Wolbachia Frequencies Research Articles

Further evidence of low infection frequencies of Wolbachia in soil arthropod communities

Endosymbiotic Alphaproteobacteria of the genus Wolbachia are exclusively transferred maternally from mother to offspring, but horizontal transfer across species boundaries seems to be frequent as well. However, the (ecological) mechanisms of how these bacteria are transferred between distantly related arthropod hosts remain unclear. Based on the observation that species that are part of the same ecological community often also share similar Wolbachia strains, host ecology has been hypothesized as an important factor enabling transmission and a key factor in explaining the global distribution of Wolbachia lineages.In this study, we focus on the diversity and abundance of Wolbachia strains in soil arthropods, a so far rather neglected community. We screened 82 arthropod morphotypes collected in the beech forest (dominated by Fagus sp.) soil in the area of Göttingen in central Germany for the presence of Wolbachia. By performing a PCR screen with Wolbachia-MLST markers (coxA, dnaA, fbpA, ftsZ, gatB, and hcpA), we found a rather low infection frequency of 12,2%. Additionally, we performed metagenomic screening of pooled individuals from the same sampling site and could not find evidence that this low infection frequency is an artefact due to PCR-primer bias. Phylogenetic analyses of the recovered Wolbachia strains grouped them in three known supergroups (A, B, and E), with the first report of Wolbachia in Protura (Hexapoda). Moreover, Wolbachia sequences from the pseudoscorpion Neobisium carcinoides cluster outside the currently known supergroup diversity. Our screening supports results from previous studies that the prevalence of Wolbachia infections seems to be lower in soil habitats than in above-ground terrestrial habitats. The reasons for this pattern are not completely understood but might stem from the low opportunity of physical contact and the prevalence of supergroups that are less suited for horizontal transfer.

The Diversity of Wolbachia and Other Bacterial Symbionts in Spodoptera frugiperda.

Bacterial symbionts associated with insects can be crucial in insect nutrition, metabolism, immune responses, development, and reproduction. However, the bacterial symbionts of the fall armyworm Spodoptera frugiperda remain unclear. S. frugiperda is an invasive polyphagous pest that severely damages many crops, particularly maize and wheat. Here, we investigated the infection, composition, abundance, and diversity of bacterial symbionts, especially Wolbachia, in different tissues of S. frugiperda female adults. The infection prevalence frequencies of Wolbachia in five provinces of China, namely Pu'er, Yunnan; Nanning, Guangxi; Sanya, Hainan; Yunfu, Guangdong; and Nanping, Fujian, were assessed. The results indicated that Proteobacteria, Firmicutes, and Bacteroidetes were the three most dominant bacterial phyla in S. frugiperda adults. At the genus level, the abundant microbiota, which included Enterobacter and Enterococcus, varied in abundance between tissues of S. frugiperda. Wolbachia was found in the ovaries and salivary glands of S. frugiperda adults, and was present in 33.33% of the Pu'er, Yunnan, 23.33% of the Nanning, Guangxi, and 13.33% of the Sanya, Hainan populations, but Wolbachia was absent in the Yunfu, Guangdong and Nanping, Fujian populations. Further phylogenetic analyses revealed that all of the Wolbachia strains from the different S. frugiperda populations belonged to the supergroup B and were named the wFru strain. Since there were Wolbachia strains inducing cytoplasmic incompatibility in supergroup B, these findings may provide a foundation for developing potential biocontrol techniques against S. frugiperda.

Cardinium symbionts are pervasive in Iranian populations of the spider mite Panonychus ulmi despite inducing an infection cost and no demonstrable reproductive phenotypes when Wolbachia is a symbiotic partner.

