Design of ELISA-based diagnostic system for detection of enterohaemorrhagic Escherichia coli
-
Mohammad Javad Rezaei
,
Maryam Eidi
,
Seyed Ali Mirhosseini
,
Rouhollah Kazemi
,
Mohammad Javad Motamedi
,
Soghra Khani
,
Jafar Amani
Abstract
Background and Objectives: Escherichia coli (E. coli) O157:H7 is an intestinal pathogen of humans and animals, which causes serious gastrointestinal, urinary tract infection and hemolytic uremic syndrome. Connecting to the host cell is important in pathogenesis. EspA, Intimin and Tir proteins (EIT) are the most important bacterial features in the process of binding. These antigens can be very useful in detecting these bacteria. The aim of this study was to produce recombinant EspA, Intimin and Tir proteins (rEIT) to detect pathogenic E. coli O157:H7 by means of ELISA method.
Materials and Methods: The eit recombinant gene was expressed using IPTG in E. coli BL21 (DE3) and evaluated by western blotting. The purified rEIT protein was injected to rabbits and mice subcutaneously. Purified antibody was evaluated using indirect, competitive and sandwich ELISA confirming the precise detection of E. coli O157: H7.
Results: Indirect, competitive and sandwich ELISA specifically detected E. coli O157:H7 and each methods had the ability to identify more than 104, 104, 103 bacteria. The specificity of this method was evaluated by Entroheamoragic E. coli, enterotoxygenic E. coli, Klebsiella pneumoniae, Vibrio cholera and Acinetobacter.
Conclusion: These methods are the fastest, most accurate and cost effective methods for diagnosis of E. coli O157: H7, comparing to the conventional methods.
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19. Zhu J, Yang Q, Cao L, Dou X, Zhao J, Zhu W, et al. Development of porcine rotavirus vp6 protein based ELISA for differentiation of this virus and other viruses. Virol J 2013; 10: 91.
20. Jalallou N, Bandehpour M, Khazan H, Haghighi A, Kazemi B. Evaluation of recombinant SAG1 protein for detection of toxoplasma gondii specific immunoglobulin M by ELISA test. Iran J Parasitol 2012; 7: 17-21.
21. Byer JD, Struger J, Sverko E, Klawunn P, Todd A. Spatial and seasonal variations in atrazine and metolachlor surface water concentrations in Ontario (Canada) using ELISA. Chemosphere 2011; 82: 1155-1160.
22. Strachan NJ, Ogden ID. A sensitive microsphere coagulation ELISA for Escherichia coli O157: H7 using Russell's viper venom. FEMS Microbiol Lett 2000; 186: 79-84.
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24. Mirhosseini SA, Fooladi AAI, Amani J, Sedighian H. Production of recombinant flagellin to develop ELISA-based detection of Salmonella Enteritidis. Braz J Microbiol 2017; 48: 774-781.
25. Karimi Rahjerdi A, Jafari M, Motamedi MJ, Amani J, Salmanian AH. Immunogenic evaluation of bivalent vaccine Candidate against Enterohemorrhagic and Enterotoxigenic Escherichia coli. Iran J Immunol 2019; 16: 200-211.
26. Karami MS, Maraghi S, Rafiei A, Latifi SM, Kaydani GA. Comparison of sensitivity and specificity of native ELISA Test (Prepared in Khouzestan, Iran) and commercial ELISA Kit in the Diagnosis of human hydatidosis. Zahedan J Res Med Sci 2019; 21(4): e91416.
27. Parma YR, Chacana PA, Lucchesi PM, Roge A, Granobles Velandia CV, Krüger A, et al. Detection of Shiga toxin-producing Escherichia coli by sandwich enzyme-linked immunosorbent assay using chicken egg yolk IgY antibodies. Front Cell Infect Microbiol 2012; 2: 84.
28. Shen Z, Hou N, Jin M, Qiu Z, Wang J, Zhang B, et al. A novel enzyme-linked immunosorbent assay for detection of Escherichia coli O157: H7 using immunomagnetic and beacon gold nanoparticles. Gut Pathog 2014; 6: 14.
