An Appropriate Cutoff Value for Determining the Colonization of Helicobacter pylori by the Pyrosequencing Method: Comparison with Conventional Methods
An Appropriate Cutoff Value for Determining the Colonization of Helicobacter pylori by the Pyrosequencing Method: Comparison with Conventional Methods
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DOI：
10.1111/hel.12214
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发表时间：
2015-10-01
期刊：
影响因子：
4.4
通讯作者：
Jung, Hyun Chae
中科院分区：
文献类型：
--
作者：
Kim, Jaeyeon;Kim, Nayoung;Jung, Hyun Chae
背景：16S核糖体RNA(RRNA)基因的测序改善了微生物群落的特征。它使得即使在用传统方法发现幽门螺杆菌阴性的受试者中也能检测到低丰度的胃幽门螺杆菌序列。本研究的目的是通过焦磷酸测序法获得幽门螺杆菌在胃粘膜标本中定植的临界值。材料和方法：收集63例胃粘膜活检标本，通过血清学、快速尿素酶试验、培养和组织学相结合的方法确定幽门螺杆菌感染情况。用通用细菌引物进行聚合酶链式反应，从粘膜标本中扩增出微生物DNA。对16S rDNA扩增产物进行焦解测序。用ROC曲线分析焦磷酸测序确定幽门螺杆菌定植的临界值。结果：在63例受试者中，所有(28/28)常规幽门螺杆菌阳性样本和60%(21/35)幽门螺杆菌阴性样本均检出幽门螺杆菌序列。在幽门螺杆菌阴性组中，每个样本中幽门螺杆菌的平均读数百分比为0.67+/-1.09%。根据ROC曲线分析，临床阳性幽门螺杆菌状态的临界值约为1.22%(auc=0.957；p
Background: 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.Materials and Methods: 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.Results: 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