Intraspecific variation and molecular regulation of a multipartite defensive symbiosis

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Symbiotic microorganisms can protect animal hosts against pathogens by producing antimicrobial compounds, but the selective pressures shaping the defensive chemistry under lab and field conditions and the molecular factors underlying symbiosis establishment remain poorly understood. While our project of the first funding phase was targeted towards unraveling the selective pressures shaping antimicrobial defense cocktails under controlled laboratory conditions, the current proposal aims to characterize the variation in a symbiont-produced chemical defense in nature (both qualitatively and quantitatively), and to identify the symbiont genes that are necessary for establishing the association with its insect host. Using the Lagria beetle-Burkholderia symbiosis as an experimentally and genetically tractable system, we aim to (1) characterize the natural intra- and interspecific variation in microbial communities and defensive chemistry in two Lagria species and (2) unravel the molecular factors underlying the symbionts’ dual lifestyle, switching between plant pathogenicity and insect mutualism. To this end, we will collect beetles of two Lagria species in the field and characterize their microbial community profiles by high-throughput sequencing, while at the same time analyzing the symbiont-produced defensive chemistry via HPLC-MS/MS. The results will provide valuable insights into the natural variation in defensive chemistry and microbial community profiles between individuals, populations, and species of Lagria beetles. For objective 2, we will test candidate symbiont colonization factors from a previous transposon-insertion sequencing experiment by generating targeted mutants and assessing their colonization ability in the beetle. As the beetle symbionts can also opportunistically infect the beetle’s food plant, we will perform a transposon-seq experiment to elucidate candidate colonization factors for infecting the plant. The results will yield insights into the molecular factors underlying defensive symbiosis establishment as well as the life-style switch between insect mutualism and plant pathogenicity.