The chimeric UreB, FliD and Omp18 proteins for a sensitive and specific diagnosis of Helicobacter pylori infections
,
Abstract
Background and Objectives: Helicobacter pylori is known as the main cause of gastrointestinal diseases including gastritis, gastric ulcer and stomach cancer. Serodiagnosis of H. pylori infection is a noninvasive and rapid method but the efficiency of this method is highly dependent to the antigens used. This study evaluated the efficacy of recombinant UreB-Omp18 and FliD for serodiagnosis of H. pylori infection.
Materials and Methods: The genes encoding for fliD, ureB, and omp18 was amplified by PCR and cloned into pET-22b and pET-28a vectors. The constructs were expressed in E. coli BL21 and purified by affinity chromatography. The antigenic properties and diagnostic potential of the recombinant proteins were analysed by immunoblotting and ELISA, respectively.
Results: The recombinant UreB-Omp18 and FliD with molecular weights of 48 kDa and 25 kDa were observed on SDS-PAGE and purified by the Ni-NTA column. The ELISA results showed that the sensitivity and specificity of recombinant UreB-Omp18 protein in serodiagnosis of H. pylori infection were 89% and 83%, respectively. Also, the sensitivity and specificity of the recombinant FliD protein were calculated to be 91% and 76%, respectively.
Conclusion: The results indicated that the recombinant UreB-Omp18 and FliD could diagnose H. pylori infection with high sensitivity and specificity.
1. Alam J, Maiti S, Ghosh P, De R, Chowdhury A, Das S, et al. Significant association of the dupA gene of Helicobacter pylori with duodenal ulcer development in a South-east Indian population. J Med Microbiol 2012; 61: 1295-1302.
2. Khalilpour A, Santhanam A, Wei LC, Saadatnia G, Velusamy N, Osman S, et al. Antigenic proteins of Helicobacter pylori of potential diagnostic value. Asian Pac J Cancer Prev 2013; 14: 1635-1642.
3. Sukri A, Hanafiah A, Mohamad Zin N, Kosai NR. Epidemiology and role of Helicobacter pylori virulence factors in gastric cancer carcinogenesis. APMIS 2020; 128: 150-161.
4. Schumann C, Triantafilou K, Rasche FM, Möricke A, Vogt K, Triantafilou M, et al. Serum antibody positivity for distinct Helicobacter pylori antigens in benign and malignant gastroduodenal disease. Int J Med Microbiol 2006; 296: 223-228.
5. Khashei R, Dara M, Bazargani A, Bagheri Lankarani K, Taghavi A, Moeini M, et al. High rate of A2142G point mutation associated with clarithromycin resistance among Iranian Helicobacter pylori clinical isolates. APMIS 2016; 124: 787-793.
6. Ueda J, Okuda M, Nishiyama T, Lin Y, Fukuda Y, Kikuchi S. Diagnostic accuracy of the E-plate serum antibody test kit in detecting Helicobacter pylori infection among Japanese children. J Epidemiol 2014; 24: 47-51.
7. Marchildon PA, Sugiyama T, Fukada Y, Peacock JS, Asaka M, Shimoyama T, et al. Evaluation of the effects of strain-specific antigen variation on the accuracy of serologic diagnosis of Helicobacter pylori infection. J Clin Microbiol 2003; 41: 1480-1485.
8. Khalifeh Gholi M, Kalali B, Formichella L, Göttner G, Shamsipour F, Zarnani AH, et al. Helicobacter pylori FliD protein is a highly sensitive and specific marker for serologic diagnosis of H. pylori infection. Int J Med Microbiol 2013; 303: 618-623.
9. Pan KF, Formichella L, Zhang L, Zhang Y, Ma JL, Li ZX, et al. Helicobacter pylori antibody responses and evolution of precancerous gastric lesions in a Chinese population. Int J Cancer 2014; 134: 2118-2125.
