Loo, H. Q. | bioRxiv Science

bioRxiv2024

Intraspecific Diversity of Staphylococcus aureus Populations Isolated from Cystic Fibrosis Respiratory Infections

Chronic bacterial infections are often polymicrobial, comprising multiple bacterial species or variants of the same species. Because chronic infections may last for decades, they have the potential to generate high levels of intraspecific variation through within-host diversification over time, and the potential for superinfections to occur through the introduction of multiple pathogen populations to the ongoing infection. Traditional methods for identifying infective agents generally involve isolating one single colony from a given sample, usually after selecting for a specific pathogen or antibiotic resistance profile. Isolating a recognized virulent or difficult to treat pathogen is an important part of informing clinical treatment and correlative research; however, these reductive methods alone, do not provide researchers or healthcare providers with the potentially important perspective on the true pathogen population structure and dynamics over time. To begin to address this limitation, in this study, we compare findings on Staphylococcus aureus single colonies versus and pools of colonies taken from fresh sputum samples from three patients with cystic fibrosis to isolates collected from the same sputum samples and processed by the clinical microbiology laboratory. Phenotypic and genotypic analysis of isolated S. aureus populations revealed coexisting lineages in two of three sputum samples as well as population structures that were not reflected in the single colony isolates. Altogether, our observations presented here demonstrate that clinically relevant diversity can be missed with standard sampling methods when assessing chronic infections. More broadly, this work outlines the potential impact that comprehensive population-level sampling may have for both research efforts and more effective treatment practices. Data SummaryThe authors confirm all supporting data, code and protocols have been provided within the article.

microbiology↗

bioRxiv2021

Species-wide phylogenomics of the Staphylococcus aureus agr operon reveals convergent evolution of frameshift mutations

Staphylococcus aureus is a prominent nosocomial pathogen that causes several life-threatening diseases such as pneumonia and bacteremia. S. aureus modulates expression of its arsenal of virulence factors through sensing and integrating responses to environmental signals. The agr (accessory gene regulator) quorum sensing (QS) system is a major regulator of virulence phenotypes in S. aureus. There are four agr specificity groups each with a different autoinducer peptide sequence (encoded by the agrD gene). Though agr is critical for expression of many toxins, paradoxically, S. aureus strains often have non-functional agr activity due to loss-of-function mutations in the four-gene agr operon. To understand patterns in agr variability across S. aureus, we undertook a species-wide genomic investigation. We developed a software tool (AgrVATE; https://github.com/VishnuRaghuram94/AgrVATE) for typing and detecting frameshift mutations in the agr operon. In an analysis of over 40,000 S. aureus genomes, we showed close association between agr type and S. aureus clonal complex. We also found strong linkage between agrBDC alleles (encoding the peptidase, the autoinducing peptide itself, and the peptide sensor respectively) but not agrA (encoding the -response regulator). More than five percent of genomes were found to have frameshift mutations in the agr operon. Though most mutations occur only once in the entire species, we observed a small number of recurring mutations evolving convergently across different clonal lineages. Phylogenetic patterns suggested that strains with agr frameshifts were evolutionary dead ends. Overall, genomic analysis of agr operon suggests evolution through multiple processes with functional consequences that are not fully understood. ImportanceStaphylococcus aureus is a globally pervasive pathogen that produces a plethora of toxic molecules that can degrade, evade, or inhibit the host immune cells. Production of these toxins is mainly controlled by an active agr quorum sensing system, which senses and responds to bacterial cell density. However, there are many reports of S. aureus strains with genetic changes leading to impaired agr activity, often found during chronic bloodstream infections, and may be associated with increased disease severity. We developed an open-source software called AgrVATE to type agr systems and identify putative frameshifts. We used AgrVATE for a species-wide genomic survey of S. aureus, finding that more than 5 % of strains in the public database had non-functional agr systems. We also provide new insights into the evolution of these genetic mutations in the agr system. Overall, this study contributes to our understanding of a common but relatively understudied means of virulence regulation in S. aureus.

genomics↗