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 11 articles
Establishment and application of a TaqMan-based quantitative PCR assay for simultaneous detection of bovine Brucella spp. and Mycobacterium spp
Provisionally accepted
Shuai Zhang1*
Hui Zhao1Qiuju Guo1
Ruixue Xue2Zixin Jiang2Wenduo Jiang2Linlin Xing2Xinhui Wei1
208 Lab Diao1
YI TANG3Zouran Lan2Yue 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
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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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