The recent association of Zika virus with cases of microcephaly has sparked a global health crisis and highlighted the need for mechanisms to combat the Zika vector, Aedes aegypti mosquitoes. Wolbachia pipientis, a bacterial endosymbiont of insect, has recently garnered attention as a mechanism for arbovirus control. Here we report that Aedes aegypti harboring Wolbachia are highly resistant to infection with two currently circulating Zika virus isolates from the recent Brazilian epidemic. Wolbachia-harboring mosquitoes displayed lower viral prevalence and intensity and decreased disseminated infection and, critically, did not carry infectious virus in the saliva, suggesting that viral transmission was blocked. Our data indicate that the use of Wolbachia-harboring mosquitoes could represent an effective mechanism to reduce Zika virus transmission and should be included as part of Zika control strategies.
Wolbachia pipientis is an endosymbiotic bacterium estimated to chronically infect between 40-75% of all arthropod species. Aedes aegypti, the principle mosquito vector of dengue virus (DENV), is not a natural host of Wolbachia. The transinfection of Wolbachia strains such as wAlbB, wMel and wMelPop-CLA into Ae. aegypti has been shown to significantly reduce the vector competence of this mosquito for a range of human pathogens in the laboratory. This has led to wMel-transinfected Ae. aegypti currently being released in five countries to evaluate its effectiveness to control dengue disease in human populations. Here we describe the generation of a superinfected Ae. aegypti mosquito line simultaneously infected with two avirulent Wolbachia strains, wMel and wAlbB. The line carries a high overall Wolbachia density and tissue localisation of the individual strains is very similar to each respective single infected parental line. The superinfected line induces unidirectional cytoplasmic incompatibility (CI) when crossed to each single infected parental line, suggesting that the superinfection would have the capacity to replace either of the single constituent infections already present in a mosquito population. No significant differences in fitness parameters were observed between the superinfected line and the parental lines under the experimental conditions tested. Finally, the superinfected line blocks DENV replication more efficiently than the single wMel strain when challenged with blood meals from viremic dengue patients. These results suggest that the deployment of superinfections could be used to replace single infections and may represent an effective strategy to help manage potential resistance by DENV to field deployments of single infected strains.
Wolbachia pipientis is a widespread intracellular bacterial symbiont of arthropods and is common in insects. One of their more exotic and unexpected hosts is the filarial nematodes, notable for the parasites responsible for onchocerciasis (river blindness), lymphatic filariasis (elephantiasis) and dirofilariasis (heartworm). Wolbachia are only present in a sub-group of the filarial nematodes and do not extend to other groups of nematodes either parasitic or free-living. In the medically and veterinary important species that host Wolbachia, the symbiont has become an essential partner to key biological processes in the life of the nematode to the point where antibiotic elimination of the bacteria leads to a potent and effective anti-filarial drug treatment. We review the cellular and molecular basis of Wolbachia-filarial interactions and highlight the key processes provided by the endosymbiont upon which the nematodes have become entirely dependent. This dependency is primarily restricted to periods of the life-cycle with heavy metabolic demands including growth and development of larval stages and embryogenesis in the adult female. Also, the longevity of filarial parasites is compromised following depletion of the symbiont, which for the first time has delivered a safe and effective treatment to kill adult parasites with antibiotics.
In a placebo controlled field trial, the effects of doxycycline (200mg/day) for 23 days followed by doxycycline (200mg/day) in combination with albendazole (ABZ) (400mg/day) for 7 days on depletion of Wolbachia endobacteria from Wuchereria bancrofti and microfilaricidal activity were studied in 68 patients (34 males and 34 females) from West Bengal, India. The drugs in combination (i.e., doxycycline+ABZ) provided the best efficacy by totally eliminating the circulating microfilaria (mf) (in 42% cases) on day 365 with (99.8%, P<0.05) suppression even on day 365 post-treatment compared to both exclusive doxycycline (69%, P<0.05) and ABZ (89%, P<0.05) groups. Thus, our results have established that a 30-day course of doxycycline in combination with a 7-day course of ABZ is sufficient to ensure long-term reduction in mf level by depleting Wolbachia from worm tissues. Doxycycline combined with ABZ led to a greater reduction in mf density in blood at 4 months (post-treatment) in comparison to doxycycline or ABZ alone. There were significant differences between the three treatments after 12 months (post-treatment). Further, the impact of a 7-day regimen of ABZ was surprisingly good in reducing mf compared to doxycycline-alone group. Adverse reactions were mild. A 30-day course of doxycycline and ABZ in combination is a safe and well-tolerated treatment for lymphatic filariasis with significant activity against microfilaremia.
Aedes aegypti mosquitoes infected with Wolbachia bacteria are currently being released for arbovirus suppression around the world. Their potential to invade populations and persist will depend on interactions with environmental conditions, particularly as larvae are often exposed to fluctuating and extreme temperatures in the field. We reared Ae. aegypti larvae infected with different types of Wolbachia (wMel, wAlbB and wMelPop-CLA) under diurnal cyclical temperatures. Rearing wMel and wMelPop-CLA-infected larvae at 26-37°C reduced the expression of cytoplasmic incompatibility, a reproductive manipulation induced by Wolbachia. We also observed a sharp reduction in the density of Wolbachia in adults. Furthermore, the wMel and wMelPop-CLA infections were not transmitted to the next generation when mosquitoes were exposed to 26-37°C across all life stages. In contrast, the wAlbB infection was maintained at a high density, exhibited complete cytoplasmic incompatibility, and was transmitted from mother to offspring with a high fidelity under this temperature cycle. These findings have implications for the potential success of Wolbachia interventions across different environments and highlight the importance of temperature control in rearing.
