Parasitism causes changes in caterpillar odours and associated bacterial communities with consequences for host-location by a hyperparasitoid

Microorganisms living in and on macroorganisms may produce microbial volatile compounds (mVOCs) that characterise organismal odours. The mVOCs might thereby provide a reliable cue to carnivorous enemies in locating their host or prey. Parasitism by parasitoid wasps might alter the microbiome of their caterpillar host, affecting organismal odours and interactions with insects of higher trophic levels such as hyperparasitoids. Hyperparasitoids parasitise larvae or pupae of parasitoids, which are often concealed or inconspicuous. Odours of parasitised caterpillars aid them to locate their host, but the origin of these odours and its relationship to the caterpillar microbiome are unknown. Here, we analysed the odours and microbiome of the large cabbage white caterpillar Pieris brassicae in relation to parasitism by its endoparasitoid Cotesia glomerata. We identified how bacterial presence in and on the caterpillars is correlated with caterpillar odours and tested the attractiveness of parasitised and unparasitised caterpillars to the hyperparasitoid Baryscapus galactopus. We manipulated the presence of the external microbiome and the transient internal microbiome of caterpillars to identify the microbial origin of odours. We found that parasitism by C. glomerata led to the production of five characteristic volatile products and significantly affected the internal and external microbiome of the caterpillar, which were both found to have a significant correlation with caterpillar odours. The preference of the hyperparasitoid was correlated with the presence of the external microbiome. Likely, the changes in external microbiome and body odour after parasitism were driven by the resident internal microbiome of caterpillars, where the bacterium Wolbachia sp. was only present after parasitism. Micro-injection of Wolbachia in unparasitised caterpillars increased hyperparasitoid attraction to the caterpillars compared to untreated caterpillars, while no differences were found compared to parasitised caterpillars. In conclusion, our results indicate that host-parasite interactions can affect multi-trophic interactions and hyperparasitoid olfaction through alterations of the microbiome. Author summaryBacteria living in and on macroorganisms produce the majority of their odours and thereby play an essential role in species interactions. For example, the hyperparasitoid enemies of parasitic wasps can reliably use odours of parasitised caterpillars to find the parasitic wasp larvae developing inside the caterpillar. We studied whether parasitism of caterpillars caused changes in caterpillar odours and associated internal and external bacterial communities to elucidate whether specific bacteria are responsible for the odours emitted by parasitised caterpillars. We identified which bacteria corresponded with odour production and whether hyperparasitoids could still find parasitised caterpillars that had their bacterial communities experimentally disrupted. We found that parasitised caterpillars have different odour profiles compared to unparasitised caterpillars and that bacteria in and on caterpillars correspond with these changes. The hyperparasitoid was less attracted to parasitised caterpillars with a disrupted skin microbiome. A characteristic bacterium, Wolbachia sp., which was only found inside parasitised caterpillars, may have initiated changes in the external microbiome and effects on odour emission. Furthermore, we show that micro-injection of Wolbachia in unparasitized caterpillars enhanced hyperparasitoid attraction. Hence, our results indicate that parasitoid wasps indirectly reveal themselves to their hyperparasitoid natural enemies by alteration of the bacterial communities of their host.

Automated summary

Microorganisms living in and on macroorganisms can produce volatile compounds that can help carnivorous enemies locate their host or prey. Parasitism by parasitoid wasps can alter the microbiome and odour of their caterpillar host, which can affect interactions with other insects. This study analyzed the odours and microbiome of cabbage white caterpillars in relation to parasitism by endoparasitoid wasps and found that parasitism led to the production of characteristic volatile compounds and significant changes in the caterpillar's microbiome. The preference of a hyperparasitoid was correlated with the presence of the external microbiome.