Field evaluations of disposable sticky lures for surveillance of Aedes aegypti (Stegomyia aegypti) and Culex quinquefasciatus in Jakarta.

From December 1997 to April 1998, disposable sticky lures (1608 lure days) were trialled in homes in north Jakarta, Indonesia as surveillance tools for Aedes aegypti (Stegomyia aegypti) (Diptera: Culicidae) and Culex quinquefasciatus (Diptera: Culicidae), referenced to indoor resting adult collections (92 × 10 min). The lures collected 89.4% of the total of 1339 Ae. aegypti and 92.1% of the total of 1272 Cx. quinquefasciatus collected by all methods. Because there were no significant differences with respect to numbers collected in bedrooms, living rooms and kitchens, bedrooms were selected for subsequent trials for reasons of convenience. The main trials involved a replicated complete block design with L-lysine and sodium carbonate. Lures without attractant or with four different dilutions of L-lysine collected 3.4-8.5 times more Ae. aegypti and 4.2-8.1 times more Cx. quinquefasciatus than were collected by mouth aspirator. Lures with or without dilutions of sodium carbonate collected 2.7-5.0 times more Ae. aegypti and 1.8-4.2 times more Cx. quinquefasciatus than aspirator collections. The precision associated with catches of sticky lures was better than that for aspirator collections. Although olfactants generally improved the numbers of mosquitoes collected, the differences in catch between lures with and without attractants were usually non-significant. Any deficit in catch may be offset by increasing the surveillance period to ≥30 days to detect all four dengue serotypes from infected mosquitoes.

Evaluations of mosquito age grading techniques based on morphological changes.

Evaluations were made of the accuracy and practicality of mosquito age grading methods based on changes to mosquito morphology; including the Detinova ovarian tracheation, midgut meconium, Polovodova ovariole dilatation, ovarian injection, and daily growth line methods. Laboratory maintained Aedes vigilax (Skuse) and Culex annulirostris (Skuse) females of known chronological and physiological ages were used for these assessments. Application of the Detinova technique to laboratory reared Ae. vigilax females in a blinded trial enabled the successful identification of nulliparous and parous females in 83.7-89.8% of specimens. The success rate for identifying nulliparous females increased to 87.8-98.0% when observations of ovarian tracheation were combined with observations of the presence of midgut meconium. However, application of the Polovodova method only enabled 57.5% of nulliparous, 1-parous, 2-parous, and 3-parous Ae. vigilax females to be correctly classified, and ovarian injections were found to be unfeasible. Poor correlation was observed between the number of growth lines per phragma and the calendar age of laboratory reared Ae. vigilax females. In summary, morphological age grading methods that offer simple two-category predictions (ovarian tracheation and midgut meconium methods) were found to provide high-accuracy classifications, whereas methods that offer the separation of multiple age categories (ovariolar dilatation and growth line methods) were found to be extremely difficult and of low accuracy. The usefulness of the morphology-based methods is discussed in view of the availability of new mosquito age grading techniques based on cuticular hydrocarbon and gene transcription changes.

Sublethal effects of mosquito larvicides on swimming performance of larvivorous fish Melanotaenia duboulayi (Atheriniformes: Melanotaeniidae).

Laboratory studies were conducted to determine the sublethal effects of exposure to selected larvicides on the critical swimming speed (Ucrit) of crimson-spotted rainbowfish, Melanotaenia duboulayi (Castlenau). This native fish is common throughout southeastern Queensland, and it is increasingly being distributed as a biological control agent of mosquitoes. The selected larvicides included, two organophosphate (OP) compounds (temephos and pirimiphos-methyl), two microbial larvicides (Bacillus thuringiensis spp. israelensis [Bti] de Barjac and Bacillus sphaericus [Bs] Neide), and an insect growth regulator (IGR) (s-methoprene). Exposure to the OP temephos at 10 times the effective field concentration (EFC; 0.33 mg/liter), and OP pirimiphos-methyl at the EFC (0.50 mg/liter), resulted in a significant reduction in the Ucrit of M. duboulayi under controlled conditions. Conversely, exposure to the microbial (Bti and Bs) and IGR (s-methoprene) larvicides at 10 times the EFC had no effect on the Ucrit of M. duboulayi. Accordingly, these products are suitable for integrated pest management programs in Australia.

