Identification of murine Helicobacters by PCR and restriction enzyme analyses
Identification of murine Helicobacters by PCR and restriction enzyme analyses
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DOI：
10.1128/jcm.34.4.942-946.1996
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发表时间：
1996-04-01
影响因子：
9.4
通讯作者：
BeschWilliford, C
中科院分区：
文献类型：
--
作者：
Riley, LK;Franklin, CL;BeschWilliford, C
最近发现了三种鼠螺杆菌：肝螺杆菌、鼠螺杆菌和比尔螺杆菌。肝螺杆菌和比尔螺杆菌感染与肝炎和肝肿瘤有关。在这项研究中，寡核苷酸引物是根据幽门螺杆菌属成员中保守的 16S rRNA 基因区域设计的。该检测扩增了所有三种啮齿动物螺杆菌的预期 373 bp 产物，并且能够检测到少至 5 pg 的肝螺杆菌。 H. bills 或 H. muridarum DNA。通过测试未感染小鼠和有螺杆菌感染记录的小鼠的盲肠 DNA 以及测试其他细菌属的 DNA 来确定反应的特异性。使用感染螺杆菌的小鼠的盲肠 DNA 生成了预期大小的产物，但未使用未感染小鼠的 DNA 生成了预期大小的产物。除了与螺杆菌属密切相关的“Flexispira rappini”外，其他细菌属的 DNA 均未用螺杆菌属特异性引物扩增。扩增产物的 MboI、MaeI 和 HhaI 限制性内切酶分析能够区分鼠螺杆菌种类，但无法区分 H、bills 和 ''F。拉皮尼。为了区分 H. bilis，设计了基于 H. bilis 16S rRNA 序列的反向引物，使用 H. bilis 特异性反向引物 (Hbr) 和螺杆菌属特异性正向引物 (H276f) 进行 PCR 扩增 H. bills DNA，但不扩增来自 ''F. bills 的 DNA。 Rappini'' 或其他啮齿动物螺杆菌。通过 PCR 检测和限制性内切酶分析的结合对大量小鼠盲肠组织进行的检查表明，肝幽门螺杆菌和嘴幽门螺杆菌感染在实验室小鼠和大鼠群体中广泛存在。
Three murine helicobacter species have recently been identified: Helicobacter hepaticus, Helicobacter muridarum, and Helicobacter bills. Infections with H. hepaticus and H. bills have been associated with hepatitis and hepatic neoplasia. In this study, oligonucleotide primers were designed from regions of the 16S rRNA gene that are conserved among members of the He[icobacter genus. The assay amplified the expected 373-bp product from all three rodent Helicobacter species and was able to detect as little as 5 pg of H. hepaticus. H. bills, or H. muridarum DNA. The specificity of the reaction was determined by testing cecal DNA from uninfected mice and mice with documented Helicobacter infections and by testing DNA from other bacterial genera. A product of the expected size was generated with cecal DNA from Helicobacter-infected mice but not with DNA from uninfected mice, With the exception of that of ''Flexispira rappini,'' which is closely related to the Helicobacter genus, DNA from other bacterial genera was not amplified with the Helicobacter genus-specific primers. MboI, MaeI, and HhaI restriction enzyme analyses of the amplified product were able to differentiate among the murine Helicobacter species but could not differentiate H, bills from ''F. rappini. To distinguish H. bilis, a reverse primer based on H. bilis 16S rRNA sequence was designed, PCR with the H. bilis-specific reverse primer (Hbr) and the Helicobacter genus-specific forward primer (H276f) amplified H. bills DNA but not DNA from ''F. rappini'' or other rodent helicobacters. Examination of a large number of murine cecal tissues with this combination of PCR assays and restriction enzyme analyses indicated that H. hepaticus and H. bills infections are widespread in laboratory mouse and rat colonies.