Maternally transmitted symbionts such as Cardinium and Wolbachia are widespread in arthropods. Both Cardinium and Wolbachia can cause cytoplasmic incompatibility, a reproductive phenotype that interferes with the development of uninfected eggs that are fertilized by infected sperm. In haplodiploid hosts, these symbionts can also distort sex allocation to facilitate their spread through host populations. Without other fitness effects, symbionts that induce strong reproductive phenotypes tend to spread to high and stable infection frequencies, whereas variants that induce weak reproductive phenotypes are typically associated with intermediate and variable frequencies. To study the spread of Cardinium in a haplodiploid host, we sampled Iranian populations of the economically important spider mite Panonychus ulmi in apple orchards. Within several field populations, we also studied the Wolbachia infection frequencies. All P. ulmi field populations carried a Cardinium infection and exhibited high infection frequencies. In contrast, Wolbachia frequency ranged between ca. 10% and ca. 70% and was only found in co-infected mites. To test whether Cardinium induce reproductive phenotypes in P. ulmi, a Cardinium-cured derived line was generated by antibiotic treatment from a co-infected field population. Genetic crosses indicated that Cardinium do not induce demonstrable levels of cytoplasmic incompatibility and sex allocation distortion in co-infected P. ulmi. However, Cardinium infection was associated with a longer developmental time and reduced total fecundity for co-infected females. We hypothesize that Cardinium spread through P. ulmi populations via uncharacterized fitness effects and that co-infection with Wolbachia might impact these drive mechanisms.

Kajian aspek keamanan nyamuk Aedes aegypti Linnaeus ber-Wolbachia di Yogyakarta, Indonesia

Dengue prevention efforts are limited to the control strategies of its vector and the management of breeding sites. New alternatives for dengue vector control that are sustainable and more environmentally friendly are needed to complement the government’s current efforts. Research on Wolbachia-infected Aedes aegypti Linnaeus mosquitoes as an alternative biocontrol strategy has been performed in Yogyakarta City. However, one of the concerns of the community members and stakeholders about this technology is the safety aspect regarding the transmission of Wolbachia to other species and the possibility that humans will contract Wolbachia. This study aimed to address these concerns, namely to find out whether horizontal transmission of Wolbachia occurred from A. aegypti that were released to other species and whether residents living in the released areas were infected with Wolbachia. The research was conducted in Dusun Nogotirto and Dusun Kronggahan (Sleman Regency), as well as in Dusun Jomblangan and Dusun Singosaren (Bantul Regency), Yogyakarta Special Province. Wolbachia qPCR screening using the target gene WD0513 was performed on 922 Culex quinquefasciatus Say and 331 Aedes albopictus (Skuse). ELISA test was carried out on 190 pairs of plasma samples, namely the sample before the Wolbachia frequency was established (still <80%) and the sample after it was established (>80%). The results showed no evidence of Wolbachia transfer from Wolbachia-infected A. aegypti to other mosquito species coexisting in the same habitat or to humans. This study corroborates the safety evidence of Wolbachia-infected A. aegypti technology as an alternative to control dengue virus transmission

Spatial Distribution and Long-Term Persistence of Wolbachia-Infected Aedes aegypti in the Mentari Court, Malaysia.

Dengue is endemic in Malaysia, and vector control strategies are vital to reduce dengue transmission. The Wolbachia strain wAlbB carried by both sexes of Ae. aegypti was released in Mentari Court, a high-rise residential site, in October 2017 and stopped after 20 weeks. Wolbachia frequencies are still being monitored at multiple traps across this site, providing an opportunity to examine the spatiotemporal distribution of Wolbachia and mosquito density with respect to year, residential block, and floor, using spatial interpolation in ArcGIS, GLMs, and contingency analyses. In just 12 weeks, Wolbachia-infected mosquitoes were established right across the Mentari Court site with an overall infection frequency of >90%. To date, the Wolbachia frequency of Ae. aegypti has remained high in all areas across the site despite releases finishing four years ago. Nevertheless, the Wolbachia invaded more rapidly in some residential blocks than others, and also showed a relatively higher frequency on the eighth floor. The Ae. aegypti index tended to differ somewhat between residential blocks, whilst the Ae. albopictus index was relatively higher at the top and bottom floors of buildings. In Mentari Court, only a short release period was required to infiltrate Wolbachia completely and stably into the natural population. The results inform future releases in comparable sites in a dengue control programme.