Genetic-modification strategies are currently being developed to reduce the transmission of vector-borne diseases, including African trypanosomiasis. For tsetse, the vector of African trypanosomiasis, a paratransgenic strategy is being considered: this approach involves modification of the commensal symbiotic bacteria Sodalis to express trypanosome-resistance-conferring products. Modified Sodalis can then be driven into the tsetse population by cytoplasmic incompatibility (CI) from Wolbachia bacteria. To evaluate the effectiveness of this paratransgenic strategy in controlling African trypanosomiasis, we developed a three-species mathematical model of trypanosomiasis transmission among tsetse, humans, and animal reservoir hosts. Using empirical estimates of CI parameters, we found that paratransgenic tsetse have the potential to eliminate trypanosomiasis, provided that any extra mortality caused by Wolbachia colonization is low, that the paratransgene is effective at protecting against trypanosome transmission, and that the target tsetse species comprises a large majority of the tsetse population in the release location.
Lymphatic filariasis and onchocerciasis are two important neglected tropical diseases (NTDs) that cause severe disability. Control efforts are hindered by the lack of a safe macrofilaricidal drug. Targeting the Wolbachia bacterial endosymbionts in these parasites with doxycycline leads to a macrofilaricidal outcome, but protracted treatment regimens and contraindications restrict its widespread implementation. The Anti-Wolbachia consortium aims to develop improved anti-Wolbachia drugs to overcome these barriers. We describe the first screening of a large, diverse compound library against Wolbachia. This whole-organism screen, streamlined to reduce bottlenecks, produced a hit rate of 0.5%. Chemoinformatic analysis of the top 50 hits led to the identification of six structurally diverse chemotypes, the disclosure of which could offer interesting avenues of investigation to other researchers active in this field. An example of hit-to-lead optimization is described to further demonstrate the potential of developing these high-quality hit series as safe, efficacious, and selective anti-Wolbachia macrofilaricides.
The endosymbiotic bacteria, Wolbachia, induce neutrophilic responses to the human helminth pathogen Onchocerca volvulus. The formation of Neutrophil Extracellular Traps (NETs), has been implicated in anti-microbial defence, but has not been identified in human helminth infection. Here, we demonstrate NETs formation in human onchocerciasis. Extracellular NETs and neutrophils were visualised around O. volvulus in nodules excised from untreated patients but not in nodules from patients treated with the anti-Wolbachia drug, doxycycline. Whole Wolbachia or microspheres coated with a synthetic Wolbachia lipopeptide (WoLP) of the major nematode Wolbachia TLR2/6 ligand, peptidoglycan associated lipoprotein, induced NETosis in human neutrophils in vitro. TLR6 dependency of Wolbachia and WoLP NETosis was demonstrated using purified neutrophils from TLR6 deficient mice. Thus, we demonstrate for the first time that NETosis occurs during natural human helminth infection and demonstrate a mechanism of NETosis induction via Wolbachia endobacteria and direct ligation of Wolbachia lipoprotein by neutrophil TLR2/6.
In many regions of the world, mosquito-borne viruses pose a growing threat to human health. As an alternative to traditional control measures, the bacterial symbiont Wolbachia has been transferred from Drosophila into the mosquito Aedes aegypti, where it can block the transmission of dengue and Zika viruses. A recent paper has reported large-scale releases of Wolbachia-infected Ae. aegypti in the city of Cairns, Australia. Wolbachia, which is maternally transmitted, invaded and spread through the populations due to a sperm-egg incompatibility called cytoplasmic incompatibility. Over a period of 2 years, a wave of Wolbachia infection slowly spread out from 2 release sites, demonstrating that it will be possible to deploy this strategy in large urban areas. In line with theoretical predictions, Wolbachia infection at a third, smaller release site collapsed due to the immigration of Wolbachia-free mosquitoes from surrounding areas. This remarkable field experiment has both validated theoretical models of Wolbachia population dynamics and demonstrated that this is a viable strategy to modify mosquito populations.
Endosymbiosis is a unique form of interaction between organisms, with one organism dwelling inside the other. One of the most widespread endosymbionts is Wolbachia pipientis, a maternally transmitted bacterium carried by insects, crustaceans, mites and filarial nematodes. Though candidate proteins have been identified that contribute to maternal transmission, the molecular basis for maternal Wolbachia transmission remains largely unknown. To investigate transmission-related processes that respond to Wolbachia infection, ovarian proteomes were analyzed from Wolbachia-infected D. melanogaster and D. simulans Endogenous and variant host-strain combinations were investigated. Significant and differentially abundant ovarian proteins were detected, indicating substantial regulatory changes in response to Wolbachia Variant Wolbachia strains were associated with broader impact on the ovary proteome than endogenous Wolbachia infections. The D. melanogaster ovarian environment also exhibited a higher diversity of proteomic responses to Wolbachia than D. simulans Overall, many Wolbachia-responsive ovarian proteins detected in this study were consistent with expectations from experimental literature. This suggests that context-specific changes in protein abundance contribute to Wolbachia manipulation of transmission-related mechanisms in oogenesis.