Wolbachia Bacteria Research Articles

The Evolution and Biogeography of Wolbachia in Ants (Hymenoptera: Formicidae)

Wolbachia bacteria are widely distributed across invertebrate taxa, including ants, but several aspects of this host-associated interaction are still poorly explored, especially with regard to the ancestral state association, origin, and dispersion patterns of this bacterium. Therefore, in this study, we explored the association of Wolbachia with Formicidae in an evolutionary context. Our data suggest that supergroup F is the ancestral character state for Wolbachia infection in ants, and there is only one transition to supergroup A, and once ants acquired infection with supergroup A, there have been no other strains introduced. Our data also reveal that the origin of Wolbachia in ants likely originated in Asia and spread to the Americas, and then back to Asia. Understanding the processes and mechanisms of dispersion of these bacteria in Formicidae is a crucial step to advance the knowledge of this symbiosis and their implications in an evolutionary context.

Base Pair: 28 Years of Sustained High School Biomedical Research Mentorship Driving Health Sciences Career Progression

Base Pair is an ongoing (1992-present) collaboration between University of Mississippi Medical Center (UMMC) faculty mentors and Mississippi high school students and teachers. UMMC faculty and Base Pair teachers designed a course which has allowed curriculum innovations by teachers. The student component engages 15 trainees annually in laboratory experiences under direct faculty mentorship while receiving high school graduation credit. Students have a 100% high school graduation rate (vs. ~88% nationally) and 99% college entrance. These students have co-authored/ presented over 450 times in UMMC, state or national venues. Of 220 graduates, roughly 63% are in/have completed post-baccalaureate training. Of these, at least 49 have gained a terminal degree in science or law, while another 35 students have attained a Master’s degree. Over 100 teachers have been mentored in grant-writing exercises and curriculum development, resulting in over 158 applications for external funding of inquiry-based classroom activities of which 78% were funded. Teachers have also implemented several innovative STEM curricula, reaching thousands more students. These include a student team-based competition that fuses citizen science and molecular biology to assess insect infection with Wolbachia bacteria. Base Pair bolsters student engagement, promotes STEM/health career advancement for students, and enhances teacher job satisfaction.

Stable Introduction of Plant-Virus-Inhibiting Wolbachia into Planthoppers for Rice Protection

Progress has been made in developing the maternally inherited endosymbiotic bacterium Wolbachia as a tool for protecting humans from mosquito-borne diseases. In contrast, Wolbachia-based approaches have not yet been developed for the protection of plants from insect pests and their associated diseases, with a major challenge being the establishment of artificial Wolbachia infections expressing desired characteristics in the hemipterans that transmit the majority of plant viruses. Here, we report stable introduction of Wolbachia into the brown planthopper, Nilaparvata lugens, the most destructive rice pest that annually destroys millions of hectares of staple crops. The Wolbachia strain wStri from the small brown planthopper, Laodelphax striatellus, was transferred to this new host, where it showed high levels of cytoplasmic incompatibility, enabling rapid invasion of laboratory populations. Furthermore, wStri inhibited infection and transmission of Rice ragged stunt virus and mitigated virus-induced symptoms in rice plants, opening up the development of Wolbachia-based strategies against major agricultural pests and their transmitted pathogens. VIDEO ABSTRACT.

A feedback control perspective on biological control of dengue vectors by Wolbachia infection

Controlling diseases such as dengue fever, chikungunya and zika fever by introduction of the intracellular parasitic bacterium Wolbachia in mosquito populations which are their vectors, is presently quite a promising tool to reduce their spread. While description of the conditions of such experiments has received ample attention from biologists, entomologists and applied mathematicians, the issue of effective scheduling of the releases remains an interesting problem. Having in mind the important uncertainties present in the dynamics of the two populations in interaction, we attempt here to identify general ideas for building feedback-based release strategies, enforceable to a variety of models and situations. These principles are exemplified by several feedback control laws whose stabilizing properties are demonstrated, illustrated numerically and compared, when applied to a model retrieved from [P.-A. Bliman et al., Ensuring successful introduction of Wolbachia in natural populations of Aedes aegypti by means of feedback control. J. of Math. Bio. 76(5):1269–1300, 2018]. The contribution is believed to be also of potential interest to tackle other important issues related to the biological control of vectors and pests. A crucial use of the theory of monotone dynamical systems is made in the derivations.