Why Wolbachia-induced cytoplasmic incompatibility is so common

Cytoplasmic incompatibility (CI) is the most common reproductive manipulation produced by Wolbachia, obligately intracellular alphaproteobacteria that infect approximately half of all insect species. Once infection frequencies within host populations approach 10%, intense CI can drive Wolbachia to near fixation within 10 generations. However, natural selection among Wolbachia variants within individual host populations does not favor enhanced CI. Indeed, variants that do not cause CI but increase host fitness or are more reliably maternally transmitted are expected to spread if infected females remain protected from CI. Nevertheless, approximately half of analyzed Wolbachia infections cause detectable CI. Why? The frequency and persistence of CI are more plausibly explained by preferential spread to new host species (clade selection) rather than by natural selection among variants within host populations. CI-causing Wolbachia lineages preferentially spread into new host species because 1) CI increases equilibrium Wolbachia frequencies within host populations, and 2) CI-causing variants can remain at high frequencies within populations even when conditions change so that initially beneficial Wolbachia infections become harmful. An epidemiological model describing Wolbachia acquisition and loss by host species and the loss of CI-induction within Wolbachia lineages yields simple expressions for the incidence of Wolbachia infections and the fraction of those infections causing CI. Supporting a determinative role for differential interspecific spread in maintaining CI, many Wolbachia infections were recently acquired by their host species, many show evidence for contemporary spatial spread or retreat, and rapid evolution of CI-inducing loci, especially degradation, is common.

Sensitivity of wMel and wAlbB Wolbachia infections in Aedes aegypti Puducherry (Indian) strains to heat stress during larval development

BackgroundICMR-Vector Control Research Centre, Puducherry, India, developed two colonies of Aedes aegypti infected with wMel and wAlbB Wolbacia strains called Ae. aegypti (Pud) lines for dengue control. The sensitivity of wMel and wAlbB strains in Ae. aegypti (Pud) lines to heat stress was studied.MethodswMel and wAlbB infected and uninfected Ae. aegypti larvae (first to fourth instars) were reared in the laboratory to adults at 26 °C, 30 °C, 36 °C and 40 °C constant temperatures and also 26–30 °C, 26–36 °C and 26–40 °C diurnal cyclic temperatures. The adults were tested for Wolbachia infection. Experiments were also carried out rearing the larvae under simulated field conditions in summer (April and June) under sunlight using fully open and half open bowls and also under sunlight and natural shade.ResultsAt 36 °C and 40 °C constant temperatures, complete larval mortality was observed. At 30 °C and 26 °C, no larval mortality occurred, but Wolbachia density was relatively low in wMel infected males compared to control (maintained at 26 ± 1 °C). At diurnal cyclic temperature of 26–40 °C, Wolbachia density was reduced in males of both the (Pud) lines, but not in females. At 26–36 °C, reduction in Wolbachia density was observed in wMel males but not in wAlbB males. At 26–30 °C, no significant reduction in Wolbachia density was observed with wMel and wAlbB strains. In simulated field conditions (April), under sunlight, the daytime water temperature reached a maximum of 35.7 °C in both full and half open bowls. No larval mortality occurred. Wolbachia frequency and density was reduced in wMel-infected Ae. aegypti (Pud) males from both type of bowls and in females from full open bowls, and in wAlbB males from half open bowls. In June, rearing of larvae under sunlight, the first-instar larvae experienced a maximum daytime water temperature of > 38 °C that caused complete mortality. No larval mortality was observed in bowls kept under shade (< 32 °C).ConclusionsExposure of larvae to higher rearing temperatures in the laboratory and simulated-field conditions reduced the densities of wMel and wAlbB strains particularly in males, but the impact was more pronounced for wMel strain. The actual effect of heat stress on the stability of these two Wolbachia strains needs to be tested under natural field conditions.Graphical

Wolbachia in the spittlebug Prosapia ignipectus: Variable infection frequencies, but no apparent effect on host reproductive isolation.

