Viral metagenomics analysis of planktonic viruses in East Lake, Wuhan, China

East Lake (Lake Donghu), located in Wuhan, China, is a typical city freshwater lake that has been experiencing eutrophic conditions and algal blooming during recent years. Marine and fresh water are considered to contain a large number of viruses. However, little is known about their genetic diversity because of the limited techniques for culturing viruses. In this study, we conducted a viral metagenomic analysis using a high-throughput sequencing technique with samples collected from East Lake in Spring, Summer, Autumn, and Winter. The libraries from four samples each generated 234,669, 71,837, 12,820, and 34,236 contigs (> 90 bp each), respectively. The genetic structure of the viral community revealed a high genetic diversity covering 23 viral families, with the majority of contigs homologous to DNA viruses, including members of Myoviridae, Podoviridae, Siphoviridae, Phycodnaviridae, and Microviridae, which infect bacteria or algae, and members of Circoviridae, which infect invertebrates and vertebrates. The highest viral genetic diversity occurred in samples collected in August, then December and June, and the least diversity in March. Most contigs have low-sequence identities with known viruses. PCR detection targeting the conserved sequences of genes (g20, psbA, psbD, and DNApol) of cyanophages further confirmed that there are novel cyanophages in the East Lake. Our viral metagenomic data provide the first preliminary understanding of the virome in one freshwater lake in China and would be helpful for novel virus discovery and the control of algal blooming in the future.     

Previously unknown and highly divergent ssDNA viruses populate the oceans

Single-stranded DNA (ssDNA) viruses are economically important pathogens of plants and animals, and are widespread in oceans; yet, the diversity and evolutionary relationships among marine ssDNA viruses remain largely unknown. Here we present the results from a metagenomic study of composite samples from temperate (Saanich Inlet, 11 samples; Strait of Georgia, 85 samples) and subtropical (46 samples, Gulf of Mexico) seawater. Most sequences (84%) had no evident similarity to sequenced viruses. In total, 608 putative complete genomes of ssDNA viruses were assembled, almost doubling the number of ssDNA viral genomes in databases. These comprised 129 genetically distinct groups, each represented by at least one complete genome that had no recognizable similarity to each other or to other virus sequences. Given that the seven recognized families of ssDNA viruses have considerable sequence homology within them, this suggests that many of these genetic groups may represent new viral families. Moreover, nearly 70% of the sequences were similar to one of these genomes, indicating that most of the sequences could be assigned to a genetically distinct group. Most sequences fell within 11 well-defined gene groups, each sharing a common gene. Some of these encoded putative replication and coat proteins that had similarity to sequences from viruses infecting eukaryotes, suggesting that these were likely from viruses infecting eukaryotic phytoplankton and zooplankton.     

Comparison of Viromes in Ticks from Different Domestic Animals in China

Ticks are involved in the transmission of various arboviruses and some tick-borne viruses pose significant threats to the health of humans or livestock. This study aimed to investigate the geographical distribution of tick species and tickassociated viruses in central and eastern China. Total 573 ticks from domestic animals including dogs, sheep and cattle were collected in 2017. Two genera of ticks were identified including Rhipicephalus and Haemaphysalis. Sequencing was performed on Miseq Illumina platform to characterize the tick viromes from the four different sampling locations.Following trimming, 13,640 reads were obtained and annotated to 19 virus families. From these sequences, above 37.74% of the viral reads were related to the RNA viruses. Virome comparison study revealed that the tick viral diversity was considerably different in the two identified tick genera. The viral diversity of R. microplus was significantly different from that of other Rhipicephalus species. On the other hand, substantial overlap in viral species was observed between the same genera. In addition, we found no evidence that the natural host played a major role in shaping virus diversity based on the comparison of their viromes. Rather, the geographic location seems to significantly influence the viral families. Phylogenetic study indicated that the novel negative-sense RNA viruses identified in this study was closely related to Bole tick virus 1 and 3 viruses. In conclusion, the present study provides a baseline for comparing viruses detected in ticks, according to species, natural hosts, and geographic locations.     

