Lactiplantibacillus plantarum are probiotic bacteria, maintaining the integrity of the gastrointestinal epithelial barrier, and preventing the infection of pathogenic bacteria. Exopolysaccharides (EPSs) are often involved in the probiotic property of L. plantarum. Here, we identified a new EPS-synthetic gene cluster, cpsWc, carrying 13 genes, laid on a large plasmid in a well-characterized probiotic L. plantarum strain LTC-113. The cpsWc gene cluster was horizontally acquired from Weissella cibaria, enhancing the biofilm formation ability of the host strain and its tolerance to harsh environmental stresses, including heat, acid, and bile. Transfer of cpsWc also conferred the probiotic properties to other L. plantarum strains. Moreover, cpsWc strengthened the adhesion of LTC-113 to intestinal epithelial cells. Both the cpsWc-carrying LTC-113 and its EPSs per se effectively attenuated the LPS-induced pro-inflammatory effect of intestinal epithelial cells, and inhibited the adhesion of pathogenic bacteria, such as S. typhimurium and E. coli by exclusion and competition. The newly identified cpsWc gene cluster emphasized the contribution of mobile EPS-synthetic element on the probiotic activity of L. plantarum, and shed a light on the engineering of probiotic bacteria.

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Background Antibiotic tolerance allows bacteria to overcome antibiotic treatment transiently and potentially accelerates the emergence of resistance. However, our understanding of antibiotic tolerance at the genetic level during adaptive evolution of Staphylococcus aureus remains incomplete. We sought to identify the mutated genes and verify the role of these genes in the formation of vancomycin tolerance in S. aureus. Methods Vancomycin-susceptible S. aureus strain Newman was used to induce vancomycin-tolerant isolates in vitro by cyclic exposure under a high concentration of vancomycin (20× MIC). WGS and Sanger sequencing were performed to identify the genetic mutations. The function of mutated genes in vancomycin-tolerant isolates were verified by gene complementation. Other phenotypes of vancomycin-tolerant isolates were also determined, including mutation frequency, autolysis, lysostaphin susceptibility, cell wall thickness and cross-tolerance. Results A series of vancomycin-tolerant S. aureus (VTSA) strains were isolated and 18 mutated genes were identified by WGS. Among these genes, pbp4, htrA, stp1, pth and NWMN_1068 were confirmed to play roles in VTSA formation. Mutation of mutL promoted the emergence of VTSA. All VTSA showed no changes in growth phenotype. Instead, they exhibited reduced autolysis, decreased lysostaphin susceptibility and thickened cell walls. In addition, all VTSA strains were cross-tolerant to antibiotics targeting cell wall synthesis but not to quinolones and lipopeptides. Conclusions Our results demonstrate that genetic mutations are responsible for emergence of phenotypic tolerance and formation of vancomycin tolerance may lie in cell wall changes in S. aureus.

Phenotypic heterogeneity within a bacterial population may confer new functionality and allow microorganisms to adapt to fluctuating environments. Previous work has suggested that Staphylococcus aureus could form small colony variants to avoid elimination by therapeutic antibiotics and host immunity systems. Here we show that a reversible non-pigment large colony morphology (Mu50∆lcpA-LC) was observed in S. aureus Mu50 after knocking out lcpA, coding for the LytR-CpsA-Psr family A protein. Mu50∆lcpA-LC increased resistance to β-lactam antibiotics, in addition, the enlarged cell size, enhanced spreading ability on solid medium, and reduced biofilm formation, suggesting better abilities for bacterial expansion. Moreover, the expression of spa encoding protein A was significantly increased in Mu50∆lcpA-LC. This study shows that besides the small colony variants, S. aureus could fight against antibiotics and host immunity through phenotype switching into a large colony variant.