we collected and tested ticks for Heartland virus (HRTV), a recently described human pathogen in the genus Phlebovirus (Bunyaviridae), from six sites in northwestern Missouri. Five sites were properties owned by HRTV patients, and the sixth was a conservation area that yielded virus in ticks during 2012. We collected 39,096 ticks representing five species; however, two species, Amblyomma americanum (L.) (97.6%) and Dermacentor variabilis (Say) (2.3%), accounted for nearly all ticks collected. We detected 60 HRTV-positive tick pools and all were composed of A. americanum: 53 pools of nymphs, six pools of male adults, and one pool of female adults. This is the first record of HRTV in adult ticks. Virus was detected at five properties that yielded A. americanum ticks, including properties owned by four of five patients. Virus was detected at two sites that yielded virus in 2012. Detection of virus in multiple years indicates that the virus persists in ticks within a relatively small geographic area, although infection rates (IR) may vary greatly among sites and between years at a site. IR per 1,000 A. americanumin northwestern Missouri during the April-July 2013 study period were as follows: all adults, IR = 1.13; adult females, IR = 0.33; adult males, IR = 1.90; and nymphs, IR = 1.79. The IR in nymphs, the stage with the largest data set, corresponds to 1/559 infected ticks. Having robust estimates of IR in various stages for A. americanum should lead to more accurate public health messaging and a better understanding of virus transmission. |
Temporal and spatial variability of entomological risk indices for West Nile virus infection in northern Colorado: 2006-2013
Fauver JR , Pecher L , Schurich JA , Bolling BG , Calhoon M , Grubaugh ND , Burkhalter KL , Eisen L , Andre BG , Nasci RS , LeBailly A , Ebel GD , Moore CG . J Med Entomol 2015 53 (2) 425-34 West Nile virus (WNV) is enzootic in northern Colorado. Annual surveillance activities in Fort Collins, CO, include collecting female Culex mosquitoes and testing them for the presence of WNV RNA in order to calculate 1) Culex female abundance, 2) WNV infection rate, and 3) the vector index (VI). These entomological risk indices inform public policy regarding the need for emergency adulticiding. Currently, these are calculated on a city-wide basis. In this study, we present descriptive data from historical surveillance records spanning 2006-2013 to discern seasonal and yearly patterns of entomological risk for WNV infection. Also, we retrospectively test the hypothesis that entomological risk is correlated with human transmission risk and is heterogeneous within the City of Fort Collins. Four logistically relevant zones within the city were established and used to test this hypothesis. Zones in the eastern portion of the city consistently had significantly higher Culex abundance and VI compared with zones in the west, leading to higher entomological risk indicators for human WNV infection in the east. Moreover, the relative risk of a reported human case of WNV infection was significantly higher in the eastern zones of the city. Our results suggest that a more spatially targeted WNV management program may better mitigate human risk for WNV infection in Fort Collins, and possibly other cities where transmission is enzootic, while at the same time reducing pesticide use. |
Laboratory validation of the sand fly fever virus antigen assay
Reeves WK , Szymczak MS , Burkhalter KL , Miller MM . J Am Mosq Control Assoc 2015 31 (4) 380-3 Sandfly fever group viruses in the genus Phlebovirus (family Bunyaviridae) are widely distributed across the globe and are a cause of disease in military troops and indigenous peoples. We assessed the laboratory sensitivity and specificity of the Sand Fly Fever Virus Antigen Assay, a rapid dipstick assay designed to detect sandfly fever Naples virus (SFNV) and Toscana virus (TOSV) against a panel of phleboviruses. The assay detected SFNV and TOSV, as well as other phleboviruses including Aguacate, Anahanga, Arumowot, Chagres, and Punta Toro viruses. It did not detect sandfly fever Sicilian, Heartland, Rio Grande, or Rift Valley fever viruses. It did not produce false positive results in the presence of uninfected sand flies (Lutzomyia longipalpis) or Cache Valley virus, a distantly related bunyavirus. Results from this laboratory evaluation suggest that this assay may be used as a rapid field-deployable assay to detect sand flies infected with TOSV and SFNV, as well as an assortment of other phleboviruses. |
Evaluating the use of commercial West Nile Virus antigens as positive controls in the Rapid Analyte Measurement Platform West Nile virus assay
Burkhalter KL , Savage HM . J Am Mosq Control Assoc 2015 31 (4) 371-4 We evaluated the utility of 2 types of commercially available antigens as positive controls in the Rapid Analyte Measurement Platform (RAMP(R)) West Nile virus (WNV) assay. Purified recombinant WNV envelope antigens and whole killed virus antigens produced positive RAMP results and either type would be useful as a positive control. Killed virus antigens provide operational and economic advantages and we recommend their use over purified recombinant antigens. We also offer practical applications for RAMP positive controls and recommendations for preparing them. |
Serological investigation of heartland virus (Bunyaviridae: Phlebovirus) exposure in wild and domestic animals adjacent to human case sites in Missouri 2012-2013
