Characterization of two virulence-associated S. aureus sRNAs

Project Summary Staphylococcus aureus (S. aureus) infections, including those caused by antibiotic-resistant strains (MRSA), are increasing globally, posing significant public health challenges. While hospital-acquired infections have stabilized, community-acquired infections have shown a steep increase in recent years. Understanding the pathophysiology of these infections is crucial for developing effective treatment strategies. Non-coding RNAs (ncRNAs) in S. aureus play pivotal roles in regulating virulence but remain relatively understudied. These ncRNAs, including small- RNAs (sRNAs), are integral components of complex regulatory networks that respond to environmental cues. However, only a small fraction of S. aureus ncRNAs have been characterized, and even fewer have been studied in vivo. The Staphylococcal Regulatory RNAs Database (SRD) catalogs predicted and confirmed ncRNAs across S. aureus lineages, underscoring their importance in S. aureus physiology and pathogenesis. Researchers utilized a pangenome of S. aureus updated with ncRNA locations to analyze S. aureus transcriptomes from cystic fibrosis sputum samples (CF) and human chronic wounds (HCW), identifying two highly expressed uncharacterized sRNAs, rsaX20 and S1077. By analyzing publicly available RNA sequencing data and performing Northern blots, researchers observed that both sRNAs were induced during stress and low metal environments. Moreover, using our established murine chronic wound model, researchers observed that rsaX20 and S1077 reached similar levels of expression as in human samples. The overarching hypothesis is that S1077 and rsaX20 encode sRNAs that post- transcriptionally modulate functions critical for S. aureus chronic wound infection Researchers will test this hypothesis in two Aims: Aim 1 will elucidate the mechanisms controlling expression of these sRNAs, assess heterogeneity in expression at the single-cell level, and identify their targets; Aim 2 will assess the importance of S1077 and rsaX20 for S. aureus virulence using a murine surgical wound infection model. The results from these experiments will provide new insights into the genetic targets and roles in virulence of two sRNAs of unknown function that are highly expressedduring human chronic infection.