title: 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.
authors.affiliation: Microbiology Branch, Dental and Craniofacial Trauma Research and Tissue Regeneration Directorate, Institute of Surgical Research, JBSA Fort Sam Houston, TX, USA.
authors.forename: Christine L
authors.identifier: 0000-0002-8166-9843
authors.initials: CL
authors.lastname: Miller
authors.affiliation: Microbiology Branch, Dental and Craniofacial Trauma Research and Tissue Regeneration Directorate, Institute of Surgical Research, JBSA Fort Sam Houston, TX, USA.
authors.forename: Tricia A
authors.initials: TA
authors.lastname: Van Laar
authors.affiliation: The Forsyth Institute, Cambridge, MA, USA.
authors.forename: Tsute
authors.initials: T
authors.lastname: Chen
authors.affiliation: Microbiology Branch, Dental and Craniofacial Trauma Research and Tissue Regeneration Directorate, Institute of Surgical Research, JBSA Fort Sam Houston, TX, USA.
authors.forename: S L Rajasekhar
authors.initials: SLR
authors.lastname: Karna
authors.affiliation: Microbiology Branch, Dental and Craniofacial Trauma Research and Tissue Regeneration Directorate, Institute of Surgical Research, JBSA Fort Sam Houston, TX, USA.
authors.forename: Ping
authors.initials: P
authors.lastname: Chen
authors.affiliation: Microbiology Branch, Dental and Craniofacial Trauma Research and Tissue Regeneration Directorate, Institute of Surgical Research, JBSA Fort Sam Houston, TX, USA.
authors.forename: Tao
authors.initials: T
authors.lastname: You
authors.affiliation: Microbiology Branch, Dental and Craniofacial Trauma Research and Tissue Regeneration Directorate, Institute of Surgical Research, JBSA Fort Sam Houston, TX, USA.
authors.forename: Kai P
authors.initials: KP
authors.lastname: Leung
mesh_terms: D018441:Biofilms / Q000254:growth & development*; D020869:Gene Expression Profiling; D055624:Methicillin-Resistant Staphylococcus aureus / Q000235:genetics / Q000502:physiology*; D056265:Microbial Interactions; D011550:Pseudomonas aeruginosa / Q000235:genetics / Q000502:physiology*; D012329:RNA, Bacterial / Q000096:biosynthesis* / Q000235:genetics; D017423:Sequence Analysis, RNA
chemical_list: D012329:RNA, Bacterial
keywords: 
  Pseudomonas aeruginosa
  ; 
  Staphylococcus aureus
  ; 
  sRNA
  ; transcriptome