Biofilms Up Close: Many bacterial infections involve biofilm formation. Cells within a biofilm are significantly more resistant to immune clearance and antibiotics compared to unattached, planktonic cells. Berk et al. (p. 236 ) applied superresolution optical methods to image living bacteria with nanometer-scale precision as they form a biofilm. Vibrio cholerae biofilms were observed to have three distinct levels of spatial organization: cells, clusters of cells, and collections of clusters. Each cell cluster was wrapped in a flexible, elastic envelope. Several V. cholerae matrix proteins played complementary architectural roles during biofilm development. RbmA provided cell-cell adhesion, Bap1 allowed the developing biofilm to adhere to surfaces, and heterogeneous mixtures of VPS, RbmC, and Bap1 formed the dynamic, flexible, and ordered envelopes that encase the cell clusters.

In many bacteria, including Vibrio cholerae, cyclic dimeric guanosine monophosphate (c-di-GMP) controls the motile to biofilm life style switch. Yet, little is known about how this occurs. In this study, we report that changes in c-di-GMP concentration impact the biosynthesis of the MshA pili, resulting in altered motility and biofilm phenotypes in V. cholerae. Previously, we reported that cdgJ encodes a c-di-GMP phosphodiesterase and a ΔcdgJ mutant has reduced motility and enhanced biofilm formation. Here we show that loss of the genes required for the mannose-sensitive hemagglutinin (MshA) pilus biogenesis restores motility in the ΔcdgJ mutant. Mutations of the predicted ATPase proteins mshE or pilT, responsible for polymerizing and depolymerizing MshA pili, impair near surface motility behavior and initial surface attachment dynamics. A ΔcdgJ mutant has enhanced surface attachment, while the ΔcdgJmshA mutant phenocopies the high motility and low attachment phenotypes observed in a ΔmshA strain. Elevated concentrations of c-di-GMP enhance surface MshA pilus production. MshE, but not PilT binds c-di-GMP directly, establishing a mechanism for c-di-GMP signaling input in MshA pilus production. Collectively, our results suggest that the dynamic nature of the MshA pilus established by the assembly and disassembly of pilin subunits is essential for transition from the motile to sessile lifestyle and that c-di-GMP affects MshA pilus assembly and function through direct interactions with the MshE ATPase. The human pathogen Vibrio cholerae causes the debilitating disease cholera through ingestion of contaminated food and water. V. cholerae is a natural inhabitant of aquatic environments. Transmission of V. cholerae to the human host is dependent on survival of the pathogen in aquatic reservoirs where it is challenged with many stressors, including changes in the physiochemical parameters of environments and predation by protozoa and phages. One method utilized to endure these assaults is to form a multicellular community called a biofilm. The signaling molecule cyclic dimeric guanosine monophosphate (c-di-GMP) is utilized to induce biofilm formation in many bacteria, including V. cholerae. We demonstrate that c-di-GMP promotes the production of a cell surface structure called MshA pili by binding the molecular motor responsible for polymerizing pilus subunits. These pili are adhesive appendages that are essential for attachment to surfaces. This study identifies a novel mechanism for c-di-GMP regulation of pilus production through interactions with the molecular motor responsible for pilus assembly. Since many bacteria utilize pili for attachment to surfaces and c-di-GMP as a pro-biofilm signaling molecule, the mechanism for pilus regulation and biofilm formation described here may be widespread among many pathogens.

Infectious diseases kill nearly 9 million people annually. Bacterial pathogens are responsible for a large proportion of these diseases, and the bacterial agents of pneumonia, diarrhea, and tuberculosis are leading causes of death and disability worldwide. Increasingly, the crucial role of nonhost environments in the life cycle of bacterial pathogens is being recognized. Heightened scrutiny has been given to the biological processes impacting pathogen dissemination and survival in the natural environment, because these processes are essential for the transmission of pathogenic bacteria to new hosts. This chapter focuses on the model environmental pathogen Vibrio cholerae to describe recent advances in our understanding of how pathogens survive between hosts and to highlight the processes necessary to support the cycle of environmental survival, transmission, and dissemination. We describe the physiological and molecular responses of V. cholerae to changing environmental conditions, focusing on its survival in aquatic reservoirs between hosts and its entry into and exit from human hosts.