Animals serve as hosts for complex communities of microorganisms, including endosymbionts that live inside their cells. Wolbachia bacteria are perhaps the most common endosymbionts, manipulating host reproduction to propagate. Many Wolbachia cause cytoplasmic incompatibility (CI), which results in reduced egg hatch when uninfected females mate with infected males. Wolbachia that cause intense CI spread to high and relatively stable frequencies, while strains that cause weak or no CI tend to persist at intermediate, often variable, frequencies. Wolbachia could also contribute to host reproductive isolation (RI), although current support for such contributions is limited to a few systems. To test for Wolbachia frequency variation and effects on host RI, we sampled several local Prosapia ignipectus (Fitch) (Hemiptera: Cercopidae) spittlebug populations in the northeastern United States over two years, including closely juxtaposed Maine populations with different monomorphic color forms, “black” and “lined.” We discovered a group‐B Wolbachia (wPig) infecting P. ignipectus that diverged from group‐A Wolbachia—like model wMel and wRi strains in Drosophila—6 to 46 MYA. Populations of the sister species Prosapia bicincta (Say) from Hawaii and Florida are uninfected, suggesting that P. ignipectus acquired wPig after their initial divergence. wPig frequencies were generally high and variable among sites and between years. While phenotyping wPig effects on host reproduction is not currently feasible, the wPig genome contains three divergent sets of CI loci, consistent with high wPig frequencies. Finally, Maine monomorphic black and monomorphic lined populations of P. ignipectus share both wPig and mtDNA haplotypes, implying no apparent effect of wPig on the maintenance of this morphological contact zone. We hypothesize P. ignipectus acquired wPig horizontally as observed for many Drosophila species, and that significant CI and variable transmission produce high but variable wPig frequencies.

Wolbachia in mosquitoes from the Central Valley of California, USA

BackgroundWolbachia bacteria are widely distributed throughout terrestrial arthropod species. These bacteria can manipulate reproduction and influence the vector competence of their hosts. Recently, Wolbachia have been integrated into vector control programmes for mosquito management. A number of supergroups and strains exist for Wolbachia, and they have yet to be characterized for many mosquito species. In this study, we examined Wolbachia prevalence and their phylogenetic relationship to other Wolbachia, using mosquitoes collected in Merced County in the Central Valley of California.MethodsAdult mosquitoes were collected from 85 sites in Merced County, California in 2017 and 2018. Traditional and quantitative PCR were used to investigate the presence or absence and the density of Wolbachia, using Wolbachia-specific 16S rRNA and Wolbachia-surface protein (wsp) genes. The supergroup of Wolbachia was determined, and Multilocus Sequence Typing (MLST) by sequencing five housekeeping genes (coxA, gatB, ftsZ, hcpA and fbpA) was also used to determine Wolbachia supergroup as well as strain.ResultsOver 7100 mosquitoes of 12 species were collected: Aedesmelanimon, Ae. nigromaculis, Ae. vexans, Ae. aegypti, Culex pipiens, Cx. stigmatosoma, Cx. tarsalis, Anopheles franciscanus, An. freeborni, An. punctipennis, Culiseta incidens and Cs. inornata. Eight showed evidence of Wolbachia. To our knowledge, this study is the first to report detection of Wolbachia in five of these species (Ae. melanimon, Cx. stigmatosoma, Cx. tarsalis, Cs. incidens and Cs. inornata). Culex pipiens and Cx. stigmatosoma had a high frequency and density of Wolbachia infection, which grouped into supergroup B; Cs. inornata clustered with supergroup A. MLST comparisons identified Cx. pipiens and Cx. stigmatosoma as wPip strain type 9 supergroup B. Six species had moderate to low (< 14%) frequencies of Wolbachia. Four species were negative, Ae. nigromaculis, An. franciscanus, An. freeborni and Ae. aegypti.ConclusionsNew records of Wolbachia detection were found in mosquitoes from Merced County, California. Culex stigmatosoma and Cs. inornata were new records for Wolbachia supergroup B and A, respectively. Other species with Wolbachia occurred with low frequency and low density. Detection of Wolbachia in mosquitoes can be used to inform potential vector control applications. Future study of Wolbachia within Cx. stigmatosoma and Cs. inornata in California and through the range of these species could further explore Wolbachia infection in these two species.

