Kim Jaeyeon, Kim Nayoung, Jo Hyun Jin, Park Ji Hyun, Nam Ryoung Hee, Seok Yeong-Jae, Kim Yeon-Ran, Kim Joo Sung, Kim Jung Mogg, Kim Jung Min, Lee Dong Ho, Jung Hyun Chae
Department of Internal Medicine, Seoul National University Bundang Hospital, Seongnam, Gyeonggi-do, South Korea.
Department of Internal Medicine, Liver Research Institute, Seoul National University College of Medicine, Seoul, South Korea.
Helicobacter. 2015 Oct;20(5):370-80. doi: 10.1111/hel.12214. Epub 2015 Feb 9.
Sequencing of 16S ribosomal RNA (rRNA) gene has improved the characterization of microbial communities. It enabled the detection of low abundance gastric Helicobacter pylori sequences even in subjects that were found to be H. pylori negative with conventional methods. The objective of this study was to obtain a cutoff value for H. pylori colonization in gastric mucosa samples by pyrosequencing method.
Gastric mucosal biopsies were taken from 63 subjects whose H. pylori status was determined by a combination of serology, rapid urease test, culture, and histology. Microbial DNA from mucosal samples was amplified by PCR using universal bacterial primers. 16S rDNA amplicons were pyrosequenced. ROC curve analysis was performed to determine the cutoff value for H. pylori colonization by pyrosequencing. In addition, temporal changes in the stomach microbiota were observed in eight initially H. pylori-positive and eight H. pylori-negative subjects at a single time point 1-8 years later.
Of the 63 subjects, the presence of H. pylori sequences was detected in all (28/28) conventionally H. pylori-positive samples and in 60% (21/35) of H. pylori-negative samples. The average percent of H. pylori reads in each sample was 0.67 ± 1.09% in the H. pylori-negative group. Cutoff value for clinically positive H. pylori status was approximately 1.22% based on ROC curve analysis (AUC = 0.957; p < .001). Helicobacter pylori was successfully eradicated in five of seven treated H. pylori-positive subjects (71.4%), and the percentage of H. pylori reads in these five subjects dropped from 1.3-95.18% to 0-0.16% after eradication.
These results suggest that the cutoff value of H. pylori sequence percentage for H. pylori colonization by pyrosequencing could be set at approximately 1%. It might be helpful to analyze gastric microbiota related to H. pylori sequence status.
16S核糖体RNA（rRNA）基因测序改善了微生物群落的特征描述。它能够检测到低丰度的胃幽门螺杆菌序列，即使在那些用传统方法检测为幽门螺杆菌阴性的受试者中。本研究的目的是通过焦磷酸测序法获得胃黏膜样本中幽门螺杆菌定植的临界值。
从63名受试者中采集胃黏膜活检组织，这些受试者的幽门螺杆菌状态通过血清学、快速尿素酶试验、培养和组织学联合确定。使用通用细菌引物通过PCR扩增黏膜样本中的微生物DNA。对16S rDNA扩增子进行焦磷酸测序。进行ROC曲线分析以确定通过焦磷酸测序法检测幽门螺杆菌定植的临界值。此外，在8名初始幽门螺杆菌阳性和8名幽门螺杆菌阴性受试者中，在1 - 8年后的单个时间点观察胃微生物群的时间变化。
在63名受试者中，在所有（28/28）传统方法检测为幽门螺杆菌阳性的样本以及60%（21/35）的幽门螺杆菌阴性样本中检测到幽门螺杆菌序列。幽门螺杆菌阴性组中每个样本中幽门螺杆菌读数的平均百分比为0.67±1.09%。基于ROC曲线分析，临床幽门螺杆菌阳性状态的临界值约为1.22%（AUC = 0.957；p <.001）。7名接受治疗的幽门螺杆菌阳性受试者中有5名（71.4%）成功根除了幽门螺杆菌，这5名受试者中幽门螺杆菌读数的百分比在根除后从1.3 - 95.18%降至0 - 0.16%。
这些结果表明，通过焦磷酸测序法检测幽门螺杆菌定植的幽门螺杆菌序列百分比临界值可设定为约1%。分析与幽门螺杆菌序列状态相关的胃微生物群可能会有所帮助。
