Gradients1-KOK1606-DiazotrophAbundance_2020-03-04_v1.2

Cruise Name: Gradients 1.0 / KOK1606 Project Name: SIMONS Foundation NPSG Gradient At select stations and depths (ranging from 5 to 45m), the entire volume of a Niskin bottle was drained directly into clean ~10L carboys fitted with spigots. Carboys were covered in dark cloth and the volume was gravity filtered through a 47-mm diameter, 3 micron pore size and 10 micron pore size, black polycarbonate filter with a polyester drain disk as a backing filter. If the volume was not filtered after 2 hours, filtration was terminated and the remaining volume of the carboy was measured in order to calculate volume filtered. Following filtration, filter holders were fit with a short section of tubing and a syringe luer fitting on once side and a 2-way valve on the outflow side. For each filter, 5-ml of 2% glutaraldehyde was slowly injected onto the filter and samples were allowed to fix for 30 minutes. Fixative was drained after this time and 60ml of air was used to flush all filters. Polycarbonate filters were then mounted onto 3x2” glass slides with immersion oil, cover slides were added and the edges of each cover slip was sealed with quick dry nail polish. All slides were stored at -20°C and counted within 30 days. Enumeration of diazotrophic taxa was performed using epifluorescence microscopy.The entire slide was counted for DDAs and Trichodesimum abundance. Endosymbiont bearing diatoms of the following genus were enumerated: Rhizosolenia, Hemiaulus, Climacodium and Chaetoceras. Free Richelia intracellularis were also counted. Trichodesmium filaments were counted and the length of each filament was recorded. Trichodesmium cell number was then calculated by dividing the filament length by the mean cell length (9.9 micron ±2.5 micron).

航次名称：Gradients 1.0 / KOK1606；项目名称：SIMONS基金会NPSG梯度项目（SIMONS Foundation NPSG Gradient）。在选定的站位与水深范围（5米至45米）内，将尼斯金采水器（Niskin bottle）内的全部水样直接注入带有旋塞的洁净10升储液瓶（carboys）。储液瓶以黑布遮盖，水样经重力过滤：采用直径47毫米、孔径3微米与10微米的黑色聚碳酸酯滤膜（polycarbonate filter），以聚酯引流盘作为支撑滤膜。若水样在2小时内未完成过滤，则终止过滤操作，并测量储液瓶剩余体积以计算已过滤水样的总体积。过滤完成后，滤器支架加装一段短连接管路（tubing），一侧配备注射器鲁尔接头（syringe luer fitting），流出侧加装二通阀。针对每一片滤膜，缓慢注入5毫升2%戊二醛（glutaraldehyde）固定液，静置固定30分钟。固定完成后排去固定液，使用60毫升空气吹扫所有滤膜。随后将聚碳酸酯滤膜安装于3×2英寸的载玻片（glass slides）上，滴加浸油，加盖盖玻片，并用快干指甲油封固盖玻片边缘。所有玻片均保存于-20℃环境中，并在30天内完成镜检计数。固氮类群的计数采用落射荧光显微镜（epifluorescence microscopy）完成：对全部玻片进行计数以获取DDAs与束毛藻（Trichodesmium）的丰度。对以下属的携带内共生体的硅藻进行计数：根管藻属（Rhizosolenia）、半管藻属（Hemiaulus）、梭形藻属（Climacodium）以及角毛藻属（Chaetoceras）。同时计数游离的胞内列氏藻（Richelia intracellularis）。记录每一根束毛藻丝状体的长度，通过将丝状体长度除以平均细胞长度（9.9微米±2.5微米）计算束毛藻的细胞总数。