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Aedes aegypti is the principal mosquito vector of dengue, yellow fever, Zika and chikungunya viruses. The wMel strain of the endosymbiotic bacteria Wolbachia pipientis was introduced into the vector as a novel biocontrol strategy to stop transmission of these viruses. Mosquitoes with Wolbachia have been released in the field in Northern Queensland, Australia since 2011, at various locations and over several years, with populations remaining stably infected. Wolbachia infection is known to alter gene expression in its mosquito host, but whether (and how) this changes over the long-term in the context of field releases remains unknown. We sampled mosquitoes from Wolbachia-infected populations with three different release histories along a time gradient and performed RNA-seq to investigate gene expression changes in the insect host. We observed a significant impact on gene expression in Wolbachia-infected mosquitoes versus uninfected controls. Fewer genes had significantly upregulated expression in mosquitoes from the older releases (512 and 486 from the 2011 and 2013/14 release years, respectively) versus the more recent releases (1154 from the 2017 release year). Nonetheless, a fundamental signature of Wolbachia infection on host gene expression was observed across all releases, comprising upregulation of immunity (e.g. leucine-rich repeats, CLIPs) and metabolism (e.g. lipid metabolism, iron transport) genes. There was limited downregulation of gene expression in mosquitoes from the older releases (84 and 71 genes from the 2011 and 2013/14 release years, respectively), but significantly more in the most recent release (509 from the 2017 release year). Our findings indicate that at > 8 years post-introgression into field populations, Wolbachia continues to profoundly impact expression of host genes, such as those involved in insect immune response and metabolism. If Wolbachia-mediated virus blocking is underpinned by these differential gene expression changes, our res

With global expansion of the two main vectors of dengue, Aedes aegypti (Linnaeus, Diptera: Culicidae) and Aedes albopictus (Skuse, Diptera: Culicidae), there is a need to further develop cost-effective and user-friendly surveillance tools to monitor the population dynamics of these species. The abundance of Ae. aegypti and Ae. Albopictus, and associated bycatch captured by Male Aedes Sound Traps (MASTs) and BG-Sentinel (BGS) traps that were unbaited or baited with BG-Lures were compared in Cairns, Australia and Madang, Papua New Guinea. Mean male Ae. aegypti and Ae. albopictus catch rates in MASTs did not significantly differ when deployed with BG-Lures. Similarly, males of both these species were not sampled at statistically different rates in BGS traps with or without BG-Lures. However, MASTs with BG-Lures caught significantly less male Ae. aegypti than BGS traps baited with BG-Lures in Cairns, and MASTs without BG-Lures caught significantly more male Ae. albopictus than BGS traps without BG-Lures in Madang. Additionally, BG-Lures significantly increased female Ae. aegypti catch rates in BGS traps in Cairns. Lastly, bycatch capture rates in BGS traps were not significantly influenced by the addition of the BG-Lures. While this study provides useful information regarding the surveillance of Ae. aegypti and Ae. albopictus in these locations, further development and investigation is required to successfully integrate an olfactory lure into the MAST system.

Aedes aegypti and Aedes albopictus vector dengue, chikungunya and Zika viruses. With both species expanding their global distributions at alarming rates, developing effective surveillance equipment is a continuing priority for public health researchers. Sound traps have been shown, in limited testing, to be highly species-specific when emitting a frequency corresponding to a female mosquito wingbeat. Determining male mosquito capture rates in sound traps based on lure frequencies in endemic settings is the next step for informed deployment of these surveillance tools. We field-evaluated Male Aedes Sound Traps (MASTs) set to either 450 Hz, 500 Hz, 550 Hz or 600 Hz for sampling Aedes aegypti and/or Aedes albopictus and compared catch rates to BG-Sentinel traps within Pacific (Madang, Papua New Guinea) and Latin American (Molas, Mexico and Orange Walk Town, Belize) locations. MASTs set to 450-550 Hz consistently caught male Ae. aegypti at rates comparable to BG-Sentinel traps in all locations. A peak in male Ae. albopictus captures in MASTs set at 550 Hz was observed, with the lowest mean abundance recorded in MASTs set to 450 Hz. While significantly higher abundances of male Culex were sampled in MASTs emitting lower relative frequencies in Molas, overall male Culex were captured in significantly lower abundances in the MASTs, relative to BG-Sentinel traps within all locations. Finally, significant differences in rates at which male Aedes and Culex were positively detected in trap-types per weekly collections were broadly consistent with trends in abundance data per trap-type. MASTs at 550 Hz effectively captured both male Ae. aegypti and Ae. albopictus while greatly reducing bycatch, especially male Culex, in locations where dengue transmission has occurred. This high species-specificity of the MAST not only reduces staff-time required to sort samples, but can also be exploited to develop an accurate smart-trap system-both outcomes potentially reducing public health

