Global discovery of colonization determinants in the squid symbiontVibrio fischeri

Abstract

Animal epithelial tissue becomes reproducibly colonized by specific environmental bacteria. The bacteria (microbiota) perform critical functions for the host's tissue development, immune system development, and nutrition; yet the processes by which bacterial diversity in the environment is selected to assemble the correct communities in the host are unclear. To understand the molecular determinants of microbiota selection, we examined colonization of a simplified model in which the light organ of Euprymna scolopes squid is colonized exclusively by Vibrio fischeri bacteria. We applied highthroughput insertion sequencing to identify which bacterial genes are required during host colonization. A library of over 41,000 unique transposon insertions was analyzed before and after colonization of 1,500 squid hatchlings. Mutants that were reproducibly depleted following squid colonization represented 380 genes, including 37 that encode known colonization factors. Validation of select mutants in defined competitions against the wild-type strain identified nine mutants that exhibited a reproducible colonization defect. Some of the colonization factors identified included genes predicted to influence copper regulation and secretion. Other mutants exhibited defects in biofilm development, which is required for aggregation in host mucus and initiation of colonization. Biofilm formation in culture and in vivo was abolished in a strain lacking the cytoplasmic chaperone DnaJ, suggesting an important role for protein quality control during the elaboration of bacterial biofilm in the context of an intact host immune system. Overall these data suggest that cellular stress responses and biofilm regulation are critical processes underlying the reproducible colonization of animal hosts by specific microbial symbionts.H umans and other animals are often sterile before birth, from which point they immediately proceed to acquire environmental bacteria (1). The bacteria that reproducibly colonize animal hosts are critical for host tissue development, immune system development, and nutrient acquisition. The selection process by which the functional symbionts take residence in the animal from among the great diversity of environmental microbes is poorly understood, so model systems have been especially valuable to examine how specific patterns of colonization are shaped by the genetic makeup of the bacteria and the host environment (2).The light organ of the Hawaiian bobtail squid, Euprymna scolopes, is colonized exclusively by the Gram-negative luminous bacterium Vibrio fischeri. The host inhabits seawater containing 10 6 bacteria per milliliter, with V. fischeri comprising at most 0.02% of the environmental population (3). E. scolopes hatch without symbionts, but then rapidly acquire environmental bacteria and proceed to select for V. fischeri in a "winnowing" process that ensures colonization by only the specific symbiont (4). The squid-Vibrio system thus presents an opportunity to investigate the processes that underlie acquisition of s...

Niche-Specific Impact of a Symbiotic Function on the Persistence of Microbial Symbionts within a Natural Host

“…For each trial, the mean CFU competitive index value (test CFU/control CFU) was not significantly different from the hypothetical mean of 1.0 (Table 1), which is similar to previous reports of competitions between isogenic test and control strains (18,31). By examining the corresponding sets of light organ images, we counted the number of crypts colonized with the control strain, the test strain, or both strains.…”

Genome-wide screen identifies host colonization determinants in a bacterial gut symbiont

“…Some critical mechanisms for S. alvi colonization of the bee gut, including LPS modifications, membrane integrity components, the stringent response, proteases, and DNA break repair, are also important for infection by human pathogens (41)(42)(43). Similarly, colonization of light-producing organs by the squid symbiont Vibrio fisheri depends on genes dictating biofilm formation and cellular stress responses (44). In aquatic Vibrio colonizing zebra fish intestines, genes underlying cell envelope, chemosensory and motility functions seem crucial (45).…”

TfoX-Based Genetic Mapping Identifies Vibrio fischeri Strain-Level Differences and Reveals a Common Lineage of Laboratory Strains

“…For that purpose, we constructed a mariner transposition vector, termed pMarVF1, that transposes efficiently to generate single stable insertions in V. fischeri ES114 and other V. fischeri strains. We generated a library of over 40,000 unique erythromycin-resistant insertions (Tnerm) in strain ES114 (28), and this served as the donor library in the mapping.…”