An Expanded Transposon Mutant Library Reveals that Vibrio fischeri δ-Aminolevulinate Auxotrophs Can Colonize Euprymna scolopes.

摘要: Libraries of defined mutants are valuable research tools but necessarily lack gene knockouts that lethal under the conditions used in library construction. In this study, we augmented a Vibrio fischeri mutant generated on rich medium (LBS, which contains [per liter] 10 g tryptone, 5 yeast extract, 20 NaCl, and 50 mM Tris [pH 7.5]) by selecting transposon insertion supplemented LBS screening for those unable to grow LBS. We isolated strains with insertions alr, glr (murI), glmS, several heme biosynthesis genes, ftsA, as well disrupted 14 bp upstream ftsQ Mutants ftsA or were recovered addition Mg2+ LBS, their cell morphology motility affected. The was more strongly affected formed cells chains appeared wind back themselves helically. Growth N-acetylglucosamine (NAG), d-alanine, d-glutamate, respectively. hypothesized NAG, d-glutamate might be available V. Euprymna scolopes light organ; however, none these colonized host effectively. contrast, hemA hemL mutants, auxotrophic δ-aminolevulinate (ALA), at wild-type levels, although later biosynthetic pathway severely impaired colonize. Our findings parallel observations legume hosts provide Bradyrhizobium symbionts ALA, they contrast virulence phenotypes some pathogens. results further inform our understanding symbiotic organ environment.IMPORTANCE By supplementing yeast-based medium, able recover conditionally essential characterization provided new insights into bacterium's environment. Most notably, show evidence squid can enough ALA support its growth organ, paralleling finding legumes nodules. Taken together, how simple method augmenting already media expand reach utility libraries.