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Mosquito Isolates of Ross River Virus from Cairns, Queensland, Australia

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Mosquito Isolates of Ross River Virus from Cairns, Queensland, Australia Am. J. Trop. Med. Hyg., 62(5), 2000, pp. 561–565 Copyright ᭧ 2000 by The American Society of Tropical Medicine and Hygiene MOSQUITO ISOLATES OF ROSS RIVER VIRUS FROM CAIRNS, QUEENSLAND, AUSTRALIA DAVID HARLEY, SCOTT RITCHIE, DEBRA PHILLIPS, AND ANDREW VAN DEN HURK Australian Centre for International and Tropical Health and Nutrition, The University of Queensland, Medical School, Herston Road, Herston, Queensland, 4006, Australia; Tropical Public Health Unit, Cairns, Queensland, Australia; Centre for Public Health Sciences, 39 Kessels Road, Coopers Plains, Queensland, 4108, Australia; Department of Microbiology, The University of Queensland, St. Lucia, Queensland, 4072, Australia Abstract. During 1996–1998 60,619 mosquitoes were collected around Cairns, Australia and processed for Al- phavirus isolation. Thirty-three isolates of Ross River (RR) virus were made from 9 species, Aedes imprimens, Aedes kochi, Aedes notoscriptus, Aedes vigilax, Culex annulirostris, Culex gelidus, Mansonia septempunctata, Verrallina (formerly Aedes) carmenti, and Verrallina lineatus. Attempts to isolate RR virus from 121 Aedes aegypti were unsuccessful. Twenty-six (79%) of the isolates came from within 1 km of a colony of spectacled flying-foxes, Pteropus conspicillatus. The minimum infection rate for these mosquitoes was 1.0 compared with 0.2 per 1,000 for mosquitoes trapped at all other sites. Ross River virus has not previously been isolated from Ae. imprimens, Cx. gelidus, Ma. septempunctata, Ve. carmenti, or Ve. lineatus. This is also the first isolation of an arbovirus from Cx. gelidus in Australia. In conclusion, the vector status of Ve. carmenti, Ae. aegypti and Ma. septempunctata warrants further study. This study also provides evidence that P. conspicillatus may be a reservoir host. Ross River (RR) virus is a mosquito-borne Alphavirus that this paper were 1. to determine what mosquito species are occurs in Australia, Papua New Guinea, and the Solomon infected with RR virus in the Cairns region, and 2. to com- Islands.1–3 Human infection may cause arthralgia and arthri- pare the minimum infection rates for mosquitoes collected tis, possibly persisting for long periods.4,5 The average num- Յ 1 km and Ͼ 1 km from a spectacled flying-fox (Pteropus ber of notified cases in Australia during 1991–1996 was conspicillatus) camp. 4,800 with a maximum of 7,823 in 1996 and a minimum of 2,602 in 1995. The majority of notifications come from MATERIALS AND METHODS Queensland,6 especially north Queensland. During 1989– 1992 the incidences in Cairns and Townsville, provincial cit- Mosquito collections. Centers for Disease Control and ies in the north, ranged from 131 to 233 and 150 to 367, Prevention (CDC) traps27 were set between 3.45 and 6.25 respectively, while the incidence in Brisbane, the state cap- PM, and collected between 7.45 and 9.30 AM. Traps were ital in the southeast, ranged from 17 to 96 per 100,000 per baited with 1-octen-3-ol (release rate 5 mg/hr)28 and 500 gm annum.7 In Cairns the majority of cases of human disease of dry ice. Twenty trapping sites were used, 14 within the occur during February to April (Tulip F, unpublished data). city of Cairns. Trapping was on 12 nights during 1996 (early In Australia the natural reservoir hosts for RR virus are February to late March), 6 nights during 1997 (early Feb- kangaroos and wallabies but other species, including horses, ruary to early April), and 1 night during 1998 (mid Febru- may act as urban reservoirs for human infection.3,8–10 Sero- ary) for a total 60 trap-nights. Two trap-nights were within logical surveys and virus isolation from mosquitoes trapped suburban yards, otherwise trapping was in Melaleuca near a flying fox camp suggested flying foxes might be res- swamps and other natural habitats in and around Cairns. ervoir hosts.8,11,12 However, Ryan and others concluded that One of the trapping sites contained a flying-fox camp with the gray-headed flying-fox, Pteropus poliocephalus, was not about 15,000 spectacled flying-fox, P. conspicillatus (Olson an important reservoir host because only 10 of 510 (2%) A, unpublished data). In the camp the swamp canopy is Aedes vigilax that fed on infected flying-foxes were infected dominated by the paperbark Melaleuca quinquenervia, with with RR virus after an extrinsic incubation period, and be- Pandanus sp., and Archontophoenix alexandrae (Warming- cause RR virus could not be detected in any of 122 blood- ton D, unpublished