29. Pang B, Zhao C, Li L, Song X, Xu K, Wang J, et al. Development of a low-cost paper-based ELISA method for rapid Escherichia coli O157: H7 detection. Anal Biochem 2018; 542: 58-62.
30. Zhang X, Li M, Zhang B, Chen K, He K. Development of a Sandwich ELISA for EHEC O157: H7 Intimin γ1. PLoS One 2016; 11(9): e0162274.
2. Abadias M, Usall J, Anguera M, Solsona C, Viñas I. Microbiological quality of fresh, minimally-processed fruit and vegetables, and sprouts from retail establishments. Int J Food Microbiol 2008; 123: 121-129.
3. Wu C-J, Hsueh P-R, Ko W-C. A new health threat in Europe: Shiga toxin–producing Escherichia coli O104: H4 infections. J Microbiol Immunol Infect 2011; 44: 390-393.
4. Blanco JE, Blanco M, Alonso MP, Mora A, Dahbi G, Coira MA, et al. Serotypes, virulence genes, and intimin types of Shiga toxin (verotoxin)-producing Escherichia coli isolates from human patients: prevalence in Lugo, Spain, from 1992 through 1999. J Clin Microbiol 2004; 42: 311-319.
5. Yazdanparast A, Mousavi SL, Rasooli I, Amani J, Jalalinadoushan M. Immunogenical study of chimeric recombinant intimin-Tir of Escherichia coli O157: H7 in mice. Arch Clin Infect Dis 2012; 7: 45-51.
6. Chaisri U, Nagata M, Kurazono H, Horie H, Tongtawe P, Hayashi H, et al. Localization of Shiga toxins of enterohaemorrhagic Escherichia coli in kidneys of paediatric and geriatric patients with fatal haemolytic uraemic syndrome. Microb Pathog 2001; 31: 59-67.
7. te Loo DM, Monnens LA, van der Velden TJ, Vermeer MA, Preyers F, Demacker PN, et al. Binding and transfer of verocytotoxin by polymorphonuclear leukocytes in hemolytic uremic syndrome. Blood 2000; 95: 3396-3402.
8. Jawetz M. Adelberg, Arthropod borne and Rodent borne viral diseases In: Medical Microbiology 28e, Chapter 38. Singapore, The Mc Graw Hill Companies; 2004.
9. Li Y, Frey E, Mackenzie AM, Finlay BB. Human response to Escherichia coli O157: H7 infection: Antibodies to secreted virulence factors. Infect Immun 2000; 68: 5090-5095.
10. Gray MD, Lampel KA, Strockbine NA, Fernandez RE, Melton-Celsa AR, Maurelli AT. Clinical isolates of Shiga toxin 1a–producing Shigella flexneri with an epidemiological link to recent travel to Hispañiola. Emerg Infect Dis 2014; 20: 1669-1677.
11. Türütoğlu H, Öztürk D, Güler L, Pehlivanoğlu F. Presence and characteristics of sorbitol-negative Escherichia coli O157 in healthy sheep faeces. Vet Med 2007; 52: 301-307.
12. McKee R, Madden RH, Gilmour A. Occurrence of verocytotoxin-producing Escherichia coli in dairy and meat processing environments. J Food Prot 2003; 66: 1576-1580.
13. Clarke SC, Haigh RD, Freestone PP, Williams PH. Virulence of enteropathogenic Escherichia coli, a global pathogen. Clin Microbiol Rev 2003; 16: 365-378.
14. Clarke SC. Diarrhoeagenic Escherichia coli--an emerging problem? Diagn Microbiol Infect Dis 2001; 41: 93-98.
15. Wong LY, Cao Y, Balachandran P, Zoder P, Furtado MR, Petrauskene OV, et al. Validation of the applied biosystems MicroSEQ® real-time PCR system for detection of E. coli O157: H7 in food. J AOAC Int 2012; 95: 1495-1504.
16. Wasilenko JL, Fratamico PM, Narang N, Tillman GE, Ladely S, Simmons M, et al. Influence of primer sequences and DNA extraction method on detection of non-O157 Shiga toxin–producing Escherichia coli in ground beef by real-time PCR targeting the eae, stx, and serogroup-specific genes. J Food Prot 2012; 75: 1939-1950.