The influence of a dissolved inorganic nitrogen gradient on phytoplankton community dynamics in a chain of lakes

In a chain of lakes along which nutrient availability varies in a gradient, we performed factorial nutrient enrichment experiments to determine if nitrogen limitation was the principal factor controlling the differences in phytoplankton biomass, photosynthetic productivity, diversity, and species composition among two of the lakes in the chain. In the least productive lake, East Graham Lake, P and C enrichments (in the absence of N enrichment) had no effect on biomass and diversity, whereas within two weeks the N enrichments (alone or in any combination with P and/or C) increased the biomass and decreased the diversity of East Graham Lake phytoplankton to levels similar or identical to those in more productive Shoe Lake. Short-term ¹⁴C photosynthetic rates in East Graham Lake water also responded only to N in the third week. However, photosynthesis was stimulated by P in the first week, and a few species did increase in numbers with P enrichment, suggesting that some degree of P limitation remains in addition to the strong N limitation in East Graham Lake. A number of species responded individually to the enrichments in a manner similar to that of the overall community, and a strong overlapping of discriminant analysis scores for N-enriched East Graham Lake with those of Shoe Lake was consistent with our prediction that the community structure of N-enriched East Graham Lake water would shift toward that of Shoe Lake. However, many species did not respond consistently with these results, and the nutrients tested were clearly not a major factor in the differences in abundance of those species among the two lakes. The results support the argument that overall biomass production and diversity of the phytoplankton community in a lake can be a relatively simple function of a single most-limiting nutrient. However, many of the species responses also confirm that, while nutrient availability is an important factor in the control of the species composition of the community, other factors are likely to prevent reliable predictions of all species effects on the basis of nutrient availability alone.     

中国部分地区蝙蝠携带病毒的宏基因组学分析

蝙蝠携带有60多种病毒,其中许多对人有高度致病性.为了解中国蝙蝠携带病毒的自然本底、蝙蝠病毒的多样性和挖掘潜在的病毒病原,通过基于Solexa高通量测序的病毒宏基因组学技术对从吉林、云南、湖南采集的蝙蝠组织进行病毒组学研究,获得了11 644 232条读长(Reads),并拼接出44 872条重叠序列(Contig).通过核酸序列注释发现,其中8.2％(4 002/44 872)的重叠序列与病毒相关,能进一步注释到36个病毒科,包括19种脊椎动物病毒、6种植物病毒、4种昆虫病毒和4种噬菌体.通过对重叠序列的遗传进化分析、多序列比对显示,被注释为细小病毒、腺联病毒、博卡病毒、腺病毒、小双节RNA病毒等的重叠序列与已知病毒相似,部分序列却又呈现出明显的序列差异.通过对腺病毒和博卡病毒进一步的PCR扩增证实了此研究方法可靠.旨在了解我国蝙蝠携带病毒组的构成,对建立高效的野生动物源人兽共患病的监测方法提供参考.     

Plant community dynamics in a chain of lakes: principal factors in the decline of rooted macrophytes with eutrophication

Shoe Lake and East Graham Lake, part of a small chain of lakes in southeastern Michigan, USA, differ in nutrient loading and in the structure and productivity of their aquatic plant communities. A comparative study of species frequency and biomass distributions, nutrient contents, and responses to experimental nutrient enrichment and shading, was conducted to determine the principal factors controlling the macrophyte dynamics. A central objective was to address the question of why rooted macrophyte growth declines with eutrophication, and to test existing models designed to explain this phenomenon. In the more eutrophic Shoe Lake, diversity and productivity of rooted macrophytes were relatively low, restricted primarily by combined shading of phytoplankton, periphyton, and non-rooted macrophytes (principally Ceratophyllum demersum, along with Utricularia vulgaris and Cladophora fracta). In the less eutrophic East Graham Lake, lower nitrogen availability restricted the growth of all of these shading components, resulting in clearer water and higher productivity and diversity of rooted macrophytes. The macrophytes did not allelopathically suppress the phytoplankton in East Graham Lake. The results supported a direct relationship between nutrient loading, increasing growth of phytoplankton, periphyton and non-rooted macrophytes, and decline of rooted macrophytes.     