A dengue suppression strategy based on release of Aedes aegypti mosquitoes infected with the bacterium Wolbachia pipientis is being trialed in many countries. Wolbachia inhibits replication and transmission of dengue viruses. Questions remain regarding the long-term stability of virus-suppressive effects. We sequenced the Wolbachia genome and analyzed Ae. aegypti mitochondrial DNA markers isolated from mosquitoes sampled 2-8 years after releases in the greater Cairns region, Australia. Few changes were detected when Wolbachia genomes of field mosquitoes were compared with Wolbachia genomes of mosquitoes obtained soon after initial releases. Mitochondrial variants associated with the initial Wolbachia release stock are now the only variants found in release sites, highlighting maternal leakage as a possible explanation for rare Wolbachia-negative mosquitoes and not migration from non-release areas. There is no evidence of changes in the Wolbachia genome that indicate selection against its viral-suppressive effects or other phenotypes attributable to infection with the bacterium.

Background The Wolbachia incompatible insect technique (IIT) shows promise as a method for eliminating populations of invasive mosquitoes such as Aedes aegypti (Linnaeus) (Diptera: Culicidae) and reducing the incidence of vector-borne diseases such as dengue, chikungunya and Zika. Successful implementation of this biological control strategy relies on high-fidelity separation of male from female insects in mass production systems for inundative release into landscapes. Processes for sex-separating mosquitoes are typically error-prone and laborious, and IIT programmes run the risk of releasing Wolbachia-infected females and replacing wild mosquito populations. Results We introduce a simple Markov population process model for studying mosquito populations subjected to a Wolbachia-IIT programme which exhibit an unstable equilibrium threshold. The model is used to study, in silico, scenarios that are likely to yield a successful elimination result. Our results suggest that elimination is best achieved by releasing males at rates that adapt to the ever-decreasing wild population, thus reducing the risk of releasing Wolbachia-infected females while reducing costs. Conclusions While very high-fidelity sex separation is required to avoid establishment, release programmes tend to be robust to the release of a small number of Wolbachia-infected females. These findings will inform and enhance the next generation of Wolbachia-IIT population control strategies that are already showing great promise in field trials.

Aedes aegypti mosquitoes infected with the wMel strain of Wolbachia are being released into natural mosquito populations in the tropics as a way of reducing dengue transmission. High temperatures adversely affect wMel, reducing Wolbachia density and cytoplasmic incompatibility in some larval habitats that experience large temperature fluctuations. We monitored the impact of a 43.6°C heatwave on the wMel infection in a natural population in Cairns, Australia, where wMel was first released in 2011 and has persisted at a high frequency. Wolbachia infection frequencies in the month following the heatwave were reduced to 83% in larvae sampled directly from field habitats and 88% in eggs collected from ovitraps, but recovered to be near 100% four months later. Effects of the heatwave on wMel appeared to be stage-specific and delayed, with reduced frequencies and densities in field-collected larvae and adults reared from ovitraps but higher frequencies in field-collected adults. Laboratory experiments showed that the effects of heatwaves on cytoplasmic incompatibility and density are life stage-specific, with first instar larvae being the most vulnerable to temperature effects. Our results indicate that heatwaves in wMel-infected populations will have only temporary effects on Wolbachia frequencies and density once the infection has established in the population. Our results are relevant to ongoing releases of wMel-infected Ae. aegypti in several tropical countries.

