Vibrio parahaemolyticus T6SS2 effector repertoires.

All strains of the marine bacterium Vibrio parahaemolyticus harbor a type VI secretion system (T6SS) named T6SS2, suggesting that this system plays an important role in the life cycle of this emerging pathogen. Although T6SS2 was recently shown to play a role in interbacterial competition, its effector repertoire remains unknown. Here, we employed proteomics to investigate the T6SS2 secretome of two V. parahaemolyticus strains, and we identified several antibacterial effectors encoded outside of the main T6SS2 gene cluster. We revealed two T6SS2-secreted proteins that are conserved in this species, indicating that they belong to the core secretome of T6SS2; other identified effectors are found only in subsets of strains, suggesting that they comprise an accessory effector arsenal of T6SS2. Remarkably, a conserved Rhs repeat-containing effector serves as a quality control checkpoint and is required for T6SS2 activity. Our results reveal effector repertoires of a conserved T6SS, including effectors that have no known activity and that have not been previously associated with T6SSs.

A Rapid Fluorescence-Based Screen to Identify Regulators and Components of Interbacterial Competition Mechanisms in Bacteria.

Contact-dependent antibacterial mechanisms enhance bacterial fitness as they enable bacteria to outcompete their rivals and thrive in diverse environments. Such systems also allow pathogenic bacteria to establish a niche inside a host, where they must compete with commensal microflora. In many cases, antibacterial systems are tightly regulated by complex sensor and signal transduction networks. Deciphering these regulatory networks, as well as identifying functional components of antibacterial mechanisms, are valuable objectives since essential regulators and components present possible targets for developing antivirulence therapies. Here we describe Bacterial Competition Fluorescence (BaCoF), a methodology that relies on a fluorescence signal to determine the outcome of bacterial competitions. This methodology enables screening of mutant libraries to identify genes that are essential for activating a contact-dependent antibacterial system of interest. Thus, this methodology can be applied to reveal essential regulators and components of antibacterial systems in bacterial pathogens.