Original Articles

Determination of Immunological Properties of Pseudomonas aeroginosa PA103, by Serum Bactericidal Assay & ELISA

Abstract

Background:   Exotoxin A (ExoA) is one of the most known and important virulence factors of Pseudomonas aeruginosa. This toxin is the cause of ADP-ribosylation in eukaryutic Elongation Factor-2 (EF-2), which results in the protein synthesis inhibition. Recent studies had shown immunostimulational characteristics of detoxified the ExoA. Our aim in this study was to evaluate the immunological properties of detoxified ExoA by Serum Bactericidal assay, in comparison with ELISA.

Methods:   The production of ExoA was done on the ExoA-producing strain, Pseudomonas aeruginosa PA103, provided by Pasteur institute of Iran. After culture in semi-industrial scale, it was detoxified and purified by dialysis. The dialysate was injected to mice and rabbit. After 3weeks, the total sera were collected. Serum bactericidal assay & ELISA were performed.

Results:   The results had shown a significant increase of antibodies against detoxified ExoA of 1/16 and antibody in ELISA method. Also, this has shown more antigenecity & immunogenosity by SBA method.

Conclusion:   To conclude the study that has been done here, SBA has much higher advantages than ELISA, in determining the immunological properties of Exo A. Furthermore, since Exo A could act as a hapten, many other subunits from other bacteria or viruses could be conjugated on Exo A and form multi-target vaccines.

1. Tang Y, Ali Z, Zou J, et al. Detection methods for Pseudomonas aeruginosa: history and future perspective. Rsc Advances 2017; 7(82):51789-800.
2. Baltch A, Franke M, Smith R, et al. Serum antibody concentrations of cytotoxin, exotoxin A, lipopolysaccharide, protease, and elastase and survival of patients with Pseudomonas aeruginosa bacteremia. Clin Infect Dis 1996; 23(5):1109-16.
3. Humada TA, Abdullah HI. Quantitative determination of endogenic human Pseudomonas aeruginosa exotoxin A (PEA) concentrations in serum of renal failure patients by ELISA. Tikrit Med J 2015; 20(1).
4. Schultz WW, Phipps TJ, Pollack M. Enzyme-linked immunosorbent assay for Pseudomonas aeruginosa exotoxin A. J Clin Microb 1979; 9(6):705-8.
5. Khan A, Tauseef I, Aalia B, et al. Age-related variations in the in vitro bactericidal activity of human sera against Pseudomonas aeruginosa. Central Eur J Immunol 2018; 43(1):18-25.
6. Morlon-Guyot J, Méré J, Bonhoure A, et al. Processing of Pseudomonas aeruginosa exotoxin A is dispensable for cell intoxication. Infect Immun 2009; 77(7):3090-9.
7. Farrant KV, Spiga L, Davies JC, et al. Response of Pseudomonas aeruginosa to the innate immune system-derived oxidants hypochlorous acid and hypothiocyanous acid. J Bacteriol 2020; 203(2):10.1128/jb. 00300-20.
8. Alhazmi A. NOD-like receptor (s) and host immune responses with Pseudomonas aeruginosa infection. Inflam Res 2018; 67:479-93.
9. Cigana C, Lorè NI, Riva C, et al. Tracking the immunopathological response to Pseudomonas aeruginosa during respiratory infections. Sci Rep 2016; 6(1):21465.
10. Priebe GP, Goldberg JB. Vaccines for Pseudomonas aeruginosa: a long and winding road. Exp Rev Vaccines 2014; 13(4):507-19.
11. Ryskjær J, Deacon CF, Carr RD, et al. Plasma dipeptidyl peptidase-IV activity in patients with type-2 diabetes mellitus correlates positively with HbAlc levels, but is not acutely affected by food intake. Eur J Endocrinol 2006; 155(3):485-93.
12. Tramper-Stranders GA, van der Ent CK, Slieker MG, et al. Diagnostic value of serological tests against Pseudomonas aeruginosa in a large cystic fibrosis population. Thorax 2006; 61(8):689-93.
13. Gotschlich E. Development of polysaccharide vaccines for the prevention of meningococcal diseases. Monogr Allergy 1975; 9:245-58.
14. Galloway D, Hedstrom R, Pavlovskis O. Production and characterization of monoclonal antibodies to exotoxin A from Pseudomonas aeruginosa. Infect Immun 1984; 44(2):262-7.
15. Mauch RM, Jensen PØ, Moser C, Levy CE, Høiby N. Mechanisms of humoral immune response against Pseudomonas aeruginosa biofilm infection in cystic fibrosis. J Cystic Fibrosis 2018; 17(2):143-52.
16. Mahmood AS. DNA extraction protocol for rapid pcr detection of pathogenic Pseudomonas aeruginosa and detection CD4 and CD8 in Iraqi burn case. Biochem Cell Archives 2019; 19(1).
17. Grumelli S. Host-pathogen interaction in the lung of patients infected with Pseudomonas aeruginosa. Pseudomonas aeruginosa-an armory Within: London, IntechOpen; 2019.
18. Nabonee MA, Rahman A, Mishu NJ, et al. Immunization with formaldehyde inactivated whole cell vaccine against multidrug resistant Pseudomonas aeruginosa produces protective IgG antibodies in mice model. Arch Microbiol Immunol 2023; 7:230-5.
19. Lovewell RR, Patankar YR, Berwin B. Mechanisms of phagocytosis and host clearance of Pseudomonas aeruginosa. Am J Physiology 2014; 306(7):L591-L603.
20. Szu SC, Taylor DN, Trofa AC, et al. Laboratory and preliminary clinical characterization of Vi capsular polysaccharide-protein conjugate vaccines. Infect Immun 1994; 62(10):4440-4.
21. Holst J, Feiring B, Fuglesang J, et al. Serum bactericidal activity correlates with the vaccine efficacy of outer membrane vesicle vaccines against Neisseria meningitidis serogroup B disease. Vaccine 2003; 21(7-8):734-7.
22. Stratton CW. Serum bactericidal test. Clin Microbiol Rev 1988; 1(1):19-26.
Files
IssueVol 13 No 1 (2025) QRcode
SectionOriginal Articles
DOI https://doi.org/10.18502/jmb.v13i1.18046
Keywords
Assay Bactericidal Exotoxin A Pseudomonas aeruginosa Serum.
Rights and permissions
Creative Commons License This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.
How to Cite
1.
Aliniay Sharafshadehi S, shadvar N, Seify SN, Mohammadi MR, Taher Sarjam M, Najafi S, Fathi J, S.Sadati M, Hadi N. Determination of Immunological Properties of Pseudomonas aeroginosa PA103, by Serum Bactericidal Assay & ELISA. J Med Bacteriol. 2025;13(1):40-45.