Ross River virus disease clusters and spatial relationship with mosquito biting exposure in Redland Shire, southern Queensland, Australia.

The spatial heterogeneity in the risk of Ross River virus (family Togaviridae, genus Alphavirus, RRV) disease, the most common mosquito-borne disease in Australia, was examined in Redland Shire in southern Queensland, Australia. Disease cases, complaints from residents of intense mosquito biting exposure, and human population data were mapped using a geographic information system. Surface maps of RRV disease age-sex standardized morbidity ratios and mosquito biting complaint morbidity ratios were created. To determine whether there was significant spatial variation in disease and complaint patterns, a spatial scan analysis method was used to test whether the number of cases and complaints was distributed according to underlying population at risk. Several noncontiguous areas in proximity to productive saline water habitats of Aedes vigilax (Skuse), a recognized vector of RRV, had higher than expected numbers of RRV disease cases and complaints. Disease rates in human populations in areas which had high numbers of adult Ae. vigilax in carbon dioxide- and octenol-baited light traps were up to 2.9 times those in areas that rarely had high numbers of mosquitoes. It was estimated that targeted control of adult Ae. vigilax in these high-risk areas could potentially reduce the RRV disease incidence by an average of 13.6%. Spatial correlation was found between RRV disease risk and complaints from residents of mosquito biting. Based on historical patterns of RRV transmission throughout Redland Shire and estimated future human population growth in areas with higher than average RRV disease incidence, it was estimated that RRV incidence rates will increase by 8% between 2001 and 2021. The use of arbitrary administrative areas that ranged in size from 4.6 to 318.3 km2 has the potential to mask any small scale heterogeneity in disease patterns. With the availability of georeferenced data sets and high-resolution imagery, it is becoming more feasible to undertake spatial analyses at relatively small scales.

Field evaluation of the establishment potential of wMelPop Wolbachia in Australia and Vietnam for dengue control.

BACKGROUND: Introduced Wolbachia bacteria can influence the susceptibility of Aedes aegypti mosquitoes to arboviral infections as well as having detrimental effects on host fitness. Previous field trials demonstrated that the wMel strain of Wolbachia effectively and durably invades Ae. aegypti populations. Here we report on trials of a second strain, wMelPop-PGYP Wolbachia, in field sites in northern Australia (Machans Beach and Babinda) and central Vietnam (Tri Nguyen, Hon Mieu Island), each with contrasting natural Ae. aegypti densities. METHODS: Mosquitoes were released at the adult or pupal stages for different lengths of time at the sites depending on changes in Wolbachia frequency as assessed through PCR assays of material collected through Biogents-Sentinel (BG-S) traps and ovitraps. Adult numbers were also monitored through BG-S traps. Changes in Wolbachia frequency were compared across hamlets or house blocks. RESULTS: Releases of adult wMelPop-Ae. aegypti resulted in the transient invasion of wMelPop in all three field sites. Invasion at the Australian sites was heterogeneous, reflecting a slower rate of invasion in locations where background mosquito numbers were high. In contrast, invasion across Tri Nguyen was relatively uniform. After cessation of releases, the frequency of wMelPop declined in all sites, most rapidly in Babinda and Tri Nguyen. Within Machans Beach the rate of decrease varied among areas, and wMelPop was detected for several months in an area with a relatively low mosquito density. CONCLUSIONS: These findings highlight challenges associated with releasing Wolbachia-Ae. aegypti combinations with low fitness, albeit strong virus interference properties, as a means of sustainable control of dengue virus transmission.

Vaccines against arthropods.

The possibility of vaccinating hosts against blood-feeding arthropods using antigens derived from salivary gland, gut, and other tissues is reviewed. These vaccines directed against vector arthropods also have the potential to effect the arthropods capacity to transmit pathogens, and this is distinct from transmission-blocking vaccines that use antigens derived from pathogens. Antigen extracts have been used in attempts to vaccinate against fleas, lice, keds, flies, mosquitoes, and a number of tick species. A vaccine against the cattle tick, Boophilus microplus (Canestrini), using a recombinant antigen, has been tested under field conditions. Ticks feeding on vaccinated hosts are damaged by an immune response directed against their gut cells. Some die on the host, others engorge but their fecundity is reduced. The Commonwealth Scientific Industrial Research Organization-Biotechnology Australia tick vaccine against B. microplus is cited as a model for the development of other vaccines. It is suggested that the weaker effects of vaccines against insects as compared with ticks are related to the different structure and physiologies of the gut rather than being related to time spent on the vertebrate host. These differences in the effects of vaccines on insects may favor vaccines which block the passage of pathogens into vector insects. Vaccines against mosquitoes have been shown to reduce susceptibility of mosquitoes to arboviruses. The potential of the different vaccines is discussed.

