Volume 27, Issue 2 (Iran South Med J 2024)
Iran South Med J 2024, 27(2): 123-137 |
Back to browse issues page
Download citation:
BibTeX | RIS | EndNote | Medlars | ProCite | Reference Manager | RefWorks
Send citation to:
BibTeX | RIS | EndNote | Medlars | ProCite | Reference Manager | RefWorks
Send citation to:
Samiei-Abianeh H, Nazarian S, Resaei A, Hadi N, Kayvan shokooh K. Production of Chicken Egg Yolk Immuno-globulin (IgY) against Recombinant TolC Pro-tein From Shigella flexneri Bacteria. Iran South Med J 2024; 27 (2) :123-137
URL: http://ismj.bpums.ac.ir/article-1-2011-en.html
URL: http://ismj.bpums.ac.ir/article-1-2011-en.html
1- Department of Medical Biotechnology and Nanotechnology, School of Medicine، Mashhad University of Medical Sciences Mashhad, Iran
Department of Biology, School of Basic Sciences, Imam Hossein University, Tehran, Iran
2- Department of Biology, School of Basic Sciences, Imam Hossein University, Tehran, Iran ,nazarian@ihu. ac.ir
3- Department of Biology, School of Basic Sciences, Imam Hossein University, Tehran, Iran
4- Department of Bacteriology and Virology, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran
5- Department of Microbiology, School of Advanced Science and Technology, Tehran Medical Science, Islamic Azad University, Tehran, Iran
Department of Biology, School of Basic Sciences, Imam Hossein University, Tehran, Iran
2- Department of Biology, School of Basic Sciences, Imam Hossein University, Tehran, Iran ,
3- Department of Biology, School of Basic Sciences, Imam Hossein University, Tehran, Iran
4- Department of Bacteriology and Virology, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran
5- Department of Microbiology, School of Advanced Science and Technology, Tehran Medical Science, Islamic Azad University, Tehran, Iran
Abstract: (810 Views)
Background: Growing antibiotic resistance is a global concern for the treatment of shigellosis. The development of efflux pumps such as the AcrAB-TolC pump has increased antibiotic resistance in Shigella flexneri strains. The aim of this study was to produce IgY against the recombinant immunogen TolC and evaluate the binding of the antibody to the recombinant immunogen
Materials and Methods: A construct containing immunogen regions of TolC protein was designed and synthesized; then, the recombinant plasmid pET28a-tolC was transferred to E. coli BL21 (DE3) competent cells. Expression of the recombinant protein in E. coli was induced with 1 mM IPTG. The recombinant protein was purified using affinity chromatography by Nickel-Nitrilotriacetic acid (Ni-NTA) resin and evaluated by western blotting. Leghorn chickens were immunized with 200 micrograms of the recombinant protein. IgY was purified using PEG precipitation, and the antibody titer was determined in immunized chicken’s serum and egg yolk by ELISA method compared to the control group.
Results: Recombinant protein TolC was expressed with a molecular weight of 40 kDa and an expression yield of 10 mg/l of the culture media. The IgY concentration was determined to be 9 mg/ml of egg yolk. The antibody titer in serum against the recombinant protein after the third immunization was 1: 6400 (P<0.05). In the ELISA method, at least seven nanograms of antibody reacted with the recombinant protein.
Conclusion: Antibodies against TolC recombinant immunogen can be investigated as a source of passive immunity in the disease caused by Shigella flexneri.
Materials and Methods: A construct containing immunogen regions of TolC protein was designed and synthesized; then, the recombinant plasmid pET28a-tolC was transferred to E. coli BL21 (DE3) competent cells. Expression of the recombinant protein in E. coli was induced with 1 mM IPTG. The recombinant protein was purified using affinity chromatography by Nickel-Nitrilotriacetic acid (Ni-NTA) resin and evaluated by western blotting. Leghorn chickens were immunized with 200 micrograms of the recombinant protein. IgY was purified using PEG precipitation, and the antibody titer was determined in immunized chicken’s serum and egg yolk by ELISA method compared to the control group.
