A novel hemA mutation is responsible for a small-colony-variant phenotype in Escherichia coli

Abstract

We identified a small colony variant (SCV) of an amoxicillin/clavulanic acid-resistant derivative of a clinical isolate of Escherichia coli from Malawi, which was selected for in vitro in a subinhibitory concentration of gentamicin. The SCV was auxotrophic for hemin and had impaired biofilm formation compared to the ancestral isolates. A single novel nucleotide polymorphism (SNP) in hemA, which encodes a glutamyl-tRNA reductase that catalyses the initial step of porphyrin biosynthesis leading to the production of haem, was responsible for the SCV phenotype. We showed the SNP in hemA resulted in a significant fitness cost to the isolate, which persisted even in the presence of hemin. However, the phenotype quickly reverted during sequential sub-culturing in liquid growth media. As hemA is not found in mammalian cells, and disruption of the gene results in a significant fitness cost, it represents a potential target for novel drug development specifically for the treatment of catheter-associated urinary tract infections caused by E. coli .

Involvement of Small Colony Variant-Related Heme Biosynthesis Genes in Staphylococcus aureus Persister Formation in vitro

“…In addition to the 260 clones that were found to have decreased survival after 6 days of levofloxacin exposure in the previous study ( Wang et al, 2015 ), we identified seven clones with deficiency in persister formation that had a transposon insertion between nucleotides 854 and 855 of hemA ( Figure 1A ; Supplementary Figure S1 ). And mutant in hemA resulting in SCV has been reported before ( Curtis et al, 2016 ; Hubbard et al, 2021 ).…”

“…To examine whether the heme biosynthesis pathway is associated with persister formation, we knocked out hemA and the downstream hemB gene by allelic gene replacement from S. aureus USA500 wildtype ( Figure 1B ). When either of these two genes was knocked out, the strains displayed a typical SCV phenotype ( Figures 1C , D ; Jonsson et al, 2003 ; Garcia et al, 2012 ; Proctor et al, 2014 ; Hubbard et al, 2021 ). After complementing the knockout genes separately, the phenotype was both reverted and the reversion was complete ( Figures 1E , F ).…”

Microbial Musings – November 2020

“…Here the group isolate an SCV from a clinical isolate of E. coli from Malawi by selecting for isolates that are more tolerant to the aminoglycoside gentamicin [20]. By genome sequencing the resistant isolate, they identify a single-nucleotide polymorphism (SNP) in the hemA gene, which results in the Phe to Leu alternation in the resulting protein that likely reduces function.…”

Microbial Musings – October 2021

“…They were able to recapitulate the phenotypes of Gotz’s group of reduced colony size (the small-colony variant, SCV) and also a growth defect at atmospheric CO 2 levels that could be complemented by 5 % CO 2 . A known phenotype of the SCVs is reduced membrane potential, which they also observed, seeing similar levels to a hemB mutant, which reminded me of a paper we published last year by Alastair Hubbard ( @dralhubb ) and colleagues on a novel hemA mutant that also induces SCV formation by a similar mechanism [ 7 ]. So why is the presence of MpsB making the cells more sensitive to serum?…”