Leucine-responsive regulatory protein (Lrp) is an essential transcriptional regulator in prokaryotes. However, the regulatory role of lrp in Vibrio alginolyticus has still not been studied in detail. In this study, an lrp mutant strain was constructed to gain insight into the role of lrp in Vibrio alginolyticus. The absence of lrp significantly enhances swarming motility, biofilm formation, extracellular protease secretion activity, and tolerance to copper ions. The cumulative mortality of zebrafish (Danio rerio) challenged by intraperitoneal injection against the lrp mutant strain reached 68.89%, significantly higher than the 40.00% suffered by fish injected with the wild-type strain. The expression levels of lrp decreased gradually with increasing culture time under the influence of various physicochemical factors. The expression level of lrp was significantly increased after two hours of culture at pH 5, 22°C, 5% NaCl, the presence of 1 mM Cu2+, 1/4/7 mM ferric citrate, 0.1 mg/L NaNO3, and 0.1 mg/L KH2PO4. The mRNA level of lrp decreased significantly after six hours of culture at 37°C, 1% and 5% NaCl, and the presence of 1/7 mM ferric citrate, 0.1/5 mg/L NaNO3, and 0.1/0.5/2.0 mg/L KH2PO4. The expression of lrp increased after ten hours of culture at pH 5/9, 22°C, 1% NaCl, and the presence of 1 mM Cu2+, 7 mM ferric citrate, 5 mg/L NaNO3, and 0.1 mg/L KH2PO4. Overall, this study indicates that lrp negatively controls the virulence of V. alginolyticus, probably by reducing its swarming motility, biofilm formation, extracellular protease secretion activity, and tolerance to copper ions, and that the expression of lrp is affected by numerous physicochemical factors, and is especially up-regulated after 2 hours of culture in bacterial growth.

Conjugation is the most common horizontal gene transfer (HGT) process that can be affected by environmental change and promote bacterial virulence and drug resistance. However, it is unknown whether environmental changes can influence the conjugation ability of the marine fish pathogen Vibrio harveyi, thereby affecting its pathogenicity and drug resistance. This study systematically analyzes the effect of environmental stress on the ability of V. harveyi to obtain shuttle plasmids from Escherichia coli during conjugation. The results indicate that V. harveyi cannot receive shuttle plasmid pMMB207 without exposure to stress. However, certain stress exposure (37–46°C, 4%–16% ethanol, 0.14–0.56 mM SDS, 0.04~0.05 M NaOH, and 0.012–0.024 M HCl for 5–60 minutes) in the log phase of V. harveyi before conjugation successfully induces the fertility of the V. harveyi recipient in intergeneric mating with E. coli. In particular, ethanol and heat stress showed strong induction with up to 2.5 × 105 and 5.3 × 103 transconjugants when exposed to 16% ethanol for 10 minutes and 40°C for 60 minutes, respectively. Additionally, appropriate levels of NaOH (0.05 M, 10 minutes), SDS (0.42 mM, 5 minutes), and HCl (0.024 M, 5 minutes) lead to 2.3 × 103, 4.5 × 102, and 1.8 × 102 transconjugants, respectively. These results will help establish homologous recombination gene knockout technology and greatly advance molecular theoretical research on V. harveyi. They will also support the establishment of disease prevention and control strategies based on the interruption of the HGT process by environmental regulation.