A promising strategy for reducing the transmission of dengue and other arboviral human diseases by Aedes aegypti mosquito vector populations involves field introductions of the endosymbiotic bacteria Wolbachia. Wolbachia infections inhibit viral transmission by the mosquito, and can spread between mosquito hosts to reach high frequencies in the vector population. Wolbachia spreads by maternal transmission, and spread dynamics can be variable and highly dependent on natural mosquito population dynamics, population structure and fitness components. We develop a mathematical model of an A. aegypti metapopulation that incorporates empirically validated relationships describing density‐dependent mosquito fitness components. We assume that density dependent relationships differ across subpopulations, and construct heterogeneous landscapes for which model‐predicted patterns of variation in mosquito abundance and demography approximate those observed in field populations. We then simulate Wolbachia release strategies similar to that used in field trials. We show that our model can produce rates of spatial spread of Wolbachia similar to those observed following field releases. We then investigate how different types of spatio‐temporal variation in mosquito habitat, as well as different fitness costs incurred by Wolbachia on the mosquito host, influence predicted spread rates. We find that fitness costs reduce spread rates more strongly when the habitat landscape varies temporally due to stochastic and seasonal processes. Synthesis and applications: Our empirically based modelling approach represents effects of environmental heterogeneity on the spatial spread of Wolbachia. The models can assist in interpreting observed spread patterns following field releases and in designing suitable release strategies for targeting spatially heterogeneous vector populations.

Wolbachia bacteria are now being introduced into Aedes aegypti mosquito populations for dengue control. When Wolbachia infections are at a high frequency, they influence the local transmission of dengue by direct virus blocking as well as deleterious effects on vector mosquito populations. However, the effectiveness of this strategy could be influenced by environmental temperatures that decrease Wolbachia density, thereby reducing the ability of Wolbachia to invade and persist in the population and block viruses. We reared wMel-infected Ae. aegypti larvae in the field during the wet season in Cairns, North Queensland. Containers placed in the shade produced mosquitoes with a high Wolbachia density and little impact on cytoplasmic incompatibility. However, in 50% shade where temperatures reached 39°C during the day, wMel-infected males partially lost their ability to induce cytoplasmic incompatibility and females had greatly reduced egg hatch when crossed to infected males. In a second experiment under somewhat hotter conditions (>40°C in 50% shade), field-reared wMel-infected females had their egg hatch reduced to 25% when crossed to field-reared wMel-infected males. Wolbachia density was reduced in 50% shade for both sexes in both experiments, with some mosquitoes cleared of their Wolbachia infections entirely. To investigate the critical temperature range for the loss of Wolbachia infections, we held Ae. aegypti eggs in thermocyclers for one week at a range of cyclical temperatures. Adult wMel density declined when eggs were held at 26-36°C or above with complete loss at 30-40°C, while the density of wAlbB remained high until temperatures were lethal. These findings suggest that high temperature effects on Wolbachia are potentially substantial when breeding containers are exposed to partial sunlight but not shade. Heat stress could reduce the ability of Wolbachia infections to invade mosquito populations in some locations and may compromise the ability of Wolbachi

BACKGROUND: Malaria is the most important vector-borne disease in the world. Epidemiological and ecological studies of malaria traditionally utilize detection of Plasmodium sporozoites in whole mosquitoes or salivary glands by microscopy or serological or molecular assays. However, these methods are labor-intensive, and can over- or underestimate mosquito transmission potential. To overcome these limitations, alternative sample types have been evaluated for the study of malaria. It was recently shown that Plasmodium could be detected in saliva expectorated on honey-soaked cards by Anopheles stephensi, providing a better estimate of transmission risk. We evaluated whether excretion of Plasmodium falciparum nucleic acid by An. stephensi correlates with expectoration of parasites in saliva, thus providing an additional sample type for estimating transmission potential. Mosquitoes were exposed to infectious blood meals containing cultured gametocytes, and excreta collected at different time points post-exposure. Saliva was collected on honey-soaked filter paper cards, and salivary glands were dissected and examined microscopically for sporozoites. Excreta and saliva samples were tested by real time polymerase chain reaction (RT-rtPCR). RESULTS: Plasmodium falciparum RNA was detected in mosquito excreta as early as four days after ingesting a bloodmeal containing gametocytes. Once sporogony (the development of sporozoites) occurred, P. falciparum RNA was detected concurrently in both excreta and saliva samples. In the majority of cases, no difference was observed between the Ct values obtained from matched excreta and saliva samples, suggesting that both samples provide equally sensitive results. A positive association was observed between the molecular detection of the parasites in both samples and the proportion of mosquitoes with sporozoites in their salivary glands from each container. No distinguishable parasites were observed when excreta samples were stained and m

