High-throughput transposon mutagenesis in the family Enterobacteriaceae reveals core essential genes and rapid turnover of essentiality

Abstract

The Enterobacteriaceae are a scientifically and medically important clade of bacteria, containing the gut commensal and model organism Escherichia coli, as well as several major human pathogens including multiple serovars of Salmonella enterica and Klebsiella pneumoniae. Essential gene sets have been determined for several members of the Enterobacteriaceae, with the E. coli Keio single-gene deletion library often regarded as a gold standard for gene essentiality studies. However, it remains unclear how gene essentiality varies between related strains and species. To investigate this, we have assembled a collection of thirteen sequenced high-density transposon mutant libraries from five genera within the Enterobacteriaceae. We first benchmark a number of gene essentiality prediction approaches, investigate the effects of transposon density on essentiality prediction, and identify biases in transposon insertion sequencing data. Based on these investigations we develop a new classifier for gene essentiality. Using this new classifier, we define a core essential genome in the Enterobacteriaceae of 201 universally essential genes, and reconstruct an ancestral essential gene set of 296 genes. Despite the presence of a large cohort of variably essential genes, surprisingly we find an absence of evidence for genus-specific essential genes. A clear example of this sporadic essentiality is given by the set of genes regulating the σE extracytoplasmic stress response, which appears to have independently become essential multiple times in the Enterobacteriaceae. Finally, we compare our essential gene sets to the natural experiment of gene loss in obligate insect endosymbionts that have emerged from within the Enterobacteriaceae. This isolates a remarkably small set of genes absolutely required for survival, and uncovers several instances of essential stress responses masked by redundancy in free-living bacteria.

Genome-wide fitness analysis identifies genes required for in vitro growth and macrophage infection by African and global epidemic pathovariants of Salmonella enterica Enteritidis

“…We found that a total of 207 genes were required across S. enterica Enteritidis, S. enterica Typhimurium, S. enterica Typhi and E. coli (Fig. 2b); a recent comparative TraDIS study also identified a similar number of 201 universally essential genes in Enterobacteriaceae [83]. The 207 required genes mainly encode the basic cellular machinery (e.g.…”

“…enterica Typhi and E. coli (); a recent comparative TraDIS study also identified a similar number of 201 universally essential genes in Enterobacteriaceae [83]. The 207 required genes mainly encode the basic cellular machinery (e.g.…”

The PitA protein contributes to colistin susceptibility in Pseudomonas aeruginosa

“…Only an insertion near the middle of pitA resulted in an increase in MIC 50 , from 0.5 mg/L to 4 mg/L ( Fig 2A ). A second insertion near the start of the pitA gene did not impact susceptibility, likely because expression of the remaining portion of pitA was sufficient to retain protein function [ 36 ]. Transposon mutations in phoQ , pmrB , eno , carB , speE2 and PA5005 did not increase colistin MIC 50 , and mutants were not available for pmrA and speD2 .…”