Aedes aegypti survival and dispersal estimated by mark-release-recapture in northern Australia.

The survival and dispersal of adult Aedes aegypti were estimated in northern Australia where sporadic outbreaks of Ae. aegypti-borne dengue viruses have occurred in recent years. Standard mark-release-recapture methods were used. In addition, a new sticky trap was used to capture the mosquitoes. Prior to the field study, the survival and effect of marking Ae. aegypti with fluorescent powder were determined in the laboratory. Mortality was age-dependent and the marked cohorts had higher survival rates than the untreated cohorts. Recapture rates of 13.0% and 3.6% over a seven-day period were achieved for two batches of marked Ae. aegypti released simultaneously at the field site. More males than females were recaptured although the proportion of females increased with time. The probability of daily survival was 0.91 and 0.86 for the blue- and pink-marked females, respectively, and 0.57 and 0.70 for the blue- and pink-marked and males, respectively. The mean distance traveled of recaptured Ae. aegypti was 56 m and 35 m for females and males, respectively. The maximum observed distance traveled of 160 m was the same for both sexes. The warm to hot and dry climatic conditions may have restricted the dispersal of released mosquitoes in this study. The frequency of recaptures at certain trap locations suggested that shade, wind, and the availability of hosts affected the distribution of Ae. aegypti within the study site.

The Premise Condition Index: a tool for streamlining surveys of Aedes aegypti.

Premise inspections and treatment of the larval habitats of container-breeding Aedes aegypti are extremely labor intensive. Since this means of control is the only one available in relation to dengue fever, this report presents an effective approach for streamlining premise surveys in north Queensland, Australia. From a survey of 877 premises in Townsville, Charters Towers, and Mingela/Ravenswood in 1990, occupier and premise variables were collected to examine any relationships with the presence of Ae. aegypti. Statistical modeling of these parameters using multiple and simple Poisson regression indicated that for both adjusted and unadjusted models respectively, the degree of shade and tidiness of the yard had strong correlations with both the proportion of positive premises and the numbers of infested containers therein. Condition of the house was also a significant variable in the unadjusted model. On this basis, the Premise Condition Index is proposed as a rapid assessment tool that can increase efficiency of detecting positive premises and containers by 270-370%. Although habitat selection by Aedes varies throughout the world, this approach could be used as a model for control of vectors of both dengue and yellow fever.

Understanding productivity, a key to Aedes aegypti surveillance.

The objective of this work was to define criteria that could be applied to achieve faster, more economical, and accurate assessment of vector populations for control of dengue viruses. During 1989-1990, 1,349 premises were surveyed in Townsville, Charters Towers and Mingela/Ravenswood, Queensland, Australia. In each locality, 1.9-8.4% of premises contained three or more containers with Aedes aegypti immature forms and were designated as key premises. Comparison of surveys in Townsville from 1989 to 1990 indicated that positive premises (i.e., those with at least one container with Ae. aegypti present) were 3.22 times more likely to remain positive than negative houses to become positive the following year. The Ae. aegypti population in Townsville was seen to be totally associated with garden receptacles, discarded household items, and trash but one well and one rainwater tank were responsible for 28% of all immature forms recorded in the 1,349 premises inspected. These breeding sites of high productivity were designated as key containers. At Charters Towers, Mingela, and Ravenswood, rainwater tanks were seen as the most important key container because although they constituted 13-29% of positive containers, they supported 60-63% of the immature forms. This study demonstrates that there is a certain degree of stability with regard to positive premises and that some of these, or some container types, contribute disproportionately to the Ae. aegypti population. Control programs could be made more efficient if efforts were concentrated on these sites of key vector productivity.

