Optimizing microbiome reference databases with PacBio full-length 16S rRNA sequencing for enhanced taxonomic classification and biomarker discovery
BackgroundThe study of the human microbiome is crucial for understanding disease mechanisms, identifying biomarkers, and guiding preventive measures. Advances in sequencing platforms, particularly 16S rRNA sequencing, have revolutionized microbiome research. Despite the benefits, large microbiome re...
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Frontiers Media S.A.
2024-11-01
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| Series: | Frontiers in Microbiology |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fmicb.2024.1485073/full |
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| author | Hyejung Han Yoon Hee Choi Si Yeong Kim Jung Hwa Park Jin Chung Hee Sam Na |
| author_facet | Hyejung Han Yoon Hee Choi Si Yeong Kim Jung Hwa Park Jin Chung Hee Sam Na |
| author_sort | Hyejung Han |
| collection | DOAJ |
| description | BackgroundThe study of the human microbiome is crucial for understanding disease mechanisms, identifying biomarkers, and guiding preventive measures. Advances in sequencing platforms, particularly 16S rRNA sequencing, have revolutionized microbiome research. Despite the benefits, large microbiome reference databases (DBs) pose challenges, including computational demands and potential inaccuracies. This study aimed to determine if full-length 16S rRNA sequencing data produced by PacBio could be used to optimize reference DBs and be applied to Illumina V3-V4 targeted sequencing data for microbial study.MethodsOral and gut microbiome data (PRJNA1049979) were retrieved from NCBI. DADA2 was applied to full-length 16S rRNA PacBio data to obtain amplicon sequencing variants (ASVs). The RDP reference DB was used to assign the ASVs, which were then used as a reference DB to train the classifier. QIIME2 was used for V3-V4 targeted Illumina data analysis. BLAST was used to analyze alignment statistics. Linear discriminant analysis Effect Size (LEfSe) was employed for discriminant analysis.ResultsASVs produced by PacBio showed coverage of the oral microbiome similar to the Human Oral Microbiome Database. A phylogenetic tree was trimmed at various thresholds to obtain an optimized reference DB. This established method was then applied to gut microbiome data, and the optimized gut microbiome reference DB provided improved taxa classification and biomarker discovery efficiency.ConclusionFull-length 16S rRNA sequencing data produced by PacBio can be used to construct a microbiome reference DB. Utilizing an optimized reference DB can increase the accuracy of microbiome classification and enhance biomarker discovery. |
| format | Article |
| id | doaj-art-09a560aaa02c4f9ca80fe6bb679efda4 |
| institution | Kabale University |
| issn | 1664-302X |
| language | English |
| publishDate | 2024-11-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Microbiology |
| spelling | doaj-art-09a560aaa02c4f9ca80fe6bb679efda42024-11-25T06:24:11ZengFrontiers Media S.A.Frontiers in Microbiology1664-302X2024-11-011510.3389/fmicb.2024.14850731485073Optimizing microbiome reference databases with PacBio full-length 16S rRNA sequencing for enhanced taxonomic classification and biomarker discoveryHyejung Han0Yoon Hee Choi1Si Yeong Kim2Jung Hwa Park3Jin Chung4Hee Sam Na5Department of Oral Microbiology, School of Dentistry, Pusan National University, Yangsan, Republic of KoreaDepartment of Internal Medicine, Dongnam Institute of Radiological and Medical Sciences, Busan, Republic of KoreaDepartment of Oral Microbiology, School of Dentistry, Pusan National University, Yangsan, Republic of KoreaDepartment of Oral Microbiology, School of Dentistry, Pusan National University, Yangsan, Republic of KoreaDepartment of Oral Microbiology, School of Dentistry, Pusan National University, Yangsan, Republic of KoreaDepartment of Oral Microbiology, School of Dentistry, Pusan National University, Yangsan, Republic of KoreaBackgroundThe study of the human microbiome is crucial for understanding disease mechanisms, identifying biomarkers, and guiding preventive measures. Advances in sequencing platforms, particularly 16S rRNA sequencing, have revolutionized microbiome research. Despite the benefits, large microbiome reference databases (DBs) pose challenges, including computational demands and potential inaccuracies. This study aimed to determine if full-length 16S rRNA sequencing data produced by PacBio could be used to optimize reference DBs and be applied to Illumina V3-V4 targeted sequencing data for microbial study.MethodsOral and gut microbiome data (PRJNA1049979) were retrieved from NCBI. DADA2 was applied to full-length 16S rRNA PacBio data to obtain amplicon sequencing variants (ASVs). The RDP reference DB was used to assign the ASVs, which were then used as a reference DB to train the classifier. QIIME2 was used for V3-V4 targeted Illumina data analysis. BLAST was used to analyze alignment statistics. Linear discriminant analysis Effect Size (LEfSe) was employed for discriminant analysis.ResultsASVs produced by PacBio showed coverage of the oral microbiome similar to the Human Oral Microbiome Database. A phylogenetic tree was trimmed at various thresholds to obtain an optimized reference DB. This established method was then applied to gut microbiome data, and the optimized gut microbiome reference DB provided improved taxa classification and biomarker discovery efficiency.ConclusionFull-length 16S rRNA sequencing data produced by PacBio can be used to construct a microbiome reference DB. Utilizing an optimized reference DB can increase the accuracy of microbiome classification and enhance biomarker discovery.https://www.frontiersin.org/articles/10.3389/fmicb.2024.1485073/fulloral microbiomegut microbiomePacBioIlluminanext generation sequencingreference database |
| spellingShingle | Hyejung Han Yoon Hee Choi Si Yeong Kim Jung Hwa Park Jin Chung Hee Sam Na Optimizing microbiome reference databases with PacBio full-length 16S rRNA sequencing for enhanced taxonomic classification and biomarker discovery Frontiers in Microbiology oral microbiome gut microbiome PacBio Illumina next generation sequencing reference database |
| title | Optimizing microbiome reference databases with PacBio full-length 16S rRNA sequencing for enhanced taxonomic classification and biomarker discovery |
| title_full | Optimizing microbiome reference databases with PacBio full-length 16S rRNA sequencing for enhanced taxonomic classification and biomarker discovery |
| title_fullStr | Optimizing microbiome reference databases with PacBio full-length 16S rRNA sequencing for enhanced taxonomic classification and biomarker discovery |
| title_full_unstemmed | Optimizing microbiome reference databases with PacBio full-length 16S rRNA sequencing for enhanced taxonomic classification and biomarker discovery |
| title_short | Optimizing microbiome reference databases with PacBio full-length 16S rRNA sequencing for enhanced taxonomic classification and biomarker discovery |
| title_sort | optimizing microbiome reference databases with pacbio full length 16s rrna sequencing for enhanced taxonomic classification and biomarker discovery |
| topic | oral microbiome gut microbiome PacBio Illumina next generation sequencing reference database |
| url | https://www.frontiersin.org/articles/10.3389/fmicb.2024.1485073/full |
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