Morotomi M, Hoshina S, Green P, Neu H C, LoGerfo P, Watanabe I, Mutai M, Weinstein I B
Department of Surgery, Columbia-Presbyterian Medical Center of Columbia University, New York, New York 10032.
J Clin Microbiol. 1989 Dec;27(12):2652-5. doi: 10.1128/jcm.27.12.2652-2655.1989.
We have developed a novel and practical DNA-RNA hybridization assay for the detection and identification of Campylobacter pylori in the gastric mucosa. This technique utilizes a [32P]ddATP-labeled synthetic oligonucleotide probe complementary to a nucleotide sequence present in C. pylori 16S rRNA. This probe is very sensitive and reacted with all 23 strains of C. pylori tested. It is also highly specific, since there was no cross-reactivity with the heterologous organisms Campylobacter coli, C. fetus subsp. fetus, C. jejuni, and C. laridis or with Escherichia coli. Hybridization of the oligonucleotide probe with C. pylori RNA was completely inhibited by treatment of the membrane filters with RNase but not DNase. Although a gastric mucosa tissue homogenate slightly inhibited the hybridization, as few as 10(4) C. pylori cells could be detected even in the presence of 5 mg of gastric mucosa. Gastric biopsy specimens obtained from patients referred for upper gastrointestinal tract endoscopy were tested for C. pylori infection by direct oligonucleotide hybridization, and the results were compared with those of bacteriological cultures, the urease test, and histological observations. A comparison of the urease test and the oligonucleotide hybridization results showed an excellent correlation between the two methods. The clinical usefulness of this oligonucleotide-RNA hybridization method is discussed.
我们开发了一种新颖实用的DNA-RNA杂交检测方法，用于检测和鉴定胃黏膜中的幽门螺杆菌。该技术利用一种与幽门螺杆菌16S rRNA中存在的核苷酸序列互补的[32P]ddATP标记的合成寡核苷酸探针。该探针非常灵敏，与所有测试的23株幽门螺杆菌均发生反应。它也具有高度特异性，因为与异源生物结肠弯曲菌、胎儿弯曲菌胎儿亚种、空肠弯曲菌和拉氏弯曲菌或大肠杆菌没有交叉反应。用核糖核酸酶处理膜滤器可完全抑制寡核苷酸探针与幽门螺杆菌RNA的杂交，但用脱氧核糖核酸酶处理则不能。尽管胃黏膜组织匀浆会轻微抑制杂交，但即使存在5毫克胃黏膜，也能检测到低至10(4)个幽门螺杆菌细胞。对因上消化道内镜检查而转诊的患者获取的胃活检标本进行直接寡核苷酸杂交检测幽门螺杆菌感染，并将结果与细菌培养、尿素酶试验和组织学观察结果进行比较。尿素酶试验和寡核苷酸杂交结果的比较显示这两种方法之间具有极好的相关性。本文讨论了这种寡核苷酸-RNA杂交方法的临床实用性。