Wolbachia Genome Stability and mtDNA Variants in Aedes aegypti Field Populations Eight Years after Release.

SummaryA 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.

Phylogeny and Density Dynamics of Wolbachia Infection of the Health Pest Paederus fuscipes Curtis (Coleoptera: Staphylinidae).

Simple SummaryWolbachia pipientis is a maternally inherited endosymbiont of arthropods and filarial nematodes, and was reported to occur in Paederus fuscipes, a beetle that causes dermatitis linearis and conjunctivitis in humans when they come in contact with skin. In this study, we report the phylogenetic position and density dynamics of Wolbachia in P. fuscipes. The phylogeny of Wolbachia, based on an analysis of MLST genotyping, showed that Wolbachia from P. fuscipes belongs to supergroup B. Quantitative PCR indicated that the infection density in adults was higher than in any other life stage (egg, larva or pupa), and that reproductive tissue in adults had the highest infection densities, with similar densities in the sexes. These findings provide a starting point for understanding Wolbachia infection dynamics in P. fuscipes, and interactions with other components of the microbiota.The maternally inherited obligate intracellular bacteria Wolbachia infects the reproductive tissues of a wide range of arthropods and affects host reproduction. Wolbachia is a credible biocontrol agent for reducing the impact of diseases associated with arthropod vectors. Paederus fuscipes is a small staphylinid beetle that causes dermatitis linearis and conjunctivitis in humans when they come into contact with skin. Wolbachia occur in this beetle, but their relatedness to other Wolbachia, their infection dynamics, and their potential host effects remain unknown. In this study, we report the phylogenetic position and density dynamics of Wolbachia in P. fuscipes. The phylogeny of Wolbachia based on an analysis of MLST genotyping showed that the bacteria from P. fuscipes belong to supergroup B. Quantitative PCR indicated that the infection density in adults was higher than in any other life stage (egg, larva or pupa), and that reproductive tissue in adults had the highest infection densities, with similar densities in the sexes. These findings provide a starting point for understanding the Wolbachia infection dynamics in P. fuscipes, and interactions with other components of the microbiota.

English

Wuchereria bancrofti has been reported to cause 90% of all cases of lymphatic filariasis in Africa. wolbachia endosymbiot infect a wide range of insects and nematodes. This study was conducted in 48 settlements (8 from each division). A total of 2003 mosquitoes was pooled into 120 clusters from the 48 settlements in The Gambia. Polymerase chain reaction (PCR) was used to detect the presence of Wolbachia and filarial nematode and further differentiate Wolbachia into super groups among the positive samples. The level of association between Wolbachia and the genera of anthropophilic mosquitoes in The Gambia was also determined. Microscopic results showed 64.9% Anopheles, 32.0% Culex, 3.0% Aedes. PCR showed that, 34.17% of Wolbachia in the mosquito obtained varied among the three mosquito genera, with highest being among Anopheles. Only Wolbachia super group B was identified in Culex and Aedes. The purpose of this study carried out in The Gambia was to describe the most common mosquito species and to identify, by means of PCR, their association with the Wolbachia bacteria and the presence of nematodes responsible for filiariasis, in order to understand the role that this bacterium plays in the chain of filarial transmission. Key words: Wuchereria bancrofti, Filariasis, Anopheles gambiae, Culex quinquefasciatus, elephantiasis.

