Wietz Matthias, Metfies Katja, Bienhold Christina, Wolf Christian, Janssen Felix, Salter Ian, Boetius Antje
Deep-Sea Ecology and Technology, Alfred Wegener Institute Helmholtz Centre for Polar and Marine Research, Bremerhaven, Germany.
Max Planck Institute for Marine Microbiology, Bremen, Germany.
Front Microbiol. 2022 Sep 7;13:999925. doi: 10.3389/fmicb.2022.999925. eCollection 2022.
Automated sampling technologies can enhance the temporal and spatial resolution of marine microbial observations, particularly in remote and inaccessible areas. A critical aspect of automated microbiome sampling is the preservation of nucleic acids over long-term autosampler deployments. Understanding the impact of preservation method on microbial metabarcoding is essential for implementing genomic observatories into existing infrastructure, and for establishing best practices for the regional and global synthesis of data. The present study evaluates the effect of two preservatives commonly used in autosampler deployments (mercuric chloride and formalin) and two extraction kits (PowerWater and NucleoSpin) on amplicon sequencing of 16S and 18S rRNA gene over 50 weeks of sample storage. Our results suggest the combination of mercuric chloride preservation and PowerWater extraction as most adequate for 16S and 18S rRNA gene amplicon-sequencing from the same seawater sample. This approach provides consistent information on species richness, diversity and community composition in comparison to control samples (nonfixed, filtered and frozen) when stored up to 50 weeks at temperature. Preservation affects the recovery of certain taxa, with specific OTUs becoming overrepresented (SAR11 and diatoms) or underrepresented ( and pico-eukaryotes) after preservation. In case eukaryotic sequence information is the sole target, formalin preservation and NucleoSpin extraction performed best. Our study contributes to the design of long-term autonomous microbial observations in remote ocean areas, allowing cross-comparison of microbiome dynamics across sampling devices (e.g., water and particle samplers) and marine realms.
自动化采样技术可以提高海洋微生物观测的时间和空间分辨率，特别是在偏远和难以到达的地区。自动化微生物组采样的一个关键方面是在长期自动采样器部署过程中保存核酸。了解保存方法对微生物代谢条形码的影响对于将基因组观测站纳入现有基础设施以及建立区域和全球数据综合的最佳实践至关重要。本研究评估了自动采样器部署中常用的两种防腐剂（氯化汞和福尔马林）和两种提取试剂盒（PowerWater和NucleoSpin）对样品储存50周后16S和18S rRNA基因扩增子测序的影响。我们的结果表明，氯化汞保存和PowerWater提取的组合最适合从同一海水样品中进行16S和18S rRNA基因扩增子测序。与对照样品（未固定、过滤和冷冻）相比，这种方法在4℃储存长达50周时，能提供关于物种丰富度、多样性和群落组成的一致信息。保存会影响某些分类群的回收率，特定的操作分类单元在保存后会过度代表（SAR11和硅藻）或代表性不足（ 和微微真核生物）。如果真核生物序列信息是唯一目标，福尔马林保存和NucleoSpin提取效果最佳。我们的研究有助于设计偏远海洋区域的长期自主微生物观测，允许跨采样设备（如水和颗粒采样器）和海洋领域进行微生物组动态的交叉比较。
