Zhang Shaohua, Zhao Yeping, Liang Weijia, Wang Shuai, Cui Xiu, Zhu Haohan, Zhang Yueyue, Liu Xiaolei, Li Huimin, Mu Wenjie, Guo Aijiang
State Key Laboratory for Animal Disease Control and Prevention, Key Laboratory of Veterinary Parasitology of Gansu Province, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, PR China; State Key Laboratory for Diagnosis and Treatment of Severe Zoonotic Infectious Diseases/Key Laboratory for Zoonosis Research of the Ministry of Education, College of Veterinary Medicine, Jilin University, Changchun, PR China.
State Key Laboratory for Animal Disease Control and Prevention, Key Laboratory of Veterinary Parasitology of Gansu Province, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, PR China; State Key Laboratory for Diagnosis and Treatment of Severe Zoonotic Infectious Diseases/Key Laboratory for Zoonosis Research of the Ministry of Education, College of Veterinary Medicine, Jilin University, Changchun, PR China.
Vet Parasitol. 2025 Oct;339:110582. doi: 10.1016/j.vetpar.2025.110582. Epub 2025 Aug 15.
Monieziasis is a prevalent issue in small ruminant husbandry, primarily caused by Moniezia expansa and M. benedeni in China. There is a critical need for highly sensitive methods for disease surveillance and prevention. In this study, we developed a visual assay combining recombinase polymerase amplification (RPA) with a lateral flow strip (RPA-LFD) for the rapid detection of Moniezia spp. in sheep fecal samples. Seven primer/probe sets were designed based on a specific fragment of the M. expansa β-tubulin gene (MTUB). The RPA-LFD assay performed optimally under reaction conditions of 39 ℃ for 15 min, with a primer-to-probe ratio of 4:0.6. The optimized method demonstrated high specificity for Moniezia spp., with no cross-reactivity with genomic DNA from other common gastrointestinal parasites in ruminant livestock. The detection limit was 10 copies/μL of plasmid DNA or 10 pg/μL of M. expansa genomic DNA per reaction. When compared to PCR using clinical and sheep-simulated samples, the RPA-LFD assay exhibited equivalent detection capability, achieving 95.7 % consistency (K value = 0.939, p < 0.001). These results suggest that the RPA-LFD method is a reliable and portable diagnostic tool for routine screening, monitoring, and rapid confirmation of monieziasis in sheep flocks, particularly in endemic areas and remote regions.
莫尼茨绦虫病是小型反刍动物养殖中的一个普遍问题，在中国主要由扩展莫尼茨绦虫和贝氏莫尼茨绦虫引起。迫切需要高度灵敏的疾病监测和预防方法。在本研究中，我们开发了一种将重组酶聚合酶扩增（RPA）与侧向流动试纸条（RPA-LFD）相结合的可视化检测方法，用于快速检测绵羊粪便样本中的莫尼茨绦虫属。基于扩展莫尼茨绦虫β-微管蛋白基因（MTUB）的特定片段设计了7对引物/探针组。RPA-LFD检测在39℃反应15分钟、引物与探针比例为4:0.6的条件下表现最佳。优化后的方法对莫尼茨绦虫属具有高度特异性，与反刍家畜其他常见胃肠道寄生虫的基因组DNA无交叉反应。检测限为每个反应10拷贝/μL质粒DNA或10 pg/μL扩展莫尼茨绦虫基因组DNA。与使用临床样本和模拟绵羊样本的PCR相比，RPA-LFD检测表现出相当的检测能力，一致性达到95.7%（K值 = 0.939，p < 0.001）。这些结果表明，RPA-LFD方法是一种可靠且便于携带的诊断工具，可用于绵羊群莫尼茨绦虫病的常规筛查、监测和快速确诊，特别是在流行地区和偏远地区。
