Heterogenous Susceptibility to R-Pyocins in Populations of Pseudomonas aeruginosa Sourced from Cystic Fibrosis Lungs

Abstract

Bacteriocins are proteinaceous antimicrobials produced by bacteria that are active against other strains of the same species. R-type pyocins are phage tail-like bacteriocins produced by Pseudomonas aeruginosa. Due to their antipseudomonal activity, R-pyocins have potential as therapeutics in infection. P. aeruginosa is a Gram-negative opportunistic pathogen and is particularly problematic for individuals with cystic fibrosis (CF). P. aeruginosa organisms from CF lung infections develop increasing resistance to antibiotics, making new treatment approaches essential. P. aeruginosa populations become phenotypically and genotypically diverse during infection; however, little is known of the efficacy of R-pyocins against heterogeneous populations. R-pyocins vary by subtype (R1 to R5), distinguished by binding to different residues on the lipopolysaccharide (LPS). Each type varies in killing spectrum, and each strain produces only one R-type. To evaluate the prevalence of different R-types, we screened P. aeruginosa strains from the International Pseudomonas Consortium Database (IPCD) and from our biobank of CF strains. We found that (i) R1-types were the most prevalent R-type among strains from respiratory sources, (ii) a large number of strains lack R-pyocin genes, and (iii) isolates collected from the same patient have the same R-type. We then assessed the impact of intrastrain diversity on R-pyocin susceptibility and found a heterogenous response to R-pyocins within populations, likely due to differences in the LPS core. Our work reveals that heterogeneous populations of microbes exhibit variable susceptibility to R-pyocins and highlights that there is likely heterogeneity in response to other types of LPS-binding antimicrobials, including phage. IMPORTANCE R-pyocins have potential as alternative therapeutics against Pseudomonas aeruginosa in chronic infection; however, little is known about the efficacy of R-pyocins in heterogeneous bacterial populations. P. aeruginosa is known to become resistant to multiple antibiotics and to evolve phenotypic and genotypic diversity over time; thus, it is particularly difficult to eradicate in chronic cystic fibrosis (CF) lung infections. In this study, we found that P. aeruginosa populations from CF lungs maintain the same R-pyocin genotype but exhibit heterogeneity in susceptibility to R-pyocins from other strains. Our findings suggest there is heterogeneity in response to other types of LPS-binding antimicrobials, such as phage, highlighting the necessity of further studying the potential of LPS-binding antimicrobial particles as alternative therapies in chronic infections.

Genetic and Transcriptomic Characteristics of RhlR-Dependent Quorum Sensing in Cystic Fibrosis Isolates of Pseudomonas aeruginosa

“…We also found common mutations in genes coding for S-pyocin production, a bacteriocin mechanism that allows P. aeruginosa to toxify neighboring nonkin strains using proteinaceous antimicrobial peptides internalized via siderophore receptors such as FptA ( 43 , 44 ). Divergence of genes coding for S-pyocins may reflect the pressures of interbacterial competition within infections, as selection may also target other similar competitive mechanisms, such as the phage tail-like R-pyocins or T4SS, in evolving infection isolates ( 45 ). The regional isolation of specific lineages of P. aeruginosa found in a study of explanted CF lungs provides additional support for this view of isolate pathoadaptation; as bacteria become isolated to a specific long-term niche in the lung, selection may purify mechanisms associated with competitive exclusion ( 46 ).…”

O-Specific Antigen-Dependent Surface Hydrophobicity Mediates Aggregate Assembly Type in Pseudomonas aeruginosa

“…Despite a large body of work showing how P. aeruginosa adapts to the CF lung environment ( 19 – 21 , 33 , 47 , 48 ) and population heterogeneity during chronic infection ( 20 , 21 , 23 , 49 , 50 ), little is known about the impact of this intraspecific heterogeneity on the formation of P. aeruginosa aggregates. To test whether genetic heterogeneity impacts aggregation, we investigated the aggregate formation of selected evolved isolates from a previous 50-day biofilm evolution experiment of PAO1 ( 23 ) in SCFM2 ( 24 ).…”

SOS-Independent Pyocin Production in P. aeruginosa Is Induced by XerC Recombinase Deficiency

“…This finding raises the possibility that chemical inhibitors of XerC might sensitize P. aeruginosa to fluoroquinolone antibiotics via increased production of pyocins. Notably, recent work indicates that many clinical isolates are sensitive to pyocins and/or produce pyocins themselves ( 3 , 4 ). Hence, we envision an exciting prospect for combination treatment with a pyocin-stimulating XerC inhibitor plus a fluoroquinolone.…”

“…For instance, P. aeruginosa secretes soluble antibiotics like pyocyanin, enacts contact-mediated toxin injection using type VI secretion, and can produce phage tail-like bacteriocins (PTLBs) ( 2 ). Bacteriocins are typically named according to the producing species; in P. aeruginosa , they are termed pyocins and are thought to primarily enable intraspecies competition ( 3 – 5 ). There is also limited evidence that they may have activity against other species ( 6 , 7 ).…”