Zhao Mengfei, Wu Wenqing, Song Wenbo, Yang Hao, Liu Hanyuan, Xie Rui, Huang Xi, Huang Jingwen, Hua Lin, Chen Huanchun, Wu Bin, Peng Zhong
National Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, 430070, China; Hubei Hongshan Laboratory, Wuhan, 430070, China; Frontiers Science Center for Animal Breeding and Sustainable Production, The Cooperative Innovation Center for Sustainable Pig Production, Wuhan, 430070, China.
National Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, 430070, China; Hubei Hongshan Laboratory, Wuhan, 430070, China; Frontiers Science Center for Animal Breeding and Sustainable Production, The Cooperative Innovation Center for Sustainable Pig Production, Wuhan, 430070, China.
Microb Pathog. 2025 Oct;207:107889. doi: 10.1016/j.micpath.2025.107889. Epub 2025 Jul 8.
Pasteurella multocida is a zoonotic pathogen with the ability to infect a diverse range of hosts, including humans through contact with pets. Due to its diverse capsular serogroups, rapid and accurate identification, as well as serotyping, are essential for the effective prevention and control of infections caused by this bacterium. This study developed a quadruplex quantitative PCR (qPCR) method for the identification and serotyping of P. multocida, specifically differentiating capsular serogroups A, D, and F in cats and other hosts. Primers and probes targeted the kmt1 gene (species-specific), hyaD-hyaC intergenic region (serogroup A), dcbF (serogroup D), and fcbD (serogroup F). The developed method demonstrated a detection limit of 1.7 copies/μL for the identification of P. multocida and 2.03, 11.24, and 14.52 copies/μL for the serotyping of capsular serogroups A, D, and F, respectively. The assay did not display cross-reactivity with other pathogens or different serogroups of P. multocida. Both the intra-assay and inter-assay coefficients of variation are below 1 %, highlighting the excellent reproducibility and specificity of the method. Applying this method, we examined 121 respiratory swabs from domestic cats, and 96 were tested positive for P. multocida. Among these positive samples, 66 were determined as capsular serogroup A, 1 as capsular serogroup D, and 24 as capsular serogroup F. The typing results exhibited high concordance with those obtained through bacterial isolation from the positive samples. The method developed in this study offers a rapid, accurate, and high-throughput approach for the identification and serotyping of P. multocida capsular serogroups A, D, and F. It serves as a powerful tool for advancing research into the pathogenic mechanisms of P. multocida and for implementing public health control measures.
多杀性巴氏杆菌是一种人畜共患病原体，能够感染多种宿主，包括人类通过与宠物接触而感染。由于其有多种荚膜血清群，快速准确的鉴定以及血清分型对于有效预防和控制由该细菌引起的感染至关重要。本研究开发了一种四重定量PCR（qPCR）方法用于多杀性巴氏杆菌的鉴定和血清分型，特别用于区分猫及其他宿主中的荚膜血清群A、D和F。引物和探针靶向kmt1基因（种特异性）、hyaD-hyaC基因间区域（血清群A）、dcbF（血清群D）和fcbD（血清群F）。所开发的方法对多杀性巴氏杆菌鉴定的检测限为1.7拷贝/μL，对荚膜血清群A、D和F血清分型的检测限分别为2.03、11.24和14.52拷贝/μL。该检测方法与其他病原体或多杀性巴氏杆菌的不同血清群均无交叉反应。批内和批间变异系数均低于1%，突出了该方法出色的重现性和特异性。应用该方法，我们检测了121份家猫的呼吸道拭子，其中96份多杀性巴氏杆菌检测呈阳性。在这些阳性样本中，66份被确定为荚膜血清群A，1份为荚膜血清群D，24份为荚膜血清群F。分型结果与从阳性样本中进行细菌分离所获得的结果高度一致。本研究中开发的方法为多杀性巴氏杆菌荚膜血清群A、D和F的鉴定和血清分型提供了一种快速、准确且高通量的方法。它是推进多杀性巴氏杆菌致病机制研究和实施公共卫生控制措施的有力工具。
