浮游藻类环境DNA宏条形码监测引物的比较与验证

环境DNA (eDNA)宏条形码技术通量高、重复性好,在未来生态环境监测中有巨大的应用潜力。目前,浮游藻类环境DNA监测仍处在发展阶段,尚缺乏统一的浮游藻类扩增引物。利用同一个野外环境样本,比较8对通用引物在浮游藻类环境DNA监测中的差异,为初步建立规范化的浮游藻类环境DNA监测方法提供支撑。结果表明,不同引物对浮游藻类扩增存在明显偏好性,靶向扩增16S rDNA的引物主要检出硅藻,其次是隐藻和绿藻;靶向扩增18S rDNA的1391、AD3和ANF 3对引物具有较高的浮游藻类扩增效率和物种辨识度,分别检出67、62、63个浮游藻属,其检出的浮游藻类的相对丰度排序均为硅藻>绿藻>隐藻>金藻>甲藻,可以作为通用引物用于浮游藻类环境DNA宏条形码监测。     

The use of environmental DNA in invasive species surveillance of the Great Lakes commercial bait trade

Over 180 non‐native species have been introduced in the Laurentian Great Lakes region, many posing threats to native species and ecosystem functioning. One potential pathway for introductions is the commercial bait trade; unknowing or unconcerned anglers commonly release unused bait into aquatic systems. Previous surveillance efforts of this pathway relied on visual inspection of bait stocks in retail shops, which can be time and cost prohibitive and requires a trained individual that can rapidly and accurately identify cryptic species. Environmental DNA (eDNA) surveillance, a molecular tool that has been used for surveillance in aquatic environments, can be used to efficiently detect species at low abundances. We collected and analyzed 576 eDNA samples from 525 retail bait shops throughout the Laurentian Great Lake states. We used eDNA techniques to screen samples for multiple aquatic invasive species (AIS) that could be transported in the bait trade, including bighead (Hypophthalmichthys nobilis) and silver carp (H. molitrix), round goby (Neogobius melanostomus), tubenose goby (Proterorhinus marmoratus), Eurasian rudd (Scardinius erythrophthalmus), and goldfish (Carassius auratus). Twenty‐seven samples were positive for at least one target species (4.7% of samples), and all target species were found at least once, except bighead carp. Despite current regulations, the bait trade remains a potential pathway for invasive species introductions in the Great Lakes region. Alterations to existing management strategies regarding the collection, transportation, and use of live bait are warranted, including new and updated regulations, to prevent future introductions of invasive species in the Great Lakes via the bait trade. El Uso del ADN Ambiental en la Vigilancia de Especies Invasoras del Mercado de Carnada Comercial de los Grandes Lagos     

Using environmental DNA to assess population‐wide spatiotemporal reserve use

Scientists increasingly rely on protected areas to assist in biodiversity conservation, yet the efficacy of these areas is rarely systematically assessed, often because of underfunding. Still, adaptive management strategies to maximize conservation success often rely on understanding the temporal and spatial dynamism of populations therein. Examination of environmental DNA (eDNA) is a time and cost‐effective way to monitor species’ distribution, and quantitative polymerase chain reaction (qPCR) provides information on organismal abundance. To date, however, such techniques remain underused for population assessments in protected areas. We determined eDNA concentration of the critically endangered Yangtze finless porpoise (Neophocaena asiaeorientalis asiaeorientalis) to describe its occurrence, range, and use of the Tian e‐Zhou National Nature Reserve in Hubei, China, across seasons and hydrological depths. Despite the observation that total eDNA concentrations were highest in surface waters in summer, finless porpoise eDNA concentrations were significantly higher in deeper waters than in surface waters in summer. During the breeding season (spring), eDNA signals were site specific and restricted to the core area of the reserve. However, postbreeding eDNA concentrations were widespread across the reserve, encompassing sites previously thought to be unfrequented by the species. Our results suggest spatiotemporal idiosyncrasies in site, depth, and seasonal use of the reserve and a propensity for postbreeding population dispersal. With eDNA and qPCR we were able to assess an entire population's use of a protected area. Illuminating nuances in habitat use via eDNA could be valuable to set pragmatic conservation goals for this, and other, species.     

BALB／C鼠睾丸组织中冷诱导RNA结合蛋白的cDNA克隆与序列分析

冷诱导RNA结合蛋白(Cold inducible RNA-binding protein,CIRP)在多种冷应激细胞(包括重组中国仓鼠卵巢细胞)中被发现.迄今为止,冷应激对活体生物基凶表达的影响还未见报道.和细胞相比,生物体具有更加复杂的冷应激调节机制.本研究以冷处理的BALB/C鼠为实验动物,从其睾丸组织巾克隆出了CIRP的cDNA.结果表明,CIRP在生物体中能够被低温诱导,可能防止生物体遭受冷损伤.根据克隆的cDNA所推测的氨基酸序列与GenBank上公布的小鼠、大鼠、人类、牛蛙、美西螈、非洲爪蟾胚胎细胞和卵母细胞的CIRP氨基酸序列同源性分别为100%、99.40%、95.5%、67.4%、58.4%、76.9%和79.1%.这表明CIRP在生物进化过程中是高度保守的,可能具有多种生理功能.因此,这一研究将为探索人类和动物冷应激分子机制创立系统试验模型和奠定新的实践基础.图5参14     

