Kimura Shigeko, Uehara Mika, Morimoto Daichi, Yamanaka Momoko, Sako Yoshihiko, Yoshida Takashi
Graduate School of Agriculture, Kyoto University, Kyoto, Japan.
School of Environmental Science, The University of Shiga Prefecture, Hikone, Japan.
Front Microbiol. 2018 Mar 8;9:425. doi: 10.3389/fmicb.2018.00425. eCollection 2018.
The freshwater cyanobacterium frequently forms toxic massive blooms and exists in an arms race with its infectious phages in aquatic natural environments, and as a result, has evolved extremely diverse and elaborate antiviral defense systems, including the clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated genes (Cas) system. Here, to assess population dynamics associated with exogenous mobile genetic elements such as phages and plasmids, we examined the temporal variation in CRISPR genotypes (CTs) by analyzing spacer sequences detected in a natural pond between June and October 2013 when a cyanobacterial bloom occurred. A total of 463,954 high-quality leader-end CRISPR sequences were obtained and the sequences containing spacers were classified into 31 previously reported CTs and 68 new CTs based on the shared order of the leader-end spacers. CT19 was the most dominant genotype (32%) among the 16 most common CTs, followed by CT52 (14%) and CT58 (9%). Spacer repertoires of CT19 showed mainly two different types; CT19, which was identical to the CT19 spacer repertoire of previously isolated strains, and CT19, which contained a new spacer at the leader-end of the CRISPR region of CT19, which were present in almost equal abundance, accounting for up to 99.94% of CT19 sequences. Surprisingly, we observed the spacer repertoires of the second to tenth spacers of CT19 at the most leader-end of proto-genotype sequences of CT19. These were observed during the sampling in this study and our previous study at the same ecosystem in 2010 and 2011, suggesting these CTs persisted from 2011 to 2013 in spite of phage pressure. The leader-end variants were observed in other CT genotypes. These findings indicated an incomplete selective sweep of populations. We explained the phenomenon as follow; the abundance of varied seasonally and drastically, resulting that populations experience a bottleneck once a year, and thereby founder effects following a bottleneck mean that older CTs have an equal chance of increasing in prevalence as the CTs generated following acquisition of newer spacers.
淡水蓝藻经常形成有毒的大量水华，并且在水生自然环境中与其感染性噬菌体处于军备竞赛状态，因此，它进化出了极其多样和复杂的抗病毒防御系统，包括成簇规律间隔短回文重复序列（CRISPR）-CRISPR相关基因（Cas）系统。在此，为了评估与噬菌体和质粒等外源移动遗传元件相关的种群动态，我们通过分析2013年6月至10月蓝藻水华发生期间在一个天然池塘中检测到的间隔序列，研究了CRISPR基因型（CTs）的时间变化。共获得463,954条高质量的前导端CRISPR序列，根据前导端间隔序列的共享顺序，将包含间隔序列的序列分为31个先前报道的CTs和68个新的CTs。CT19是16种最常见的CTs中最主要的基因型（32%），其次是CT52（14%）和CT58（9%）。CT19的间隔序列库主要表现为两种不同类型；与先前分离菌株的CT19间隔序列库相同的CT19，以及在CT19的CRISPR区域前导端含有一个新间隔序列的CT19，它们的丰度几乎相等，占CT19序列的99.94%。令人惊讶的是，我们在CT19原基因型序列的最前导端观察到了CT19第二至第十个间隔序列的间隔序列库。这些在本研究的采样过程中以及我们之前在2010年和2011年对同一生态系统的研究中都被观察到，这表明尽管存在噬菌体压力，这些CTs在2011年至2013年期间仍然持续存在。在前导端变体在其他CT基因型中也被观察到。这些发现表明种群的选择性清除不完全。我们对这一现象的解释如下；蓝藻的丰度随季节剧烈变化，导致蓝藻种群每年经历一次瓶颈，因此瓶颈后的奠基者效应意味着较老的CTs与获得新间隔序列后产生的CTs有同等的机会在流行率上增加。