Investigation of gray-headed flying foxes (Pteropus poliocephalus) (Megachiroptera: Pteropodidae) and mosquitoes in the ecology of Ross River virus in Australia.

Entomologic and virologic factors were investigated to determine whether gray-headed flying foxes (Pteropus poliocephalus) from Indooroopilly Island, Brisbane, Australia could be vertebrate hosts of Ross River (RR) virus. Aedes funereus was the most abundant mosquito species with 6,300-38,700 females per light trap night in the flying fox camp containing gray-headed, black (P. alecto), and little red (P. scapulatus) flying foxes. Sixteen Ae. funereus blood meals from this collection were analyzed by hemoglobin electrophoresis and were found to be from P. alecto. From pledget feeding with RR virus, the infectious dose required to infect 50% of wild caught Ae. funereus was log10 4.2 50% tissue culture infectious doses per mosquito, with a transmission rate to mice of 17% at 9-10 days post infection. Experimental infection of 10 juvenile P. poliocephalus produced viremias of low titer in five animals, with a duration of 1-4 days and a mean of two days. Three percent of colonized Ae. vigilax that fed on the 10 animals during this period became infected. One of the five viremic flying foxes and two of the five aviremic animals produced a detectable immune response by either neutralization or hemagglutination-inhibition tests. Based on the low to moderate vector competence of Ae. funereus for RR virus, and evidence that P. poliocephalus is a poor vertebrate host of RR virus, it is unlikely that RR virus transmission would be maintained between these two species, but it could be maintained by other more competent vector/host pairs.

Vector competence of Culex pipiens quinquefasciatus for Murray Valley encephalitis, Kunjin, and Ross River viruses from Australia.

Australian populations of Culex pipiens quinquefasciatus Say from Brisbane, Charleville, Cairns, Kowanyama (Queensland), Darwin (Northern Territory), Mildura (Victoria), and Port Hedland (Western Australia) proved to be either poorly susceptible or refractory to oral infection with low passage level Murray Valley encephalitis, Kunjin, and Ross River viruses. With respect to past or future epidemics of Murray Valley encephalitis or Kunjin (both known to cause clinical encephalitis in man), and of epidemic polyarthritis, this mosquito is viewed as an unlikely vector.

Critical examination of Aedes aegypti indices: correlations with abundance.

The following immature stage indices for Aedes (Stegomyia) aegypti surveillance were evaluated in four north Queensland, Australia towns with respect to their relationship to immature and adult female densities: Breteau, House, Container, Larval Density, Stegomyia (and modifications thereof), and a newly created Adult Productivity Index. Spearman's correlations of indices that considered larval or immature (larvae and pupae) numbers had a better relationship with immature abundance but this was not necessarily the case against adult abundance. To examine the robustness of the indices, data from 758 premises in Townsville, Charters Towers, Ravenswood, and Mingela were pooled and 30 random subsamples, each consisting of 50 premises were taken. After each subsample was taken, the premises selected were reintroduced into the original data bank of 758 premises, and therefore, were available for further selection, i.e., sampling with replacement. Indices were calculated for each of the 30 subsamples and the coefficients of variation of each index were estimated from these. The Breteau, Adult Productivity, House, and Adult density indices proved to have the smallest coefficients compared with index size. No alternate index was regarded as being superior to the Breteau, including the Adult Productivity Index measuring both container type frequency and immature density. For this reason and in view of the labor intensiveness of estimating immature indices that incorporate productivity, it is recommended that new and cost-effective methods of adult surveillance be pursued.

Aedes aegypti control in the Lao People's Democratic Republic, with reference to copepods.

An evaluation of the Lao Aedes aegypti control program and of the predatory abilities of copepods from Vientiane, Lao People's Democratic Republic was undertaken before a field release of copepods in Thongkankam village, Vientiane. Copepods were transported to Australia for evaluation of predatory abilities and their survival under various nutrient and pH conditions. Mesocyclops guangxiensis was chosen for release over M. aspericornis due to its higher reproduction rate and its ability to survive in lower nutrient environments. Mesocyclops guangxiensis was released into 142 containers and 20 wells in a village in Vientiane. Copepods were present in 7% of the containers after one month and were absent six months postinoculation. In comparison, 100% of wells were still positive after six months, with average numbers of Ae. aegypti in the wells decreasing from 59.5 +/- 18.5 (+/- SEM) to 0 after six months. Numbers of Culex quinquefasciatus and Anopheles maculatus also decreased to 0 after six months. This study indicates that predacious copepods will be accepted by the community and could be integrated as a low-cost, persistent control agent into new strategies for improving surveillance and control of dengue vectors.

