Revealing within-species diversity in uncultured human gut bacteria with single-cell long-read sequencing
Obtaining complete and accurate bacterial genomes is vital for studying the characteristics of uncultured bacteria. Single-cell genomics is a promising approach for the culture-independent recovery of bacterial genomes from individual cells. However, single-amplified genomes (SAGs) often have fragme...
| Main Authors: | , , , , , , |
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2023-02-01
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| Series: | Frontiers in Microbiology |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fmicb.2023.1133917/full |
| _version_ | 1797895436397182976 |
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| author | Masato Kogawa Yohei Nishikawa Yohei Nishikawa Tatsuya Saeki Takuya Yoda Koji Arikawa Haruko Takeyama Haruko Takeyama Haruko Takeyama Haruko Takeyama Masahito Hosokawa Masahito Hosokawa Masahito Hosokawa Masahito Hosokawa Masahito Hosokawa |
| author_facet | Masato Kogawa Yohei Nishikawa Yohei Nishikawa Tatsuya Saeki Takuya Yoda Koji Arikawa Haruko Takeyama Haruko Takeyama Haruko Takeyama Haruko Takeyama Masahito Hosokawa Masahito Hosokawa Masahito Hosokawa Masahito Hosokawa Masahito Hosokawa |
| author_sort | Masato Kogawa |
| collection | DOAJ |
| description | Obtaining complete and accurate bacterial genomes is vital for studying the characteristics of uncultured bacteria. Single-cell genomics is a promising approach for the culture-independent recovery of bacterial genomes from individual cells. However, single-amplified genomes (SAGs) often have fragmented and incomplete sequences due to chimeric and biased sequences introduced during the genome amplification process. To address this, we developed a single-cell amplified genome long-read assembly (scALA) workflow to construct complete circular SAGs (cSAGs) from long-read single-cell sequencing data of uncultured bacteria. We used the SAG-gel platform, which is both cost-effective and high-throughput, to obtain hundreds of short-read and long-read sequencing data for specific bacterial strains. The scALA workflow generated cSAGs by repeated in silico processing for sequence bias reduction and contig assembly. From 12 human fecal samples, including two cohabitant groups, scALA generated 16 cSAGs of three specifically targeted bacterial species: Anaerostipes hadrus, Agathobacter rectalis, and Ruminococcus gnavus. We discovered strain-specific structural variations shared among cohabiting hosts, while all cSAGs of the same species showed high homology in aligned genomic regions. A. hadrus cSAGs exhibited 10 kbp-long phage insertions, various saccharide metabolic capabilities, and different CRISPR-Cas systems in each strain. The sequence similarity of A. hadrus genomes did not necessarily correspond with orthologous functional genes, while host geographical regionality seemed to be highly related to gene possession. scALA allowed us to obtain closed circular genomes of specifically targeted bacteria from human microbiota samples, leading to an understanding of within-species diversities, including structural variations and linking mobile genetic elements, such as phages, to hosts. These analyses provide insight into microbial evolution, the adaptation of the community to environmental changes, and interactions with hosts. cSAGs constructed using this method can expand bacterial genome databases and our understanding of within-species diversities in uncultured bacteria. |
| first_indexed | 2024-04-10T07:26:49Z |
| format | Article |
| id | doaj.art-d4ebadf512a94c478d636f39ea0729a2 |
| institution | Directory Open Access Journal |
| issn | 1664-302X |
| language | English |
| last_indexed | 2024-04-10T07:26:49Z |
| publishDate | 2023-02-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Microbiology |
| spelling | doaj.art-d4ebadf512a94c478d636f39ea0729a22023-02-24T07:10:36ZengFrontiers Media S.A.Frontiers in Microbiology1664-302X2023-02-011410.3389/fmicb.2023.11339171133917Revealing within-species diversity in uncultured human gut bacteria with single-cell long-read sequencingMasato Kogawa0Yohei Nishikawa1Yohei Nishikawa2Tatsuya Saeki3Takuya Yoda4Koji Arikawa5Haruko Takeyama6Haruko Takeyama7Haruko Takeyama8Haruko Takeyama9Masahito Hosokawa10Masahito Hosokawa11Masahito Hosokawa12Masahito Hosokawa13Masahito Hosokawa14Research Organization for Nano and Life Innovation, Waseda University, Tokyo, JapanResearch Organization for Nano and Life Innovation, Waseda University, Tokyo, JapanComputational Bio Big-Data Open Innovation Laboratory, National Institute of Advanced Industrial Science and