10. Oleastro M, Matos R, Cabral J, Barros R, Lopes AI, Ramalho P, et al. Evaluation of a Western blot test, Helico Blot 2.1, in the diagnosis of Helicobacter pylori infection in a pediatric population. Helicobacter 2002; 7: 210-215.
11. Formichella L, Romberg L, Bolz C, Vieth M, Geppert M, Göttner G, et al. A novel line immunoassay based on recombinant virulence factors enables highly specific and sensitive serologic diagnosis of Helicobacter pylori infection. Clin Vaccine Immunol 2013; 20: 1703-1710.
12. Raoufi E, Akrami H, Khansarinejad B, Abtahi H. Expression and antigenic evaluation of Helicobacter pylori UreB fragment. Jundishapur J Microbiol 2017; 10(5): e41645.
13. Talebkhan Y, Bababeik M, Esmaeili M, Oghalaei A, Saberi S, Karimi Z, et al. Helicobacter pylori bacterial ghost containing recombinant Omp18 as a putative vaccine. J Microbiol Methods 2010; 82: 334-337.
14. Talebkhan Y, Ebrahimzadeh F, Esmaeili M, Zamaninia L, Nahvijoo A, Khedmat H, et al. Helicobacter pylori Omp18 and its application in serologic screening of infection. Curr Microbiol 2011; 62: 325-330.
15. Mirjamali NA, Soufian S, Molaee N, Abbasian SS, Abtahi H. Cloning and expression of the enzymatic region of Streptococcal hyaluronidase. Iran J Basic Med Sci 2014; 17: 667-672.
16. Hasanzadeh L, Ghaznavi-Rad E, Soufian S, Farjadi V, Abtahi H. Expression and antigenic evaluation of VacA antigenic fragment of Helicobacter pylori. Iran J Basic Med Sci 2013; 16: 835-840.
17. McMahon BJ, Bruce MG, Koch A, Goodman KJ, Tsukanov V, Mulvad G, et al. The diagnosis and treatment of Helicobacter pylori infection in Arctic regions with a high prevalence of infection: Expert Commentary. Epidemiol Infect 2016; 144: 225-233.
18. Frydman GH, Davis N, Beck PL, Fox JG. Helicobacter pylori eradication in patients with immune thrombocytopenic purpura: A review and the role of biogeography. Helicobacter 2015; 20: 239-251.
19. Perrais M, Rousseaux C, Ducourouble MP, Courcol R, Vincent P, Jonckheere N, et al. Helicobacter pylori urease and flagellin alter mucin gene expression in human gastric cancer cells. Gastric Cancer 2014; 17: 235-246.
20. Wang Y-K, Kuo F-C, Liu C-J, Wu M-C, Shih H-Y, Wang S-S, et al. Diagnosis of Helicobacter pylori infection: Current options and developments. World J Gastroenterol 2015; 21: 11221-11235.
21. Negash M, Kassu A, Amare B, Yismaw G, Moges B. Evaluation of SD BIOLINE H. pylori Ag rapid test against double ELISA with SD H. pylori Ag ELISA and EZ-STEP H. pylori Ag ELISA tests. BMC Clin Pathol 2018; 18: 4.
22. Rastawicki W, Wiechetek-Płaza K. Seroprevalence of Helicobacter pylori infections in children and adults in Poland in the years 2020-2023. Przegl Epidemiol 2023; 77: 317-326.
23. Manes G, Zanetti MV, Piccirillo MM, Lombardi G, Balzano A, Pieramico O. Accuracy of a new monoclonal stool antigen test in post-eradication assessment of Helicobacter pylori infection: comparison with the polyclonal stool antigen test and urea breath test. Dig Liver Dis 2005; 37: 751-755.
24. Pelerito A, Oleastro M, Lopes AI, Ramalho P, Cabral J, Monteiro L. Evaluation of rapid test Assure Helicobacter pylori for diagnosis of H. pylori in pediatric population. J Microbiol Methods 2006; 66: 331-335.