Natural Wolbachia infection in field-collected Anopheles and other mosquito species from Malaysia

BackgroundThe endosymbiont bacterium Wolbachia is maternally inherited and naturally infects some filarial nematodes and a diverse range of arthropods, including mosquito vectors responsible for disease transmission in humans. Previously, it has been found infecting most mosquito species but absent in Anopheles and Aedes aegypti. However, recently these two mosquito species were found to be naturally infected with Wolbachia. We report here the extent of Wolbachia infections in field-collected mosquitoes from Malaysia based on PCR amplification of the Wolbachia wsp and 16S rRNA genes.MethodsThe prevalence of Wolbachia in Culicinae mosquitoes was assessed via PCR with wsp primers. For some of the mosquitoes, in which the wsp primers failed to amplify a product, Wolbachia screening was performed using nested PCR targeting the 16S rRNA gene. Wolbachia sequences were aligned using Geneious 9.1.6 software, analyzed with BLAST, and the most similar sequences were downloaded. Phylogenetic analyses were carried out with MEGA 7.0 software. Graphs were drawn with GraphPad Prism 8.0 software.ResultsA total of 217 adult mosquitoes representing 26 mosquito species were screened. Of these, infections with Wolbachia were detected in 4 and 15 mosquito species using wsp and 16S rRNA primers, respectively. To our knowledge, this is the first time Wolbachia was detected using 16S rRNA gene amplification, in some Anopheles species (some infected with Plasmodium), Culex sinensis, Culex vishnui, Culex pseudovishnui, Mansonia bonneae and Mansonia annulifera. Phylogenetic analysis based on wsp revealed Wolbachia from most of the mosquitoes belonged to Wolbachia Supergroup B. Based on 16S rRNA phylogenetic analysis, the Wolbachia strain from Anopheles mosquitoes were more closely related to Wolbachia infecting Anopheles from Africa than from Myanmar.ConclusionsWolbachia was found infecting Anopheles and other important disease vectors such as Mansonia. Since Wolbachia can affect its host by reducing the life span and provide resistance to pathogen infection, several studies have suggested it as a potential innovative tool for vector/vector-borne disease control. Therefore, it is important to carry out further studies on natural Wolbachia infection in vector mosquitoes’ populations as well as their long-term effects in new hosts and pathogen suppression.

Modeling the metabolic interplay between a parasitic worm and its bacterial endosymbiont allows the identification of novel drug targets.

The filarial nematode Brugia malayi represents a leading cause of disability in the developing world, causing lymphatic filariasis in nearly 40 million people. Currently available drugs are not well-suited to mass drug administration efforts, so new treatments are urgently required. One potential vulnerability is the endosymbiotic bacteria Wolbachia-present in many filariae-which is vital to the worm. Genome scale metabolic networks have been used to study prokaryotes and protists and have proven valuable in identifying therapeutic targets, but have only been applied to multicellular eukaryotic organisms more recently. Here, we present iDC625, the first compartmentalized metabolic model of a parasitic worm. We used this model to show how metabolic pathway usage allows the worm to adapt to different environments, and predict a set of 102 reactions essential to the survival of B. malayi. We validated three of those reactions with drug tests and demonstrated novel antifilarial properties for all three compounds.

Wolbachia Inhibits Binding of Dengue and Zika Viruses to Mosquito Cells.

As traditional approaches to the control of dengue and Zika are insufficient, significant efforts have been made to develop utilization of the endosymbiotic bacterium Wolbachia to reduce the ability of mosquitoes to transmit pathogens. Although Wolbachia is known to inhibit flaviviruses in mosquitoes, including dengue virus (DENV) and Zika virus (ZIKV), it remains unclear how the endosymbiont interferes with viral replication cycle. In this study, we have carried out viral binding assays to investigate the impact of the Wolbachia strain wAlbB on the attachment of DENV serotype 2 (DENV-2) and ZIKV to Aedes aegypti Aag-2 cells. RNA interference (RNAi) was used to silence a variety of putative mosquito receptors of DENV that were differentially regulated by wAlbB in Aag-2 cells, in order to identify host factors involved in the inhibition of viral binding. Our results showed that, in addition to suppression of viral replication, Wolbachia strongly inhibited binding of both DENV-2 and ZIKV to Aag-2 cells. Moreover, the expression of two putative mosquito DENV receptors – dystroglycan and tubulin – was downregulated by wAlbB, and their knock-down resulted in the inhibition of DENV-2 binding to Aag-2 cells. These results will aid in understanding the Wolbachia-DENV interactions in mosquito and the development of novel control strategies for mosquito-borne diseases.

