Identification of Novel Factors Involved in Modulating Motility of Salmonella enterica Serotype Typhimurium

Abstract

Salmonella enterica serotype Typhimurium can move through liquid using swimming motility, and across a surface by swarming motility. We generated a library of targeted deletion mutants in Salmonella Typhimurium strain ATCC14028, primarily in genes specific to Salmonella, that we have previously described. In the work presented here, we screened each individual mutant from this library for the ability to move away from the site of inoculation on swimming and swarming motility agar. Mutants in genes previously described as important for motility, such as flgF, motA, cheY are do not move away from the site of inoculation on plates in our screens, validating our approach. Mutants in 130 genes, not previously known to be involved in motility, had altered movement of at least one type, 9 mutants were severely impaired for both types of motility, while 33 mutants appeared defective on swimming motility plates but not swarming motility plates, and 49 mutants had reduced ability to move on swarming agar but not swimming agar. Finally, 39 mutants were determined to be hypermotile in at least one of the types of motility tested. Both mutants that appeared non-motile and hypermotile on plates were assayed for expression levels of FliC and FljB on the bacterial surface and many of them had altered levels of these proteins. The phenotypes we report are the first phenotypes ever assigned to 74 of these open reading frames, as they are annotated as ‘hypothetical genes’ in the Typhimurium genome.

Beyond Antimicrobial Resistance: Evidence for a Distinct Role of the AcrD Efflux Pump in Salmonella Biology

“…Recent studies have focused on finding novel genes and/or pathways which mediate swarming (60-62). A screen of a mutant library containing 1023 mutants in S. typhimurium 14028s found 21 mutants with impaired swimming and swarming, forty nine with impaired swimming, but normal swarming, and also 49 with impaired swarming but normal swimming, and 39 hyper-motile mutants (61).…”

“…A screen of a mutant library containing 1023 mutants in S. typhimurium 14028s found 21 mutants with impaired swimming and swarming, forty nine with impaired swimming, but normal swarming, and also 49 with impaired swarming but normal swimming, and 39 hyper-motile mutants (61). Further exploration of these novel motility genes identified by Bogomolnaya et al 2014 is needed.…”

Solid tumors provide niche-specific conditions that lead to preferential growth of Salmonella

“…To establish the mechanism by which Salmonella preferentially colonizes tumor tissue, we used a negative selection strategy employing a single-gene deletion mutant library, SGD-K [46–48, 69]. This strategy allowed us to identify bacterial processes that are needed for fitness in tumors of mice.…”

Strain Library Imaging Protocol for high-throughput, automated single-cell microscopy of large bacterial collections arrayed on multiwell plates

“…Advances in biotechnology have enabled the creation of large genetic libraries of bacteria, such as single nonessential gene-deletion libraries in E. coli 6 and Salmonella enterica 7 , deep mutational scanning of genes of interest 8 in a variety of organisms, and a CRISPR interference (CRISPRi)-based essential gene knockdown library in Bacillus subtilis 9 . Population-level screening of these libraries has generated useful insights into gene functions related to environmental changes 7,9,10 .…”

“…Population-level screening of these libraries has generated useful insights into gene functions related to environmental changes 7,9,10 . These libraries contain hundreds to thousands of strains, and thus screens are typically performed in 96-, 384-, or 1536-well format in multi-well liquid media or on rectangular agar plates.…”