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ORIGINAL RESEARCH article

Front. Microbiol.

Sec. Infectious Agents and Disease

Volume 16 - 2025 | doi: 10.3389/fmicb.2025.1633809

This article is part of the Research TopicRapid and Efficient Analytical Technologies for Pathogen DetectionView all 7 articles

Establishment and application of a TaqMan-based quantitative PCR assay for simultaneous detection of bovine Brucella spp. and Mycobacterium spp

Provisionally accepted
Shuai  ZhangShuai Zhang1*Hui  ZhaoHui Zhao1Qiuju  GuoQiuju Guo1Ruixue  XueRuixue Xue2Zixin  JiangZixin Jiang2Wenduo  JiangWenduo Jiang2Linlin  XingLinlin Xing2Xinhui  WeiXinhui Wei1208  Lab Diao208 Lab Diao1YI  TANGYI TANG3Zouran  LanZouran Lan2Yue  ZhangYue Zhang2
  • 1Shandong Agricultural University, Taian, China
  • 2Shandong Animal Disease Prevention and Control Centre (Shandong Zoonotic Disease Flow Monitoring Centre), Jinan, China
  • 3Beijing Animal Husbandry and Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Beijing, China

The final, formatted version of the article will be published soon.

Brucellosis and tuberculosis are two zoonotic, chronic infectious diseases caused by bacteria of the genus Brucella and Mycobacterium, respectively, which pose significant hazards to both animal husbandry and human health. Currently, mixed infections of these two pathogens are prevalent in livestock production; thus, establishing a molecular diagnostic method for the simultaneous detection and analysis of brucellosis and tuberculosis is crucial for the prevention and control of these diseases. By utilizing conserved regions within the genomes of Brucella and Mycobacterium, we designed specific primers and probes. After optimizing the developed qPCR assay conditions, we determined the lower limit of detection to be 10 copies/μL. Cross-testing with other bovine-derived pathogens demonstrated no cross-reactivity. Repeatability tests indicated that the coefficient of variation for the developed qPCR assay was less than 4.10% both within and between batches. We employed both the developed qPCR assay and a commercial qPCR assay to analyze 60 mixed infection samples of Brucella and Mycobacterium from various regions. The results revealed positivity rates of 100% and 96.67% for Brucella, and 100% and 95.00% for Mycobacterium, respectively. These findings indicate that a highly sensitive, specific, reproducible, and versatile qPCR method has been developed for the simultaneous quantitative detection of Brucella and Mycobacterium, which can be applied in studying the pathogenesis and epidemiology of these pathogens.

Keywords: Brucella, Mycobacterium, Real-Time PCR, TaqMan probe, Simultaneous detection

Received: 26 May 2025; Accepted: 31 Jul 2025.

Copyright: © 2025 Zhang, Zhao, Guo, Xue, Jiang, Jiang, Xing, Wei, Diao, TANG, Lan and Zhang. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

* Correspondence: Shuai Zhang, Shandong Agricultural University, Taian, China

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