Endosymbiotic Candidates for Parasitoid Defense in Exotic and Native New Zealand Weevils

Some insects are infected with maternally inherited bacterial endosymbionts that protect them against pathogens or parasitoids. The weevil Sitona obsoletus (=Sitona lepidus) is invasive in New Zealand, and suspected to contain such defensive symbionts, because it is particularly resistant to a Moroccan strain of the parasitoid Microctonus aethiopoides (which successfully attacks many other weevil species), and shows geographic variation in susceptibility to an Irish strain of the same parasitoid. Using 454 pyrosequencing, we investigated the bacterial community associated with S. obsoletus, two other exotic weevils (Sitona discoideus and Listronotus bonariensis) and two endemic New Zealand weevils (Irenimus aequalis and Steriphus variabilis). We found that S. obsoletus was infected by one strain of Wolbachia and two strains of Rickettsia, none of which were found in any other weevil species examined. Using diagnostic PCR, we found that S. obsoletus in the Northland region, where parasitism is highly variable, were primarily infected with Wolbachia and Rickettsia strain 2, indicating that these two symbionts should be investigated for potential defensive properties. In comparison, S. discoideus lacked any apparent maternally inherited bacterial endosymbionts. In the other weevil species, we found a different strain of Wolbachia and two different strains of Spiroplasma. Two weevil species (St. variabilis and L. bonariensis) were infected with distinct strains of Nardonella, the ancestral endosymbiont of weevils, whereas three weevil species (S. obsoletus, S. discoideus, and I. aequalis) lacked evidence for Nardonella infection. However, I. aequalis was consistently infected with a novel Enterobacteriaceae strain, suggesting that a symbiont replacement may have taken place, similar to that described for other weevil clades.