Chitin induces natural competence in Vibrio cholerae

The mosaic-structured Vibrio cholerae genome points to the importance of horizontal gene transfer (HGT) in the evolution of this human pathogen. We showed that V. cholerae can acquire new genetic material by natural transformation during growth on chitin, a biopolymer that is abundant in aquatic habitats (e.g., from crustacean exoskeletons), where it lives as an autochthonous microbe. Transformation competence was found to require a type IV pilus assembly complex, a putative DNA binding protein, and three convergent regulatory cascades, which are activated by chitin, increasing cell density, and nutrient limitation, a decline in growth rate, or stress.

Chitin colonization, chitin degradation and chitin-induced natural competence of Vibrio cholerae are subject to catabolite repression

Melanie Blokesch

Although Vibrio cholerae is a human pathogen its primary habitat are aquatic environments. In this environment, V. cholerae takes advantage of the abundance of zooplankton, whose chitinous exoskeletons provide a nutritious surface. Chitin also induces the developmental programme of natural competence in several species of the genus Vibrio. Because the chitin surface can serve as the sole carbon source for V. cholerae, the link between carbon catabolite repression and chitin-induced natural competence for transformation was investigated in this study. Provision of competing phosphoenolpyruvate: carbohydrate phosphotransferase system (PTS)-dependent carbon sources in addition to chitin significantly lowered natural transformability. These sugars are known to interfere with the accumulation of 3,5- cyclic AMP (cAMP); therefore, the contributions of the cAMP-producing enzyme, adenylate cyclase and the cAMP receptor protein (CRP) to chitin surface colonization, chitin degradation and natural transformation were also analysed. The results provided here indicate that cAMP and CRP are important in at least three interlinked areas of the chitin-induced natural competence programme. First, cAMP and CRP are required for the efficient colonization of the chitin surface; second both contribute to chitin degradation and utilization, and third, cAMP plus CRP play a role in increasing competence gene expression. These findings highlight the complex regulatory circuit of chitininduced natural competence in V. cholerae.

Wiley-Blackwell2012

Genetic manipulation of Vibrio cholerae by combining natural transformation with FLP recombination

Melanie Blokesch, Olga de Souza Silva

Even though Vibrio cholerae is a well-known human pathogen, it is also a normal member of aquatic habitats. Within this environment it often forms biofilms on the chitin-containing exoskeleton of crustaceans and their molts. Chitin not only serves as nutrient source but also induces a developmental program called natural competence. Naturally competent bacteria take up free DNA and integrate it into their genome by homologous recombination, thereby becoming naturally transformed. In this study, we made use of the knowledge on the environmental lifestyle of V. cholerae to genetically manipulate its genome. We achieved this by combining the methods of chitin-induced natural transformation and Flp recombination. Using this approach, we disrupted several genes by insertion of FRT-site-flanked antibiotic-resistance cassettes. The cassettes were subsequently excised by induction of the Flp recombinase, which acts on the FRT sites. This method represents a simplified and faster alternative to standard gene deletion techniques, which often depend on bacterial conjugation and the availability of suicide vectors.

Elsevier2010

Two nanomachines drive evolution in diverse Vibrio species

Noémie Matthey

Horizontal gene transfer (HGT) has a major impact on bacterial evolution, leading to acquisition or deletion of genes and gene clusters, including those encoding antibiotic resistances and virulence factors. HGT therefore contributes to pathogen emergence, which can have a major impact on human health. As a mode of HGT, natural competence for transformation allows bacteria to directly acquire DNA from the environment and to integrate parts of this genetic material within their genomes. The human pathogen Vibrio cholerae is naturally competent when it grows in association with the chitinous exoskeletons of zooplankton. Under this condition, the competence state is initiated by the regulatory protein TfoX, which co-induces a molecular killing device known as the type VI secretion system (T6SS). The co-regulation of the T6SS and competence results in the lysis of non-immune neighboring bacteria and the subsequent acquisition of their DNA, which fosters HGT. In V. cholerae, the T6SS is also activated by the TfoX homologue protein, named TfoY. In this study, we aimed at studying the regulatory pathways and the co-regulation of interbacterial predation and DNA uptake in order to provide new insights into the environmental lifestyle of V. cholerae and other vibrios. We first investigated the conserved function of TfoX- and TfoY in V. fischeri, V. alginolyticus, and V. parahaemolyticus using diverse methods such as qRT-PCR, interbacterial killing assays, and motility assessments. We demonstrated that TfoX-mediated T6SS and competence induction and TfoY-induced motility are conserved phenotypes in the tested Vibrio species, whereas the TfoY-mediated T6SS regulation varied. These variations might reflect diverse defense mechanisms, as these different species have adapted to cope with their environmental niches. Next, we tested the outcome of the TfoX-mediated co-regulation of the T6SS and competence in V. cholerae. Under conditions mimicking the bacteriumâs natural habitat, we showed for the first time the extent of DNA that was integrated on V. choleraeâs genome after T6SS-mediated interbacterial predation. Indeed, we demonstrated that T6SS-dependent bacterial predation not only increased the transformation rates but also fostered the exchange of multiple and huge fragments of DNA. We also showed the necessity of an exquisite co-regulation between the T6SS, the competence-related DNA uptake machinery, and the competence-repressed nuclease. Finally, we disclosed that prey-released DNA is not a private good of the attacking bacterium but also accessible to other surrounding cells that have likewise entered the competence state. Altogether, our findings shed light on the conservation of the TfoX- and TfoY-driven phenotypes in several Vibrio species. We also contributed to a better understanding of the interplay between neighbor predation and DNA uptake and the consequences that this interplay has on genome plasticity.

EPFL2019

Two nanomachines drive evolution in diverse Vibrio species

Noémie Matthey

EPFL2019