淡水富营养型湖泊沉积物亚硝酸还原酶基因(nirS)的多样性和系统发育

【目的】以亚硝酸盐还原酶基因(nirS)为分子标记,探讨富营养化湖泊武汉东湖沉积物中NirS类反硝化细菌群落的多样性及系统发育,并分析环境因子对群落分布的影响。【方法】在武汉东湖4个典型子湖郭郑湖、汤菱湖、团湖和庙湖采集沉积物样品,测定环境参数;提取沉积物中微生物群落基因组DNA,分别构建4个子湖的反硝化微生物的nirS基因文库,利用限制性片段长度的多态性分析(Restriction Fragment LengthPolymorphism,RFLP)技术初步分群,确定各群的代表菌株并测定其nirS基因序列;利用DOTUR软件计算各群落多样性和丰富度指数,以Neighbor-Joining法构建供试菌与参比菌的系统发育树。【结果】环境参数测定结果表明东湖4个子湖中庙湖沉积物总氮(TN)和氨态氮(NH 4+-N)含量最高,团湖最低,郭郑湖沉积物中NO 3-浓度最高。基于NirS序列的生物多样性和丰富度分析表明团湖生物多样性和丰富度指数最高而庙湖各项指数均较低。各子湖供试序列及其代表序列综合RFLP聚类分析表明,武汉东湖沉积物中NirS类反硝化微生物种群具有丰富的多样性。NJ系统发育分析表明东湖沉积物NirS类反硝化菌群可分成3个较大群体(群I-III)。群I占总群体的67.7%,广泛分布于不同的生态环境;来自郭郑湖代表菌的81%分布于群I,而庙湖的代表菌中65%分布于群II。比较分析发现来自于东湖和人工湿地两种生境的NirS群落间具有较高的相似性。【结论】武汉东湖淡水富营养型湖泊沉积物中亚硝酸还原酶基因(nirS)具有丰富的多样性。东湖沉积物中TN、NH 4+和NO 3-的浓度可能是影响NirS类反硝化微生物多样性和空间分布的重要因素之一。     

Complex seasonality observed amongst diverse phytoplankton viruses in the Bay of Quinte,an embayment of Lake Ontario

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Hybridization Analysis of Chesapeake Bay Virioplankton

It has been hypothesized that, by specifically lysing numerically dominant host strains, the virioplankton may play a role in maintaining clonal diversity of heterotrophic bacteria and phytoplankton populations. If viruses selectively lyse only those host species that are numerically dominant, then the number of a specific virus within the virioplankton would be expected to change dramatically over time and space, in coordination with changes in abundance of the host. In this study, the abundances of specific viruses in Chesapeake Bay water samples were monitored, using nucleic acid probes and hybridization analysis. Total virioplankton in a water sample was separated by pulsed-field gel electrophoresis and hybridized with nucleic acid probes specific to either single viral strains or a group of viruses with similar genome sizes. The abundances of specific viruses were inferred from the intensity of the hybridization signal. By using this technique, a virus comprising 1/1,000 of the total virioplankton abundance (ca. 10⁴ PFU/ml) could be detected. Titers of either a single virus species or a group of viruses changed over time, increasing to peak abundance and then declining to low or undetectable levels, and were geographically localized in the bay. Peak signal intensities, i.e., peak abundances of virus strains, were 10-fold greater than the low background level. Furthermore, virus species were found to be restricted to a particular depth, since probes specific to viruses from bottom water did not hybridize with virus genomes from surface water at the same geographical location. Overall, changes in abundances of specific viruses within the virioplankton were episodic, supporting the hypothesis that viral infection influences, if not controls, clonal diversity within heterotrophic bacteria and phytoplankton communities.     