National progress in dengue vector control in Vietnam: survey for Mesocyclops (Copepoda), Micronecta (Corixidae), and fish as biological control agents.

This paper describes the process of expanding a successful dengue control program in 3 provinces in northern Vietnam into a national one and demonstrates the presence of a rich, low-cost resource that could have similar applicability to other countries in the region. The cornerstone of the preventive strategy is larval control of Aedes aegypti (L.), the major vector, using predators such as copepods, Mesocyclops spp., aided by the corixid bug Micronecta quadristrigata Bredd, and fish in large water storage containers. From 1989 to 1998, 9 species of Mesocyclops (M. woutersi Van de Velde, M. aspericornis (Daday), M. ruttneri Kiefer, M. thermocyclopoides Harada, M. affinis Van de Velde, M. ogunnus Onabamiro, M. yenae Holynska, M. cf. pehpeiensis Hu, and M. dissimilis Defaye and Kawabata) were found in natural and artificial habitats in 26 provinces throughout Vietnam. The predatory capacities of 6 of these were evaluated in the laboratory. This indicated that daily consumption/killing averaged between 16 and 41 Ae. aegypti larvae per copepod. From detailed evaluations in 9 provinces, Mesocyclops spp. were surprisingly common in 8,413 artificial containers (concrete tanks, wells, ornamental ponds and in the south, large jars). Because of existing practices for washing and water transfer from ponds and lakes in Ha Tay and Ha Bac, Mesocyclops spp. already occurred in 60-100% of the water storage containers. When the relationship between the presence or absence of Mesocyclops and Aedes larvae in 5,111 containers was analyzed by the chi-square test, their distributions were significantly related, indicating control (odds ratio = 0.56). When 3,426 containers that did not contain Mesocyclops or fish were analyzed in relation to the distribution of Aedes larvae, those with Micronecta also had significantly less Aedes (odds ratio = 0.43). Therefore, this study demonstrates that there is an abundance of local Mesocyclops spp. in Vietnam that can be incorporated into specifically designed community-based control programs aided by Micronecta and fish.

Eradication of Aedes aegypti from a village in Vietnam, using copepods and community participation.

In northern Vietnam, copepods of the genus Mesocyclops were used for biological control of Aedes aegypti, the principal vector of dengue viruses, by inoculation into wells, large cement tanks, ceramic jars, and other domestic containers that served as Ae. aegypti breeding sites. The use of Mesocyclops was complemented by community participation with respect to recycling to eliminate unused and discarded containers that collected rainwater and provided Ae. aegypti breeding sites that could not be treated effectively with Mesocyclops. Aedes aegypti disappeared from 400 houses of the treated village in August 1994 and has not reappeared, a result of particular significance, because there are virtually no other recorded instances of eradicating this mosquito anywhere in the world during the past 25 years, and certainly not with community-based approaches. When used in combination with community recycling, Mesocyclops is an easy and inexpensive method of Ae. aegypti control that should be effective for many communities in Vietnam and elsewhere.

Experimental infection of Australian brushtail possums, Trichosurus vulpecula (Phalangeridae: Marsupialia), with Ross River and Barmah Forest viruses by use of a natural mosquito vector system.

Brushtail possums, Trichosurus vulpecula Kerr, were experimentally infected with Ross River (RR) or Barmah Forest (BF) virus by Aedes vigilax (Skuse) mosquitoes. Eight of 10 animals exposed to RR virus developed neutralizing antibody, and 3 possums developed high viremia for < 48 hr after infection, sufficient to infect recipient mosquitoes. Two of 10 animals exposed to BF virus developed neutralizing antibody. Both infected possums maintained detectable neutralizing antibody to BF for at least 45 days after infection (log neutralization index > 2.0 at 45 days). Eight possums did not develop neutralizing antibody to BF despite exposure to infected mosquitoes. These results suggest that T. vulpecula may potentially act as a reservoir species for RR in urban areas. However, T. vulpecula infected with BF do not develop viremia sufficient to infect mosquitoes and are unlikely to be important hosts for BF.

