Human tear fluid modulates the Pseudomonas aeruginosa transcriptome to alter antibiotic susceptibility

Previously, we showed that tear fluid protects corneal epithelial cells against Pseudomonas aeruginosa without suppressing bacterial viability. Here, we studied how tear fluid affects bacterial gene expression. RNA-sequencing was used to study the P. aeruginosa transcriptome after tear fluid exposur...

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Bibliographic Details
Published in:	The ocular surface Vol. 22; pp. 94 - 102
Main Authors:	Tabor, Lauren M., Grosser, Melinda R., Metruccio, Matteo M.M.E., Kumar, Naren G., Wu, Yvonne T., Nieto, Vincent, Evans, David J., Fleiszig, Suzanne M.J.
Format:	Journal Article
Language:	English
Published:	United States Elsevier Inc 01-10-2021
Subjects:	Anti-Bacterial Agents - pharmacology Antimicrobial susceptibility arn operon Bacterial Proteins - genetics Bacterial Proteins - metabolism Gene Expression Regulation, Bacterial Humans oprH-phoP/phoQ Polymyxin B Pseudomonas aeruginosa Pseudomonas aeruginosa - genetics RNA-Sequencing Tear fluid Tear-like fluid Transcriptome Tear fluid Tear-like fluid Pseudomonas aeruginosa Antimicrobial susceptibility arn operon oprH-phoP/phoQ Polymyxin B RNA-Sequencing
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Summary:	Previously, we showed that tear fluid protects corneal epithelial cells against Pseudomonas aeruginosa without suppressing bacterial viability. Here, we studied how tear fluid affects bacterial gene expression. RNA-sequencing was used to study the P. aeruginosa transcriptome after tear fluid exposure (5 h, 37 oC). Outcomes were further investigated by biochemical and physiological perturbations to tear fluid and tear-like fluid (TLF) and assessment of bacterial viability following tear/TLF pretreatment and antibiotic exposure. Tear fluid deregulated ~180 P. aeruginosa genes ≥8 fold versus PBS including downregulating lasI, rhlI, qscR (quorum sensing/virulence), oprH, phoP, phoQ (antimicrobial resistance) and arnBCADTEF (polymyxin B resistance). Upregulated genes included algF (biofilm formation) and hemO (iron acquisition). qPCR confirmed tear down-regulation of oprH, phoP and phoQ. Tear fluid pre-treatment increased P. aeruginosa resistance to meropenem ~5-fold (4 μg/ml), but enhanced polymyxin B susceptibility ~180-fold (1 μg/ml), the latter activity reduced by dilution in PBS. Media containing a subset of tear components (TLF) also sensitized bacteria to polymyxin B, but only ~22.5-fold, correlating with TLF/tear fluid Ca2+ and Mg2+ concentrations. Accordingly, phoQ mutants were not sensitized by TLF or tear fluid. Superior activity of tear fluid versus TLF against wild-type P. aeruginosa was heat resistant but proteinase K sensitive. P. aeruginosa responds to human tear fluid by upregulating genes associated with bacterial survival and adaptation. Meanwhile, tear fluid down-regulates multiple virulence-associated genes. Tears also utilize divalent cations and heat resistant/proteinase K sensitive component(s) to enhance P. aeruginosa sensitivity to polymyxin B.
Bibliography:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 Contributed equally to first authorship Present addresses: University of North Carolina, Asheville, NC USA
ISSN:	1542-0124 1937-5913
DOI:	10.1016/j.jtos.2021.07.004