鸡毒霉形体感染的PCR检测方法的建立及应用

STUDIES ON POLYMERASE CHAIN REACTION (PCR) FOR THE DETECTION OF Mycoplasma gallisepticum (MG) AND ITS CLINICAL APPLICATION

用多聚酶链反应检测了鸡毒霉形体（Ｍｙｃｏｐｌａｓｍａｇａｌｉｓｅｐｔｉｃｕｍ，ＭＧ）种内菌株的特异性ＤＮＡ片段，并用该法对人工感染鸡及野外鸡群进行了ＭＧ感染的分子流行病学调查。结果表明，通过基因扩增及电泳，仅ＭＧ出现特异性扩增条带，最低能检出１８ｐｇＭＧＤＮＡ，说明该方法具有很高的特异性和敏感性。人工感染６０只鸡，３ｄ后即可用ＰＣＲ查出ＭＧ阳性，６ｄ后１００％为阳性，对照鸡全部为阴性。与分离培养的结果完全一致。ＰＣＲ对野外鸡群的ＭＧ检出率为１０．５％～３５％，平均为２０％（１６／７９），与血清学检查结果基本一致；从ＰＣＲ阳性鸡中，有６１．５％（６／１３）能分离到ＭＧ。这说明ＰＣＲ在ＭＧ感染的分子流行病学调查中有重要的应用价值。

By using polymerase chain reaction (PCR), a species specific DNA fragment of MG strains was detected. This technique was also used to take epidemiological studies of MG infection in experimentally diseased birds and wild bird flocks. It was showed that only MG strains were amplified yielding specific products visualized by agarose gel electrophoresis, and that as low as 18 picogram of MG DNA was detected following amplification by PCR. The results showed that this method was very sensitive and specific. 3 days after challenge, PCR results could be positive among 60 experimentally diseased birds. 6 days post inoculation, PCR results were positive wholly, yet the detecting results of controlled group were all negative, which were the same as the results of culture. In wild bird flocks, MG PCR result was from 10.5 to 35 percent (on average, it was 20 percent (16/79)), which corresponded to the serological results. Among PCR positive birds, MG can be cultured by 61.5 percent (6/13). In conclusion, MG PCR should play an important role in epidemiological studies of MG infection.