Brian L. Montgomery

The wMel infection of Drosophila melanogaster was successfully transferred into Aedes aegypti mosquitoes where it has the potential to suppress dengue and other arboviruses. The infection was subsequently spread into two natural populations at Yorkeys Knob and Gordonvale near Cairns, Queensland in 2011. Here we report on the stability of the infection following introduction and we characterize factors influencing the ongoing dynamics of the infection in these two populations. While the Wolbachia infection always remained high and near fixation in both locations, there was a persistent low frequency of uninfected mosquitoes. These uninfected mosquitoes showed weak spatial structure at both release sites although there was some clustering around two areas in Gordonvale. Infected females from both locations showed perfect maternal transmission consistent with patterns previously established pre-release in laboratory tests. After >2 years under field conditions, the infection continued to show complete cytoplasmic incompatibility across multiple gonotrophic cycles but persistent deleterious fitness effects, suggesting that host effects were stable over time. These results point to the stability of Wolbachia infections and their impact on hosts following local invasion, and also highlight the continued persistence of uninfected individuals at a low frequency most likely due to immigration.

<ns4:p><ns4:bold>Background: </ns4:bold>The <ns4:italic>w</ns4:italic>Mel strain of<ns4:italic> Wolbachia</ns4:italic> has been successfully introduced into <ns4:italic>Aedes aegypti</ns4:italic> 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 <ns4:italic>Wolbachia</ns4:italic> across nearly all significant dengue transmission risk areas in Australia.</ns4:p><ns4:p> <ns4:bold>Methods: </ns4:bold>The <ns4:italic>w</ns4:italic>Mel strain of <ns4:italic>Wolbachia</ns4:italic> was backcrossed into the local <ns4:italic>Aedes aegypti</ns4:italic> 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 <ns4:italic>Wolbachia</ns4:italic>. Dengue case notifications were used to track dengue incidence in each location before and after releases.</ns4:p><ns4:p> <ns4:bold>Results: </ns4:bold>Empirical analyses of the <ns4:italic>Wolbachia</ns4:italic> mosquito releases, including data on the density, frequency and duration of <ns4:italic>Wolbachia</ns4:italic> mosquito releases, indicate that <ns4:italic>Wolbachia</ns4:italic> 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, <ns4:italic>Wolbachia</ns4:italic> 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 <ns4:italic>Wolbachia</ns4:italic> has been established. The regression model estimate of <ns4:italic>Wolbachia </ns4:italic>intervention effect from interrupted time series analyses of case notifications data prior to and after releases, indicated a 96% reduction in dengue incidence in <ns4:italic>Wolbachia </ns4:italic>treated populations (95% confidence interval: 84 – 99%).</ns4:p><ns4:p> <ns4:bold>Conclusion: </ns4:bold>Deployment of the <ns4:italic>w</ns4:italic>Mel strain of <ns4:italic>Wolbachia</ns4:italic> into local <ns4:italic>Ae. aegypti</ns4:italic> populations across the Australian regional cities of Cairns and most smaller regional communities with a past history of dengue has resulted in the reduction of local dengue transmission across all deployment areas.</ns4:p>

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-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.