17. Hayrapetyan H, Tran T, Tellez-Corrales E, Madiraju C. Enzyme-linked immunosorbent assay: types and applications. Methods Mol Biol 2023; 2612: 1-17.
18. Chunglok W, Wuragil DK, Oaew S, Somasundrum M, Surareungchai W. Immunoassay based on carbon nanotubes-enhanced ELISA for Salmonella enterica serovar Typhimurium. Biosens Bioelectron 2011; 26: 3584-3589.
19. Zhu J, Yang Q, Cao L, Dou X, Zhao J, Zhu W, et al. Development of porcine rotavirus vp6 protein based ELISA for differentiation of this virus and other viruses. Virol J 2013; 10: 91.
20. Jalallou N, Bandehpour M, Khazan H, Haghighi A, Kazemi B. Evaluation of recombinant SAG1 protein for detection of toxoplasma gondii specific immunoglobulin M by ELISA test. Iran J Parasitol 2012; 7: 17-21.
21. Byer JD, Struger J, Sverko E, Klawunn P, Todd A. Spatial and seasonal variations in atrazine and metolachlor surface water concentrations in Ontario (Canada) using ELISA. Chemosphere 2011; 82: 1155-1160.
22. Strachan NJ, Ogden ID. A sensitive microsphere coagulation ELISA for Escherichia coli O157: H7 using Russell's viper venom. FEMS Microbiol Lett 2000; 186: 79-84.
23. Amani J, Salmanian AH, Rafati S, Mousavi SL. Immunogenic properties of chimeric protein from espA, eae and tir genes of Escherichia coli O157: H7. Vaccine 2010; 28: 6923-6929.
24. Mirhosseini SA, Fooladi AAI, Amani J, Sedighian H. Production of recombinant flagellin to develop ELISA-based detection of Salmonella Enteritidis. Braz J Microbiol 2017; 48: 774-781.
25. Karimi Rahjerdi A, Jafari M, Motamedi MJ, Amani J, Salmanian AH. Immunogenic evaluation of bivalent vaccine Candidate against Enterohemorrhagic and Enterotoxigenic Escherichia coli. Iran J Immunol 2019; 16: 200-211.
26. Karami MS, Maraghi S, Rafiei A, Latifi SM, Kaydani GA. Comparison of sensitivity and specificity of native ELISA Test (Prepared in Khouzestan, Iran) and commercial ELISA Kit in the Diagnosis of human hydatidosis. Zahedan J Res Med Sci 2019; 21(4): e91416.
27. Parma YR, Chacana PA, Lucchesi PM, Roge A, Granobles Velandia CV, Krüger A, et al. Detection of Shiga toxin-producing Escherichia coli by sandwich enzyme-linked immunosorbent assay using chicken egg yolk IgY antibodies. Front Cell Infect Microbiol 2012; 2: 84.
28. Shen Z, Hou N, Jin M, Qiu Z, Wang J, Zhang B, et al. A novel enzyme-linked immunosorbent assay for detection of Escherichia coli O157: H7 using immunomagnetic and beacon gold nanoparticles. Gut Pathog 2014; 6: 14.
29. Pang B, Zhao C, Li L, Song X, Xu K, Wang J, et al. Development of a low-cost paper-based ELISA method for rapid Escherichia coli O157: H7 detection. Anal Biochem 2018; 542: 58-62.
30. Zhang X, Li M, Zhang B, Chen K, He K. Development of a Sandwich ELISA for EHEC O157: H7 Intimin γ1. PLoS One 2016; 11(9): e0162274.
| Files | ||
| Issue | Vol 17 No 2 (2025) | |
| Section | Original Article(s) | |
| DOI | https://doi.org/10.18502/ijm.v17i2.18388 | |
| Keywords | ||
| Escherichia coli O157:H7; Indirect enzyme-linked immunosorbent assay (ELISA); Sandwich ELISA; Competitive ELISA | ||
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This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License. |
How to Cite
1.
Rezaei MJ, Eidi M, Mirhosseini SA, Kazemi R, Motamedi MJ, Khani S, Amani J. Design of ELISA-based diagnostic system for detection of enterohaemorrhagic Escherichia coli. Iran J Microbiol. 2025;17(2):278-286.