Metagenomic analysis of viral community in the Yangtze River expands known eukaryotic and prokaryotic virus diversity in freshwater

Viruses in aquatic ecosystems are characterized by extraordinary abundance and diversity. Thus far, there have been limited studies focused on viral communities in river water systems. Here, we investigated the virome of the Yangtze River Delta using viral metagenomic analysis. The compositions of viral communities from six sampling sites were analyzed and compared. By using library construction and next generation sequencing, contigs and singlet reads similar to viral sequences were classified into 17 viral families, including nine dsDNA viral families, four ssDNA viral families and four RNA viral families. Statistical analysis using Friedman test suggested that there was no significant difference among the six sampling sites (P > 0.05). The viromes in this study were all dominated by the order Caudovirales, and a group of Freshwater phage uvFW species were particularly prevalent among all the samples. The virome from Nanjing presented a unique pattern of viral community composition with a relatively high abundance of family Parvoviridae. Phylogenetic analyses based on virus hallmark genes showed that the Caudovirales order and CRESS-DNA viruses presented high genetic diversity, while viruses in the Microviridae and Parvoviridae families and the Riboviria realm were relatively conservative. Our study provides the first insight into viral community composition in large river ecosystem, revealing the diversity and stability of river water virome, contributing to the proper utilization of freshwater resource.     

Seasonal variations in PCR-DGGE fingerprinted viruses infecting phytoplankton in large and deep peri-alpine lakes

Double-stranded DNA viruses infecting eukaryotic algae (e.g., phycodnaviruses) and cyanobacteria (e.g., cyanophages) are now recognized as widespread and ubiquitous in aquatic environments. However, both the diversity and functional roles of these viruses in fresh waters are still poorly understood. We conducted a year-long study in 2011 of the community structure of planktonic virus groups in the upper lit layer of two important freshwater natural ecosystems in France, Lake Annecy (oligotrophic) and Lake Bourget (oligo-mesotrophic). Using PCR-DGGE to target a number of different structural and functional signature genes, i.e.,g20, g23, psbA, polB, and mcp, the phytoplankton viruses were shown to display temporal and spatial variability. There were marked seasonal changes in community structure for all viral groups in Lake Bourget, but only for T4-like myoviruses and psbA-containing cyanophages in Lake Annecy. The multivariate statistical analyses revealed that (1) various environmental factors can directly or indirectly explain the community structure observed for each phytoplankton viral group, and (2) temporal patterns of T4-like myovirus community structure were similar between the two lakes. In general, our results (1) suggest that the observed algal virus patterns were associated with significant shifts in phytoplankton biomass and/or structure, which in turn were shaped by the abiotic environment, and (2) support the Bank model proposed by Breitbart and Rohwer (Trends Microbiol 13:278–284, 2005). This study provides new evidence that freshwater lakes contain a significant diversity of algal viruses, and that the distribution of these viruses strongly mirrors that of their hosts.     

贵州省四种蝙蝠携带病毒组

翼手目(Chiroptera)动物已被确认是人畜共患病毒的重要自然宿主。贵州省翼手目物种多样性资源丰富,包括了2亚目7科19属65种,但在其携带病毒方面的研究仍然不全面。本文基于病毒宏基因组学和s RNA病毒检测,对贵州省广泛分布的大蹄蝠(Hipposideriderosarmiger)、三叶蹄蝠(Aselliscus wheeleri)、贵州菊头蝠(Rhinolophus rex)和皮氏菊头蝠(R. pearsoni)携带的病毒进行注释及鉴定。通过分析得到所携带病毒的种类;并比较了贵州省与云南省和广西省3个地区翼手目携带病毒在种类上的差异。结果显示,在4种蝙蝠中检测出脊椎动物病毒、昆虫病毒、植物病毒、细菌病毒4大类,共计53科111属170余种病毒,其中具有公共卫生学意义病毒9科10属46种,如:人疱疹病毒1型病毒(Human herpesvirus 1)、戊型肝炎病毒(Hepatitis E virus)、人乳头瘤病毒16型(Human papillomavirustype16)等相关的病毒。贵州省与云南省和广西省3个地区的蝙蝠所携带病毒种类比较发现,只有腺病毒科(Adenoviridae)和逆转录病毒科(Retroviridae)均存在于3个省份的蝙蝠体内,说明病毒种类在不同地域分布的蝙蝠种类中存在较大的差异。贵州省分布的大蹄蝠、三叶蹄蝠、贵州菊头蝠和皮氏菊头蝠携带病毒的种类较多,其病毒种类在公共卫生学上具有一定的代表性,对开展贵州省分子流行病学调查具有重要价值。     