Environmental and Genetic Contributions to Imperfect wMel-Like Wolbachia Transmission and Frequency Variation.

Maternally transmitted Wolbachia bacteria infect about half of all insect species. They usually show imperfect maternal transmission and often produce cytoplasmic incompatibility (CI). Irrespective of CI, Wolbachia frequencies tend to increase when rare only if they benefit host fitness. Several Wolbachia, including wMel that infects Drosophila melanogaster, cause weak or no CI and persist at intermediate frequencies. On the island of São Tomé off West Africa, the frequencies of wMel-like Wolbachia infecting Drosophila yakuba (wYak) and Drosophila santomea (wSan) fluctuate, and the contributions of imperfect maternal transmission, fitness effects, and CI to these fluctuations are unknown. We demonstrate spatial variation in wYak frequency and transmission on São Tomé. Concurrent field estimates of imperfect maternal transmission do not predict spatial variation in wYak frequencies, which are highest at high altitudes where maternal transmission is the most imperfect. Genomic and genetic analyses provide little support for D. yakuba effects on wYak transmission. Instead, rearing at cool temperatures reduces wYak titer and increases imperfect transmission to levels observed on São Tomé. Using mathematical models of Wolbachia frequency dynamics and equilibria, we infer that temporally variable imperfect transmission or spatially variable effects on host fitness and reproduction are required to explain wYak frequencies. In contrast, spatially stable wSan frequencies are plausibly explained by imperfect transmission, modest fitness effects, and weak CI. Our results provide insight into causes of wMel-like frequency variation in divergent hosts. Understanding this variation is crucial to explain Wolbachia spread and to improve wMel biocontrol of human disease in transinfected mosquito systems.

Survey of Wolbachia frequency in Nashville, Tennessee Reveals Novel Infections

Wolbachia (Rickettsiales: Anaplasmataceae) are maternally transmitted intracellular bacteria that infect approximately half of all insect species. These bacteria commonly act as reproductive parasites or mutualists to enhance their transmission from mother to offspring, resulting in high prevalence among some species. Despite decades of research on Wolbachia’s global frequency, there are many arthropod families and geographic regions that have not been tested for Wolbachia. Here, arthropods were collected on the Vanderbilt University campus in Nashville, Tennessee, where Wolbachia frequency has not been previously studied. The dataset consists of 220 samples spanning 34 unique arthropod families collected on the Vanderbilt University campus. The majority of our samples were from the families Blattidae (Blattodea), Pulicidae (Siphonaptera), Dryinidae (Hymenoptera), Aphididae (Hemiptera), Paronellidae (Entomobryomorpha), Formicidae (Hymenoptera), Pseudococcidae (Hemiptera), Sphaeroceridae (Diptera), and Coccinellidae (Coleoptera). PCR-based techniques were used to assign infection states and, from these data, the first cases of Wolbachia in the Paronellidae springtails, Lithobiidae (Lithobiomorpha) centipedes, Lonchopteridae (Diptera) spear-winged flies, Sepsidae (Diptera) black scavenger flies, Cryptocercidae (Blattodea) wood roaches, and Lauxaniidae (Diptera) acalyptrate flies were identified. Within-family infection frequencies ranged from 17-100% when Wolbachia was observed; however, numerous families tested did not reveal evidence of infection. These results expand on the field’s understanding of Wolbachia’sfrequencyin Nashville, Tennessee, and among arthropod families broadly, and is the first report of Wolbachia in centipedes. KEYWORDS: Wolbachia; Infection Frequency; Endosymbiont; Tennessee; Centipede; Arthropod; Polymerase Chain Reaction; Nashville

Establishment of wMel Wolbachia in Aedes aegypti mosquitoes and reduction of local dengue transmission in Cairns and surrounding locations in northern Queensland, Australia.

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 Wolbachiawas backcrossed into the local Aedes aegyptigenotype (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 has resulted in the reduction of local dengue transmission across all deployment areas.

