Global transcriptome responses including small RNAs during mixed‐species interactions with methicillin‐resistant Staphylococcus aureus and Pseudomonas aeruginosa

Abstract

Pseudomonas aeruginosa and Staphylococcus aureus mixed‐species biofilm infections are more resilient to biocide attacks compared to their single‐species counterparts. Therefore, this study used an in vitro model recapitulating bacterial burdens seen in in vivo infections to investigate the interactions of P. aeruginosa and S. aureus in biofilms. RNA sequencing (RNA‐seq) was utilized to identify the entire genomic response, both open reading frames (ORFs) and small RNAs (sRNAs), of each species. Using competitive indexes, transposon mutants validated uncharacterized PA1595 of P. aeruginosa and Panton–Valentine leukocidin ORFs of S. aureus are required for competitive success. Assessing spent media on biofilm development determined that the effects of these ORFs are not solely mediated by mechanisms of secretion. Unlike PA1595, leukocidin (lukS‐PV) mutants of S. aureus lack a competitive advantage through contact‐mediated mechanisms demonstrated by cross‐hatch assays. RNA‐seq results suggested that during planktonic mixed‐species growth there is a robust genomic response or active combat from both pathogens until a state of equilibrium is reached during the maturation of a biofilm. In mixed‐species biofilms, P. aeruginosa differentially expressed only 0.3% of its genome, with most ORFs necessary for growth and biofilm development, whereas S. aureus modulated approximately 5% of its genome, with ORFs suggestive of a phenotype of increased virulence and metabolic quiescence. Specific expression of characterized sRNAs aligned with the genomic response to presumably coordinate the adaptive changes necessary for this homeostatic mixed‐species biofilm and sRNAs may provide viable foci for the design of future therapeutics.

Trophic cooperation promotes bacterial survival of Staphylococcus aureus and Pseudomonas aeruginosa

“…While coexistence is characterized by an absence of S. aureus growth inhibition during 8-hour co-culture (18), it appears that nutritional competition can still occur under unfavourable conditions and affect S. aureus's survival. Therefore, coexistence between the two pathogens is promoted in nutritionally rich environments, in line with previous observations (35,36). Although its survival rate is affected under adverse nutritional conditions induced by long-term culture, acetoin catabolism benefits its producer, S. aureus (Fig.…”

“…S. aureus survival thus appears to be highly affected by long-term co-culture with P. aeruginosa, even if the strains coexist stably during shorter spans of culture (18). The state of interaction with S. aureus thus seems to rely on nutritional conditions, a fact already observed elsewhere (35,36).…”

Antimicrobial and Antibiofilm Activity of Synergistic Combinations of a Commercially Available Small Compound Library With Colistin Against Pseudomonas aeruginosa

“…We used a model of biofilm formation that has been previously described (Miller et al, 2016 ). First, we adjusted log-phase cultures of P. aeruginosa to an OD 600 of 0.05 in sterile PBS (seeding solution).…”

Increased rates of genomic mutation in a biofilm co-culture model of Pseudomonas aeruginosa and Staphylococcus aureus

“…This was interesting as 514 numerous studies have shown that the viability of S. aureus is negatively affected by 515 P. aeruginosa even in planktonic culture 10,27-29 . However, Miller et al (2017) found 516 that use of a more nutrient rich medium improved S. aureus survival 30 . This 517 suggests, consistent with our results that media composition is an important 518 consideration for the development of a co-culture model.…”