Establishment of a Multiplex Real-Time TaqMan-MGB Polymerase Chain Reaction (PCR) Method for the Simultaneous Detection of Three Animal Chlamydia Species
Fuping Nie, Qian Gong, Jun Yang, Cunxian Xi, Yu Wang, Guomin Wang, Lei Zhang, Xianliang Li, Danqun Huo, Changjun Hou
College of Biomedical Engineering, Chongqing University, Chongqing, China (mainland)
Med Sci Monit 2019; 25:9369-9376
Chlamydiae are spread globally and cause infectious diseases in both humans and animals. The existing detection methods for this disease have numerous shortcomings, including low sensitivity, time consuming procedures, and high contamination vulnerability.
MATERIAL AND METHODS: To overcome shortcomings for detecting animal chlamydiosis, a multiplex quantitative polymerase chain reaction (PCR) assay was established for simultaneously detecting and differentiating 3 Chlamydia species (C. pecorum, C. abortus, and C. psittaci) by real time PCR based on TaqMan-MGB technology.
RESULTS: The limit of detection was 20.2 copies/µL for Chlamydophila (Cp.) abortus, 30.8 copies/µL for Cp. pecorum, and 16 copies/µL for Cp. psittaci. This method has good repeatability and stability as coefficients of variation range from 0.04% to 1.38%. Furthermore, compared with OIE (World Organization for Animal Health) recommended PCR assay and previously reported animal chlamydia shell PCR, this multiplex PCR assay demonstrated 99% concordance in detecting clinical samples of porcine nasal swabs and vaginal swabs.
CONCLUSIONS: The novel established method in this study was able to detect 3 types of Chlamydia species simultaneously, and had high sensitivity, strong specificity, and good stability. It provided a rapid, reliable, and convenient method for epidemiological and clinical diagnosis of chlamydiosis in animals.
Keywords: Chlamydophila, Diagnosis, Real-Time Polymerase Chain Reaction, Sensitivity and Specificity, Zoonoses