Alternative evolutionary outcomes following endosymbiont-mediated selection on male mating preference alleles.

In many arthropods, intracellular bacteria, such as those of the genus Wolbachia, may spread through host populations as a result of cytoplasmic incompatibility (CI). Here, there is sterility or reduced fertility in crosses between infected males and uninfected females. As the bacterium is maternally inherited, the reduced fertility of uninfected females increases the frequency of the infection. If the transmission fidelity of the bacterium is less than 100%, the bacterium cannot invade from a low frequency, but if its frequency exceeds a threshold, it increases to a high, stable, equilibrium frequency. We explore the expected evolutionary dynamics of mutant alleles that cause their male bearers to avoid mating with uninfected females. For alleles which create this avoidance behaviour conditional upon the male being infected, there is a wide zone of parameter space that allows the preference allele to drive Wolbachia from the population when it would otherwise stably persist. There is also a wide zone of parameter space that allows a joint stable equilibrium for the Wolbachia and a polymorphism for the preference allele. When the male's avoidance of uninfected females is unconditional, the preference allele's effect on Wolbachia frequency is reduced, but there is a narrow range of values for the transmission rate and CI fertility that allow an unconditional preference allele to drive Wolbachia from the population, in a process driven by positive linkage disequilibrium between Wolbachia and the preference allele. The possibility of the evolution of preference could hamper attempts to manipulate wild populations through Wolbachia introductions.

Endosymbiont diversity in natural populations of Tetranychus mites is rapidly lost under laboratory conditions.

Although the diversity of bacterial endosymbionts in arthropods is well documented, whether and how such diversity is maintained remains an open question. We investigated the temporal changes occurring in the prevalence and composition of endosymbionts after transferring natural populations of Tetranychus spider mites from the field to the laboratory. These populations, belonging to three different Tetranychus species (T. urticae, T. ludeni and T. evansi) carried variable infection frequencies of Wolbachia, Cardinium, and Rickettsia. We report a rapid change of the infection status of these populations after only 6 months of laboratory rearing, with an apparent loss of Rickettsia and Cardinium, while Wolbachia apparently either reached fixation or was lost. We show that Wolbachia had variable effects on host longevity and fecundity, and induced variable levels of cytoplasmic incompatibility (CI) in each fully infected population, despite no sequence divergence in the markers used and full CI rescue between all populations. This suggests that such effects are largely dependent upon the host genotype. Subsequently, we used these data to parameterize a theoretical model for the invasion of CI-inducing symbionts in haplodiploids, which shows that symbiont effects are sufficient to explain their dynamics in the laboratory. This further suggests that symbiont diversity and prevalence in the field are likely maintained by environmental heterogeneity, which is reduced in the laboratory. Overall, this study highlights the lability of endosymbiont infections and draws attention to the limitations of laboratory studies to understand host-symbiont interactions in natural populations.

Ovitraps Provide a Reliable Estimate of Wolbachia Frequency during wMelBr Strain Deployment in a Geographically Isolated Aedes aegypti Population.

Deployment of Aedes aegypti mosquitoes carrying the endosymbiont bacterium Wolbachia has been identified as a promising strategy to reduce dengue, chikungunya, and Zika transmission. We investigated whether sampling larvae from ovitraps can provide reliable estimates on Wolbachia frequency during releases, as compared to the expensive adult-based BG-Sentinel. We conducted pilot releases in a semi-field system (SFS) divided into six cages of 21 m2, each with five ovitraps. Five treatments were chosen to represent different points of a hypothetical invasion curve: 10%, 25%, 50%, 75%, and 90% of Wolbachia frequency. Collected eggs were counted and hatched, and the individuals from a net sample of 27% of larvae per treatment were screened for Wolbachia presence by RT-qPCR. Ovitrap positioning had no effect on egg hatching rate. Treatment strongly affected the number of eggs collected and also the hatching rate, especially when Wolbachia was at a 10% frequency. A second observation was done during the release of Wolbachia in Rio under a population replacement approach when bacterium frequency was estimated using 30 BG-Sentinel traps and 45 ovitraps simultaneously. By individually screening 35% (N = 3904) of larvae collected by RT-qPCR, we were able to produce a similar invasion curve to the one observed when all adults were individually screened. If sampling is reduced to 20%, monitoring Wolbachia frequency with 45 ovitraps would be roughly half the cost of screening all adult mosquitoes captured by 30 BG-Sentinels. Our findings support the scale-up of Wolbachia releases, especially in areas with limited resources to afford massive trapping with BG-Sentinel traps.

