Molecular characterization of Haemaphysalis longicornis-borne rickettsiae, Republic of Korea and China.

Haemaphysalis longicornis, the cattle tick or bush tick, has an extended distribution throughout Asia and the Pacific region, including China, Russia, the Republic of Korea (ROK), Japan, Australia, New Zealand, and the South Pacific islands. It is an obligate ectoparasite found commonly on medium to large sized wild and domestic animals, with humans as an accidental host. Haemaphysalis longicornis transmits a number of pathogens, including severe fever with thrombocytopenia syndrome and tick-borne encephalitis viruses, bacteria, helminths, and protozoans, that impact on veterinary (wild and domestic animals) and human health.

Development of conventional and real-time reverse transcription polymerase chain reaction assays to detect Tembusu virus in Culex tarsalis mosquitoes.

Tembusu virus (TMUV) is an important emerging arthropod-borne virus that may cause encephalitis in humans and has been isolated in regions of southeast Asia, including Malaysia, Thailand, and China. Currently, detection and identification of TMUV are limited to research laboratories, because quantitative rapid diagnostic assays for the virus do not exist. We describe the development of sensitive and specific conventional and real-time quantitative reverse transcription polymerase chain reaction assays for detecting TMUV RNA in infected cell culture supernatant and Culex tarsalis mosquitoes.

Field detection of Tembusu virus in western Thailand by rt-PCR and vector competence determination of select culex mosquitoes for transmission of the virus.

Tembusu virus (TMUV; Ntaya serocomplex) was detected in two pools of mosquitoes captured near Sangkhlaburi, Thailand, as well as from sera from sentinel ducks from the same area. Although TMUV has been isolated from several mosquito species in Asia, no studies have ever shown competent vectors for this virus. Therefore, we allowed mosquitoes captured near Sangkhlaburi to feed on young chickens that had been infected with TMUV.

Evaluation of a field-portable DNA microarray platform and nucleic acid amplification strategies for the detection of arboviruses, arthropods, and bloodmeals.

Highly multiplexed assays, such as microarrays, can benefit arbovirus surveillance by allowing researchers to screen for hundreds of targets at once. We evaluated amplification strategies and the practicality of a portable DNA microarray platform to analyze virus-infected mosquitoes. The prototype microarray design used here targeted the non-structural protein 5, ribosomal RNA, and cytochrome b genes for the detection of flaviviruses, mosquitoes, and bloodmeals, respectively.

Development of three quantitative real-time PCR assays for the detection of Rickettsia raoultii, Rickettsia slovaca, and Rickettsia aeschlimannii and their validation with ticks from the country of Georgia and the Republic of Azerbaijan.

A previous surveillance study of human pathogens within ticks collected in the country of Georgia showed a relatively high infection rate for Rickettsia raoultii, R. slovaca, and R. aeschlimannii.

Isolation and genomic characterization of Chaoyang virus strain ROK144 from Aedes vexans nipponii from the Republic of Korea.

During June 2003, mosquito surveillance was conducted at a US Army installation and a US Military training site 2 km south of the demilitarized zone, Republic of Korea. Mosquitoes were collected using Mosquito Magnets™, sorted to species, and assayed for the presence of arboviruses. From the 3,149 mosquitoes that were sorted into 126 pools, one Aedes vexans nipponii pool (out of 73 pools) tested positive for flavivirus RNA by reverse transcription-PCR.

Hantaan virus surveillance in small mammals at firing points 10 and 60, Yeoncheon, Gyeonggi Province, Republic of Korea.

We used epidemiological data and indirect fluorescent antibody tests to determine the Hantaan virus (HTNV) antibody-positive (Ab+) prevalence in small mammals captured at firing point 10 (FP-10) and firing point 60 (FP-60), Gyeonggi Province, near the demilitarized zone, Republic of Korea (ROK), from 2001 to 2005. We used these data, combined with the partial M segment amplified from HTNV recovered from lung tissues of Apodemus agrarius, to clarify the genetic diversity and phylogenetic relationships among HTNV strains in the ROK. Of the eight species of rodents and one insectivore species captured, A.

Ecological surveillance of small mammals at Dagmar North Training Area, Gyeonggi Province, Republic of Korea, 2001-2005.