Background: The wMel strain of Wolbachia has been successfully introduced into Aedes aegypti mosquitoes and subsequently shown in laboratory studies to reduce transmission of a range of viruses including dengue, Zika, chikungunya, yellow fever, and Mayaro viruses that cause human disease. Here we report the entomological and epidemiological outcomes of staged deployment of Wolbachia across nearly all significant dengue transmission risk areas in Australia. Methods: The wMel strain of Wolbachia was backcrossed into the local Aedes aegypti genotype (Cairns and Townsville backgrounds) and mosquitoes were released in the field by staff or via community assisted methods. Mosquito monitoring was undertaken and mosquitoes were screened for the presence of Wolbachia. Dengue case notifications were used to track dengue incidence in each location before and after releases. Results: Empirical analyses of the Wolbachia mosquito releases, including data on the density, frequency and duration of Wolbachia mosquito releases, indicate that Wolbachia can be readily established in local mosquito populations, using a variety of deployment options and over short release durations (mean release period 11 weeks, range 2-22 weeks). Importantly, Wolbachia frequencies have remained stable in mosquito populations since releases for up to 8 years. Analysis of dengue case notifications data demonstrates near-elimination of local dengue transmission for the past five years in locations where Wolbachia has been established. The regression model estimate of Wolbachia intervention effect from interrupted time series analyses of case notifications data prior to and after releases, indicated a 96% reduction in dengue incidence in Wolbachia treated populations (95% confidence interval: 84 - 99%). Conclusion: Deployment of the wMel strain of Wolbachia into local Ae. aegypti populations across the Australian regional cities of Cairns and most smaller regional communities with a past history of dengue h

Dengue-suppressing Wolbachia strains are promising tools for arbovirus control, particularly as they have the potential to self-spread following local introductions. To test this, we followed the frequency of the transinfected Wolbachia strain wMel through Ae. aegypti in Cairns, Australia, following releases at 3 nonisolated locations within the city in early 2013. Spatial spread was analysed graphically using interpolation and by fitting a statistical model describing the position and width of the wave. For the larger 2 of the 3 releases (covering 0.97 km2 and 0.52 km2), we observed slow but steady spatial spread, at about 100–200 m per year, roughly consistent with theoretical predictions. In contrast, the smallest release (0.11 km2) produced erratic temporal and spatial dynamics, with little evidence of spread after 2 years. This is consistent with the prediction concerning fitness-decreasing Wolbachia transinfections that a minimum release area is needed to achieve stable local establishment and spread in continuous habitats. Our graphical and likelihood analyses produced broadly consistent estimates of wave speed and wave width. Spread at all sites was spatially heterogeneous, suggesting that environmental heterogeneity will affect large-scale Wolbachia transformations of urban mosquito populations. The persistence and spread of Wolbachia in release areas meeting minimum area requirements indicates the promise of successful large-scale population transformation.

BACKGROUND: Arbovirus transmission by the mosquito Aedes aegypti can be reduced by the introduction and establishment of the endosymbiotic bacteria Wolbachia in wild populations of the vector. Wolbachia spreads by increasing the fitness of its hosts relative to uninfected mosquitoes. However, mosquito fitness is also strongly affected by population size through density-dependent competition for limited food resources. We do not understand how this natural variation in fitness affects symbiont spread, which limits our ability to design successful control strategies. RESULTS: We develop a mathematical model to predict A. aegypti-Wolbachia dynamics that incorporates larval density-dependent variation in important fitness components of infected and uninfected mosquitoes. Our model explains detailed features of the mosquito-Wolbachia dynamics observed in two independent experimental A. aegypti populations, allowing the combined effects on dynamics of multiple density-dependent fitness components to be characterized. We apply our model to investigate Wolbachia field release dynamics, and show how invasion outcomes can depend strongly on the severity of density-dependent competition at the release site. Specifically, the ratio of released relative to wild mosquitoes required to attain a target infection frequency (at the end of a release program) can vary by nearly an order of magnitude. The time taken for Wolbachia to become established following releases can differ by over 2 years. These effects depend on the relative fitness of field and insectary-reared mosquitoes. CONCLUSIONS: Models of Wolbachia invasion incorporating density-dependent demographic variation in the host population explain observed dynamics in experimental A. aegypti populations. These models predict strong effects of density-dependence on Wolbachia dynamics in field populations, and can assist in the effective use of Wolbachia to control the transmission of arboviruses such as dengue, chikungunya and

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.

