Rapid and convergent evolution of Wolbachia genomes in mosquito cell culture

Wolbachia are widespread bacterial symbionts of arthropods that manipulate host reproduction and can confer fitness benefits, such as protection against viral pathogens. Their self-spreading ability coupled with their antiviral effect has been harnessed in health programmes to curb the transmission of dengue virus by Aedes mosquitoes. In this context, a better understanding of evolutionary dynamics over short timescales is critical to inform and improve the efficacy of Wolbachia-based disease control interventions. Moreover, experimental evolution of Wolbachia may provide a powerful approach for generating novel symbiont strains with improved properties for disease control. Here we generated new mosquito cell lines by introducing five Wolbachia strains from Drosophila into the symbiont-free Aedes albopictus Aa23 cell line. Following transinfection, long-read whole-genome sequencing revealed multiple mutations, including structural changes, some of them being associated with major shifts in bacterial density. Signs of convergent evolution, an excess of non-synonymous mutations and comparisons with previous studies, indicate that Wolbachia evolution in cell culture is predominantly driven by selection and occurs faster than in insect hosts. Altogether, our study demonstrates that cell culture is a valuable tool to investigate short-term symbiont evolution, explore the genetic basis of bacterial density regulation and generate higher-density symbiont strains suitable for disease control.