Genome Report: Whole Genome Sequence and Annotation of the Parasitoid Jewel Wasp Nasonia giraulti Laboratory Strain RV2X[u

Jewel wasps in the genus of Nasonia are parasitoids with haplodiploidy sex determination, rapid development and are easy to culture in the laboratory. They are excellent models for insect genetics, genomics, epigenetics, development, and evolution. Nasonia vitripennis (Nv) and N. giraulti (Ng) are closely-related species that can be intercrossed, particularly after removal of the intracellular bacterium Wolbachia, which serve as a powerful tool to map and positionally clone morphological, behavioral, expression and methylation phenotypes. The Nv reference genome was assembled using Sanger, PacBio and Nanopore approaches and annotated with extensive RNA-seq data. In contrast, Ng genome is only available through low coverage resequencing. Therefore, de novo Ng assembly is in urgent need to advance this system. In this study, we report a high-quality Ng assembly using 10X Genomics linked-reads with 670X sequencing depth. The current assembly has a genome size of 259,040,977 bp in 3,160 scaffolds with 38.05% G-C and a 98.6% BUSCO completeness score. 97% of the RNA reads are perfectly aligned to the genome, indicating high quality in contiguity and completeness. A total of 14,777 genes are annotated in the Ng genome, and 72% of the annotated genes have a one-to-one ortholog in the Nv genome. We reported 5 million Ng-Nv SNPs which will facility mapping and population genomic studies in Nasonia. In addition, 42 Ng-specific genes were identified by comparing with Nv genome and annotation. This is the first de novo assembly for this important species in the Nasonia model system, providing a useful new genomic toolkit.

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.

Reduced dengue incidence following deployments of Wolbachia-infected Aedes aegypti in Yogyakarta, Indonesia: a quasi-experimental trial using controlled interrupted time series analysis

Background: Ae. aegypti mosquitoes stably transfected with the intracellular bacterium Wolbachia pipientis ( wMel strain) have been deployed for biocontrol of dengue and related arboviral diseases in multiple countries. Field releases in northern Australia have previously demonstrated near elimination of local dengue transmission from Wolbachia-treated communities, and pilot studies in Indonesia have demonstrated the feasibility and acceptability of the method. We conducted a quasi-experimental trial to evaluate the impact of scaled Wolbachia releases on dengue incidence in an endemic setting in Indonesia. Methods: In Yogyakarta City, Indonesia, following extensive community engagement, wMel Wolbachia-carrying mosquitoes were released every two weeks for 13–15 rounds over seven months in 2016–17, in a contiguous 5 km 2 area (population 65,000). A 3 km 2 area (population 34,000) on the opposite side of the city was selected a priori as an untreated control area. Passive surveillance data on notified hospitalised dengue patients was used to evaluate the epidemiological impact of Wolbachia deployments, using controlled interrupted time-series analysis. Results: Rapid and sustained introgression of wMel Wolbachia into local Ae. aegypti populations was achieved. Thirty-four dengue cases were notified from the intervention area and 53 from the control area (incidence 26 vs 79 per 100,000 person-years) during 24 months following Wolbachia deployment. This corresponded in the regression model to a 73% reduction in dengue incidence (95% confidence interval 49%,86%) associated with the Wolbachia intervention. Exploratory analysis including 6 months additional post-intervention observations showed a small strengthening of this effect (30 vs 115 per 100,000 person-years; 76% reduction in incidence, 95%CI 60%,86%). Conclusions: We demonstrate a significant reduction in dengue incidence following successful introgression of Wolbachia into local Ae. aegypti populations in an endemic setting in Indonesia. These findings are consistent with previous field trials in northern Australia, and support the effectiveness of this novel approach for dengue control.