25. Michel A, Waterboer T, Kist M, Pawlita M. Helicobacter pylori multiplex serology. Helicobacter 2009; 14: 525-535.
26. Yan J, Mao Y-F, Shao Z-X. Frequencies of the expression of main protein antigens from Helicobacter pylori isolates and production of specific serum antibodies in infected patients. World J Gastroenterol 2005; 11: 421-425.
27. Kao CY, Sheu BS, Wu JJ. Helicobacter pylori infection: An overview of bacterial virulence factors and pathogenesis. Biomed J 2016; 39: 14-23.
28. Li H, Zhang B, Hu X, Dong Y, Fan Q, Guo F, et al. Serum Helicobacter pylori FliD antibody and the risk of gastric cancer. Oncotarget 2016; 7: 22397-22408.
29. Haas G, Karaali G, Ebermayer K, Metzger WG, Lamer S, Zimny-Arndt U, et al. Immunoproteomics of Helicobacter pylori infection and relation to gastric disease. Proteomics 2002; 2: 313-324.
30. Voland P, Weeks DL, Vaira D, Prinz C, Sachs G. Specific identification of three low molecular weight membrane-associated antigens of Helicobacter pylori. Aliment Pharmacol Ther 2002; 16: 533-544.
31. Pawelec DP, Korsak D, Wyszyńska AK, Rozynek E, Popowski J, Jagusztyn-Krynicka EK. Genetic diversity of the Campylobacter genes coding immunodominant proteins. FEMS Microbiol Lett 2000; 185: 43-49.
32. Rathinavelu S, Kao JY, Zavros Y, Merchant JL. Helicobacter pylori outer membrane protein 18 (Hp1125) induces dendritic cell maturation and function. Helicobacter 2005; 10: 424-432.
33. Zhang X, Zhang J, Yang F, Wu W, Sun H, Xie Q, et al. Immunization with Heat Shock Protein A and γ-Glutamyl Transpeptidase Induces Reduction on the Helicobacter pylori Colonization in Mice. PLoS One 2015; 10(6): e0130391.
34. Zhou WY, Shi Y, Wu C, Zhang WJ, Mao XH, Guo G, et al. Therapeutic efficacy of a multi-epitope vaccine against Helicobacter pylori infection in BALB/c mice model. Vaccine 2009; 27: 5013-5019.
35. Li HB, Zhang JY, He YF, Chen L, Li B, Liu KY, et al. Systemic immunization with an epitope-based vaccine elicits a Th1-biased response and provides protection against Helicobacter pylori in mice. Vaccine 2012; 31: 120-126.
2. Khalilpour A, Santhanam A, Wei LC, Saadatnia G, Velusamy N, Osman S, et al. Antigenic proteins of Helicobacter pylori of potential diagnostic value. Asian Pac J Cancer Prev 2013; 14: 1635-1642.
3. Sukri A, Hanafiah A, Mohamad Zin N, Kosai NR. Epidemiology and role of Helicobacter pylori virulence factors in gastric cancer carcinogenesis. APMIS 2020; 128: 150-161.
4. Schumann C, Triantafilou K, Rasche FM, Möricke A, Vogt K, Triantafilou M, et al. Serum antibody positivity for distinct Helicobacter pylori antigens in benign and malignant gastroduodenal disease. Int J Med Microbiol 2006; 296: 223-228.
5. Khashei R, Dara M, Bazargani A, Bagheri Lankarani K, Taghavi A, Moeini M, et al. High rate of A2142G point mutation associated with clarithromycin resistance among Iranian Helicobacter pylori clinical isolates. APMIS 2016; 124: 787-793.
6. Ueda J, Okuda M, Nishiyama T, Lin Y, Fukuda Y, Kikuchi S. Diagnostic accuracy of the E-plate serum antibody test kit in detecting Helicobacter pylori infection among Japanese children. J Epidemiol 2014; 24: 47-51.
