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The role of type 4 pilus, flagella and type III secretion system in the interactions between Pseudomonas aeruginosa bacteria and corneal fibroblasts in human microbial keratitis

The role of type 4 pilus, flagella and type III secretion system in the interactions between Pseudomonas aeruginosa bacteria and corneal fibroblasts in human microbial keratitis
The role of type 4 pilus, flagella and type III secretion system in the interactions between Pseudomonas aeruginosa bacteria and corneal fibroblasts in human microbial keratitis
Purpose : To examine the specific individual roles of bacterial virulence factors in the interactions between Pseudomonas aeruginosa (PA) bacteria and corneal fibroblasts in human microbial keratitis.

Methods : Human corneal fibroblast (CF) were extracted from clinical samples from several patients and cultured to confluence in vitro. CFs were challenged with live wild type PAO1 bacteria and mutant strains deficient in type 4 pilus (TFP), flagella or both, and total bacterial association was quantified at 3h using the saponin-lysis and viable counting method and confirmed using laser scanning confocal microscopy (LSCM); and live wild type PA14 bacteria and mutant strains deficient in type III secretion system (TTSS) or flagella, and lactose dehydrogenase (LDH) release from CF was quantified and used as a measure of cytotoxicity.

Results : Bacterial adherence to CF was significantly less (P=0.000, ANOVA) in mutant PAO1 strains deficient in TFP and flagella compared to wild type strain as demonstrated by viable counting assays and LSCM. Bacteria-induced cytotoxicity was significantly less (P=0.000, ANOVA) in mutant strains deficient in TTSS or flagella compared to wild type strain as demonstrated by reduced LDH release.

Conclusions : We have established in vitro cell culture models of bacterial keratitis and demonstrated that PA utilises TFP and flagella to adhere to, and TTSS to induce cytotoxicity in, human CF.
0146-0404
Elsahn, Ahmad
db5d3c4e-61a6-4069-8a20-64b077127ca6
Christodoulides, Myron
eba99148-620c-452a-a334-c1a52ba94078
Hossain, Parwez
563de5fc-84ad-4539-9228-bde0237eaf51
Elsahn, Ahmad
db5d3c4e-61a6-4069-8a20-64b077127ca6
Christodoulides, Myron
eba99148-620c-452a-a334-c1a52ba94078
Hossain, Parwez
563de5fc-84ad-4539-9228-bde0237eaf51

Elsahn, Ahmad, Christodoulides, Myron and Hossain, Parwez (2017) The role of type 4 pilus, flagella and type III secretion system in the interactions between Pseudomonas aeruginosa bacteria and corneal fibroblasts in human microbial keratitis. Investigative Ophthalmology & Visual Science, 58 (8).

Record type: Meeting abstract

Abstract

Purpose : To examine the specific individual roles of bacterial virulence factors in the interactions between Pseudomonas aeruginosa (PA) bacteria and corneal fibroblasts in human microbial keratitis.

Methods : Human corneal fibroblast (CF) were extracted from clinical samples from several patients and cultured to confluence in vitro. CFs were challenged with live wild type PAO1 bacteria and mutant strains deficient in type 4 pilus (TFP), flagella or both, and total bacterial association was quantified at 3h using the saponin-lysis and viable counting method and confirmed using laser scanning confocal microscopy (LSCM); and live wild type PA14 bacteria and mutant strains deficient in type III secretion system (TTSS) or flagella, and lactose dehydrogenase (LDH) release from CF was quantified and used as a measure of cytotoxicity.

Results : Bacterial adherence to CF was significantly less (P=0.000, ANOVA) in mutant PAO1 strains deficient in TFP and flagella compared to wild type strain as demonstrated by viable counting assays and LSCM. Bacteria-induced cytotoxicity was significantly less (P=0.000, ANOVA) in mutant strains deficient in TTSS or flagella compared to wild type strain as demonstrated by reduced LDH release.

Conclusions : We have established in vitro cell culture models of bacterial keratitis and demonstrated that PA utilises TFP and flagella to adhere to, and TTSS to induce cytotoxicity in, human CF.

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More information

Published date: June 2017

Identifiers

Local EPrints ID: 448087
URI: http://eprints.soton.ac.uk/id/eprint/448087
ISSN: 0146-0404
PURE UUID: 3b5da129-f75b-4cb5-aa96-e9de686a1310
ORCID for Myron Christodoulides: ORCID iD orcid.org/0000-0002-9663-4731
ORCID for Parwez Hossain: ORCID iD orcid.org/0000-0002-3131-2395

Catalogue record

Date deposited: 01 Apr 2021 15:58
Last modified: 13 Apr 2021 01:41

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