Dynamics of a two-sex model for the population ecology of dengue mosquitoes in the presence of Wolbachia

The release of Wolbachia-infected mosquitoes into the population of wild mosquitoes is one of the promising biological control method for combating the population abundance of mosquitoes that cause deadly diseases, such as dengue. In this study, a new two-sex mathematical model for the population ecology of dengue mosquitoes and disease is designed and used to assess the population-level impact of the periodic release of Wolbachia-infected mosquitoes. Rigorous analysis of the model, which incorporates many of the lifecycle features of dengue disease and the cytoplasmic incompatibility property of Wolbachia bacterium in mosquitoes, reveal that the disease-free equilibrium of the model is locally-asymptotically stable whenever a certain epidemiological threshold, known as the reproduction number of the model (denoted by R0W), is less than unity. The model is shown, using centre manifold theory, to undergo the phenomenon of backward bifurcation at R0W=1. The consequence of this bifurcation is that Wolbachia may not persist, or dengue disease may not be effectively-controlled, when R0W is less than unity. Such persistence and elimination will depend on the initial sizes of the sub-populations of the model. Two mechanisms were identified for which the backward bifurcation phenomenon can be removed. When backward bifurcation does not occur, the associated non-trivial disease-free equilibrium is shown to be globally-asymptotically stable when the reproduction number of the model is less than unity. Numerical simulations, using data relevant to dengue transmission dynamics in northern Queensland, Australia, shows that releasing Wolbachia-infected mosquitoes every three weeks, for a one-year duration, can lead to the effective control of the population abundance of the local wild mosquitoes, and that such effective control increases with increasing number of Wolbachia-infected mosquitoes released (resulting in the reduction of over 90% of the wild mosquito population from their baseline values). Furthermore, simulations show that releasing only adult male Wolbachia-infected mosquitoes provide more beneficial population-level impact (in terms of reducing the population abundance of the wild mosquitoes), in comparison to releasing adult female Wolbachia-infected mosquitoes. Increasing the frequency of Wolbachia release (e.g., from the default release frequency of every three weeks to weekly) does not significantly affect the effectiveness of the Wolbachia-based control program in curtailing the local abundance of the wild mosquitoes. Finally, it was shown that the cytoplasmic incompatibility property of Wolbachia bacterium does not significantly affect the effectiveness of the Wolbachia-based mosquito control strategy implemented in the community.

Wolbachia Inter-Strain Competition and Inhibition of Expression of Cytoplasmic Incompatibility in Mosquito

Successful field trials have been reported as part of the effort to develop the maternally transmitted endosymbiontic bacteria Wolbachia as an intervention agent for controlling mosquito vectors and their transmitted diseases. In order to further improve this novel intervention, artificially transinfected mosquitoes must be optimized to display maximum pathogen blocking, the desired cytoplasmic incompatibility (CI) pattern, and the lowest possible fitness cost. Achieving such optimization, however, requires a better understanding of the interactions between the host and various Wolbabachia strains and their combinations. Here, we transferred the Wolbachia wMel strain by embryonic microinjection into Aedes albopictus, resulting in the successful establishment of a transinfected line, HM (wAlbAwAlbBwMel), with a triple-strain infection comprising wMel, wAlbA, and wAlbB. Surprisingly, no CI was induced when the triply infected males were crossed with the wild-type GUA females or with another triply infected HC females carrying wPip, wAlbA, and wAlbB, but specific removal of wAlbA from the HM (wAlbAwAlbBwMel) line resulted in the expression of CI after crosses with lines infected by either one, two, or three strains of Wolbachia. The transinfected line showed perfect maternal transmission of the triple infection, with fluctuating egg hatch rates that improved to normal levels after repeated outcrosses with GUA line. Strain-specific qPCR assays showed that wMel and wAlbB were present at the highest densities in the ovaries and midguts, respectively, of the HM (wAlbAwAlbBwMel) mosquitoes. These finding suggest that introducing a novel strain of Wolbachia into a Wolbachia-infected host may result in complicated interactions between Wolbachia and the host and between the various Wolbachia strains, with competition likely to occur between strains in the same supergroup.