中国淡水大型底栖无脊椎动物条形码数据库构建

大型底栖无脊椎动物是得到广泛应用的水质生物监测和评价指示生物,但在实际应用过程中,受类群多样性高、形态鉴定专业性强、鉴定资料不全等影响,难以将其精确鉴定到种.新兴的DNA条形码技术能够快速、精确地鉴定物种,在生物多样性调查和监测等领域被寄予厚望.该方法的有效性与准确性依赖于完整、全面的参考数据库,然而现有公共数据库无法...     

基于宏条形码技术的白洋淀水华藻类识别及其驱动因子分析

浮游藻类是引起水华暴发的主要原因.为筛选潜在水华藻类,评估白洋淀水华风险区域,于2020年8月对白洋淀373点位展开浮游藻类调查.利用宏条形码技术分析,解析水华藻类群落组成,同时采用显微镜计数法统计藻密度.根据总藻密度对白洋淀不同区域的水华程度进行评估,同时进一步针对水华藻类群落,耦合淀区水质条件,探究白洋淀不同区域水华藻类群落空间差异驱动因子,以甄别影响水华藻类群落结构关键环境因子.结果表明,95%以上采样区域无水华风险(藻类密度<2×10⁶个·L^-1),仅5个样点存在轻微水华风险.但水华藻类群落分析共检测到了90种水华藻类,其中优势水华藻种有20种,隶属于以绿藻门、蓝藻门和裸藻门为主.水华藻类群落结构在不同区域上具有显著空间异质性(P<0.05).关键驱动因子解析结果表明,总磷(TP)、总氮(TN)和氨氮(NH⁺₄-N)是造成水华藻类群落结构差异的关键因子.其中,门水平上,蓝藻门水华藻类与以上关键因子显著正相关;种水平上,硅藻门和绿藻门水华藻类与关键因子响应更显著.因此,水华藻类群落...     

基于eDNA的硅藻群落时空异质性及生态健康评价

近年来,环境DNA技术广泛应用于水生态生物多样性监测,如何基于环境DNA数据建立河流生态健康评价方法是当前研究的热点.拟开发基于环境DNA的分子硅藻指数来指示人类活动影响下的河流生态健康状况.首先通过环境DNA技术监测了沙颍河流域春秋两季硅藻群落组成和结构变化,解析了春、秋两季群落的驱动环境因子;进一步比较了4种基于环境DNA宏条形码数据的计算策略(OTU-分类、OTU-无分类、ASV-分类和ASV-无分类),构建了适合于评估沙颍河流域生态健康状况的分子硅藻指数.结果表明:①硅藻群落结构存在季节性差异,其中Discostella pseudostelligera、Nitzschia amphibia和Diatoma vulgaris等类群是区分春、秋季硅藻群落季节差异的主要因素;②重金属Mn、Fe和TN (总氮)是影响春季硅藻群落结构变化的主要环境因子,COD和Cu是影响秋季硅藻群落结构变化的主要环境因子;③在4种计算策略中,基于OTU-无分类数据计算的硅藻指数更好地反映了环境梯度变化;硅藻指数显示,沙颍河流域生态健康状况在时间上秋季优于春季,而在空间上上游优于下游.综上,通过eDNA技术监测了沙颍河春、秋两季硅藻群落,构建了沙颍河流域分子硅藻指数,促进了环境DNA技术在河流生态健康评价中的应用.     

长江流域河流和湖库的浮游细菌群落差异

在长江流域自江源区至入海口的干流重点区域、八大一级支流和多个重点湖库设置了177个样点开展浮游细菌调查,采用基于细菌16S rRNA基因的单分子实时测序技术研究浮游细菌群落的空间分布差异,期望在更广的调查范围和更高的物种注释精度上阐明长江流域浮游细菌群落结构的分布特征.首先分析了各采样点浮游细菌的α多样性状况,发现长江流域河流区域具有更高的物种丰富度,在多样性上与湖库区域差异显著.在此基础上开展了长江流域各采样点的β多样性分析,发现长江流域河流区域和湖库区域的浮游细菌群落组成差异显著,其中温度是河流浮游细菌群落分布的最大影响因素,湖库的pH是浮游细菌群落分布的最大影响因素.为探究河流区域和湖库区域浮游细菌群落结构差异在生态环境上的影响,在物种差异、功能差异和群落结构差异上开展了进一步研究.结果表明,对于河流区域而言,长江干流自西向东营养盐水平逐渐升高导致了部分物种相对丰度的逐渐增加;而湖库较河流有更高的蓝藻水华暴发风险,其中条件致病菌不动杆菌属在丹江口水库(南水北调水源区)具有较高相对丰度,存在着一定的生态风险,需要加强关注.物种组成上,河流区域和湖库区域绝大多数的物种(84%的OTU...