DataSheet_1_Metabarcoding the eukaryotic community of a threatened, iconic Mediterranean habitat: Posidonia oceanica seagrass meadows.pdf

Against the accelerating pace of worldwide species extinction, reliable biodiversity assessments are critical, both as baselines and to track potential declines. DNA metabarcoding techniques allow for fast and comprehensive assessment of biodiversity in both terrestrial and marine habitats. However, these methods need to be adapted and standardised for each ecosystem in order to be effective. Seagrass meadows are among the most diverse marine habitats and are irreplaceable in terms of the ecosystem services they provide, yet metabarcoding has never been implemented for these systems. In this study, we developed and tested a protocol for metabarcoding the eukaryotic community of meadows of the iconic species, Posidonia oceanica L. (Delile). This seagrass is the main habitat-forming species in Mediterranean coastal waters and is known for its high diversity due to the structural complexity of its canopy and rhizome structures. This habitat is experiencing a range-wide retreat, and there is an urgent need for fast and efficient methods for its biomonitoring and detection of early changes. Our proposed method involves direct sampling of the community, collecting and processing the leaves and rhizome strata separately. To test the utility of the method in distinguishing between different meadow conditions, we sampled two distinct meadows that differ in their prevailing wind and surge conditions, and a nearby rocky reef for comparison. We then adapted a method and pipeline for COI metabarcoding using generalist primers that target the eukaryote diversity present. We detected a high diversity in the two meadows analysed (3,350 molecular operational taxonomic units, dominated by Metazoa and Archaeplastida) and a clear differentiation of the seagrass samples from those of the nearby rocky reefs. The leaves and rhizomes harboured clearly distinct assemblages, and differences were also detected between the two meadows sampled. This new tool has the potential to deliver big biodiversity data for seagrass habitats in a fast and efficient way, which is crucial for the implementation of protection and management measures for this key coastal habitat.

面对全球范围内日益加快的物种灭绝步伐，可靠的生物多样性评估无论是作为基准基线，还是用于追踪潜在的种群衰退，都至关重要。DNA元条形码（DNA metabarcoding）技术能够实现陆地与海洋生境中生物多样性的快速且全面评估。然而，此类方法需针对每一类生态系统进行适配与标准化，方能发挥实效。海草床是物种最为丰富的海洋生境之一，其所提供的生态系统服务具有不可替代性，但目前尚未有元条形码技术应用于该类生境的相关研究。本研究针对标志性物种波西多尼亚海草（Posidonia oceanica L. (Delile)）的海草床真核生物群落，开发并测试了一套元条形码采样方案。该海草是地中海近岸水域的主要生境构建物种，因其冠层与根状茎结构的复杂性而拥有极高的物种多样性。当前，此类生境正经历全域性的衰退，亟需快速高效的方法对其进行生物监测并及早发现群落变化。我们提出的方案采用直接采样策略，分别采集并处理叶片与根状茎层样品。为验证该方法区分不同海草床状态的能力，我们选取了两处受主流风浪条件影响不同的典型海草床，以及一处邻近的岩礁生境作为对照进行采样。随后，我们针对群落中存在的真核生物多样性，采用通用引物优化了细胞色素c氧化酶亚基I（COI）元条形码的实验方法与分析流程。经检测，所分析的两处海草床展现出极高的物种多样性（共检出3350个分子操作分类单元（molecular operational taxonomic units，MOTUs），优势类群为后生动物（Metazoa）与古质体类（Archaeplastida）），且海草床样品与邻近岩礁样品的群落结构呈现显著分化。研究发现，叶片与根状茎的生物群落组成存在明显差异，且两处采样海草床之间的群落结构亦存在显著区别。这套全新的工具能够快速高效地获取海草生境的大规模生物多样性数据，这对于该关键近岸生境的保护与管理措施的落地实施具有重要意义。