Metagenomic analysis of the viral flora of pine marten and European badger feces

A thorough understanding of the diversity of viruses in wildlife provides epidemiological baseline information about potential pathogens. Metagenomic analysis of the enteric viral flora revealed a new anellovirus and bocavirus species in pine martens and a new circovirus-like virus and geminivirus-related DNA virus in European badgers. In addition, sequences with homology to viruses from the families Paramyxo- and Picornaviridae were detected.     

应用多种生物类群评价白洋淀水环境质量变化

白洋淀是华北平原最大的浅水湖泊。近几十年来,白洋淀上游耗水增加,天然入淀水量急剧减少,使白洋淀水位明显降低;同时,随着淀泊周边农村生活污水的排入,白洋淀发生了严重的富营养化。2017年雄安新区设立,白洋淀环境治理工作的重要性空前提高。近几年通过上游水库和跨流域生态补水,白洋淀水位明显抬升,同时淀中和淀边农村实施了生活污水收集。为了解白洋淀当前水环境状况,对前期补水和治污的效果进行评估,2019年8月对白洋淀水体理化指标及水生生物进行调查,采用水质、浮游植物、浮游动物、底栖动物指标评价白洋淀水环境质量,并与2010年的调查数据进行对比,辅以沉水植物指标评价水体生态状况。结果表明: 白洋淀2019年总磷达到优于Ⅲ类水平,总氮优于Ⅱ类水平,水环境状况比2010年有较大改善。其中,总磷浓度下降了88.6%,总氮浓度下降了83.9%,叶绿素a浓度下降了47.8%,透明度增大了43.4%;浮游动物、底栖动物等生物多样性明显提高,浮游植物密度显著降低,优势种从普遍的重富营养型转变为富营养型;沉水植物清洁型种类分布范围缩小。这说明沉水植物物种评价法不适于白洋淀水环境质量评价,浮游动物多样性评价方法也不适用于当前白洋淀水环境质量评价。早期,由于污染源集中在西部,白洋淀西部水质差于东部;现在,由于自西、北、南多方向补水,淀区的水动力条件发生显著改变,加之西部实施治污工程,淀区水质呈现空间均一化的特点。     

珊瑚礁生态系统病毒研究进展

病毒对珊瑚礁生态系统中的生物进化、生物地球化学循环、珊瑚疾病等方面具有重要的生态影响。随着珊瑚礁的全球性退化，病毒在珊瑚礁生态系统中的功能与危害日益显现。综述了珊瑚礁生态系统中病毒的研究现状与进展，包括：（1）珊瑚礁病毒的多样性与分布特征（水体、宿主、核心病毒组）；（2）珊瑚礁病毒的生态功能（感染方式、促进生物进化、生物地球化学循环）；（3）珊瑚礁病毒对全球气候变化的响应（热压力、珊瑚疾病）。总体而言，珊瑚礁生态系统具有极高的病毒多样性，所发现的60个科占已知所有病毒科数量的58%。珊瑚的核心病毒组主要由双链DNA病毒、单链DNA病毒、单链逆转录病毒所组成，珊瑚黏液层对病毒具有富集作用。"Piggyback-the-Winner"（依附-胜利）是病毒在珊瑚礁中主要的生物动力学模式，其可通过水平基因迁移的方式促进礁区生物进化。病毒可通过裂解细菌与浮游藻类的途径参与珊瑚礁的生物地球化学循环，尤其是碳循环与氮循环过程。此外，病毒还具有介导珊瑚热白化与直接引发珊瑚疾病的能力，这会影响珊瑚礁生态系统应对气候变化的适应性与恢复力。基于国际上的研究进展综述，结合南海珊瑚礁生态现状提出以下研究方向，以期促进我国珊瑚礁病毒学的发展：（1）开展南海珊瑚礁中病毒多样性的识别及其时-空分布特征研究；（2）探索病毒对南海珊瑚热白化、珊瑚疾病的介导作用及其与气候变化的关系；（3）揭示病毒对南海珊瑚礁生物地球化学循环的贡献。     

