Center for Evolutionary Hologenomics, Globe Institute, University of Copenhagen, Copenhagen, Denmark.
Section of Microbiology, Department of Biology, University of Copenhagen, Copenhagen, Denmark.
Microbiol Spectr. 2024 Apr 2;12(4):e0359023. doi: 10.1128/spectrum.03590-23. Epub 2024 Mar 7.
Shotgun metagenomics enables the reconstruction of complex microbial communities at a high level of detail. Such an approach can be conducted using both short-read and long-read sequencing data, as well as a combination of both. To assess the pros and cons of these different approaches, we used 22 fecal DNA extracts collected weekly for 11 weeks from two respective lab mice to study seven performance metrics over four combinations of sequencing depth and technology: (i) 20 Gbp of Illumina short-read data, (ii) 40 Gbp of short-read data, (iii) 20 Gbp of PacBio HiFi long-read data, and (iv) 40 Gbp of hybrid (20 Gbp of short-read +20 Gbp of long-read) data. No strategy was best for all metrics; instead, each one excelled across different metrics. The long-read approach yielded the best assembly statistics, with the highest N50 and lowest number of contigs. The 40 Gbp short-read approach yielded the highest number of refined bins. Finally, the hybrid approach yielded the longest assemblies and the highest mapping rate to the bacterial genomes. Our results suggest that while long-read sequencing significantly improves the quality of reconstructed bacterial genomes, it is more expensive and requires deeper sequencing than short-read approaches to recover a comparable amount of reconstructed genomes. The most optimal strategy is study-specific and depends on how researchers assess the trade-off between the quantity and quality of recovered genomes.IMPORTANCEMice are an important model organism for understanding the gut microbiome. When studying these gut microbiomes using DNA techniques, researchers can choose from technologies that use short or long DNA reads. In this study, we perform an extensive benchmark between short- and long-read DNA sequencing for studying mice gut microbiomes. We find that no one approach was best for all metrics and provide information that can help guide researchers in planning their experiments.
shotgun 宏基因组学能够以高细节水平重建复杂的微生物群落。这种方法可以使用短读长和长读长测序数据以及两者的组合来进行。为了评估这些不同方法的优缺点，我们使用了从两只分别的实验鼠收集的每周 11 周的 22 份粪便 DNA 提取物，研究了四个测序深度和技术组合下的七个性能指标：（i）20 Gbp 的 Illumina 短读长数据，（ii）40 Gbp 的短读长数据，（iii）20 Gbp 的 PacBio HiFi 长读长数据，以及（iv）20 Gbp 的混合（20 Gbp 的短读长+20 Gbp 的长读长）数据。没有一种策略在所有指标上都是最佳的；相反，每种策略在不同的指标上表现出色。长读长方法产生了最佳的组装统计数据，具有最高的 N50 和最低的 contig 数量。40 Gbp 的短读长方法产生了最多的精细 bin。最后，混合方法产生了最长的组装和最高的细菌基因组映射率。我们的结果表明，虽然长读长测序显著提高了重建细菌基因组的质量，但它比短读长方法更昂贵，需要更深的测序才能恢复可比数量的重建基因组。最优化的策略是特定于研究的，取决于研究人员如何评估回收基因组的数量和质量之间的权衡。
小鼠是理解肠道微生物组的重要模型生物。当使用 DNA 技术研究这些肠道微生物组时，研究人员可以从使用短或长 DNA 读长的技术中进行选择。在这项研究中，我们对短读长和长读长 DNA 测序进行了广泛的基准测试，以研究小鼠肠道微生物组。我们发现，没有一种方法在所有指标上都是最佳的，并提供了可以帮助指导研究人员规划实验的信息。
