Modulation of innate stomatal defense response in plants by Salmonella enterica under gravistimulation

Abstract

Spaceflight presents plants and microorganisms with the evolutionarily novel environment of microgravity. Plant responses to gravistimulation occur at the molecular and physiological level. Studies suggest plant susceptibility to phytopathogens is increased under spaceflight microgravity. Human pathogens, particularly enteric pathogens of the Gram-negative family Enterobacteriaceae, act as cross-kingdom pathogens by suppressing and evading plant innate immunity to colonize and persist in intracellular spaces. As humans venture deeper into space for greater amounts of time, the production of nutritious and safe food during spaceflight is essential to protecting human health. Spacecraft are closed systems in which a microbiome can be introduced via cargo deliveries or from the human crew. There exists an alarming knowledge gap regarding plant interactions with human foodborne pathogens in microgravity. Despite stringent pre-launch biosafety precautions, several genera of foodborne pathogens persist aboard the International Space Station (ISS). Space-grown plants on the ISS have also been colonized by a diverse community of bacteria and fungi, including pathogen-containing genera. Elucidation of interactions between plants and human pathogens under altered gravistimulation could highlight potential food safety risks. The purpose of this study was to determine if gravistimulation by vertical clinorotation alters susceptibility of lettuce (Lactuca sativa L.) to suppression of defensive stomatal closure and colonization by Salmonella enterica serovar Typhimurium. A slow-rotating 2-D clinostat (0-4 RPM) was used to gravistimulate inoculated in-vitro plants. Confocal microscopy was used to measure width of stomatal apertures at 0, 3, 6, and 9 hours post inoculation. Confocal Z-stacking was used to measure the depth of bacterial ingression within stomata. We found 1) different rates of clinorotation dramatically influenced stomatal width in uninoculated plants. 2) clinorotation significantly enhanced Salmonella ability to suppress stomatal closure, particularly at 2 RPM. 3) Bacteria could be observed deeper within stomatal cavities in rotated plants compared to unrotated.