青海湖国家级自然保护区水鸟的多样性及分布

青海湖国家级自然保护区为国际重要湿地,其野生候鸟种群曾于2005年感染高致病性禽流感病毒H5N1,对当地野生鸟类种群及牛态环境造成很大威胁.作者于2007年3-12月.采用直接计数法,对该保护区鸟类资源进行凋查.本次考察共记录到鸟类57种,隶属8目14科.其中留鸟2种,迷鸟3种,夏候鸟7种,迁徙路过鸟41种,冬候鸟4种,其中54种鸟类为水鸟.     

New archaeal viruses discovered by metagenomic analysis of viral communities in enrichment cultures

Viruses infecting hyperthermophilic archaea of the phylum Crenarchaeota display enormous morphological and genetic diversity, and are classified into 12 families. Eight of these families include only one or two species, indicating sparse sampling of the crenarchaeal virus diversity. In an attempt to expand the crenarchaeal virome, we explored virus diversity in the acidic, hot spring Umi Jigoku in Beppu, Japan. Environmental samples were used to establish enrichment cultures under conditions favouring virus replication. The host diversity in the enrichment cultures was restricted to members of the order Sulfolobales. Metagenomic sequencing of the viral communities yielded seven complete or near-complete double-stranded DNA virus genomes. Six of these genomes could be attributed to polyhedral and filamentous viruses that were observed by electron microscopy in the enrichment cultures. Two icosahedral viruses represented species in the family Portogloboviridae. Among the filamentous viruses, two were identified as new species in the families Rudiviridae and Lipothrixviridae, whereas two other formed a group seemingly distinct from the known virus genera. No particle morphotype could be unequivocally assigned to the seventh viral genome, which apparently represents a new virus type. Our results suggest that filamentous viruses are globally distributed and are prevalent virus types in extreme geothermal environments.     

Temporal and spatial dynamics of phytoplankton diversity in the East China Sea near Jeju Island (Korea): A pyrosequencing-based study

The East China Sea (ECS) has long been considered an important monitoring site for oceanic ecosystem changes because many water currents and river discharges constantly influence this area. In this study, the community structure and diversity of phytoplankton in the northern part of the ECS adjacent to Jeju Island were explored using small subunit ribosomal RNA (SSU) pyrosequencing. We analysed samples collected from four stations from the surface and at 30-m and 50-m depths during April and September 2011. We observed spatial and temporal variations in the phytoplankton community. Among phytoplankton, diatoms and dinoflagellates constituted a major portion at all stations (60–90%). However, comparison of the April and September samples showed seasonal variation and shifts in the dispersion of diatom and dinoflagellates among stations. Among stations, diatoms dominated St. 1 and others were dominated by dinoflagellates. Furthermore, phylotypes of potentially toxin-producing genera such as Karlodinium, Heterocapsa, Gymnodinium, Gyrodinium, and Pseudo-nitzschia were dominant in this area.     

Diversity of the major capsid genes (g23) of T4-like bacteriophages in the eutrophic Lake Kotokel in East Siberia, Russia

Numerous studies revealed high diversity of T4-like bacteriophages in various environments, but so far, little is known about T4-like virus diversity in freshwater bodies, particularly in eutrophic lakes. The present study was aimed at elucidating molecular diversity of T4-like bacteriophages in eutrophic Lake Kotokel located near Lake Baikal by partial sequencing of the major capsid genes (g23) of T4-like bacteriophages. The majority of g23 fragments from Lake Kotokel were most similar to those from freshwater lakes and paddy fields. Despite the proximity and direct water connection between Lake Kotokel and Lake Baikal, g23 sequence assemblages from two lakes were different. UniFrac analysis showed that uncultured T4-like viruses from Lake Kotokel tended to cluster with those from the distant lake of the same trophic status. This fact suggested that the trophic conditions affected the formation of viral populations, particularly of T4-like viruses, in freshwater environments.