data). Trapping was performed in this fed Ae. vigilax immediately after feeding on infected P. po- camp in early February 1996 and mid February to early liocephalus.13 April 1997 for a total of 9 trap-nights. Trapping was also The major vectors of RR virus in Australia are considered performed in a swampy area approximately 600 meters from to be Culex annulirostris, Ae. vigilax, and Aedes camptor- the camp on nights in mid March 1996 and early February hynchus.9,14–18 Ross River virus has been isolated from 27 to mid March 1997 for a total of 11 trap-nights. In order to mosquito species in Australia comprising 19 Aedes, 2 compare RR virus isolation rates with other trapping sites Anopheles, 1 Coquilletidia, 5 Culex, 1 Mansonia, 3 undes- results from these 2 flying-fox camp associated sites were cribed species, and an unidentified species of Triptero- pooled. ides.12,16,19–21 However, the vector status of most of these is On 21 days during mid February to early May 1997 day- unknown.14 Aedes polynesiensis and Aedes aegypti may have time sampling for Ae. aegypti was conducted in, around or transmitted RR virus in a large epidemic in the South Pacific under houses using either a hand-held battery-powered as- in the late 1970s and early 1980s.22,23 There is laboratory pirator29 or a sweep-net. evidence that Ae. aegypti can be infected with and transmit Virus isolation. Sweep-net, aspirator, and CDC trapped RR virus, however RR virus has not been isolated from this mosquitoes were identified by species, pooled in lots of up species in the field.24–26 The goals of the study reported in to 100 individuals, and stored at Ϫ70ЊC prior to transport on 561 562 HARLEY AND OTHERS TABLE 1 Ross River virus isolates from mosquitoes collected in Cairns, Queensland, Australia from February 1996–February 1998 No. No. of No. of MIR/1,000 Mosquito species processed pools isolates mosquitoesa Aedes aegypti 121 18 0 0.0 Aedes alboscutellatus 423 16 0 0.0 Aedes alternans 2 2 0 0.0 Aedes aurantius 50 8 0 0.0 Aedes imprimensc 99 19 1 10.3 Aedes kochi 11,405 176 2 0.2 Aedes lineatopennis 1 1 0 0.0 Aedes littlechildi 3 1 0 0.0 Aedes normanensis 2 1 0 0.0 Aedes notoscriptus 637 46 1 1.6 Aedes palmarum 37 17 0 0.0 Aedes quasirubrithorax 3 2 0 0.0 Aedes tremulus 45 22 0 0.0 Aedes tremulus (male) 15 5 0 0.0 Aedes vigilax 3,308 103 1 0.3 Aedes vittiger 11 3 0 0.0 Anopheles annulipes 4 2 0 0.0 Anopheles bancroftii 24 8 0 0.0 Anopheles farauti 402 31 0 0.0 Bironella simmondsi 8 5 0 0.0 Coquillettidia crassipes 22 12 0 0.0 Culex annulirostris 30,541 378 9 0.3 Culex annulirostris (male) 5 3 0 0.0 Culex bitaeniorhynchus 4 1 0 0.0 Culex cubiculi 31 8 0 0.0 Culex gelidusc 257 17 1 4.0 Culex hilli 15 3 0 0.0 Culex pullus 37 14 0 0.0 Culex quinquefasciatus 50 13 0 0.0 Culex sitiens 250 21 0 0.0 Culex starckeae 2 2 0 0.0 Mansonia septempunctatac 913 44 3 5.8 Mansonia uniformis 52 13 0 0.0 Mansonia uniformis (male) 1 1 0 0.0 Tripteroides magnesianus 1 1 0 0.0 Tripteroides sp. 10 2 0 0.0 Uranotaenia pygmaea 5 1 0 0.0 Uranotaenia sp. 35 5 0 0.0 Verrallina carmentib,c 6,146 124 14 2.4 Verrallina funereus 975 42 0 0.0 Verrallina lineatusc 4,644 110 1 0.2 Verrallina lineatus (male) 1 1 0 0.0 Unidentified 22 4 0 0.0 Total 60,619 1,306 33 0.6 a Minimum infection rate (MIR) after Chiang and Reeves.30 b Includes 6 isolates from a preliminary study.39 c First recorded isolate from this species. dry ice to the Centre for Public Health Sciences in Brisbane, were Alphavirus non-reactive were discarded. Repeat isola- Queensland, Australia. Blood-fed mosquitoes were not pro- tions were performed for confirmation. cessed for virus isolation. The method used to isolate virus Minimum infection rates (MIRs) per 1,000 mosquitoes was described by Ritchie and others.12 Pools of up to 25 were calculated using the method of Chiang and Reeves.30 mosquitoes were homogenized by hand in 2 mL cold RPMI- 1640 (Roswell Park Memorial Institute medium) containing RESULTS 0.2% bovine serum albumin. The homogenates were then centrifuged; 100 ␮L of supernatant was inoculated onto con- A total of 60,619 mosquitoes encompassing 8 genera and fluent monolayers of C6–36 (Aedes albopictus) cells in 25 35 species were processed for virus isolation. Most (60,473) cm2 tissue culture flasks and incubated at 28ЊC. Day 3–5 were from CDC traps and the remainder (121 Ae. aegypti post-inoculation cells were scraped from the flask onto mi- and 25 Culex quinquefasciatus) were from household sweep- croscope slides and air-dried. The cells were examined by net and aspirator sampling. A total of 33 isolates, all RR indirect immunofluorescence using the following monoclo- virus, were obtained (Table 1).
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