A seasonal rodent-borne disease surveillance program was established at Dagmar North Training Area located near the demilitarized zone, Republic of Korea, from 2001 through 2005. Selected habitats surveyed included earthen banks separating rice paddies, fighting positions along a 5 m rock-faced earthen berm, and extensive tall grasses with various degrees of herbaceous and scrub vegetation associated with dirt roads, rice paddies, ditches, ponds, or the Imjin River. Of the nine species of small mammals captured, the striped field mouse (Apodemus agrarius), the primary reservoir for Hantaan virus, was the most frequently collected, representing 92.

Sequential Rift Valley fever outbreaks in eastern Africa caused by multiple lineages of the virus.

Background: During the Rift Valley fever (RVF) epidemic of 2006-2007 in eastern Africa, spatial mapping of the outbreaks across Kenya, Somalia, and Tanzania was performed and the RVF viruses were isolated and genetically characterized.

Methods: Following confirmation of the RVF epidemic in Kenya on 19 December 2006 and in Tanzania on 2 February 2007, teams were sent to the field for case finding. Human, livestock, and mosquito specimens were collected and viruses isolated.

Detection of Crimean-Congo hemorrhagic fever, Hanta, and sandfly fever viruses by real-time RT-PCR.

The development of sensitive and specific nucleic acid diagnostic assays for viral pathogens is essential for proper medical intervention. This chapter describes four fluorescence-based PCR assays to detect the Crimean-Congo Hemorrhagic Fever (CCHFV), Andes (ANDV), Hantaan (HANV), and Sandfly Fever Sicilian (SFSV) Viruses. These assays are based on species-specific hydrolysis probes targeting the nucleocapsid protein gene for CCHFV and SFSV and the glycoprotein gene for ANDV and HANV.

Hemorrhagic fever with renal syndrome in 4 US soldiers, South Korea, 2005.

Four US soldiers acquired hemorrhagic fever with renal syndrome while training near the Demilitarized Zone, South Korea, in 2005. Hantaan virus sequences were amplified by reverse transcription-PCR from patient serum samples and from lung tissues of striped field mice (Apodemus agrarius) captured at training sites. Epidemiologic investigations specified the ecology of possible sites of patient infection.

Development of field-based real-time reverse transcription-polymerase chain reaction assays for detection of Chikungunya and O'nyong-nyong viruses in mosquitoes.

Chikungunya (CHIK) and O'nyong-nyong (ONN) are important emerging arthropod-borne diseases. Molecular diagnosis of these two viruses in mosquitoes has not been evaluated, and the effects of extraneous mosquito tissue on assay performance have not been tested. Additionally, no real-time reverse transcription-polymerase chain reaction (RT-PCR) assay exists for detecting ONN virus (ONNV) RNA.

Use of vector diagnostics during military deployments: recent experience in Iraq and Afghanistan.

Vector-borne diseases such as malaria, dengue, and leishmaniasis are a threat to military forces deployed outside of the United States. The availability of specific information on the vector-borne disease threat (e.g.

Impact of phlebotomine sand flies on U.S. military operations at Tallil Air Base, Iraq: 4. Detection and identification of leishmania parasites in sand flies.

Sand flies collected between April 2003 and November 2004 at Tallil Air Base, Iraq, were evaluated for the presence of Leishmania parasites using a combination of a real-time Leishmania-generic polymerase chain reaction (PCR) assay and sequencing of a 360-bp fragment of the glucose-6-phosphate-isomerase (GPI) gene. A total of 2,505 pools containing 26,574 sand flies were tested using the real-time PCR assay. Leishmania DNA was initially detected in 536 pools; however, after extensive retesting with the real-time PCR assay, a total of 456 pools were considered positive and 80 were considered indeterminate.

Serological surveillance of scrub typhus, murine typhus, and leptospirosis in small mammals captured at firing points 10 and 60, Gyeonggi province, Republic of Korea, 2001-2005.

Soldiers from the Republic of Korea and the United States conducting peacetime military operations at various training sites and multiple range complexes located near the demilitarized zone separating North and South Korea are exposed to rodents and their potentially disease-carrying ectoparasites. These diseases include scrub typhus, murine typhus, and leptospirosis. Many of the training sites are rural or semi-rural, surrounded or co-located with various forms of agriculture, and are infested with rodents and insectivores (as well as their ectoparasites), which are commonly found in association with unmanaged tall grasses, scrub, and crawling vegetation habitats.

Characterization of Imjin virus, a newly isolated hantavirus from the Ussuri white-toothed shrew (Crocidura lasiura).

