PAKISTAN VETERINARY JOURNAL

Pak Vet J, 2020, 40(3): 307-312

ESBL-Producing Escherichia coli from Bovine Mastitis Induced Apoptosis of Bovine Mammary Epithelial Cells Via Alteration of ROS/MMP/bax/bcl-2 Signaling Pathway

Yuxiang Shi^1,2^§, Peng Chen¹^§, Wenlin Huo¹, Xi Chen¹, Herman W. Barkema³, Jeroen De Buck³, Muhammad Shahid^1,4, Bo Han^1* and Jian Gao^1*

¹Department of Clinical Veterinary Medicine, College of Veterinary Medicine, China Agricultural University, Beijing 100193, China; ²College of Life Sciences and Food Engineering, Hebei University of Engineering, Handan, Hebei 056038, P.R. China; ³Department of Production Animal Health, Faculty of Veterinary Medicine, University of Calgary, Calgary, AB, Canada, T2N 4N1; ⁴Center of Microbiology and Biotechnology, Veterinary Research Institute, Peshawar, Pakistan
*Corresponding author: hanbo@cau.edu.cn (Bo Han); gaojian2016@cau.edu.cn (Jian Gao)
^§Contribute as First author

Abstract

Bovine mastitis is a common disease which causes important economic losses for dairy industry. Escherichia coli (E. coli) is one of major pathogens for both clinical and subclinical bovine mastitis. The persistent use and unwarranted application of antimicrobials has induced dynamic and continuous production and mutation of β-lactamases in extended-spectrum β-lactamase producing E. coli (ESBLEC). However, the pathogenic pathway of ESBLEC on bovine mammary epithelial cells (bMECs) is unknown. Aim of the present study was therefore to determine the apoptotic effects and mechanism of bovine ESBLEC on bMECs. Fifty-seven (14%) E. coli strains were isolated and identified from 400 clinical mastitic cows, and 20 (35%) strains of ESBLEC were confirmed by double-disc synergy testing and PCR. The apoptosis of bMECs infected with E. coli was analyzed with Annexin V/PI detection kit and TUNEL assay. Intracellular ROS generation and MMP were detected with commercial kits, respectively. The protein analysis of bax, bcl-2 and cleaved caspase-3 were determined by western blotting. Morphological changes were observed by TEM. After infection with ESBLEC, the apoptosis of bMECs increased significantly (P<0.05) as compared with the blank control group (bMECs were cultured without bacteria), evidenced by the decreased MMP, ROS level overproduction, upregulated expression of bax/bcl-2 ratio, cleaved caspase-3, and increased TUNEL positive cells. In addition, bMECs infected with ESBLEC manifested specific ultrastructure features like cytoplasmic cavitation, swollen mitochondria, cytomembrane disruption, and appearance of apoptotic bodies. Thus, ESBLEC has the capability to impart significant damages to the bMECs resulting in apoptosis via ROS/MMP/bax/bcl-2 pathway.

To Cite This Article: Shi Y, P Chen, W Huo, X Chen, HW Barkema, JD Buck, M Shahid, B Han and J Gao, 2020. ESBL-producing Escherichia coli from bovine mastitis induced apoptosis of bovine mammary epithelial cells via alteration of ROS/MMP/bax/bcl-2 signaling pathway. Pak Vet J, 40(3): 307-312. http://dx.doi.org/10.29261/pakvetj/2020.038

Abstract

ISSN 0253-8318 (Print)ISSN 2074-7764 (Online)

ISSN 0253-8318 (Print)
ISSN 2074-7764 (Online)