7. Marchildon PA, Sugiyama T, Fukada Y, Peacock JS, Asaka M, Shimoyama T, et al. Evaluation of the effects of strain-specific antigen variation on the accuracy of serologic diagnosis of Helicobacter pylori infection. J Clin Microbiol 2003; 41: 1480-1485.
8. Khalifeh Gholi M, Kalali B, Formichella L, Göttner G, Shamsipour F, Zarnani AH, et al. Helicobacter pylori FliD protein is a highly sensitive and specific marker for serologic diagnosis of H. pylori infection. Int J Med Microbiol 2013; 303: 618-623.
9. Pan KF, Formichella L, Zhang L, Zhang Y, Ma JL, Li ZX, et al. Helicobacter pylori antibody responses and evolution of precancerous gastric lesions in a Chinese population. Int J Cancer 2014; 134: 2118-2125.
10. Oleastro M, Matos R, Cabral J, Barros R, Lopes AI, Ramalho P, et al. Evaluation of a Western blot test, Helico Blot 2.1, in the diagnosis of Helicobacter pylori infection in a pediatric population. Helicobacter 2002; 7: 210-215.
11. Formichella L, Romberg L, Bolz C, Vieth M, Geppert M, Göttner G, et al. A novel line immunoassay based on recombinant virulence factors enables highly specific and sensitive serologic diagnosis of Helicobacter pylori infection. Clin Vaccine Immunol 2013; 20: 1703-1710.
12. Raoufi E, Akrami H, Khansarinejad B, Abtahi H. Expression and antigenic evaluation of Helicobacter pylori UreB fragment. Jundishapur J Microbiol 2017; 10(5): e41645.
13. Talebkhan Y, Bababeik M, Esmaeili M, Oghalaei A, Saberi S, Karimi Z, et al. Helicobacter pylori bacterial ghost containing recombinant Omp18 as a putative vaccine. J Microbiol Methods 2010; 82: 334-337.
14. Talebkhan Y, Ebrahimzadeh F, Esmaeili M, Zamaninia L, Nahvijoo A, Khedmat H, et al. Helicobacter pylori Omp18 and its application in serologic screening of infection. Curr Microbiol 2011; 62: 325-330.
15. Mirjamali NA, Soufian S, Molaee N, Abbasian SS, Abtahi H. Cloning and expression of the enzymatic region of Streptococcal hyaluronidase. Iran J Basic Med Sci 2014; 17: 667-672.
16. Hasanzadeh L, Ghaznavi-Rad E, Soufian S, Farjadi V, Abtahi H. Expression and antigenic evaluation of VacA antigenic fragment of Helicobacter pylori. Iran J Basic Med Sci 2013; 16: 835-840.
17. McMahon BJ, Bruce MG, Koch A, Goodman KJ, Tsukanov V, Mulvad G, et al. The diagnosis and treatment of Helicobacter pylori infection in Arctic regions with a high prevalence of infection: Expert Commentary. Epidemiol Infect 2016; 144: 225-233.
18. Frydman GH, Davis N, Beck PL, Fox JG. Helicobacter pylori eradication in patients with immune thrombocytopenic purpura: A review and the role of biogeography. Helicobacter 2015; 20: 239-251.
19. Perrais M, Rousseaux C, Ducourouble MP, Courcol R, Vincent P, Jonckheere N, et al. Helicobacter pylori urease and flagellin alter mucin gene expression in human gastric cancer cells. Gastric Cancer 2014; 17: 235-246.
20. Wang Y-K, Kuo F-C, Liu C-J, Wu M-C, Shih H-Y, Wang S-S, et al. Diagnosis of Helicobacter pylori infection: Current options and developments. World J Gastroenterol 2015; 21: 11221-11235.
21. Negash M, Kassu A, Amare B, Yismaw G, Moges B. Evaluation of SD BIOLINE H. pylori Ag rapid test against double ELISA with SD H. pylori Ag ELISA and EZ-STEP H. pylori Ag ELISA tests. BMC Clin Pathol 2018; 18: 4.
