title: The importance of the Pseudomonas aeruginosa type III secretion system in epithelium traversal depends upon conditions of host susceptibility.
abstract: Pseudomonas aeruginosa is invasive or cytotoxic to host cells, depending on the type III secretion system (T3SS) effectors encoded. While the T3SS is known to be involved in disease in vivo, how it participates remains to be clarified. Here, mouse models of superficial epithelial injury (tissue paper blotting with EGTA treatment) and immunocompromise (MyD88 deficiency) were used to study the contribution of the T3SS transcriptional activator ExsA to epithelial traversal. Corneas of excised eyeballs were inoculated with green fluorescent protein (GFP)-expressing PAO1 or isogenic exsA mutants for 6 h ex vivo before bacterial traversal and epithelial thickness were quantified by using imaging. In the blotting-EGTA model, exsA mutants were defective in capacity for traversal. Accordingly, an ∼16-fold variability in exsA expression among PAO1 isolates from three sources correlated with epithelial loss. In contrast, MyD88-/- epithelia remained susceptible to P. aeruginosa traversal despite exsA mutation. Epithelial lysates from MyD88-/- mice had reduced antimicrobial activity compared to those from wild-type mice with and without prior antigen challenge, particularly 30- to 100-kDa fractions, for which mass spectrometry revealed multiple differences, including (i) lower baseline levels of histones, tubulin, and lumican and (ii) reduced glutathione S-transferase, annexin, and dermatopontin, after antigen challenge. Thus, the importance of ExsA in epithelial traversal by invasive P. aeruginosa depends on the compromise enabling susceptibility, suggesting that strategies for preventing infection will need to extend beyond targeting the T3SS. The data also highlight the importance of mimicking conditions allowing susceptibility in animal models and the need to monitor variability among bacterial isolates from different sources, even for the same strain.
authors: School of Optometry, University of California, Berkeley, California, USA.
authors: Aaron B
authors: AB
authors: Sullivan
authors: School of Optometry, University of California, Berkeley, California, USA.
authors: K P Connie
authors: KP
authors: Tam
authors: School of Optometry, University of California, Berkeley, California, USA.
authors: Matteo M E
authors: MM
authors: Metruccio
authors: School of Optometry, University of California, Berkeley, California, USA College of Pharmacy, Touro University California, Vallejo, California, USA.
authors: David J
authors: DJ
authors: Evans
authors: School of Optometry, University of California, Berkeley, California, USA College of Pharmacy, Touro University California, Vallejo, California, USA Graduate Groups in Vision Science, Microbiology, and Infectious Diseases and Immunity, University of California, Berkeley, California, USA fleiszig@berkeley.edu.
authors: Suzanne M J
authors: SM
authors: Fleiszig
mesh_terms: D000818:Animals; D017302:Annexins / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics*; D058947:Bacterial Secretion Systems / Q000502:physiology*; D011508:Chondroitin Sulfate Proteoglycans / Q000378:metabolism; D065306:Corneal Injuries / Q000382:microbiology*; D019573:Epithelium, Corneal / Q000378:metabolism / Q000382:microbiology*; D016326:Extracellular Matrix Proteins / Q000378:metabolism; D015818:Eye Infections, Bacterial / Q000382:microbiology; D005982:Glutathione Transferase / Q000378:metabolism; D049452:Green Fluorescent Proteins; D006657:Histones / Q000378:metabolism; D054884:Host-Pathogen Interactions / Q000276:immunology*; D007632:Keratan Sulfate / Q000378:metabolism; D000071234:Lumican; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D053594:Myeloid Differentiation Factor 88 / Q000235:genetics*; D011552:Pseudomonas Infections / Q000382:microbiology; D011550:Pseudomonas aeruginosa / Q000472:pathogenicity*; D011993:Recombinant Fusion Proteins / Q000235:genetics; D015534:Trans-Activators / Q000235:genetics*; D014404:Tubulin / Q000378:metabolism
chemical_list: D017302:Annexins; D001426:Bacterial Proteins; D058947:Bacterial Secretion Systems; D011508:Chondroitin Sulfate Proteoglycans; C504453:Dpt protein, mouse; C095093:ExsA protein, Pseudomonas aeruginosa; D016326:Extracellular Matrix Proteins; D006657:Histones; C000604307:Lum protein, mouse; D000071234:Lumican; C507780:Myd88 protein, mouse; D053594:Myeloid Differentiation Factor 88; D011993:Recombinant Fusion Proteins; D015534:Trans-Activators; D014404:Tubulin; D049452:Green Fluorescent Proteins; D007632:Keratan Sulfate; D005982:Glutathione Transferase