Until recently, the single known exception to the rodent-hantavirus association was Thottapalayam virus (TPMV), a long-unclassified virus isolated from the Asian house shrew (Suncus murinus). Robust gene amplification techniques have now uncovered several genetically distinct hantaviruses from shrews in widely separated geographic regions. Here, we report the characterization of a newly identified hantavirus, designated Imjin virus (MJNV), isolated from the lung tissues of Ussuri white-toothed shrews of the species Crocidura lasiura (order Soricomorpha, family Soricidae, subfamily Crocidurinae) captured near the demilitarized zone in the Republic of Korea during 2004 and 2005.

Ecological surveillance of small mammals at Firing Points 10 and 60, Gyeonggi Province, Republic of Korea, 2001-2005.

Throughout Korea, small mammals are hosts to a number of disease-causing agents that pose a health threat to U.S. and Korean military forces while they conduct field-training exercises.

Genetic analysis of South American eastern equine encephalomyelitis viruses isolated from mosquitoes collected in the Amazon Basin region of Peru.

Identifying viral isolates from field-collected mosquitoes can be difficult and time-consuming, particularly in regions of the world where numerous closely related viruses are co-circulating (e.g., the Amazon Basin region of Peru).

Endemic eastern equine encephalitis in the Amazon region of Peru.

Eastern equine encephalitis virus (EEEV) causes severe neurologic disease in North America, but only two fatal human cases have been documented in South America. To test the hypothesis that alphavirus heterologous antibodies cross-protect, animals were vaccinated against other alphaviruses and challenged up to 3 months later with EEEV. Short-lived cross-protection was detected, even in the absence of cross-neutralizing antibodies.

Tick-borne rickettsial pathogens in ticks and small mammals in Korea.

In order to investigate the prevalence of tick-borne infectious agents among ticks, ticks comprising five species from two genera (Hemaphysalis spp. and Ixodes spp.) were screened using molecular techniques.

Impact of phlebotomine sand flies on U.S. Military operations at Tallil Air Base, Iraq: 1. background, military situation, and development of a "Leishmaniasis Control Program".

One of the most significant modern day efforts to prevent and control an arthropod-borne disease during a military deployment occurred when a team of U.S. military entomologists led efforts to characterize, prevent, and control leishmaniasis at Tallil Air Base (TAB), Iraq, during Operation Iraqi Freedom.

Detection of Bartonella species from ticks, mites and small mammals in Korea.

We investigated the prevalence of Bartonella infections in ticks, mites and small mammals (rodents, insectivores and weasels) collected during 2001 through 2004, from various military installations and training sites in Korea, using PCR and sequence analysis of 16S rRNA, 23S rRNA and groEL heat shock protein genes. The prevalence of Bartonella spp. was 5.

An update on the potential of north American mosquitoes (Diptera: Culicidae) to transmit West Nile Virus.

ABSTRACT Since first discovered in the New York City area in 1999, West Nile virus (WNV) has become established over much of the continental United States and has been responsible for >10,000 cases of severe disease and 400 human fatalities, as well as thousands of fatal infections in horses. To develop appropriate surveillance and control strategies, the identification of which mosquito species are competent vectors and how various factors influence their ability to transmit this virus must be determined. Therefore, we evaluated numerous mosquito species for their ability to transmit WNV under laboratory conditions.

Endemic Venezuelan equine encephalitis in northern Peru.

Since Venezuelan equine encephalitis virus (VEEV) was isolated in Peru in 1942, >70 isolates have been obtained from mosquitoes, humans, and sylvatic mammals primarily in the Amazon region. To investigate genetic relationships among the Peru VEEV isolates and between the Peru isolates and other VEEV strains, a fragment of the PE2 gene was amplified and analyzed by single-stranded conformation polymorphism. Representatives of seven genotypes underwent sequencing and phylogenetic analysis.

Field detection of eastern equine encephalitis virus in the Amazon Basin region of Peru using reverse transcription-polymerase chain reaction adapted for field identification of arthropod-borne pathogens.

In support of efforts to develop rapid diagnostic assays for use in the field, reverse transcription-polymerase chain reaction (RT-PCR) assays were developed to detect arboviruses circulating in the Amazon Basin region of Peru. Previous knowledge of arthropod/pathogen relationships allowed a focused evaluation to be conducted in November 2000 that assessed the feasibility and reliability of a mobile, rapid, field-expedient RT-PCR diagnostic system aimed at detecting eastern equine encephalitis virus (EEEV) in Culex (Melanoconion) pedroi mosquitoes. Modifications were made to a commercially available mobile molecular laboratory kit and assay procedures were tailored for use under harsh environmental conditions with field-collected and field-processed mosquitoes.