22. Rastawicki W, Wiechetek-Płaza K. Seroprevalence of Helicobacter pylori infections in children and adults in Poland in the years 2020-2023. Przegl Epidemiol 2023; 77: 317-326.
23. Manes G, Zanetti MV, Piccirillo MM, Lombardi G, Balzano A, Pieramico O. Accuracy of a new monoclonal stool antigen test in post-eradication assessment of Helicobacter pylori infection: comparison with the polyclonal stool antigen test and urea breath test. Dig Liver Dis 2005; 37: 751-755.
24. Pelerito A, Oleastro M, Lopes AI, Ramalho P, Cabral J, Monteiro L. Evaluation of rapid test Assure Helicobacter pylori for diagnosis of H. pylori in pediatric population. J Microbiol Methods 2006; 66: 331-335.
25. Michel A, Waterboer T, Kist M, Pawlita M. Helicobacter pylori multiplex serology. Helicobacter 2009; 14: 525-535.
26. Yan J, Mao Y-F, Shao Z-X. Frequencies of the expression of main protein antigens from Helicobacter pylori isolates and production of specific serum antibodies in infected patients. World J Gastroenterol 2005; 11: 421-425.
27. Kao CY, Sheu BS, Wu JJ. Helicobacter pylori infection: An overview of bacterial virulence factors and pathogenesis. Biomed J 2016; 39: 14-23.
28. Li H, Zhang B, Hu X, Dong Y, Fan Q, Guo F, et al. Serum Helicobacter pylori FliD antibody and the risk of gastric cancer. Oncotarget 2016; 7: 22397-22408.
29. Haas G, Karaali G, Ebermayer K, Metzger WG, Lamer S, Zimny-Arndt U, et al. Immunoproteomics of Helicobacter pylori infection and relation to gastric disease. Proteomics 2002; 2: 313-324.
30. Voland P, Weeks DL, Vaira D, Prinz C, Sachs G. Specific identification of three low molecular weight membrane-associated antigens of Helicobacter pylori. Aliment Pharmacol Ther 2002; 16: 533-544.
31. Pawelec DP, Korsak D, Wyszyńska AK, Rozynek E, Popowski J, Jagusztyn-Krynicka EK. Genetic diversity of the Campylobacter genes coding immunodominant proteins. FEMS Microbiol Lett 2000; 185: 43-49.
32. Rathinavelu S, Kao JY, Zavros Y, Merchant JL. Helicobacter pylori outer membrane protein 18 (Hp1125) induces dendritic cell maturation and function. Helicobacter 2005; 10: 424-432.
33. Zhang X, Zhang J, Yang F, Wu W, Sun H, Xie Q, et al. Immunization with Heat Shock Protein A and γ-Glutamyl Transpeptidase Induces Reduction on the Helicobacter pylori Colonization in Mice. PLoS One 2015; 10(6): e0130391.
34. Zhou WY, Shi Y, Wu C, Zhang WJ, Mao XH, Guo G, et al. Therapeutic efficacy of a multi-epitope vaccine against Helicobacter pylori infection in BALB/c mice model. Vaccine 2009; 27: 5013-5019.
35. Li HB, Zhang JY, He YF, Chen L, Li B, Liu KY, et al. Systemic immunization with an epitope-based vaccine elicits a Th1-biased response and provides protection against Helicobacter pylori in mice. Vaccine 2012; 31: 120-126.
| Files | ||
| Issue | Vol 16 No 3 (2024) | |
| Section | Original Article(s) | |
| DOI | https://doi.org/10.18502/ijm.v16i3.15768 | |
| Keywords | ||
| Helicobacter pylori; UreB; Omp18; FliD protein; Chimeric antigen | ||
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This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License. |
How to Cite
1.
Seyyedhamzeh H, Farajnia S, Kargar M, Baradaran B, Kafilzadeh F. The chimeric UreB, FliD and Omp18 proteins for a sensitive and specific diagnosis of Helicobacter pylori infections. Iran J Microbiol. 2024;16(3):357-365.
