CAREER: Experimental manipulation of host-microbe associations to reveal key features promoting symbiosis

Many animals rely on microbial symbionts to help them gain sufficient nutrients, provide protection from predators and pathogens, or allow them to live in otherwise inhospitable environments. Studying these relationships can be difficult as it is often impossible to separate the microbes from their host. As a result, understanding of the mechanisms that underly these relationships has been limited by the inability to manipulate both host and symbiont independently. Kissing bugs are insects that feed exclusively on vertebrate blood and serve as host to symbiotic bacteria, that help them successfully develop and reproduce. These bacteria also can be cleared from the kissing bugs and infected with new bacteria. The symbiont bacteria can also be grown outside of the insect and can be genetically altered. This system, thus, provides a unique opportunity to experimentally manipulate the host and symbiont, to identify and characterize factors that support the symbiosis. The research will focus on genes involved in symbiont colonization of the host, host and symbiont genes involved in the exchange of nutrients, and interactions of the host immune system and the symbiont. Given the ubiquity of animal-microbe symbioses, this research will illuminate mechanisms that may govern host-microbe interactions in other less-tractable systems. Along with the proposed research, this project will also provide research experience and training to high school teachers. This experience will be paired with curriculum development to incorporate host-microbe interactions into the teachers’ classrooms. Experimental manipulation of host-microbe systems has been limited by inability to separate highly integrated partners or difficulties disentangling the effects of individual members in highly complex communities. Triatomine kissing bugs harbor essential symbionts in their gut which are environmentally acquired each generation, allowing for generation of bacteria-free, axenic insects, which can then be experimentally inoculated with bacteria. This work leverages the unique features of this system – ability to generate axenic and gnotobiotic insects, low-complexity microbiomes, and molecular tools – to delineate the mechanistic basis of symbiosis. Transposon mutagenesis will be used to identify genes in the symbiont Rhodococcus rhodnii that are essential for symbiosis followed by generation of knockout strains of R. rhodnii lacking symbiosis-promoting genes. Knockout strains will be used for bioassays to characterize the role of the gene-of-interest in the relationship. The role of the host immune system in regulating bacterial abundance will be assessed, examining both humoral and cellular immune responses to symbionts. Lipid metabolism genes including fatty acid synthases and fatty acid CoA reductases have been identified as differentially expressed in axenic and gnotobiotic insects. Liquid chromatography and mass spectroscopy will be applied to understand how symbiotic bacteria shape the lipidome of kissing bugs while silencing of specific lipid genes will reveal the role of these genes in lipid metabolism and host fitness. The proposed work will shed new light on the evolution of host-symbiont interactions and serve as a framework for exploring systems which are less amenable to experimental manipulation. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.