Heatwaves cause fluctuations in wMel Wolbachia densities and frequencies in Aedes aegypti.

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.

Establishment of wMel Wolbachia in Aedes aegypti mosquitoes and reduction of local dengue transmission in Cairns and surrounding locations in northern Queensland, Australia

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 has resulted in the reduction of local dengue transmission across all deployment areas.

Aedes aegypti insecticide resistance underlies the success (and failure) of Wolbachia population replacement

Mosquitoes that carry Wolbachia endosymbionts may help control the spread of arboviral diseases, such as dengue, Zika and chikungunya. Wolbachia frequencies systematically increase only when the frequency-dependent advantage due to cytoplasmic incompatibility exceeds frequency-independent costs, which may be intrinsic to the Wolbachia and/or can be associated with the genetic background into which Wolbachia are introduced. Costs depend on field conditions such as the environmental pesticide load. Introduced mosquitoes need adequate protection against insecticides to ensure survival after release. We model how insecticide resistance of transinfected mosquitoes determines the success of local Wolbachia introductions and link our theoretical results to field data. Two Ae. aegypti laboratory strains carrying Wolbachia were released in an isolated district of Rio de Janeiro, Brazil: wMelBr (susceptible to pyrethroids) and wMelRio (resistant to pyrethroids). Our models elucidate why releases of the susceptible strain failed to result in Wolbachia establishment, while releases of the resistant strain led to Wolbachia transforming the native Ae. aegypti population. The results highlight the importance of matching insecticide resistance levels in release stocks to those in the target natural populations during Wolbachia deployment.

Establishment of wMel Wolbachia in Aedes aegypti mosquitoes and reduction of local dengue transmission in Cairns and surrounding locations in northern Queensland, Australia.

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 Wolbachiawas backcrossed into the local Aedes aegyptigenotype (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 has resulted in the reduction of local dengue transmission across all deployment areas.

Scaled deployment of Wolbachia to protect the community from dengue and other Aedes transmitted arboviruses.

Background: A number of new technologies are under development for the control of mosquito transmitted viruses, such as dengue, chikungunya and Zika that all require the release of modified mosquitoes into the environment. None of these technologies has been able to demonstrate evidence that they can be implemented at a scale beyond small pilots. Here we report the first successful citywide scaled deployment of Wolbachia in the northern Australian city of Townsville. Methods: The wMel strain of Wolbachia was backcrossed into a local Aedes aegypti genotype and mass reared mosquitoes were deployed as eggs using mosquito release containers (MRCs). In initial stages these releases were undertaken by program staff but in later stages this was replaced by direct community release including the development of a school program that saw children undertake releases. Mosquito monitoring was undertaken with Biogents Sentinel (BGS) traps and individual mosquitoes were screened for the presence of Wolbachia with a Taqman qPCR or LAMP diagnostic assay. Dengue case notifications from Queensland Health Communicable Disease Branch were used to track dengue cases in the city before and after release. Results: Wolbachia was successfully established into local Ae. aegypti mosquitoes across 66 km 2 in four stages over 28 months with full community support. A feature of the program was the development of a scaled approach to community engagement. Wolbachia frequencies have remained stable since deployment and to date no local dengue transmission has been confirmed in any area of Townsville after Wolbachia has established, despite local transmission events every year for the prior 13 years and an epidemiological context of increasing imported cases. Conclusion: Deployment of Wolbachia into Ae. aegypti populations can be readily scaled to areas of ~60km 2 quickly and cost effectively and appears in this context to be effective at stopping local dengue transmission.