Hjalmarsson Sandra, Alderborn Anders, Fock Caroline, Muldin Ingrid, Kling Helene, Uhlén Mathias, Engstrand Lars
Department of Medical Sciences, Clinical Bacteriology, Uppsala University, Uppsala, Sweden.
Helicobacter. 2004 Apr;9(2):138-45. doi: 10.1111/j.1083-4389.2004.00210.x.
Genetic information is becoming increasingly important in diagnosis and prognosis of infectious diseases. In this study we investigated the possibility of using a single technology, the Pyrosequencing trade mark technology (Biotage AB, Uppsala, Sweden), to gather several kinds of important genetic information from the human pathogen Helicobacter pylori, as well as from the carrier of the H. pylori infection.
DNA from 87 clinical isolates of H. pylori, 50 isolates from H. pylori-infected transgenic mice and nine gastric biopsies from H. pylori-infected patients was analyzed for targets in the 16S rRNA, 23S rRNA and cytotoxin associated gene A (cagA) genes to determine species identity, clarithromycin susceptibility and virulence level, respectively. In addition, three single nucleotide polymorphisms in the human interleukin-1B (IL-1B) gene, reported to affect the risk of developing gastric cancer, were analyzed in the gastric biopsy samples.
All DNA targets were processed and analyzed in parallel, enabling convenient genetic characterization of both pathogen and host. All genotypes were easily and accurately assigned. In the 16S rRNA analysis, 99.83% of the bases were correctly called.
We conclude that genetic analysis using Pyrosequencing trade mark technology was nonlaborious, and gave highly accurate data for different kinds of target. We therefore believe that this technology has the potential to complement or in the future substitute the time-consuming traditional microbial identification and typing methods, as well as enabling rapid typing of relevant host genetic markers.
遗传信息在传染病的诊断和预后中变得越来越重要。在本研究中，我们调查了使用单一技术——焦磷酸测序技术（瑞典乌普萨拉的Biotage AB公司），从人类病原体幽门螺杆菌以及幽门螺杆菌感染携带者中获取多种重要遗传信息的可能性。
分析了87株幽门螺杆菌临床分离株、50株来自幽门螺杆菌感染转基因小鼠的分离株以及9例幽门螺杆菌感染患者的胃活检组织中的DNA，分别检测16S rRNA、23S rRNA和细胞毒素相关基因A（cagA）基因中的靶点，以确定物种身份、克拉霉素敏感性和毒力水平。此外，还对胃活检样本中人类白细胞介素-1B（IL-1B）基因中报道的影响患胃癌风险的三个单核苷酸多态性进行了分析。
所有DNA靶点均并行处理和分析，从而能够方便地对病原体和宿主进行遗传特征分析。所有基因型都能轻松且准确地确定。在16S rRNA分析中，99.83%的碱基被正确识别。
我们得出结论，使用焦磷酸测序技术进行遗传分析并不费力，并且能为不同类型的靶点提供高度准确的数据。因此，我们认为该技术有潜力补充或在未来替代耗时的传统微生物鉴定和分型方法，同时还能实现相关宿主遗传标记的快速分型。
