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Host response and bacterial virulence factor expression in Pseudomonas aeruginosa and Streptococcus pneumoniae corneal ulcers.

ABSTRACT: P. aeruginosa and S. pneumoniae are major bacterial causes of corneal ulcers in industrialized and in developing countries. The current study examined host innate immune responses at the site of infection, and also expression of bacterial virulence factors in clinical isolates from patients in south India. Corneal ulcer material was obtained from 49 patients with confirmed P. aeruginosa and 27 patients with S. pneumoniae, and gene expression of Toll Like Receptors (TLR), cytokines and inflammasome proteins was measured by quantitative PCR. Expression of P. aeruginosa type III secretion exotoxins and S. pneumoniae pneumolysin was detected by western blot analysis. We found that neutrophils comprised >90% cells in corneal ulcers, and that there was elevated expression of TLR2, TLR4, TLR5 and TLR9, the NLRP3 and NLRC4 inflammasomes and the ASC adaptor molecule. IL-1? IL-1? and IFN-? expression was also elevated; however, there was no significant difference in expression of any of these genes between corneal ulcers from P. aeruginosa and S. pneumoniae infected patients. We also show that 41/49 (84%) of P. aeruginosa clinical isolates expressed ExoS and ExoT, whereas 5/49 (10%) of isolates expressed ExoS, ExoT and ExoU with only 2/49 isolates expressing ExoT and ExoU. In contrast, all 27 S. pneumoniae clinical isolates produced pneumolysin. Taken together, these findings demonstrate that ExoS/T expressing P. aeruginosa and pneumolysin expressing S. pneumoniae predominate in bacterial keratitis. While P. aeruginosa strains expressing both ExoU and ExoS are usually rare, these strains actually outnumbered strains expressing only ExoU in the current study. Further, as neutrophils are the predominant cell type in these corneal ulcers, they are the likely source of cytokines and of the increased TLR and inflammasome expression.

SUBMITTER: Karthikeyan RS

PROVIDER: S-EPMC3672173 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Host response and bacterial virulence factor expression in Pseudomonas aeruginosa and Streptococcus pneumoniae corneal ulcers.

Karthikeyan Rajapandian SivaGanesa RS Priya Jeganathan Lakshmi JL Leal Sixto M SM Toska Jonida J Rietsch Arne A Prajna Venkatesh V Pearlman Eric E Lalitha Prajna P

PloS one 20130604 6

P. aeruginosa and S. pneumoniae are major bacterial causes of corneal ulcers in industrialized and in developing countries. The current study examined host innate immune responses at the site of infection, and also expression of bacterial virulence factors in clinical isolates from patients in south India. Corneal ulcer material was obtained from 49 patients with confirmed P. aeruginosa and 27 patients with S. pneumoniae, and gene expression of Toll Like Receptors (TLR), cytokines and inflammaso ...[more]

PMID: 23750216

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Project description:The Steroids for Corneal Ulcers Trial (SCUT) was a multicenter, international study of bacterial keratitis in which 101 Pseudomonas aeruginosa infections were treated. Twenty-two of 101 P. aeruginosa isolates collected had a colony morphology characteristic of a loss-of-function mutation in lasR, the gene encoding a quorum-sensing master regulator. Ulcers caused by these 22 strains were associated with larger areas of corneal opacification, worse vision, and a lower rate of vision recovery in response to treatment than ulcers caused by the other isolates. The lasR sequences from these isolates each contained one of three nonsynonymous substitutions, and these strains were deficient in production of LasR-regulated protease and rhamnolipids. Replacement of lasR with either of the two most common lasR alleles from the SCUT isolates was sufficient to decrease protease and rhamnolipid production in PA14. Loss of LasR function is associated with increased production of CupA fimbriae, and the LasR-defective isolates exhibited higher production of CupA fimbriae than LasR-intact isolates. Strains with the same lasR mutation were of the same multilocus sequence type, suggesting that LasR-deficient, environmental P. aeruginosa strains were endemic to the area, and infections caused by these strains were associated with worse patient outcomes in the SCUT study. (This study has been registered at ClinicalTrials.gov under registration no. NCT00324168.) IMPORTANCE The LasR transcription factor is an important regulator of quorum sensing in P. aeruginosa and positively controls multiple virulence-associated pathways. The emergence of strains with lasR loss-of-function alleles in chronic disease is well described and is thought to represent a specific adaptation to the host environment. However, the prevalence and virulence of these strains in acute infections remain unclear. This report describes observations revealing that lasR mutants were common among isolates from a large, multicenter clinical study of keratitis and were associated with worse clinical outcomes than LasR-intact strains despite reduced production of LasR-regulated factors. Additionally, these lasR mutants were closely related strains or clones, as determined by molecular analysis. Because bacterial keratitis is community acquired, these data indicate infection by endemic, LasR-deficient strains in the environment. These results suggest that the conventional paradigm regarding the role for LasR-mediated regulation of virulence is more complex than previously appreciated.

Dataset Information

Host response and bacterial virulence factor expression in Pseudomonas aeruginosa and Streptococcus pneumoniae corneal ulcers.

Publications

Host response and bacterial virulence factor expression in Pseudomonas aeruginosa and Streptococcus pneumoniae corneal ulcers.

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