Laboratory Transmission of Japanese Encephalitis, West Nile, and Getah Viruses by Mosquitoes (Diptera: Culicidae) Collected near Camp Greaves, Gyeonggi Province, Republic of Korea, 2003

SHORT COMMUNICATION Laboratory Transmission of Japanese Encephalitis, West Nile, and Getah Viruses by Mosquitoes (Diptera: Culicidae) Collected near Camp Greaves, Gyeonggi Province, Republic of Korea, 2003 MICHAEL J. TURELL, CHRISTOPHER N. MORES,1 DAVID J. DOHM, WON-JA LEE,2 HEUNG-CHUL KIM,3 AND TERRY A. KLEIN4 Virology Division, U.S. Army Medical Research Institute of Infectious Diseases, Fort Detrick, MD 21702Ð5011 KEY WORDS virus, mosquitoes, Korea, Japanese encephalitis, West Nile, Getah Japanese encephalitis virus (JEV) and West Nile virus (WNV) are both members of the JEV serogroup (family Flaviviridae, genus Flavivirus). Although most infections in humans with either of these viruses produce little or no clinical illness, infection with either of these viruses can cause life-threatening encephalitis (Burke and Leake 1988, Mackenzie et al. 2004). Japanese encephalitis virus is enzootic in Asia from western Nepal to Korea and Japan and has been responsible for outbreaks of encephalitis in humans, with thousands of cases being reported each year (Burke and Leake 1988, Sohn 2000, Vaughn and Hoke 1992). In the Republic of Korea (ROK), a mandatory vaccination program was initiated in the early 1980s, and this program has signiÞcantly decreased the numbers The mention of trade names or commercial products does not constitute endorsement or recommendation for use by the Department of Defense or the Korean National Institute of Health. Opinions, interpretations, conclusions, and recommendations are those of the authors and are not necessarily endorsed by the U.S. Army. 1 Current Address: University of Florida, Florida Medical Entomology Laboratory, 200 9th Street SE, Vero Beach, FL 32962. 2 Korean Center for Disease Control and Prevention, National Institute of Health, Þve Nokbun-Dong, Eunpyeong-Gu, Seoul 122701, Republic of Korea. 3 5th Medical Detachment, 168th Medical Battalion (Area Support), 18th Medical Command, Unit 15247, APO AP 96205Ð5247. 4 Force Health Protection, 18th Medical Command, Unit 15281, APO AP 96205Ð5281. of human cases. The incidence of Japanese encephalitis was reduced from ⬎18.5/100,000 people in 1964 to ⬍0.02/100,000 people since 1985 (Sohn 2000). Although not responsible for as many deaths, WNV is known to occur over a wide geographic area. This virus is an emerging pathogen and was documented for the Þrst time in the American continent in 1999 (CDC 1999, Lanciotti et al. 1999). Since 1999, infection with WNV has been responsible for ⬎15,000 human infections and ⬎500 human fatalities in North America (CDC 2002, 2003, 2004). Both of these viruses are transmitted by mosquitoes and use birds as amplifying hosts (Burke and Leake 1988, Hayes 1989). Despite the small number of recent human cases of JE in the ROK, the detection of JEV in Culex tritaeniorhynchus Giles and Getah virus (family Togaviridae, genus Alphavirus, GETV) (a virus known to cause illness in horses and humans; Lundstrom 1999) in Aedes vexans (Meigen) mosquitoes captured in Gyeonggi Province, ROK, in 2000 (Turell et al. 2003), indicates a potential risk for human exposure to these viruses in the ROK. Although Cx. tritaeniorhynchus is the known vector of JEV throughout much of its range and is the expected vector in the ROK (Baik and Joo 1991), Cx. tritaeniorhynchus from Korea have not been tested for their ability to transmit JEV, WNV, or GETV. Numerous studies have shown that the ability of a mosquito J. Med. Entomol. 43(5): 1076Ð1081 (2006) ABSTRACT We conducted experimental studies to evaluate mosquitoes captured in Paju County, Gyeonggi Province, Republic of Korea, for their ability to transmit West Nile virus (family Flaviviridae, genus Flavivirus, WNV), Japanese encephalitis virus (family Flaviviridae, genus Flavivirus, JEV), and Getah virus (family Togaviridae, genus Alphavirus, GETV) under laboratory conditions. Both Culex pipiens pallens Coquillett and Culex tritaeniorhynchus Giles were highly susceptible to infection with WNV, with infection rates ⬎65% when allowed to feed on chickens with viremias of ⬇107 plaqueforming units (PFU) of virus/ml blood. In contrast, Cx. tritaeniorhynchus were signiÞcantly more susceptible to JEV or GETV (infection rate 100%) than were the Cx. p. pallens (infection rate 3% for JEV and 0% for GETV) captured in the same area when allowed to feed on chickens with viremias of ⬇105 PFU of virus/ml blood. The detection of JEV in Þeld-collected Cx. tritaeniorhynchus in Gyeonggi Province in 2000 and the demonstrated ability of this species to transmit this virus support the importance of the continued vaccination of Koreans against JEV and indicate a risk of infection for nonvaccinated individuals. September 2006 TURELL ET AL.: TRANSMISSION OF JEV AND WNV BY KOREAN MOSQUITOES species to transmit a particular virus can vary greatly for different geographical populations (Gubler and Rosen 1976, Hardy et al. 1976, Takahashi 1980). Similarly, although members of the genus Culex have been incriminated as vectors of WNV (Hayes 1989, Hubalek and Halouzka 1999), Korean Culex pipiens pallens Coquillett have never been evaluated for their ability to transmit this virus. Therefore, we evaluated Cx. tritaeniorhynchus and Cx. p. pallens captured in Gyeonggi Province, ROK, for their ability to transmit WNV, GETV, and JEV under laboratory conditions. Materials and Methods PFU/ml 2Ð3 d after infection (M.J.T., unpublished data). Therefore, mosquitoes were allowed to feed on 2- to 4-d-old Leghorn chickens that had been inoculated with 102Ð3 PFU of JEV, GETV, or WNV 2Ð3 d earlier. Immediately after mosquito feeding, 0.1 ml of blood was obtained from the jugular vein of each chicken. This was added to 0.9 ml of heparinized diluent, and the blood suspensions were frozen at ⫺70⬚C until tested for virus by plaque assay to determine the viremias at the time of mosquito feeding. After exposure to the viremic chickens, engorged mosquitoes were transferred to 3.8-liter screen-topped cardboard cages held at 26⬚C at a photoperiod of 16:8 (L:D) h. After an incubation period of ⱖ12 d, most mosquitoes were allowed to refeed on 1- to 2-d-old chickens either individually or in small groups to determine whether they could transmit virus by bite. Immediately after the transmission attempt, the mosquitoes were killed by freezing, identiÞed to species, feeding status determined, and their legs and bodies triturated separately in 1 ml of diluent. Infection was determined by recovery of virus from the mosquito tissue suspension. If virus was recovered from its body, but not its legs, the mosquito was considered to have a nondisseminated infection limited to its midgut. In contrast, if virus was recovered from both the body and leg suspensions, the mosquito was considered to have a disseminated infection (Turell et al. 1984). We deÞned the infection and dissemination rates as the percentages of mosquitoes tested that contained virus in their body or legs, respectively. Chicken (...truncated)