Dengue viruses cause more human morbidity and mortality than any other arthropod-borne virus. Dengue prevention relies mainly on vector control; however, the failure of traditional methods has promoted the development of novel entomological approaches. Although use of the intracellular bacterium wolbachia to control mosquito populations was proposed 50 years ago, only in the past decade has its use as a potential agent of dengue control gained substantial interest. Here, we review evidence that supports a practical approach for dengue reduction through field release of wolbachia-infected mosquitoes and discuss the additional studies that have to be done before the strategy can be validated and implemented. A crucial next step is to assess the efficacy of wolbachia in reducing dengue virus transmission. We argue that a cluster randomised trial is at this time premature because choice of wolbachia strain for release and deployment strategies are still being optimised. We therefore present a pragmatic approach to acquiring preliminary evidence of efficacy through various complementary methods including a prospective cohort study, a geographical cluster investigation, virus phylogenetic analysis, virus surveillance in mosquitoes, and vector competence assays. This multipronged approach could provide valuable intermediate evidence of efficacy to justify a future cluster randomised trial.

The endosymbiotic bacteria Wolbachia pipientis (wMel strain) has been successfully established in several populations of Aedes aegypti, the primary dengue vector. The virulent Wolbachia strain wMelPop is known to cause several pathological impacts (increased egg mortality, life shortening, etc.) reducing overall fitness in the mosquito Ae. aegypti. Increased egg mortality could substantially reduce egg banks in areas with a lengthy monsoonal dry season, and be employed to eliminate local populations. We tested this application under semi-field cage conditions. First, we determined that wMelPop infection significantly reduced the survival of desiccation-resistant eggs of the dengue vector Ae. aegypti, with shade and temperature having a significant impact; nearly all wMelPop-infected eggs failed to hatch after 6 and 10 weeks in summer and winter conditions, respectively. In laboratory selection experiments we found that egg desiccation resistance can be increased by selection, and that this effect of wMelPop infection is due to the nuclear background of the host rather than Wolbachia. We then conducted an invasion of wMelPop within a semi-field cage using sustained weekly releases of wMelPop infected mosquitoes, with fixation achieved after 9 weeks. The egg populations wMelPop infected and an uninfected control were then subjected to a simulated prolonged monsoonal dry season (2.5 months) before flooding to induce hatching. The wMelPop infected eggs suffered significantly greater mortality than the controls, with only 0.67% and 4.35% of respective infected and uninfected eggs held in 99% shade hatching after 80 days. These studies suggest that wMelPop could be used to locally eliminate populations of Ae. aegypti that are exposed to prolonged dry conditions, particularly if combined with vector control.

BACKGROUND: Elimination of lymphatic filariasis (LF) in Samoa continues to be challenging despite multiple annual mass drug campaigns aimed at stopping transmission by reducing the prevalence and density of microfilaraemia. The persistence of transmission may be partly related to the highly efficient Aedes vectors. The assessment of pathogen transmission by mosquito vectors and of vector control relies on the ability to capture mosquitoes efficiently. The aims of this study are to compare trapping methods to capture LF-infected mosquitoes and determine the role in transmission of the species of Aedes mosquitoes in the area. METHODS: Fasitoo-Tai village was the chosen site because of persistent transmission despite annual mass drug administration. Sampling methods included BioGents Sentinel (BGS) trap, human-baited collections (HBC) and the Centers for Disease Control (CDC) trap. BGS and CDC traps were baited with BG-lure, CO2, and/or octenol. Individual trap locations were geo-located and efficiency of sampling methods was evaluated using a randomized Latin-square design in two locations. Number of mosquitoes collected (male and female), as well as species for each trapping method were determined. Additionally, Ae. polynesiensis and Ae. (Finlaya) spp. females were pooled by trap method and analysed for filarial DNA. Infection prevalence was estimated using the PoolScreen software. RESULTS: The BGS trap with any type of bait collected more mosquitoes compared to both the CDC trap and the HBC. The BGS trap baited with BG-lure collected more mosquitoes than with CO2 and octenol. There were no significant differences between trapping methods in terms of proportions of infected females collected. The prevalence of filarial infection in Ae. polynesiensis and Ae. (Finlaya) spp. was estimated at 4.7% and 0.67% respectively. CONCLUSIONS: This study supports the use of the BGS trap for research on and surveillance of the mosquito vectors of LF in Samoa. The BGS trap is a suit