Isolation of arboviruses (Kemerovo group, Sakhalin group) from Ixodes uriae collected at Macquarie Island, Southern ocean.

Eighty-nine pools of ticks (Ixodes [Ceratixodes] uriae White) collected at Macquarie Island (54 degrees 30' S, 159 degrees E) in the Southern Ocean yielded 16 strains of 2 viruses, 1 a "new" member of the Kemerovo group and 1 a "new" member of the Sakhalin group. The names "Nugget" and "Taggert" are proposed for them. Antigenically-related viruses have therefore now been isolated from I. uriae in both subarctic and subantarctic regions.

Identification of Australian arboviruses in inoculated cell cultures using monoclonal antibodies in ELISA.

An ELISA using a panel of specific monoclonal antibodies was developed to identify all alpha and flaviviruses isolated from mosquitoes caught throughout Australia. This technique is sensitive and rapid and is more specific than the traditional methods used to identify flaviviruses. The ability to identify unknown virus isolates from field-caught mosquitoes quickly and accurately improves the efficiency of arbovirus surveillance programs and allows health authorities to give an early warning of an increased health risk from a mosquito-borne virus in a particular region.

Vector competence of Aedes aegypti, Culex sitiens, Culex annulirostris, and Culex quinquefasciatus (Diptera: Culicidae) for Barmah Forest virus.

Aedes aegypti (L.), Culex sitiens Weidemann, Culex annulirostris Skuse, and Culex quinquefasciatus Say mosquitoes colonized at the Queensland Institute of Medical Research, Brisbane Australia, were fed on blood containing Barmah Forest virus (BF). Only Cx. annulirostris was susceptible to infection, with a median cell culture infectious dose (CCID50) of 10(3.36) per mosquito. Ae. aegypti and Cx. quinquefasciatus were infected experimentally, but at rates of < 9%. Cx. sitiens did not become infected. Infection rates for Cx. annulirostris fed 10(3.5) CCID50 of virus per mosquito, varied from 9 to 50% between 2 and 13 d after infection. Virus transmission to suckling mice by Cx. annulirostris occurred from 2 d after infection. Transmission of BF virus by Cx. annulirostris was 10% at 2 d after infection and did not exceed 8% thereafter. Although Cx. annulirostris may be infected and is able to transmit BF virus to suckling mice, it is nonetheless a relatively inefficient vector of the virus.

Mosquito (Diptera: Culicidae) dispersal: implications for the epidemiology of Japanese and Murray Valley encephalitis viruses in Australia.

One hypothesis to explain the southern extension of Japanese encephalitis (JE) virus from Papua New Guinea into the Torres Strait islands in 1995 and to mainland Australia in 1998 is the dispersal of infected mosquitoes, particularly Culex annulirostris Skuse from which JE virus has been isolated repeatedly. To investigate whether this species disperses in this manner, mosquitoes were identified from 368 aerial kite trap collections operated at 50-310 m (altitude) at inland New South Wales between November 1979 to December 1984. Forty samples (9 during daylight and 31 at night) contained mosquitoes, of which 221 could be identified as Culex australicus Dobrotworsky & Drummond (58.8%), Culex annulirostris (21.3%), Anopheles annulipes Walker s.l. (10.4%), Aedes theobaldi (Taylor) (7.2%), Aedes rubrithorax (Macquart) (1.4%), and Aedes sagax (Skuse) (< 0.9%). During the night, mosquitoes were found in 22.6% of the collections at a mean density (+/- SD) of 91.3 +/- 151.7/10(6) m3 of air sampled. During the day, only 3.8% were positive at a mean density 125.3 +/- 152.1. When examined in relation to possible flying time and wind speed, mean +/- SD dispersal distances by day and night were 23.9 +/- 15.3 km and 152.4 +/- 116.3 km, respectively. These data provide circumstantial evidence that aerial carriage southward approximately 200 km from Papua New Guinea to Cape York peninsula is feasible, but that southern dispersal of Murray Valley encephalitis virus infected mosquitoes from tropical to temperate Australia is unlikely.