Plant‐ and alga‐associated bacterial communities are generally described via 16S rDNA metabarcoding using universal primers. As plastid genomes encode 16S rDNA related to cyanobacteria, these data sets frequently contain >90% plastidial sequences, and the bacterial diversity may be under‐sampled. To overcome this limitation we evaluated in silico the taxonomic coverage for four primer combinations targeting the 16S rDNA V3‐V4 region. They included a forward primer universal to Bacteria (S‐D‐Bact‐0341‐b‐S‐17) and four reverse primers designed to avoid plastid DNA amplification. The best primer combination (NOCHL) was compared to the universal primer set in the wet lab using a synthetic community and samples from three macroalgal species. The proportion of plastid sequences was reduced by 99%–100% with the NOCHL primers compared to the universal primers, irrespective of algal hosts, sample collection and extraction protocols. Additionally, the NOCHL primers yielded a higher richness while maintaining the community structure. As Planctomycetes, Verrucomicrobia and Cyanobacteria were underrepresented (70%–90%) compared to universal primers, combining the NOCHL set with taxon‐specific primers may be useful for a complete description of the alga‐associated bacterial diversity. The NOCHL primers represent an innovation to study algal holobionts without amplifying host plastid sequences and may further be applied to other photosynthetic hosts.

中文翻译：

---

在藻类相关细菌群落的16S rDNA元条形码分析中评估一种新的引物组合以最大程度地减少质体污染。

植物和藻类相关的细菌群落通常通过使用通用引物的16S rDNA元条形码进行描述。由于质体基因组编码与蓝细菌有关的16S rDNA，因此这些数据集通常包含> 90％的质体序列，细菌多样性可能被欠采样。为克服此限制，我们在计算机上评估了针对16S rDNA V3-V4区域的四种引物组合的分类学覆盖率。其中包括细菌通用的正向引物（S-D-Bact-0341-b-S-17）和四个反向引物，可避免质体DNA扩增。使用合成群落和来自三个大型藻类的样品，将最佳引物组合（NOCHL）与湿实验室中的通用引物组进行比较。使用NOCHL引物，与通用引物相比，质体序列的比例降低了99％–100％，无论藻类宿主，样品收集和提取方案如何。另外，NOCHL引物在保持群落结构的同时产生了更高的丰度。作为扁平菌，Verrucomicrobia和蓝细菌与通用引物相比，它们的代表性不足（70％–90％），将NOCHL集与分类群特异性引物结合使用可能有助于完整描述藻类相关细菌的多样性。NOCHL引物代表着一项创新，可在不扩增宿主质体序列的情况下研究藻类全盐菌，并可进一步应用于其他光合作用宿主。