Technology, Tokyo, JapanbitBiome, Inc., Tokyo, JapanbitBiome, Inc., Tokyo, JapanbitBiome, Inc., Tokyo, JapanResearch Organization for Nano and Life Innovation, Waseda University, Tokyo, JapanComputational Bio Big-Data Open Innovation Laboratory, National Institute of Advanced Industrial Science and Technology, Tokyo, JapanDepartment of Life Science and Medical Bioscience, Waseda University, Tokyo, JapanInstitute for Advanced Research of Biosystem Dynamics, Waseda Research Institute for Science and Engineering, Tokyo, JapanResearch Organization for Nano and Life Innovation, Waseda University, Tokyo, JapanComputational Bio Big-Data Open Innovation Laboratory, National Institute of Advanced Industrial Science and Technology, Tokyo, JapanbitBiome, Inc., Tokyo, JapanDepartment of Life Science and Medical Bioscience, Waseda University, Tokyo, JapanInstitute for Advanced Research of Biosystem Dynamics, Waseda Research Institute for Science and Engineering, Tokyo, JapanObtaining complete and accurate bacterial genomes is vital for studying the characteristics of uncultured bacteria. Single-cell genomics is a promising approach for the culture-independent recovery of bacterial genomes from individual cells. However, single-amplified genomes (SAGs) often have fragmented and incomplete sequences due to chimeric and biased sequences introduced during the genome amplification process. To address this, we developed a single-cell amplified genome long-read assembly (scALA) workflow to construct complete circular SAGs (cSAGs) from long-read single-cell sequencing data of uncultured bacteria. We used the SAG-gel platform, which is both cost-effective and high-throughput, to obtain hundreds of short-read and long-read sequencing data for specific bacterial strains. The scALA workflow generated cSAGs by repeated in silico processing for sequence bias reduction and contig assembly. From 12 human fecal samples, including two cohabitant groups, scALA generated 16 cSAGs of three specifically targeted bacterial species: Anaerostipes hadrus, Agathobacter rectalis, and Ruminococcus gnavus. We discovered strain-specific structural variations shared among cohabiting hosts, while all cSAGs of the same species showed high homology in aligned genomic regions. A. hadrus cSAGs exhibited 10 kbp-long phage insertions, various saccharide metabolic capabilities, and different CRISPR-Cas systems in each strain. The sequence similarity of A. hadrus genomes did not necessarily correspond with orthologous functional genes, while host geographical regionality seemed to be highly related to gene possession. scALA allowed us to obtain closed circular genomes of specifically targeted bacteria from human microbiota samples, leading to an understanding of within-species diversities, including structural variations and linking mobile genetic elements, such as phages, to hosts. These analyses provide insight into microbial evolution, the adaptation of the community to environmental changes, and interactions with hosts. cSAGs constructed using this method can expand bacterial genome databases and our understanding of within-species diversities in uncultured bacteria.https://www.frontiersin.org/articles/10.3389/fmicb.2023.1133917/fullgutmicrobiomesingle-cell genomicslong-read sequencinggenome |
| spellingShingle | Masato Kogawa Yohei Nishikawa Yohei Nishikawa Tatsuya Saeki Takuya Yoda Koji Arikawa Haruko Takeyama Haruko Takeyama Haruko Takeyama Haruko Takeyama Masahito Hosokawa Masahito Hosokawa Masahito Hosokawa Masahito Hosokawa Masahito Hosokawa Revealing within-species diversity in uncultured human gut bacteria with single-cell long-read sequencing Frontiers in Microbiology gut microbiome single-cell genomics long-read sequencing genome |
| title | Revealing within-species diversity in uncultured human gut bacteria with single-cell long-read sequencing |
| title_full | Revealing within-species diversity in uncultured human gut bacteria with single-cell long-read sequencing |
| title_fullStr | Revealing within-species diversity in uncultured human gut bacteria with single-cell long-read sequencing |
| title_full_unstemmed | Revealing within-species diversity in uncultured human gut bacteria with single-cell long-read sequencing |
| title_short | Revealing within-species diversity in uncultured human gut bacteria with single-cell long-read sequencing |
| title_sort | revealing within species diversity in uncultured human gut bacteria with single cell long read sequencing |
| topic | gut microbiome single-cell genomics long-read sequencing genome |
| url | https://www.frontiersin.org/articles/10.3389/fmicb.2023.1133917/full |
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