The rise of antibiotic-resistant bacteria has reached an alarming level. The recently discovered type VI secretion system (T6SS) appears to be a bacterial defense mechanism that could potentially become involved with addressing this concern. Previous studies suggest Vibrio spp. may express T6SS. In this study, Vibrio parahaemolyticus, a common foodborne pathogen, is used as a model to study T6SS. The objectives of this study were to evaluate culturing conditions that would stimulate swarming motility in strains of V. parahaemolyticus and to detect for the presence of T6SS based on phenotypic criteria. Different media (BHI, TSAS), agar concentration (0.7%, 1%, 2%) and incubation temperature (25°C and 37°C) were tested to determine the optimal environment that induces swarming in strains of V. parahaemolyticus. The diameter of growth from the center of a 150-mm agar plate was measured in centimeters to quantify swarming in varying conditions. The greatest extent of swarming was observed when V. parahaemolyticus strains (n=15) were incubated in BHI medium containing 1% agar at 25°C. In these conditions, the strains swarm the surface of an entire 150-mm agar plate within 3 days of incubation. When two T6SS-positive strains of the same species swarm towards each other, instead of forming a continuous mass, a Dienes line forms at the junction indicating competition between the two strains. Thus, the next step was to detect Dienes line formation. Pairwise testing was performed from the 15 strains, with each pair inoculated 0.5 cm apart on a 150-mm agar plate. Pairwise testing of identical strains served as the negative control while strains of Proteus mirabilis served as the positive control. Eight strains (53%) of V. parahaemolyticus were positive for Dienes line formation under the test conditions. PCR was conducted to confirm the presence/absence of T6SS genes in the sample genome. RT-qPCR method was optimized.
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