Haramoto Eiji, Kitajima Masaaki, Katayama Hiroyuki, Asami Mari, Akiba Michihiro, Kunikane Shoichi
Interdisciplinary Graduate School of Medicine and Engineering, University of Yamanashi, 4-3-11 Takeda, Kofu, Yamanashi, Japan.
Water Res. 2009 Aug;43(15):3759-64. doi: 10.1016/j.watres.2009.05.043. Epub 2009 Jun 6.
Genotyping of F-specific RNA phages is currently one of the most promising approaches to differentiate between human and animal fecal contamination in aquatic environments. In this study, a total of 18 river water and sediment samples were collected from the Tonegawa River basin, Japan, in order to describe the genogroup distribution of F-specific RNA and DNA phages using genogroup-specific real-time PCR assays. F-specific phages were detected in nine (100%) river water and six (67%) sediment samples. Eighty-five phage plaques were isolated from these samples and subjected to real-time PCR assays specific for the phages. F-specific RNA phages of human genogroups (II and III) were detected in 32 (38%) plaques, whereas those of animal genogroups (I and IV) were detected in 17 (20%) plaques. No correlation was observed between the genogroup distribution of F-specific RNA phages and the occurrence of human adenovirus genomes, suggesting that genotyping of the phages alone is inadequate for the evaluation of the occurrence of viruses in aquatic environments. SYBR Green-based real-time PCR assay revealed the presence of F-specific DNA phages in four (5%) plaques, which were further classified into two genogroups (fd- and f1-like phages) by sequence analysis. Thirty-two (38%) plaques were not classified as the F-specific phage genogroups, indicating the limited applicability of these real-time PCR assays to a wide range of aquatic environmental samples worldwide.
F特异性RNA噬菌体的基因分型目前是区分水生环境中人类和动物粪便污染的最有前景的方法之一。在本研究中，从日本利根川流域总共采集了18份河水和沉积物样本，以便使用基因群特异性实时PCR检测来描述F特异性RNA和DNA噬菌体的基因群分布。在9份（100%）河水和6份（67%）沉积物样本中检测到了F特异性噬菌体。从这些样本中分离出85个噬菌斑，并对其进行噬菌体特异性实时PCR检测。在32个（38%）噬菌斑中检测到了人类基因群（II和III）的F特异性RNA噬菌体，而在17个（20%）噬菌斑中检测到了动物基因群（I和IV）的F特异性RNA噬菌体。未观察到F特异性RNA噬菌体的基因群分布与人类腺病毒基因组的出现之间存在相关性，这表明仅对噬菌体进行基因分型不足以评估水生环境中病毒的出现情况。基于SYBR Green的实时PCR检测在4个（5%）噬菌斑中检测到了F特异性DNA噬菌体，通过序列分析将其进一步分为两个基因群（fd样和f1样噬菌体）。32个（38%）噬菌斑未被归类为F特异性噬菌体基因群，这表明这些实时PCR检测在全球范围内广泛的水生环境样本中的适用性有限。