BACKGROUND: Recent releases have been carried out with Aedes aegypti mosquitoes infected with the wMelPop mosquito cell-line adapted (wMelPop-CLA) strain of Wolbachia. This infection introduced from Drosophila provides strong blockage of dengue and other arboviruses but also has large fitness costs in laboratory tests. The releases were used to evaluate the fitness of released infected mosquitoes, and (following termination of releases) to test for any effects of wMelPop-CLA on wing size and shape when mosquitoes were reared under field conditions. METHODS: We monitored gravid females via double sticky traps to assess the reproductive success of wMelPop-CLA-infected females and also sampled the overall mosquito population post-release using Biogent Sentinel traps. Morphometric analyses were used to evaluate infection effects on wing shape as well as size. RESULTS: Oviposition success as assessed through double sticky traps was unrelated to size of released mosquitoes. However, released mosquitoes with lower wing loading were more successful. Furthermore, wMelPop-CLA-infected mosquitoes had 38.3% of the oviposition success of uninfected mosquitoes based on the predicted infection frequency after release. Environmental conditions affected wing shape and particularly size across time in uninfected mosquitoes, but not in naturally-reared wMelPop-CLA-infected mosquitoes. Although the overall size and shape do not differ between naturally-reared wMelPop-CLA-infected and uninfected mosquitoes, the infected mosquitoes tended to have smaller wings than uninfected mosquitoes during the cooler November in comparison to December. CONCLUSION: These results confirm the lower fitness of wMelPop-CLA infection under field conditions, helping to explain challenges associated with a successful invasion by this strain. In the long run, invasion may depend on releasing strains carrying insecticide resistance or egg desiccation resistance, combined with an active pre-release population s

BACKGROUND: The bacterial endosymbiont Wolbachia blocks the transmission of dengue virus by its vector mosquito Aedes aegypti, and is currently being evaluated for control of dengue outbreaks. Wolbachia induces cytoplasmic incompatibility (CI) that results in the developmental failure of offspring in the cross between Wolbachia-infected males and uninfected females. This increases the relative success of infected females in the population, thereby enhancing the spread of the beneficial bacterium. However, Wolbachia spread via CI will only be feasible if infected males are sufficiently competitive in obtaining a mate under field conditions. We tested the effect of Wolbachia on the competitiveness of A. aegypti males under semi-field conditions. METHODOLOGY/PRINCIPAL FINDINGS: In a series of experiments we exposed uninfected females to Wolbachia-infected and uninfected males simultaneously. We scored the competitiveness of infected males according to the proportion of females producing non-viable eggs due to incompatibility. We found that infected males were equally successful to uninfected males in securing a mate within experimental tents and semi-field cages. This was true for males infected by the benign wMel Wolbachia strain, but also for males infected by the virulent wMelPop (popcorn) strain. By manipulating male size we found that larger males had a higher success than smaller underfed males in the semi-field cages, regardless of their infection status. CONCLUSIONS/SIGNIFICANCE: The results indicate that Wolbachia infection does not reduce the competitiveness of A. aegypti males. Moreover, the body size effect suggests a potential advantage for lab-reared Wolbachia-males during a field release episode, due to their better nutrition and larger size. This may promote Wolbachia spread via CI in wild mosquito populations and underscores its potential use for disease control.