Results: Recombinant protein TolC was expressed with a molecular weight of 40 kDa and an expression yield of 10 mg/l of the culture media. The IgY concentration was determined to be 9 mg/ml of egg yolk. The antibody titer in serum against the recombinant protein after the third immunization was 1: 6400 (P<0.05). In the ELISA method, at least seven nanograms of antibody reacted with the recombinant protein.
Conclusion: Antibodies against TolC recombinant immunogen can be investigated as a source of passive immunity in the disease caused by Shigella flexneri.
Type of Study: Original |
Subject:
Biochemistry. Cell Biology and Genetics
Received: 2024/06/17 | Accepted: 2024/10/8 | Published: 2024/12/7
Received: 2024/06/17 | Accepted: 2024/10/8 | Published: 2024/12/7
References
1. Levine MM, Kotloff KL, Barry EM & et al. Clinical trials of Shigella vaccines: two steps forward and one step back on a long, hard road. Nat Rev Microbiol 2007; 5(7): 540-53. [DOI]
2. Lampel KA, Formal SB and Maurelli AT. A Brief History of Shigella. EcoSal Plus 2018; 8(1). [DOI]
3. Gentle A, Ashton PM, Dallman TJ & et al. Evaluation of Molecular Methods for Serotyping Shigella flexneri. J Clin Microbiol 2016; 54(6): 1456-61. [DOI]
4. Hosangadi, D, WHO consultation on ETEC and Shigella burden of disease, Geneva, 6-7th April 2017: Meeting report. Vaccine 2019; 37(50): 7381-90. [DOI]
5. Qiu Sh, Wang Y, Li P & et al. Multidrug-resistant atypical variants of Shigella flexneri in China. Emerg Infect Dis 2013; 19(7): 1147-50. [DOI]
6. Taneja N & Mewara A. Shigellosis: Epidemiology in India. Indian J Med Res 2016; 143(5): 565-76. [Article]
7. Hassan, K.A, et al. Homologs of the Acinetobacter baumannii AceI transporter represent a new family of bacterial multidrug efflux systems. mBio 2015; 6(1): e01982-14. [DOI]
8. Blanco P, ernando-Amado S, Reales-Calderon JA & et al. Bacterial Multidrug Efflux Pumps: Much More Than Antibiotic Resistance Determinants. Microorganisms 2016; 4(1): 14. [DOI]
9. Alav I, Kobylka J, Kuth MS & et al. Structure, Assembly, and Function of Tripartite Efflux and Type 1 Secretion Systems in Gram-Negative Bacteria. Chem Rev 2021; 121(9): 5479-5596. [DOI]
10. Klemperer F. Ueber natürliche Immunität und ihre Verwerthung für die Immunisirungstherapie. Archiv für experimentelle pathologie und pharmakologie 1893; 31: 356-382. [DOI]
11. Pereira EPV, van Tilburg MF, Florean EOPT & et al. Egg yolk antibodies (IgY) and their applications in human and veterinary health: A review. Int Immunopharmacol 2019; 73: 293-303. [DOI]
12. He F. Laemmli-SDS-PAGE. bio-protocol 2011; 1(11). [Article]
13. Kielkopf CL, Bauer W, Urbatsch IL. Bradford Assay for Determining Protein Concentration. Cold Spring Harb Protoc 2020; 2020(4): 102269. [DOI]
14. Ahmed SF, Riddle MS, Wierzba TF & et al. Epidemiology and genetic characterization of Shigella flexneri strains isolated from three paediatric populations in Egypt (2000-2004). Epidemiol Infect 2006; 134(6): 1237-48. [DOI]
15. Jolly-Ryan J. Ebolamania and Equal Protection of Health Care Workers Under Rational Basis with Bite Review. WVLR 2017; 120(2): 575-627. [Article]
16. Ali Nor BSh, Menza NCh, Musyoki AB. Multidrug‐ resistant shigellosis among children aged below five years with diarrhea at Banadir Hospital in Mogadishu, Somalia. canad J infect dis med microbiol 2021; 2021(1): 6630272. [DOI]
