Bacterial gene essentiality under modeled microgravity

The health of eukaryotic hosts is tightly connected to relationships with symbiotic microorganisms, yet how these relationships develop and evolve during long-duration spaceflight is not well understood. In this study, we asked what bacterial genes are required for growth under modeled, or simulated, microgravity conditions compared to normal gravity controls. To conduct this study, we focused on the marine bacterium Vibrio fischeri, which forms a monospecific symbiosis with the Hawaiian bobtail squid, Euprymna scolopes. The symbiosis has been studied during spaceflight and in ground-based modeled microgravity conditions. We employed a library of over 40,000 V. fischeri transposon mutants and compared the fitness of mutants in modeled microgravity compared to the gravity controls using transposon insertion sequencing (INSeq). We identified dozens of genes that exhibited fitness defects under both conditions, likely due to the controlled anaerobic environment, yet we identified relatively few genes with differential effects under modeled microgravity or gravity specifically: only mutants in rodA were more depleted under modeled microgravity, and mutants in 12 genes exhibited greater depletion under gravity conditions. We additionally compared RNA-seq and INSeq data and determined that expression under microgravity was not predictive of the essentiality of a given gene. In summary, empirical determination of conditional gene essentiality identifies few microgravity-specific genes for environmental growth of V. fischeri, suggesting that the condition of microgravity has a minimal impact on symbiont gene requirement.