The impact of large-scale deployment of Wolbachia mosquitoes on dengue and other Aedes-borne diseases in Rio de Janeiro and Niterói, Brazil: study protocol for a controlled interrupted time series analysis using routine disease surveillance data

Background: Rio de Janeiro and Niterói are neighbouring cities in southeastern Brazil which experience large dengue epidemics every 2 to 5 years, with >100,000 cases notified in epidemic years. Costs of vector control and direct and indirect costs due to the Aedes-borne diseases dengue, chikungunya and Zika were estimated to total $650 million USD in 2016, but traditional vector control strategies have not been effective in preventing mosquito-borne disease outbreaks. The Wolbachia method is a novel and self-sustaining approach for the biological control of Aedes-borne diseases, in which the transmission potential of Aedes aegypti mosquitoes is reduced by stably transfecting them with the Wolbachia bacterium (wMel strain). This paper describes a study protocol for evaluating the effect of large-scale non-randomised releases of Wolbachia­-infected mosquitoes on the incidence of dengue, Zika and chikungunya in the two cities of Niterói and Rio de Janeiro. This follows a lead-in period since 2014 involving intensive community engagement, regulatory and public approval, entomological surveys, and small-scale pilot releases. Method: The Wolbachia releases during 2017-2019 covered a combined area of 170 km2 with a resident population of 1.2 million, across Niterói and Rio de Janeiro. Untreated areas with comparable historical dengue profiles and demographic characteristics have been identified a priori as comparative control areas in each city. The proposed pragmatic epidemiological approach combines a controlled interrupted time series analysis of routinely notified suspected and laboratory-confirmed dengue and chikungunya cases, together with monitoring of Aedes-borne disease activity utilising outbreak signals routinely used in public health disease surveillance. Discussion: If the current project is successful, this model for control of mosquito-borne disease through Wolbachia releases can be expanded nationally and regionally.

Evaluation of different dosages of doxycycline during the adulticide treatment of heartworm (Dirofilaria immitis) in dogs

The endosymbiont bacteria Wolbachia plays an important role in the pathogenesis and inflammatory immune response to heartworm (Dirofilaria immitis) infection in dogs. Doxycycline is used to reduce Wolbachia from all life stages of heartworm to avoid large releases of the bacteria during the death of the worms. However, the dose and duration currently recommended have been extrapolated from the treatment of other rickettsial infections. Therefore, the aim was to study the dynamics of Wolbachia IgG antibodies in heartworm-infected dogs under adulticide treatment using different dosages of doxycycline. Forty-nine heartworm-infected dogs were recruited. On day 0 (diagnosis), monthly ivermectin (6 μg/kg) was prescribed, as well as daily doxycycline for 30 days, at 10 mg/kg/12 h (n = 13), 5 mg/kg/12 h (n = 19), and 10 mg/kg/24 h (n = 17). Dogs underwent adulticide treatment and blood samples were collected on days 0, 30, 90, and 120. All dogs had antibodies against recombinant Wolbachia surface protein (rWSP), confirming the important role of the bacteria in heartworm. No significant differences were found in anti-rWSP response by presence/absence of microfilariae, or by parasite burden on day 0. In all treated groups, the anti-rWSP antibody response was not significantly different between days 0 and 30 but was significantly lower between days 0 and 120 (p < 0.05). The results of the present study suggest that the administration of a lower dose than currently recommended is sufficient to achieve a significant reduction of Wolbachia in dogs infected by D. immitis.

The impact of large-scale deployment of Wolbachia mosquitoes on dengue and other Aedes-borne diseases in Rio de Janeiro and Niterói, Brazil: study protocol for a controlled interrupted time series analysis using routine disease surveillance data

