Characterisation of Resistance Against The Peptide Antibiotic Bacitracin in Streptomyces venezuelae

Abstract

Antimicrobial peptides (AMPs) are a promising source of antibiotics, potentially effective against multi-drug resistant bacteria. Whilst AMP resistance among low GC-content Gram-positive bacteria has been studied in detail, the situation in high-GC Gram-positives is poorly characterised. Here, we identify that Streptomyces venezuelae is highly resistant to the AMP bacitracin. Transcriptome analysis of S. venezuelae found many putative peptidase-encoding genes strongly up-regulated under bacitracin challenge, leading us to propose that AMP degradation may contribute to the observed resistance phenotype. Using HPLC quantification, we could show that cell suspensions of S. venezuelae rapidly degrade bacitracin. Subsequently, we were able to demonstrate that degradative activity resided in the culture supernatants indicating that bacitracin degradation was an extracellular phenotype of S. venezuelae. Indeed, S. venezuelae was able to protect sensitive Streptomyces species from bacitracin inhibition on solid medium, implicating bacitracin-degrading peptidases in microbial community interactions. Comparative genomics analyses of S. venezuelae peptidases up-regulated following bacitracin exposure was used to identify candidate peptidases for involvement in bacitracin degradation by attempting to identify genes correlated with the degree of bacitracin resistance in the host. Deletion of three neighbouring peptidase encoding genes resulted in only a minor 1.1-fold reduction in the bacitracin minimum inhibitory concentration (MIC) of S. venezuelae. It therefore seems likely that multiple peptidases contribute to bacitracin degradation by S. venezuelae. Two ATP-binding cassette (ABC) transporters were also up-regulated by S. venezuelae upon bacitracin challenge. These transporters contained FtsX-like transmembrane helices found in the BceAB-like transporters that confer bacitracin resistance in the low-GC Gram-positives. Deletion of genes encoding one of these transporters resulted in a 1.2-fold reduction in bacitracin MIC. We therefore propose that bacitracin resistance in S. venezuelae is multifaceted with both bacitracin degrading peptidases, and ABC transporters contributing to a high level of resistance. Extracellular degradation mediated resistance to bacitracin could be indicative of similar forms of resistance to other antibiotics that may be implicated in soil community interactions.

Date of Award	27 Mar 2024
Original language	English
Awarding Institution	University of Bath
Supervisor	Andrew Preston (Supervisor) & Brian Jones (Supervisor)