Deventer, A. T. | bioRxiv Science

bioRxiv2025

Clinical stringent response activation promotes conjugal transfer of staphylococcal resistance plasmids

Conjugative transfer of plasmids represents a major route through which antibiotic resistance genes are spread. In the case of the prevalent and deadly pathogen Staphylococcus aureus, >90% of clinical isolates carry at least one plasmid. While plasmid-encoded mechanisms (e.g. plasmid copy number) can influence conjugation frequency, host factors and environmental stimuli can also affect transmission. In particular, stress responses like the stringent response have been associated with increased movement of mobile genetic elements. We have previously shown that clinical mutations in the stringent response controller, Rel, lead to elevated levels of the alarmones (p)ppGpp and antibiotic tolerance in S. aureus. Here, we report that stringent response activation in these strains promotes the conjugal transfer of diverse staphylococcal resistance plasmids. We observed that clinical Rel mutations promote donation, but not receipt, of plasmids from the three families of staphylococcal plasmid and a mobilisable plasmid. This increased conjugation frequency could also be induced by chemical induction of the stringent response by mupirocin. Intriguingly, detailed experimental analysis revealed that the effect of elevated (p)ppGpp on plasmid donation was not due to CodY derepression, SOS response induction, increased plasmid copy number or increased expression of conjugation machinery genes. Further, transcriptomic analysis failed to identify any other putative plasmid- or host-derived mechanisms to explain this observation. Further investigations are required to explore the mechanistic link between the stringent response and conjugation, given the pervasive transcriptional and post-translational effects of (p)ppGpp. Overall, the association between Rel mutation and increased plasmid donation is alarming, especially as Rel mutations are being increasingly identified among clinical isolates.

microbiology↗

bioRxiv2024

Staphylococcus aureus COL: An Atypical Model Strain of MRSA that Exhibits Slow Growth and Antibiotic Tolerance

Methicillin-resistant Staphylococcus aureus (MRSA) has been a pathogen of global concern since its emergence in the 1960s. As one of the first MRSA strains isolated, COL has become a common model strain of S. aureus. Here we report that COL is, in fact, an atypical strain of MRSA that exhibits slow growth and multidrug tolerance. Genomic analysis identified three mutated genes in COL (rpoB, gltX and prs) with links to tolerance. Allele swapping experiments between COL and the closely related, non-tolerant Newman strain uncovered a complex interplay between these genes. However, Prs (phosphoribosyl pyrophosphate [PRPP] synthetase) accounted for most of the growth and tolerance phenotype of COL. Biochemical and transcriptomic analysis revealed that COL does not exhibit slow growth as a result of partial stringent response activation, as previously proposed. Instead, the COL Prs mutation greatly reduces the PRPP synthetase activity of the enzyme and leads to downregulation of pyrimidine, histidine and tryptophan synthesis, three pathways that rely on PRPP. Overall, our findings indicate that COL is an atypical, antibiotic-tolerant strain of MRSA whose isolation predates the previous first report of tolerance among clinical isolates. Characterisation of clinical Prs mutations and their relationship with tolerance requires further investigation.

microbiology↗