Bosco-Lauth AM , Panella NA , Root JJ , Gidlewski T , Lash RR , Harmon JR , Burkhalter KL , Godsey MS , Savage HM , Nicholson WL , Komar N , Brault AC . Am J Trop Med Hyg 2015 92 (6) 1163-7 Heartland virus (HRTV; Bunyaviridae, Phlebovirus) has recently emerged as the causative agent of human disease characterized by thrombocytopenia and leukopenia in the United States. The lone star tick (Amblyomma americanum L.) has been implicated as a vector. To identify candidate vertebrate amplification hosts associated with enzootic maintenance of the virus, sera and ticks were sampled from 160 mammals (8 species) and 139 birds (26 species) captured near two human case residences in Andrew and Nodaway Counties in northwest Missouri. HRTV-specific neutralizing antibodies were identified in northern raccoons (42.6%), horses (17.4%), white-tailed deer (14.3%), dogs (7.7%), and Virginia opossums (3.8%), but not in birds. Virus isolation attempts from sera and ticks failed to detect HRTV virus. The high antibody prevalence coupled with local abundance of white-tailed deer and raccoons indicates these species as candidate amplification hosts. |
Incrimination of Aedes (Stegomyia) hensilli Farner as an epidemic vector of chikungunya virus on Yap Island, Federated States of Micronesia, 2013
Savage HM , Ledermann JP , Yug L , Burkhalter KL , Marfel M , Hancock WT . Am J Trop Med Hyg 2014 92 (2) 429-436 Two species of Aedes (Stegomyia) were collected in response to the first chikungunya virus (CHIKV) outbreak on Yap Island: the native species Ae. hensilli Farner and the introduced species Ae. aegypti (L.). Fourteen CHIKV-positive mosquito pools were detected. Six pools were composed of female Ae. hensilli, six pools were composed of female Ae. aegypti, one pool was composed of male Ae. hensilli, and one pool contained female specimens identified as Ae. (Stg.) spp. Infection rates were not significantly different between female Ae. hensilli and Ae. aegypti. The occurrence of human cases in all areas of Yap Island and the greater number of sites that yielded virus from Ae. hensilli combined with the ubiquitous distribution of this species incriminate Ae. hensilli as the most important vector of CHIKV during the outbreak. Phylogenic analysis shows that virus strains on Yap are members of the Asia lineage and closely related to strains currently circulating in the Caribbean. |
Evaluation of a Rapid Analyte Measurement Platform and real-time reverse-transcriptase polymerase chain reaction assay West Nile virus detection system in mosquito pools
Burkhalter KL , Horiuchi K , Biggerstaff BJ , Savage HM , Nasci RS . J Am Mosq Control Assoc 2014 30 (1) 21-30 We evaluated the commercially available Rapid Analyte Measurement Platform (RAMP (R)) West Nile virus (WNV) antigen detection test for sensitivity and consistency with real-time reverse transcriptase polymerase chain reaction (RT-PCR) confirmation testing. Panels of samples consisting of WNV-spiked mosquito pools and negative control pools were sent to 20 mosquito abatement districts (MADs) that processed the pools using the RAMP assay. The samples were then sent to the reference laboratories used by the MADs for confirmation by real-time RT-PCR. Positive pools with virus titers of roughly 1-3 log(10) PFU/ml had RAMP scores above the RAMP test positive cutoff score of 30 RAMP units, but these virus-positive samples could not be reliably confirmed by real-time RT-PCR testing. Pools with virus titers >= 4 log(10) PFU/ml scored >= 50 RAMP units. Real-time RT-PCR results varied among the confirmation laboratories. With few exceptions, pools returning a RAMP score of >= 100 were confirmed with real-time RT-PCR, while pools returning a RAMP score of 50-99 appeared to be at the limit of real-time RT-PCR detection. Therefore, we recommend using a positive cutoff of 50 RAMP units with no real-time RT-PCR confirmation to maximize speed, efficiency, and economy of the RAMP assay. A more conservative approach would be to implement a "gray zone'' range of 50-100 RAMP units. Pools scoring within the gray zone could be submitted for real-time RT-PCR confirmation with the understanding that positive pools may not confirm due to the inhibitory effect of the RAMP buffer on the real-time RT-PCR assay. We also conducted a series of experiments using laboratory-prepared mosquito pools spiked with WNV to compare mosquito homogenization buffers, pool sizes, and grinding methods in order to determine how these variables affect the RAMP and real-time RT-PCR assay results. |
Co-circulation of West Nile virus variants, Arizona, USA, 2010
Plante JA , Burkhalter KL , Mann BR , Godsey MS Jr , Mutebi JP , Beasley DW . Emerg Infect Dis 2014 20 (2) 272-5 Molecular analysis of West Nile virus (WNV) isolates obtained during a 2010 outbreak in Maricopa County, Arizona, USA, demonstrated co-circulation of 3 distinct genetic variants, including strains with novel envelope protein mutations. These results highlight the continuing evolution of WNV in North America and the current complexity of WNV dispersal and transmission. |
First detection of Heartland virus (Bunyaviridae: Phlebovirus) from field collected arthropods
Savage HM , Godsey MS Jr , Lambert A , Panella NA , Burkhalter KL , Harmon JR , Lash RR , Ashley DC , Nicholson WL . Am J Trop Med Hyg 2013 89 (3) 445-452 Heartland virus (HRTV), the first pathogenic Phlebovirus (Family: Bunyaviridae) discovered in the United States, was recently described from two Missouri farmers. In 2012, we collected 56,428 ticks representing three species at 12 sites including both patients' farms. Amblyomma americanum and Dermacentor variabilis accounted for nearly all ticks collected. Ten pools composed of deplete nymphs of A. americanum collected at a patient farm and a nearby conservation area were reverse transcription-polymerase chain reaction positive, and eight pools yielded viable viruses. Sequence data from the nonstructural protein of the Small segment indicates that tick strains and human strains are very similar, ≥ 97.6% sequence identity. This is the first study to isolate HRTV from field-collected arthropods and to implicate ticks as potential vectors. Amblyomma americanum likely becomes infected by feeding on viremic hosts during the larval stage, and transmission to humans occurs during the spring and early summer when nymphs are abundant and actively host seeking. |
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