17. Ebrahimi A A, Ebrahimi S, Aghouli M. Survey of resistance rate of Shigella species isolated from children with diarrhea Fasa, Summer, 1383. Iran South Med J 2009; 12(3): 225-230. (Persian) [Article]
18. Veisi R, Nazarian Sh, Fathi J & et al. Expression and purification of TolC as a recombinant protein vaccine against Shigella flexneri and evaluation of immunogenic response in mice. Microbial Pathogenesis 2024; 188: 106539. [DOI]
19. Soto SM. Role of efflux pumps in the antibiotic resistance of bacteria embedded in a biofilm. Virulence 2013; 4(3): 223-9. [DOI]
20. Coluccia, M., et al., Role of the MDR efflux pump AcrAB in epithelial cell invasion by Shigella flexneri. Biomolecules 2023; 13(5): 823. [DOI]
21. Leow CY, Kazi A, Hisyam Ismail CM & et al. Reverse vaccinology approach for the identification and characterization of outer membrane proteins of Shigella flexneri as potential cellular-and antibody-dependent vaccine candidates. Clin Exp Vaccine Res 2020; 9(1): 15-25. [DOI]
22. Baseer S, Ahmad S, Ranaghan KE & et al. Towards a peptide-based vaccine against Shigella sonnei: A subtractive reverse vaccinology based approach. Biologicals 2017; 50: 87-99. [DOI]
23. Koronakis V. TolC – the bacterial exit duct for proteins and drugs. FEBS Lett 2003; 555(1): 66- 71. [DOI]
24. Jennison AV, Raqib R, Verma NK. Immunoproteome analysis of soluble and membrane proteins of Shigella flexneri 2457T. World J Gastroenterol 2006; 12(41): 6683-88. [DOI]
25. Costa T, Felisberto-Rodrigues C, Meir A & et al. Secretion systems in Gram-negative bacteria: structural and mechanistic insights. Nat Rev Microbiol 2015; 13(6): 343-359. [DOI]
26. Li X, Nakano T, Sunwoo HH & et al. Effects of egg and yolk weights on yolk antibody (IgY) production in laying chickens. Poult Sci 1998; 77(2): 266-270. [DOI]
27. Verdoliva A, Basile G, Fassina G. Affinity purification of immunoglobulins from chicken egg yolk using a new synthetic ligand. J Chromatogr B Biomed Sci 2000; 749(2): 233-42. [DOI]
28. Shin JH, Yang M, Nam SW & et al. Use of egg yolk-derived immunoglobulin as an alternative to antibiotic treatment for control of Helicobacter pylori infection. Clin Diagn Lab Immunol 2002; 9(5): 1061-6. [DOI]
29. Marquardt RR, Jin LZ, Kim JW & et al. Passive protective effect of egg-yolk antibodies against enterotoxigenic Escherichia coli K88+ infection in neonatal and early-weaned piglets. FEMS Immunol Med Microbiol 1999; 23(4): 283-8. [DOI]
30. Sarker SA, Casswall TH, Juneja LR & et al. Randomized, placebo-controlled, clinical trial of hyperimmunized chicken egg yolk immunoglobulin in children with rotavirus diarrhea. J Pediatr Gastroenterol Nutr 2001; 32(1): 19-25. [Article]
31. Sugita-Konishi Y, Shibata K, Yun SS & et al. Immune functions of immunoglobulin Y isolated from egg yolk of hens immunized with various infectious bacteria. Biosci Biotechnol Biochem 1996; 60(5): 886-8. [DOI]
32. Hadi N, Nazarian S, Rouhi S & et al. Production of egg yolk antibody (IgY) against a chimeric protein containing IpaD, StxB, and TolC antigens from Shigella: An investigation of its prophylactic effects against Shiga toxin (Stx) and Shigella dysenteriae in vitro and in vivo. Heliyon 2024; 10(4): e26361. [DOI]
33. Silva TO, Costa LVS, L Chagas do Nascimento & et al. The Escherichia coli TolC efflux pump protein is immunogenic and elicits protective antibodies. BioRxiv 2023: qiae201. [DOI]
Send email to the article author
| Rights and Permissions | |
 |
This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License. |