Background: Rio de Janeiro and Niterói are neighbouring cities in southeastern Brazil which experience large dengue epidemics every 2 to 5 years, with >100,000 cases notified in epidemic years. Costs of vector control and direct and indirect costs due to the Aedes-borne diseases dengue, chikungunya and Zika were estimated to total $650 million USD in 2016, but traditional vector control strategies have not been effective in preventing mosquito-borne disease outbreaks. The Wolbachia method is a novel and self-sustaining approach for the biological control of Aedes-borne diseases, in which the transmission potential of Aedes aegypti mosquitoes is reduced by stably transfecting them with the Wolbachia bacterium ( wMel strain). This paper describes a study protocol for evaluating the effect of large-scale non-randomised releases of Wolbachia­-infected mosquitoes on the incidence of dengue, Zika and chikungunya in the two cities of Niterói and Rio de Janeiro. This follows a lead-in period since 2014 involving intensive community engagement, regulatory and public approval, entomological surveys, and small-scale pilot releases. Method: The Wolbachia releases during 2017-2019 covered a combined area of 170 km 2 with a resident population of 1.2 million, across Niterói and Rio de Janeiro. Untreated areas with comparable historical dengue profiles and demographic characteristics have been identified a priori as comparative control areas in each city. The proposed pragmatic epidemiological approach combines a controlled interrupted time series analysis of routinely notified suspected and laboratory-confirmed dengue and chikungunya cases, together with monitoring of Aedes-borne disease activity utilising outbreak signals routinely used in public health disease surveillance. Discussion: If the current project is successful, this model for control of mosquito-borne disease through Wolbachia releases can be expanded nationally and regionally.

A Review: Wolbachia-Based Population Replacement for Mosquito Control Shares Common Points with Genetically Modified Control Approaches.

The growing expansion of mosquito vectors has made mosquito-borne arboviral diseases a global threat to public health, and the lack of licensed vaccines and treatments highlight the urgent need for efficient mosquito vector control. Compared to genetically modified control strategies, the intracellular bacterium Wolbachia, endowing a pathogen-blocking phenotype, is considered an environmentally friendly strategy to replace the target population for controlling arboviral diseases. However, the incomplete knowledge regarding the pathogen-blocking mechanism weakens the reliability of a Wolbachia-based population replacement strategy. Wolbachia infections are also vulnerable to environmental factors, temperature, and host diet, affecting their densities in mosquitoes and thus the virus-blocking phenotype. Here, we review the properties of the Wolbachia strategy as an approach to control mosquito populations in comparison with genetically modified control methods. Both strategies tend to limit arbovirus infections but increase the risk of selecting arbovirus escape mutants, rendering these strategies less reliable.

The impact of city-wide deployment of Wolbachia-carrying mosquitoes on arboviral disease incidence in Medellín and Bello, Colombia: study protocol for an interrupted time-series analysis and a test-negative design study.

Background: Dengue, chikungunya and Zika are viral infections transmitted by Aedes aegypti mosquitoes, and present major public health challenges in tropical regions. Traditional vector control methods have been ineffective at halting disease transmission. The World Mosquito Program has developed a novel approach to arbovirus control using Ae. aegypti stably transfected with the Wolbachia bacterium, which have significantly reduced ability to transmit dengue, Zika and chikungunya in laboratory experiments. Field releases in eight countries have demonstrated Wolbachia establishment in local Ae. aegypti populations. Methods: We describe a pragmatic approach to measuring the epidemiological impact of city-wide Wolbachia deployments in Bello and Medellín, Colombia. First, an interrupted time-series analysis will compare the incidence of dengue, chikungunya and Zika case notifications before and after Wolbachia releases, across the two municipalities. Second, a prospective case-control study using a test-negative design will be conducted in one quadrant of Medellín. Three of the six contiguous release zones in the case-control area were allocated to receive the first Wolbachia deployments in the city and three to be treated last, approximating a parallel two-arm trial for the >12-month period during which Wolbachia exposure remains discordant. Allocation, although non-random, aimed to maximise balance between arms in historical dengue incidence and demographics. Arboviral disease cases and arbovirus-negative controls will be enrolled concurrently from febrile patients presenting to primary care, with case/control status classified retrospectively following laboratory diagnostic testing. Intervention effect is estimated from an aggregate odds ratio comparing Wolbachia-exposure odds among test-positive cases versus test-negative controls. Discussion: The study findings will add to an accumulating body of evidence from global field sites on the efficacy of the Wolbachia method in reducing arboviral disease incidence, and can inform decisions on wider public health implementation of this intervention in the Americas and beyond. Trial registration: ClinicalTrials.gov: NCT03631719. Registered on 15 August 2018.