Tropical Journal of Pharmaceutical Research

Original Research Article | OPEN ACCESS

Immunoprotective evaluation of Escherichia coli outer membrane protein A against the main pathogens of animal mastitis

Chen Chen¹, Nana Wu¹, Na Rong¹, Chao Kang¹, Chunlin Chen¹, Sanqiao Wu¹, Xiang Liu¹ , Xiaoying Zhang²

¹Chinese-German Joint Institute for Natural Product Research/Shaanxi Engineering Research Center for Tall Gastrodia Tuber and Medical Dogwood/College of Biological Science and Engineering, Shaanxi University of Technology, Hanzhong 723000, China; ²Centre of Molecular and Environmental Biology University of Minho, Department of Biology, Campus de Gualtar, Braga 4710-057, Portugal.

For correspondence:- Xiang Liu Email: iuxiang888525@163.com Tel:+862987091239

Accepted: 24 December 2019 Published: 01 February 2020

Citation: Chen C, Wu N, Rong N, Kang C, Chen C, Wu S, et al. Immunoprotective evaluation of Escherichia coli outer membrane protein A against the main pathogens of animal mastitis. Trop J Pharm Res 2020; 19(1):155-162 doi: 10.4314/tjpr.v19i1.23

© 2020 The authors.
This is an Open Access article that uses a funding model which does not charge readers or their institutions for access and distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0) and the Budapest Open Access Initiative (http://www.budapestopenaccessinitiative.org/read), which permit unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited..

Abstract

Purpose: To evaluate prokaryotic expression of the Escherichia coli (E. coli) outer membrane protein A (OmpA) and its immunoprotective function against the main pathogens of animal mastitis.

Methods: A molecular cloning method was used to develop a prokaryotic strain expressing OmpA protein, which was purified by Ni-affinity chromatography. Polyclonal antiserum was generated in mice immunized with OmpA protein. Enzyme-linked immunosorbent assay (ELISA) and western blotting were used to determine the titer and verify anti-OmpA serum specificity, respectively. Interaction between OmpA antiserum and main pathogens of animal mastitis was verified by ELISA and a pull-down method. The immune protective function of OmpA protein was evaluated in mice challenged with pathogens of animal mastitis. Optimal fermentation conditions to produce OmpA protein were determined by the L9(34) orthogonal test.

Results: A prokaryotic strain expressing OmpA protein was developed, and purified OmpA was used to develop a mouse polyclonal antibody. The anti-OmpA serum exhibited high specificity and a titer of 1:1600. Anti-OmpA serum directly interacted with E. coli and Staphylococcus aureus (S. aureus). OmpA demonstrated a significant immune protective function of 58.33 % against E. coli and 46.15 % against S. aureus. The optimal conditions for expressing fermentation OmpA were a strain absorbance of 0.5 at a wavelength of 600 nm, IPTG final concentration of 0.3 mmol/L, induction time of 12 h, and induction temperature of 28 °C.

Conclusion: OmpA possesses selective immunogenicity and a significant immune protective effect against the main pathogens of animal mastitis. The results suggest that OmpA may potentially be used as a vaccine for animal mastitis.

Keywords: E. coli, OmpA protein, Immunoprotection, Animal mastitis, Protein fermentation