Dynamic Properties of Picoplankton Over the Annual Cycle in Near-Polar Lakes

9907776Button这个项目将研究近极地湖泊中微微浮游生物在年度周期中的动态特性。调查的重点将是描述在初级生产力和温度严重转变期间来自近北极淡水系统的典型细菌物种，推断其维持过程中关键有机物的存在，并量化这些大多未描述的生物对二氧化碳产生率的贡献。众所周知，研究地点哈丁湖在年度周期中有相对稳定的细菌生物量。然而，细菌的物理性质、DNA分布、动力学常数和相关氨基酸浓度以及初级生产力在夏季和冬季之间变化很大。重点放在主要的耐培养细菌上，并注意小的低DNA形式。将对这些碳源进行分离和检测，以确定它们的活性和利用各种碳源的能力，包括那些来自植物材料水解物的碳源。将使用一些新的方法。其中包括最近完善的通过流式细胞术测量种群、每细胞生物量、细胞体积、DNA含量和基因组大小的方法。染色体比率生长率方法将在低值下得到验证，温度修正将重新评估，该方法将用作原位生长速度的探针，并通过稀释培养进行验证。DAPI-DNA分析将通过同时评估单个细菌的G+C含量来修正A+T偏差，从而提高亚群分辨率。其他单细胞活性探针包括RNA、稀释培养中后续活性的还原能力和膜潜力，以及更全面地表征现场种群在年度周期中的特征。依赖于自然产生的碳源的消亡培养被用来分离细菌，而不偏向大型快速生长的形式。通过扩散膜将培养物与稻草湖水隔开，以提高可养殖性。通过结合新的公式和DGGE技术，数据被用来确定生存能力和存在的物种数量，以确定发展中种群的物种数量。根据营养改良剂后灭绝培养物的流式细胞仪模式，对有用的碳源进行了评估。使用的底物被认为是存在的指示，动力学测量表明它们的浓度。根据最近定义的特定亲和力和寡营养能力，直接比较在低浓度下使用底物的能力。分离的物种通过16-S核糖体核糖核酸分析进行鉴定，并根据活性(总减去极低的脱氧核糖核酸)生物量和相关的生长速率计算原位二氧化碳释放。在没有碳源修正的情况下，评估温度对吸收和生长的影响，以确定在低温下速率是否崩溃，或者是否保持足够的膜电位以保持所需的特定亲和力。

9907776Button This project will examine the dynamic properties of picoplankton over the annual cycle in near-polar lakes. Investigations will focus on describing typical bacterial species from a near-arctic freshwater system during the severe transition in primary productivity and temperature, infer the presence of key organics in theirmaintenance, and quantify the contribution of these mostly undescribed organisms to carbon dioxide production rates. The study site, Harding Lake, is known to have a relatively stable bacterial biomass over the annual cycle. However physical properties of the bacteria, DNA distributions, kinetic constants and associated amino acid concentrations and primary productivities varydramatically between summer and winter. Focus is on the predominatecultivation-resistant bacteria with attention to the small low-DNA forms. Thesewill be isolated and probed for their activity and ability to utilize variouscarbon sources including those expected from the hydrolysis of plant material. A number of novel approaches will be used. These include recently perfectedmeasures of population, biomass per cell, cell volume, DNA content, and genome size by flow cytometry. A chromosome-ratio method for growth rate will be verified at low values, temperature corrections reevaluated, and the method used as a probe of in situ rates of growth with verification by dilution culture.DAPI-DNA assays will be corrected for A+T bias by simultaneously evaluating G+C content of individual bacteria, thereby improving subpopulation resolution.Additional single-cell activity probes include RNA, reducing power and membrane potential for following activity in dilution cultures and more thoroughlycharacterizing in situ populations over the annual cycle. Extinction culturesthat depend on naturally produced sources of carbon are used to isolate bacteriawithout bias toward large fast-growing forms. Cultures are partitioned againstraw lakewater through diffusion membranes to improve culturability. Data areused to determine viabilities as well as the number of species present byincorporating novel formulae as well as DGGE techniques to identify the numberof species in developing populations. Useful carbon sources are evaluatedaccording to flow cytometry patterns from the extinction cultures followingnutrient amendments. Substrates used are taken as an indication of those present and kinetic measurements suggest their concentrations. Ability to use substrates at small concentrations is compared directly according to specific affinity and oligotrophic capacity as recently defined. Isolated species are identified by 16-S ribosomal RNA analysis, and in situ carbon dioxide evolution is calculated from growth rates according to the active (total less very low-DNA) biomass and associated rates of growth. Temperature effects on both uptake and growth are evaluated without carbon-source amendment to determine if rates at low temperatures collapse, or if sufficient membrane potential is sustained tomaintain the required specific affinities.