Wolbachia feminizes a spider host with assistance from co-infecting symbionts

Arthropods commonly harbor maternally-transmitted bacterial endosymbionts that manipulate host biology. Multiple heritable symbionts can co-infect the same individual, allowing these host-restricted bacteria to engage in cooperation or conflict, which can ultimately affect host phenotype. The spider Mermessus fradeorum is variably infected with up to five heritable symbionts: Rickettsiella (R), Tisiphia (T), and three strains of Wolbachia (W1-3). Quintuply infected spiders are feminized, causing genetic males to develop as phenotypic females and produce almost exclusively female offspring. By comparing feminization across nine infection combinations, we identified a feminizing strain of Wolbachia, W1. We also observed that spiders infected with both W1 and W3 produced [~]10% more females than those lacking W3. This increase in feminization rate does not seem to be due to direct changes in W1 titer, nor does W1 titer correlate with feminization rate. Instead, we observed subtle titer interactions among symbionts, with lower relative abundance of R and T symbionts in strongly feminized infections. This synergistic effect of co-infection on Wolbachia feminization may help promote the spread of all five symbionts in spider populations. These results confirm the first instance of Wolbachia-induced feminization in spiders and demonstrate that co-infecting symbionts can improve the efficacy of symbiont-induced feminization.