Double trouble: combined action of meiotic drive and Wolbachia feminization in Eurema butterflies

Arthropod sex ratios can be manipulated by a diverse range of selfish genetic elements, including maternally inherited Wolbachia bacteria. Feminization by Wolbachia is rare but has been described for Eurema mandarina butterflies. In this species, some phenotypic and functional females, thought to be ZZ genetic males, are infected with a feminizing Wolbachia strain, wFem. Meanwhile, heterogametic WZ females are not infected with wFem. Here, we establish a quantitative PCR assay allowing reliable sexing in three Eurema species. Against expectation, all E. mandarina females, including wFem females, had only one Z chromosome that was paternally inherited. Observation of somatic interphase nuclei confirmed that W chromatin was absent in wFem females, but present in females without wFem. We conclude that the sex bias in wFem lines is due to meiotic drive (MD) that excludes the maternal Z and thus prevents formation of ZZ males. Furthermore, wFem lines may have lost the W chromosome or harbour a dysfunctional version, yet rely on wFem for female development; removal of wFem results in all-male offspring. This is the first study that demonstrates an interaction between MD and Wolbachia feminization, and it highlights endosymbionts as potentially confounding factors in MD of sex chromosomes.