Davison Michelle, Treangen Todd J, Koren Sergey, Pop Mihai, Bhaya Devaki
Carnegie Institution for Science, Department of Plant Biology, Stanford, CA, 94305, United States of America.
Stanford University, Department of Biology, Stanford, CA, 94305, United States of America.
PLoS One. 2016 Sep 9;11(9):e0160574. doi: 10.1371/journal.pone.0160574. eCollection 2016.
The polymicrobial biofilm communities in Mushroom and Octopus Spring in Yellowstone National Park (YNP) are well characterized, yet little is known about the phage populations. Dominant species, Synechococcus sp. JA-2-3B'a(2-13), Synechococcus sp. JA-3-3Ab, Chloroflexus sp. Y-400-fl, and Roseiflexus sp. RS-1, contain multiple CRISPR-Cas arrays, suggesting complex interactions with phage predators. To analyze phage populations from Octopus Spring biofilms, we sequenced a viral enriched fraction. To assemble and analyze phage metagenomic data, we developed a custom module, VIRITAS, implemented within the MetAMOS framework. This module bins contigs into groups based on tetranucleotide frequencies and CRISPR spacer-protospacer matching and ORF calling. Using this pipeline we were able to assemble phage sequences into contigs and bin them into three clusters that corroborated with their potential host range. The virome contained 52,348 predicted ORFs; some were clearly phage-like; 9319 ORFs had a recognizable Pfam domain while the rest were hypothetical. Of the recognized domains with CRISPR spacer matches, was the phage endolysin used by lytic phage to disrupt cells. Analysis of the endolysins present in the thermophilic cyanophage contigs revealed a subset of characterized endolysins as well as a Glyco_hydro_108 (PF05838) domain not previously associated with sequenced cyanophages. A search for CRISPR spacer matches to all identified phage endolysins demonstrated that a majority of endolysin domains were targets. This strategy provides a general way to link host and phage as endolysins are known to be widely distributed in bacteriophage. Endolysins can also provide information about host cell wall composition and have the additional potential to be used as targets for novel therapeutics.
黄石国家公园（YNP）蘑菇泉和章鱼泉中的多微生物生物膜群落已得到充分表征，但对噬菌体种群却知之甚少。优势物种聚球藻属（Synechococcus）JA - 2 - 3B'a(2 - 13)、聚球藻属JA - 3 - 3Ab、绿弯菌属（Chloroflexus）Y - 400 - fl和玫瑰弯菌属（Roseiflexus）RS - 1含有多个CRISPR - Cas阵列，这表明它们与噬菌体捕食者之间存在复杂的相互作用。为了分析章鱼泉生物膜中的噬菌体种群，我们对一个病毒富集部分进行了测序。为了组装和分析噬菌体宏基因组数据，我们在MetAMOS框架内开发了一个定制模块VIRITAS。该模块根据四核苷酸频率、CRISPR间隔序列 - 原间隔序列匹配和开放阅读框（ORF）识别将重叠群分组。使用这个流程，我们能够将噬菌体序列组装成重叠群，并将它们分成三个簇，这与它们潜在的宿主范围相符。病毒群落包含52348个预测的开放阅读框；其中一些明显类似噬菌体；9319个开放阅读框具有可识别的Pfam结构域，其余为假设的。在与CRISPR间隔序列匹配的已识别结构域中，有裂解性噬菌体用于破坏细胞的噬菌体溶菌酶。对嗜热蓝藻噬菌体重叠群中存在的溶菌酶进行分析，发现了一部分已表征的溶菌酶以及一个以前未与已测序蓝藻噬菌体相关联的糖基水解酶108（PF05838）结构域。对所有已鉴定的噬菌体溶菌酶进行CRISPR间隔序列匹配搜索表明，大多数溶菌酶结构域都是靶点。由于已知溶菌酶广泛分布于噬菌体中，这种策略提供了一种将宿主和噬菌体联系起来的通用方法。溶菌酶还可以提供有关宿主细胞壁组成的信息，并且还有额外的潜力用作新型治疗药物的靶点。
