Institute for Biotechnology Research, Jomo Kenyatta University of Agriculture and Technology, Nairobi, Kenya.
Taita Taveta University, Taita, Kenya.
PLoS One. 2018 Mar 22;13(3):e0194554. doi: 10.1371/journal.pone.0194554. eCollection 2018.
Human pathogens can survive and grow in hot springs. For water quality assessment, Escherichia coli or Enterococci are the main thermotolerant enteric bacteria commonly used to estimate the load of pathogenic bacteria in water. However, most of the environmental bacteria are unculturable thus culture methods may cause bias in detection of most pathogens. Illumina sequencing can provide a more comprehensive and accurate insight into environmental bacterial pathogens, which can be used to develop better risk assessment methods and promote public health awareness. In this study, high-throughput Illumina sequencing was used to identify bacterial pathogens from five hot springs; Maiwooi, Akwar, Garbanabra, Elegedi and Gelti, in Eritrea. Water samples were collected from the five hot springs. Total community DNA was extracted from samples using the phenol-chloroform method. The 16S rRNA gene variable region (V4-V7) of the extracted DNA was amplified and library construction done according to Illumina sequencing protocol. The sequence reads (length >200 bp) from Illumina sequencing libraries ranged from 22,091 sequences in the wet sediment sample from Garbanabra to 155,789 sequences in the mat sample from Elegedi. Taxonomy was assigned to each OTU using BLASTn against a curated database derived from GreenGenes, RDPII, SILVA SSU Reference 119 and NCBI. The proportion of potential pathogens from the water samples was highest in Maiwooi (17.8%), followed by Gelti (16.7%), Akwar (13.6%) and Garbanabra (10.9%). Although the numbers of DNA sequence reads from Illumina sequencing were very high for the Elegedi (104,328), corresponding proportion of potential pathogens very low (3.6%). Most of the potential pathogenic bacterial sequences identified were from Proteobacteria and Firmicutes. Legionella and Clostridium were the most common detected genera with different species. Most of the potential pathogens were detected from the water samples. However, sequences belonging to Clostridium were observed more abundantly from the mat samples. This study employed high-throughput sequencing technologies to determine the presence of pathogenic bacteria in the five hot springs in Eritrea.
人类病原体可以在温泉中存活和生长。为了进行水质评估，大肠杆菌或肠球菌是常用的耐热肠杆菌，用于估计水中致病菌的负荷。然而，大多数环境细菌是不可培养的，因此培养方法可能会导致对大多数病原体的检测存在偏差。Illumina 测序可以更全面、准确地了解环境细菌病原体，从而可以开发更好的风险评估方法并提高公众健康意识。在这项研究中，使用高通量 Illumina 测序从厄立特里亚的五个温泉（Maiwooi、Akwar、Garbanabra、Elegedi 和 Gelti）中鉴定出细菌病原体。从五个温泉采集水样。使用苯酚-氯仿法从样品中提取总社区 DNA。根据 Illumina 测序方案扩增提取 DNA 的 16S rRNA 基因可变区（V4-V7）并构建文库。Illumina 测序文库的序列读长（长度>200bp）范围从 Garbanabra 湿地沉积物样本中的 22091 个序列到 Elegedi 垫样本中的 155789 个序列。使用 BLASTn 将每个 OTU 与源自 GreenGenes、RDPII、SILVA SSU Reference 119 和 NCBI 的经过精心整理的数据库进行比对，以确定分类。水样中潜在病原体的比例 Maiwooi 最高（17.8%），其次是 Gelti（16.7%）、Akwar（13.6%）和 Garbanabra（10.9%）。尽管 Elegedi 的 Illumina 测序 DNA 序列读取数非常高（104328），但相应的潜在病原体比例非常低（3.6%）。鉴定出的潜在致病细菌序列主要来自变形菌门和厚壁菌门。军团菌属和梭菌属是最常见的检测属，具有不同的物种。大多数潜在的病原体是从水样中检测到的。然而，从垫样本中观察到更多属于梭菌属的序列。本研究采用高通量测序技术确定了厄立特里亚五个温泉中致病细菌的存在。
