Anzai T, Eguchi M, Sekizaki T, Kamada M, Yamamoto K, Okuda T
Epizootic Research Station, Equine Research Institute, Japan Racing Association, Tochigi, Japan.
J Vet Med Sci. 1999 Dec;61(12):1287-92. doi: 10.1292/jvms.61.1287.
In order to establish a rapid diagnostic method for contagious equine metritis (CEM), we developed and evaluated a polymerase chain reaction (PCR) test. Species-specific PCR primer sets were derived from the DNA sequence of a cloned DNA fragment of Taylorella equigenitalis that did not hybridize with the genome of a taxomonically related species, Oligella urethralis. Single step PCR with primer set P1-N2 and two-step semi-nested PCR with primer sets P1-N2 and P2-N2 detected as low as 100 and 10 CFU of the bacteria, respectively. Single-step PCR detected T. equigenitalis from genital swabs of experimentally infected mares with sensitivity comparable to that of bacterial isolation. Furthermore, two-step PCR was more sensitive than the culture method. Upon examination of field samples, 12 out of 3,123 samples were positive by single-step PCR while only 2 were positive by bacterial culture. The 12 PCR-positive samples originated from 5 mares, of which 3 animals were considered to be carriers based on previous bacteriologic and serologic diagnoses for CEM. The PCR test described in this study would provide a specific and highly sensitive tool for the rapid diagnosis of CEM.
为建立马传染性子宫炎（CEM）的快速诊断方法，我们开发并评估了一种聚合酶链反应（PCR）检测方法。种属特异性PCR引物对源自马泰勒菌克隆DNA片段的DNA序列，该片段不与分类学上相关物种尿道寡源杆菌的基因组杂交。引物对P1-N2的一步法PCR以及引物对P1-N2和P2-N2的两步半巢式PCR分别能检测到低至100和10 CFU的细菌。一步法PCR从实验感染母马的生殖道拭子中检测出马泰勒菌，其灵敏度与细菌分离法相当。此外，两步法PCR比培养法更灵敏。在检测现场样本时，3123份样本中有12份通过一步法PCR呈阳性，而细菌培养仅2份呈阳性。这12份PCR阳性样本来自5匹母马，其中3匹根据先前CEM的细菌学和血清学诊断被认为是携带者。本研究中描述的PCR检测方法将为CEM的快速诊断提供一种特异且高度灵敏的工具。
