An indirect enzyme-linked immunosorbent assay for rapid and quantitative assessment of Type III virulence phenotypes of Pseudomonas aeruginosa isolates

doi:10.1186/1476-0711-4-22

Annals of Clinical Microbiology and Antimicrobials

Volume 4

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Ledizet M
Kar K
Koski RA
Kazmierczak BI

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Research

An indirect enzyme-linked immunosorbent assay for rapid and quantitative assessment of Type III virulence phenotypes of Pseudomonas aeruginosa isolates

Li Li¹^,2 , Michel Ledizet³ , Kalipada Kar³ , Raymond A Koski³ and Barbara I Kazmierczak¹^,2

¹Department of Internal Medicine (Infectious Diseases), Yale University School of Medicine, New Haven CT, USA

²Section of Microbial Pathogenesis, Yale University School of Medicine, New Haven CT, USA

³L²Diagnostics, LLC, New Haven CT, USA

author email corresponding author email

Annals of Clinical Microbiology and Antimicrobials 2005, 4:22doi:10.1186/1476-0711-4-22


Published:	23 December 2005

Abstract

Background

The presence of a Type III secretion system in clinical isolates of Pseudomonas aeruginosa is associated with severe disease and poor outcomes in infections caused by this pathogen. We describe an indirect enzyme-linked immunosorbent assay that rapidly and quantitatively detects two exotoxins, ExoU and ExoT, and two structural components, PopD and PcrV, of the P. aeruginosa Type III secretion system after in-vitro growth in a calcium-free minimal medium.

Methods

We used this assay to characterize the Type III secretion phenotype of 74 clinical isolates of P. aeruginosa. Findings were compared with results of standard immunoblotting and correlated with Type III secretion-dependent virulence of isolates toward cultured epithelial cells.

Results

Results of the ELISA assay were concordant with immunoblot detection of the secreted antigens for 73 of 74 isolates. The Type III secretion phenotype assessed by this immunoassay predicted bacterial virulence toward epithelial cells in vitro for all but five of the clinical isolates.

Conclusion

The availability of an ELISA assay for rapid detection of Type III secreted virulence factors will facilitate large clinical studies to examine whether the Type III secretion phenotype of a P. aeruginosa isolate predicts the course of clinical disease in a patient and should be taken into account in determining optimal treatment strategies for infected patients.