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In vitro Antibiotic Susceptibility, Virulence Genes Distribution and Biofilm Production of Staphylococcus aureus Isolates from Bovine Mastitis in the Liaoning Province of China

Authors Zhang DX, Li Y, Yang XQ, Su HY, Wang Q, Zhang ZH, Liu YC, Tian CL, Cui CC, Liu MC

Received 30 January 2020

Accepted for publication 16 April 2020

Published 11 May 2020 Volume 2020:13 Pages 1365—1375


Checked for plagiarism Yes

Review by Single anonymous peer review

Peer reviewer comments 2

Editor who approved publication: Dr Sahil Khanna

De-Xian Zhang, Yao Li, Xiao-Qing Yang, Hong-Yu Su, Qi Wang, Ze-Hui Zhang, Yao-Chuan Liu, Chun-Lian Tian, Can-Can Cui, Ming-Chun Liu

Key Laboratory of Livestock Infectious Diseases in Northeast China, Ministry of Education, College of Animal Science and Veterinary Medicine, Shenyang Agricultural University, Shenyang, People’s Republic of China

Correspondence: Ming-Chun Liu
College of Animal Science and Veterinary Medicine, Shenyang Agricultural University, Dongling Street, Shenhe District, Shenyang 110866, People’s Republic of China
Tel/ Fax +86 24 88487156

Purpose: The aim of this study was to identify the subtype, characterize the antimicrobial resistance, determine the virulence gene distribution, and analyze the biofilm production of Staphylococcus aureus isolates from bovine mastitis milk samples in the Liaoning Province of China.
Materials and Methods: In total, 56 Staph. aureus isolates were collected and identified in this study; the isolates were divided into different spa types based on the sequence of the polymorphic X region of the spa gene. Additionally, antimicrobial susceptibility was investigated using the broth microdilution method, and 18 virulence genes were detected using PCR. Biofilm formation was measured by spectrophotometry with crystal violet staining and observed using confocal laser scanning microscopy.
Results: There were 12.12% (56/462) milk samples that were positive for Staph. aureus. These isolates were nonsusceptible to sulfamethoxazole (100%), penicillin (76.9%), daptomycin (76.79%), clindamycin (69.64%), and oxacillin (60.71%); however, the majority of the isolates (80.4%) were susceptible to amoxicillin/clavulanate. The predominant virulence genes encoded the cytotoxins, hla (94.64%) and hlb (89.29%), and the adhesion factors clfA (89.29%), clfB (89.29%), and fnbB (80.36%). Comparatively, virulence genes related to other adhesion factors such as cna (8.93%) and enterotoxins, such as seg (26.79%), sea (16.07%), seb (7.14%), and sec (7.14%) were detected at relatively lower rates. The following eight spa types were identified: t267 (35.84%), t730 (22.64%), t518 (15.09%), t1190 (11.32%), t1456 (9.43%), t224 (1.88%), t9129 (1.88%), and t177 (1.88%). The highest biofilm production was observed for t267. Staph. aureus exhibited various patterns of biofilm formation, with the biofilm often being associated with a tower-shaped structure or a thicker biofilm.
Conclusion: Our results indicated that Staph. aureus isolates from dairy cows with mastitis in the Liaoning Province of China were non-susceptible to sulfamethoxazole, penicillin, daptomycin, oxacillin, and clindamycin. Additionally, the most prevalent subtype was t267, which displayed resistance to multiple antimicrobial agents and harbored several virulence genes, including clfA, clfB, fnbB, hla, and hlb.

Keywords: Staph. aureus, antimicrobial resistance, virulence factors, biofilm formation, mastitic dairy

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