Wu Z, Nagano I, Pozio E, Takahashi Y
Department of Parasitology, Gifu University School of Medicine, Japan.
Parasitology. 1999 Feb;118 ( Pt 2):211-8. doi: 10.1017/s0031182098003679.
In the present study, polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis was developed to identify 5 species (Trichinella spiralis, Trichinella britovi, Trichinella nativa, Trichinella nelsoni and Trichinella pseudospiralis) and 3 phenotypes of uncertain taxonomic status (Trichinella T5, T6, and T8). Eleven restriction endonucleases were used to restrict 3 DNA fragments (1) a 2800 bp fragment of the 43 kDa excretory-secretory (E-S) protein gene, (2) a 1250 bp fragment amplified with the primer pair SB147A and (3) a 372 bp fragment amplified with the primer pair SB372A. This RFLP method allows the identification of the 8 Trichinella phenotypes as follows: T. spiralis by the HinfI or DdeI endonuclease restriction of the 2800 bp fragment; T. nativa by the RsaI restriction of the 2800 bp fragment, or by the AluI restriction of the 1250 bp fragment; T. britovi and Trichinella T8 by the AluI restriction of the 1250 bp fragments, and can be discriminated between them by the SspI restriction of the 2800 bp fragment; T. pseudospiralis by the MspI restriction of the 372 bp fragment; T. nelsoni by the HhaI or AluI restriction of the 2800 bp fragment; Trichinella T5 by the HhaI restriction of the 2800 bp fragment; Trichinella T6 by the AluI restriction of the 1250 bp fragment; and Trichinella T8 by the SspI or RsaI restriction of the 2800 bp fragment. This study reveals also an intraspecifies polymorphism in the 2800 bp and 1250 bp fragments for T. britovi, Trichinella T5 and T6.
在本研究中，开发了聚合酶链反应-限制性片段长度多态性（PCR-RFLP）分析方法，以鉴定5种旋毛虫（旋毛形线虫、布氏旋毛虫、本地旋毛虫、纳氏旋毛虫和伪旋毛虫）以及3种分类地位不确定的表型（旋毛虫T5、T6和T8）。使用11种限制性内切酶切割3个DNA片段：（1）43 kDa排泄-分泌（E-S）蛋白基因的2800 bp片段；（2）用引物对SB147A扩增的1250 bp片段；（3）用引物对SB372A扩增的372 bp片段。这种RFLP方法可鉴定出8种旋毛虫表型，具体如下：通过HinfI或DdeI内切酶切割2800 bp片段鉴定旋毛形线虫；通过RsaI切割2800 bp片段或通过AluI切割1250 bp片段鉴定本地旋毛虫；通过AluI切割1250 bp片段鉴定布氏旋毛虫和旋毛虫T8，且可通过SspI切割2800 bp片段区分它们；通过MspI切割372 bp片段鉴定伪旋毛虫；通过HhaI或AluI切割2800 bp片段鉴定纳氏旋毛虫；通过HhaI切割2800 bp片段鉴定旋毛虫T5；通过AluI切割1250 bp片段鉴定旋毛虫T6；通过SspI或RsaI切割2800 bp片段鉴定旋毛虫T8。本研究还揭示了布氏旋毛虫、旋毛虫T5和T6在2800 bp和1250 bp片段中存在种内多态性。
