Fitness costs of Wolbachia shift in locally-adapted Aedes aegypti mosquitoes.

Aedes aegypti mosquito eggs can remain quiescent for many months before hatching, allowing populations to persist through unfavourable conditions. A. aegypti infected with the Wolbachia strain wMel have been released in tropical and subtropical regions for dengue control. wMel reduces the viability of quiescent eggs, but this physiological cost might be expected to evolve in natural mosquito populations that frequently experience stressful conditions. We found that the cost of wMel infection differed consistently between mosquitoes collected from different locations and became weaker across laboratory generations, suggesting environment-specific adaptation of mosquitoes to the wMel infection. Reciprocal crossing experiments show that differences in the cost of wMel to quiescent egg viability were mainly due to mosquito genetic background and not Wolbachia origin. wMel-infected mosquitoes hatching from long-term quiescent eggs showed partial loss of cytoplasmic incompatibility and female infertility, highlighting additional costs of long-term quiescence. Our study provides the first evidence for a shift in Wolbachia phenotypic effects following deliberate field release and establishment and it highlights interactions between Wolbachia infections and mosquito genetic backgrounds. The unexpected changes in fitness costs observed here suggest potential tradeoffs with undescribed fitness benefits of the wMel infection.

A LAMP assay for the rapid and robust assessment of Wolbachia infection in Aedes aegypti under field and laboratory conditions.

With Wolbachia-based arbovirus control programs being scaled and operationalised around the world, cost effective and reliable detection of Wolbachia in field samples and laboratory stocks is essential for quality control. Here we validate a modified loop-mediated isothermal amplification (LAMP) assay for routine scoring of Wolbachia in mosquitoes from laboratory cultures and the field, applicable to any setting. We show that this assay is a rapid and robust method for highly sensitive and specific detection of wAlbB Wolbachia infection within Aedes aegypti under a variety of conditions. We test the quantitative nature of the assay by evaluating pooled mixtures of Wolbachia-infected and uninfected mosquitoes and show that it is capable of estimating infection frequencies, potentially circumventing the need to perform large-scale individual analysis for wAlbB infection status in the course of field monitoring. These results indicate that LAMP assays are useful for routine screening particularly under field conditions away from laboratory facilities.

A comprehensive assessment of inbreeding and laboratory adaptation in Aedes aegypti mosquitoes.

Modified Aedes aegypti mosquitoes reared in laboratories are being released around the world to control wild mosquito populations and the diseases they transmit. Several efforts have failed due to poor competitiveness of the released mosquitoes. We hypothesized that colonized mosquito populations could suffer from inbreeding depression and adapt to laboratory conditions, reducing their performance in the field. We established replicate populations of Ae. aegypti mosquitoes collected from Queensland, Australia, and maintained them in the laboratory for twelve generations at different census sizes. Mosquito colonies maintained at small census sizes (≤100 individuals) suffered from inbreeding depression due to low effective population sizes which were only 25% of the census size as estimated by SNP markers. Populations that underwent full-sib mating for nine consecutive generations had greatly reduced performance across all traits measured. We compared the established laboratory populations with their ancestral population resurrected from quiescent eggs for evidence of laboratory adaptation. The overall performance of laboratory populations maintained at a large census size (400 individuals) increased, potentially reflecting adaptation to artificial rearing conditions. However, most individual traits were unaffected, and patterns of adaptation were not consistent across populations. Differences between replicate populations may indicate that founder effects and drift affect experimental outcomes. Though we find limited evidence of laboratory adaptation, mosquitoes maintained at low population sizes can clearly suffer fitness costs, compromising the success of "rear-and-release" strategies for arbovirus control.

Costs of Three Wolbachia Infections on the Survival of Aedes aegypti Larvae under Starvation Conditions.

The mosquito Aedes aegypti, the principal vector of dengue virus, has recently been infected experimentally with Wolbachia: intracellular bacteria that possess potential as dengue biological control agents. Wolbachia depend on their hosts for nutrients they are unable to synthesize themselves. Consequently, competition between Wolbachia and their host for resources could reduce host fitness under the competitive conditions commonly experienced by larvae of Ae. aegypti in the field, hampering the invasion of Wolbachia into natural mosquito populations. We assess the survival and development of Ae. aegypti larvae under starvation conditions when infected with each of three experimentally-generated Wolbachia strains: wMel, wMelPop and wAlbB, and compare their fitness to wild-type uninfected larvae. We find that all three Wolbachia infections reduce the survival of larvae relative to those that are uninfected, and the severity of the effect is concordant with previously characterized fitness costs to other life stages. We also investigate the ability of larvae to recover from extended food deprivation and find no effect of Wolbachia on this trait. Aedes aegypti larvae of all infection types were able to resume their development after one month of no food, pupate rapidly, emerge at a large size, and exhibit complete cytoplasmic incompatibility and maternal transmission. A lowered ability of Wolbachia-infected larvae to survive under starvation conditions will increase the threshold infection frequency required for Wolbachia to establish in highly competitive natural Ae. aegypti populations and will also reduce the speed of invasion. This study also provides insights into survival strategies of larvae when developing in stressful environments.