通过对编码23S rRNA的核糖体DNA进行PCR-限制性片段长度多态性分析鉴定猪肠道螺旋体。

Identification of porcine intestinal spirochetes by PCR-restriction fragment length polymorphism analysis of ribosomal DNA encoding 23S rRNA.

作者信息

Barcellos D E, de Uzeda M, Ikuta N, Lunge V R, Fonseca A S, Kader I I, Duhamel G E

机构信息

Instituto de Microbiologia 'Paulo de Góes', Universidade Federal do Rio de Janeiro, R.J., Rio de Janeiro, Brazil.

出版信息

Vet Microbiol. 2000 Jul 31;75(2):189-98. doi: 10.1016/s0378-1135(00)00212-1.

DOI:10.1016/s0378-1135(00)00212-1

PMID:10889409

Abstract

The Brachyspira (formerly Serpulina) species rrl gene encoding 23S ribosomal RNA (rRNA) was used as a target for amplification of a 517bp DNA fragment by polymerase chain reaction (PCR). The primers for PCR amplification had sequences that were conserved among Brachyspira 23S rRNA gene and were designed from nucleotide sequences of Brachyspira hyodysenteriae, Serpulina intermedia, Brachyspira innocens and Brachyspira pilosicoli available from the GenBank database. Digestion of PCR-generated products from reference and field isolates of swine intestinal spirochetes with restriction enzymes Taq I and Alu I revealed five restriction fragment length polymorphism (RFLP) patterns. Each RFLP pattern corresponded to previously established genetic groups including B. hyodysenteriae (I), S. intermedia/B. innocens (II), Brachyspira murdochii (III), B. pilosicoli (IV) and B. alvinipulli (V). The 23S rRNA PCR/RFLP provided a relatively simple genotypic method for identification of porcine pathogenic B. hyodysenteriae and B. pilosicoli.

摘要

将编码23S核糖体RNA（rRNA）的短螺旋体（以前称为蛇形螺旋体）种rrl基因用作通过聚合酶链反应（PCR）扩增517bp DNA片段的靶标。用于PCR扩增的引物具有在短螺旋体23S rRNA基因中保守的序列，并且是根据可从GenBank数据库获得的猪痢疾短螺旋体、中间蛇形螺旋体、无害短螺旋体和毛样短螺旋体的核苷酸序列设计的。用限制性内切酶Taq I和Alu I对来自猪肠道螺旋体参考菌株和现场分离株的PCR产物进行消化，揭示了五种限制性片段长度多态性（RFLP）模式。每种RFLP模式对应于先前建立的遗传群，包括猪痢疾短螺旋体（I）、中间蛇形螺旋体/无害短螺旋体（II）、默多克短螺旋体（III）、毛样短螺旋体（IV）和阿尔文短螺旋体（V）。23S rRNA PCR/RFLP为鉴定猪致病性猪痢疾短螺旋体和毛样短螺旋体提供了一种相对简单的基因型方法。