Peek R M, Miller G G, Tham K T, Pérez-Pérez G I, Cover T L, Atherton J C, Dunn G D, Blaser M J
Department of Medicine, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-2279.
J Clin Microbiol. 1995 Jan;33(1):28-32. doi: 10.1128/jcm.33.1.28-32.1995.
Mucosal and systemic immunologic recognition of cagA by Helicobacter pylori-infected individuals is associated with peptic ulcer disease; however, in the laboratory, expression of cagA is subject to artificial conditions which may not accurately reflect the conditions in host tissues. Gastric antral and body biopsy specimens and serum for anti-H. pylori immunoglobulin G serology were obtained from 42 patients. Biopsy specimens were studied by histology, culture, and reverse transcription PCR (RT-PCR). Oligonucleotide primers specific for H. pylori (16S rRNA, ureA, and cagA) were used to detect bacterial mRNA in gastric biopsy specimens. PCR was performed on DNA from corresponding H. pylori isolates to detect genomic 16S rRNA, ureA, and cagA. Of the 42 patients from whom clinical specimens were obtained, 25 were infected with H. pylori on the basis of both serology and histology or culture (i.e., tissue positive); 13 were negative by serology, histology, and culture; and 4 were positive by serology only. RT-PCR with 16S rRNA primers detected 24 of 25 tissue-positive and 0 of 17 tissue-negative patients (P < 0.001). RT-PCR with ureA primers detected 16 of 25 tissue-positive and 0 of 17 tissue-negative patients (P < 0.001). CagA mRNA was detected by RT-PCR in 14 of 25 gastric biopsy specimens in the tissue-positive group and in 0 of 17 gastric biopsy specimens in the tissue-negative group. PCR of genomic DNA for the presence of the cagA gene in the corresponding bacterial isolates correlated absolutely with cagA gene expression in gastric tissue. These results indicate that RT-PCR is a sensitive and specific method for the detection of the presence of H. pylori and the expression of H. pylori genes in human gastric tissue. Detection of H. pylori gene expression in vivo by this approach may contribute to improving the diagnosis and understanding the pathogenesis of H. pylori infections.
幽门螺杆菌感染个体对细胞毒素相关基因A（CagA）的黏膜和全身免疫识别与消化性溃疡疾病相关；然而，在实验室中，CagA的表达受到人工条件的影响，这些条件可能无法准确反映宿主组织中的情况。从42例患者中获取胃窦和胃体活检标本以及用于抗幽门螺杆菌免疫球蛋白G血清学检测的血清。活检标本通过组织学、培养和逆转录PCR（RT-PCR）进行研究。使用针对幽门螺杆菌的寡核苷酸引物（16S rRNA、脲酶A和CagA）检测胃活检标本中的细菌mRNA。对相应幽门螺杆菌分离株的DNA进行PCR，以检测基因组16S rRNA、脲酶A和CagA。在获取临床标本的42例患者中，25例基于血清学和组织学或培养结果被诊断为幽门螺杆菌感染（即组织阳性）；13例血清学、组织学和培养结果均为阴性；4例仅血清学呈阳性。用16S rRNA引物进行的RT-PCR在25例组织阳性患者中检测到24例，在17例组织阴性患者中检测到0例（P<0.001）。用脲酶A引物进行的RT-PCR在25例组织阳性患者中检测到16例，在17例组织阴性患者中检测到0例（P<0.001）。在组织阳性组的25例胃活检标本中，有14例通过RT-PCR检测到CagA mRNA，而在组织阴性组的17例胃活检标本中未检测到。相应细菌分离株基因组DNA中cagA基因的PCR检测结果与胃组织中cagA基因的表达完全相关。这些结果表明，RT-PCR是检测人胃组织中幽门螺杆菌存在及幽门螺杆菌基因表达的一种灵敏且特异的方法。通过这种方法在体内检测幽门螺杆菌基因表达可能有助于改善幽门螺杆菌感染的诊断并增进对其发病机制的理解。
