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Khokhar, Fahad; Pereira Dias, Joana; Mylona, Elli; Keane, Jacqueline; Chattaway, Marie; MacQueen, Hilary; Rigas, Sushila; Baker, Stephen and Holmes, Mark
(2023).
Abstract
Background:
Exposure to subinhibitory antibiotic concentrations can reflect the conditions that bacteria may encounter both in clinical and environmental sample types and could select for antimicrobial resistance. This exposure can induce stress and alter gene expression, promoting both bacterial survival and the emergence of resistance via mutagenic DNA repair. The purpose of this study was to determine the extent to which exposure to subinhibitory antibiotic concentrations alters gene expression in a drug-resistant isolate.
Methods:
We used an extensively-drug resistant Salmonella Typhi isolate to investigate the effects of treatment with subinhibitory concentrations of ampicillin (AMP), ceftriaxone (CEF), chloramphenicol (CHL), ciprofloxacin (CIP), co-trimoxazole (SXT) and tebipenem-pivoxil (TBP), as well as mitomycin C (MTC), on gene expression. Total RNA was extracted followed by ribosomal RNA depletion and library preparation for sequencing on a NovaSeq platform. Sequenced reads were processed using the nf-core/rnaseq pipeline followed by analysis using DESeq2 and topGO. A q-value of <0.05 was considered significant, and a log2 fold change of >2 or <-2 in gene expression levels against the no-treatment control were investigated.
Results:
Treatment with CIP, CHL, MTC and TBP significantly altered gene expression. No genes were significantly down-regulated from treatment with CIP, with the top Gene Ontology (GO) terms for the 25 up-regulated genes associated with SOS response, DNA repair and response to radiation, consistent with previously reported studies. Interestingly, CIP and MTC caused increased expression of both lexA, a main repressor for SOS genes, as well as recA, which is the main inducer of the SOS response pathway. CHL induced a total of 97 up-regulated and 54 down-regulated genes, with the majority involved in transmembrane transport, translation, and response to chemical. LysA (amino acid metabolism) and cpxP (stress response) were the only two genes up-regulated by TBP. No significant expression changes were found for treatment with AMP, CEF or SXT.
Conclusion:
These initial results replicate findings that subinhibitory concentrations of CIP as well as treatment with MTC can induce significant transcriptional up-regulation of genes in the SOS response pathway. Further investigation into the transcriptional response, specifically genes associated with the SOS pathway, is needed.