BACKGROUND: The World Health Organization estimates that the global number of dengue infections range between 80-100 million per year, with some studies estimating approximately three times higher numbers. Furthermore, the geographic range of dengue virus transmission is extending with the disease now occurring more frequently in areas such as southern Europe. Ae. aegypti, one of the most prominent dengue vectors, is endemic to the far north-east of Australia and the city of Cairns frequently experiences dengue outbreaks which sometimes lead to large epidemics. METHOD: A spatially-explicit, individual-based mathematical model that accounts for the spread of dengue infection as a result of human movement and mosquito dispersion is presented. The model closely couples the four key sub-models necessary for representing the overall dynamics of the physical system, namely those describing mosquito population dynamics, human movement, virus transmission and vector control. Important features are the use of high quality outbreak data and mosquito trapping data for calibration and validation and a strategy to derive local mosquito abundance based on vegetation coverage and census data. RESULTS: The model has been calibrated using detailed 2003 dengue outbreak data from Cairns, together with census and mosquito trapping data, and is shown to realistically reproduce a further dengue outbreak. The simulation results replicating the 2008/2009 Cairns epidemic support several hypotheses (formulated previously) aimed at explaining the large-scale epidemic which occurred in 2008/2009; specifically, while warmer weather and increased human movement had only a small effect on the spread of the virus, a shorter virus strain-specific extrinsic incubation time can explain the observed explosive outbreak of 2008/2009. CONCLUSION: The proof-of-concept simulation model described in this study has potential as a tool for understanding factors contributing to dengue spread as well as plannin

BACKGROUND: Dengue is the world's most important mosquito-borne viral illness. Successful future management of this disease requires an understanding of the population dynamics of the vector, especially in the context of changing climates. Our capacity to predict future dynamics is reflected in our ability to explain the significant historical changes in the distribution and abundance of the disease and its vector. METHODOLOGY/PRINCIPAL FINDINGS: Here we combine daily weather records with simulation modelling techniques to explain vector (Aedes aegypti (L.)) persistence within its current and historic ranges in Australia. We show that, in regions where dengue presently occurs in Australia (the Wet Tropics region of Far North Queensland), conditions are persistently suitable for year-round adult Ae. aegypti activity and oviposition. In the historic range, however, the vector is vulnerable to periodic extinction due to the combined influence of adult activity constraints and stochastic loss of suitable oviposition sites. CONCLUSIONS/SIGNIFICANCE: These results, together with changes in water-storage behaviour by humans, can explain the observed historical range contraction of the disease vector. For these reasons, future eradication of dengue in wet tropical regions will be extremely difficult through classical mosquito control methods alone. However, control of Ae. aegypti in sub-tropical and temperate regions will be greatly facilitated by government policy regulating domestic water-storage. Exploitation of the natural vulnerabilities of dengue vectors (e.g., habitat specificity, climatic limitations) should be integrated with the emerging novel transgenic and symbiotic bacterial control techniques to develop future control and elimination strategies.

Japanese encephalitis (JE) virus spread to northern Australia during the 1990s, transmitted by Culex annulirostris Skuse and other mosquitoes (Diptera: Culicidae). To determine the relative importance of various hosts for potential vectors of JE virus, we investigated the host-feeding patterns of mosquitoes in northern Australia and Western Province of Papua New Guinea, with particular attention to pigs, Sus scrofa L. - the main amplifying host of JE virus in South-east Asia. Mosquitoes were collected by CDC light traps baited with dry ice and 1-octen-3-ol, run 16.00-08.00 hours, mostly set away from human habitations, if possible in places frequented by feral pigs. Bloodmeals of 2569 mosquitoes, representing 15 species, were identified by gel diffusion assay. All species had fed mostly on mammals: only <10% of bloodmeals were from birds. The predominant species was Cx. annulirostris (88%), with relatively few (4.4%) bloodmeals obtained from humans. From all 12 locations sampled, the mean proportion of Cx. annulirostris fed on pigs (9.1%) was considerably lower than fed on other animals (90.9%). Highest rates of pig-fed mosquitoes (>30%) were trapped where domestic pigs were kept close to human habitation. From seven of eight locations on the Australian mainland, the majority of Cx. annulirostris had obtained their bloodmeals from marsupials, probably the Agile wallaby Macropus agilis (Gould). Overall proportions of mosquito bloodmeals identified as marsupial were 60% from the Gulf Plains region of Australia, 78% from the Cape York Peninsula and 64% from the Daru area of Papua New Guinea. Thus, despite the abundance of feral pigs in northern Australia, our findings suggest that marsupials divert host-seeking Cx. annulirostris away from pigs. As marsupials are poor JE virus hosts, the prevalence of marsupials may impede the establishment of JE virus in Australia.