In Silico Study Suggesting the Bias of Primers Choice in the Molecular Identification of Fungal Aerosols
This paper presents an in silico analysis to assess the current state of the fungal UNITE database in terms of the two eukaryote nuclear ribosomal regions, Internal Transcribed Spacers 1 and 2 (ITS1 and ITS2), used in describing fungal diversity. Microbial diversity is often evaluated with amplicon-...
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| Format: | Article |
| Language: | English |
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MDPI AG
2021-01-01
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| Series: | Journal of Fungi |
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| Online Access: | https://www.mdpi.com/2309-608X/7/2/99 |
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| author | Hamza Mbareche Marc Veillette Guillaume J. Bilodeau |
| author_facet | Hamza Mbareche Marc Veillette Guillaume J. Bilodeau |
| author_sort | Hamza Mbareche |
| collection | DOAJ |
| description | This paper presents an in silico analysis to assess the current state of the fungal UNITE database in terms of the two eukaryote nuclear ribosomal regions, Internal Transcribed Spacers 1 and 2 (ITS1 and ITS2), used in describing fungal diversity. Microbial diversity is often evaluated with amplicon-based high-throughput sequencing approaches, which is a target enrichment method that relies on the amplification of a specific target using particular primers before sequencing. Thus, the results are highly dependent on the quality of the primers used for amplification. The goal of this study is to validate if the mismatches of the primers on the binding sites of the targeted taxa could explain the differences observed when using either ITS1 or ITS2 in describing airborne fungal diversity. Hence, the choice of the pairs of primers for each barcode concur with a study comparing the performance of ITS1 and ITS2 in three occupational environments. The sequence length varied between the amplicons retrieved from the UNITE database using the pair of primers targeting ITS1 and ITS2. However, the database contains an equal number of unidentified taxa from ITS1 and ITS2 regions in the six taxonomic levels employed (phylum, class, order, family, genus, species). The chosen ITS primers showed differences in their ability to amplify fungal sequences from the UNITE database. Eleven taxa consisting of Trichocomaceae, Dothioraceae, Botryosphaeriaceae, Mucorales, Saccharomycetes, Pucciniomycetes, Ophiocordyceps, Microsporidia, Archaeorhizomycetes, Mycenaceae, and Tulasnellaceae showed large variations between the two regions. Note that members of the latter taxa are not all typical fungi found in the air. As no universal method is currently available to cover all the fungal kingdom, continuous work in designing primers, and particularly combining multiple primers targeting the ITS region is the best way to compensate for the biases of each one to get a larger view of the fungal diversity. |
| first_indexed | 2024-03-09T03:15:57Z |
| format | Article |
| id | doaj.art-6fbab9e8cce0406d9e94bbc92deda7ab |
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| issn | 2309-608X |
| language | English |
| last_indexed | 2024-03-09T03:15:57Z |
| publishDate | 2021-01-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Journal of Fungi |
| spelling | doaj.art-6fbab9e8cce0406d9e94bbc92deda7ab2023-12-03T15:21:25ZengMDPI AGJournal of Fungi2309-608X2021-01-01729910.3390/jof7020099In Silico Study Suggesting the Bias of Primers Choice in the Molecular Identification of Fungal AerosolsHamza Mbareche0Marc Veillette1Guillaume J. Bilodeau2Sunnybrook Research Institute, Toronto, ON M4N 3M5, CanadaCentre de Recherche de L’institut Universitaire de Cardiologie et de Pneumologie de Québec, Quebec City, QC G1V 4G5, CanadaPathogen Identification Research Lab, Canadian Food Inspection Agency (CFIA), Ottawa, ON K2H 8P9, CanadaThis paper presents an in silico analysis to assess the current state of the fungal UNITE database in terms of the two eukaryote nuclear ribosomal regions, Internal Transcribed Spacers 1 and 2 (ITS1 and ITS2), used in describing fungal diversity. Microbial diversity is often evaluated with amplicon-based high-throughput sequencing approaches, which is a target enrichment method that relies on the amplification of a specific target using particular primers before sequencing. Thus, the results are highly dependent on the quality of the primers used for amplification. The goal of this study is to validate if the mismatches of the primers on the binding sites of the targeted taxa could explain the differences observed when using either ITS1 or ITS2 in describing airborne fungal diversity. Hence, the choice of the pairs of primers for each barcode concur with a study comparing the performance of ITS1 and ITS2 in three occupational environments. The sequence length varied between the amplicons retrieved from the UNITE database using the pair of primers targeting ITS1 and ITS2. However, the database contains an equal number of unidentified taxa from ITS1 and ITS2 regions in the six taxonomic levels employed (phylum, class, order, family, genus, species). The chosen ITS primers showed differences in their ability to amplify fungal sequences from the UNITE database. Eleven taxa consisting of Trichocomaceae, Dothioraceae, Botryosphaeriaceae, Mucorales, Saccharomycetes, Pucciniomycetes, Ophiocordyceps, Microsporidia, Archaeorhizomycetes, Mycenaceae, and Tulasnellaceae showed large variations between the two regions. Note that members of the latter taxa are not all typical fungi found in the air. As no universal method is currently available to cover all the fungal kingdom, continuous work in designing primers, and particularly combining multiple primers targeting the ITS region is the best way to compensate for the biases of each one to get a larger view of the fungal diversity.https://www.mdpi.com/2309-608X/7/2/99bioaerosolsfungiITS1ITS2UNITEdatabase |
| spellingShingle | Hamza Mbareche Marc Veillette Guillaume J. Bilodeau In Silico Study Suggesting the Bias of Primers Choice in the Molecular Identification of Fungal Aerosols Journal of Fungi bioaerosols fungi ITS1 ITS2 UNITE database |
| title | In Silico Study Suggesting the Bias of Primers Choice in the Molecular Identification of Fungal Aerosols |
| title_full | In Silico Study Suggesting the Bias of Primers Choice in the Molecular Identification of Fungal Aerosols |
| title_fullStr | In Silico Study Suggesting the Bias of Primers Choice in the Molecular Identification of Fungal Aerosols |
| title_full_unstemmed | In Silico Study Suggesting the Bias of Primers Choice in the Molecular Identification of Fungal Aerosols |
| title_short | In Silico Study Suggesting the Bias of Primers Choice in the Molecular Identification of Fungal Aerosols |
| title_sort | in silico study suggesting the bias of primers choice in the molecular identification of fungal aerosols |
| topic | bioaerosols fungi ITS1 ITS2 UNITE database |
| url | https://www.mdpi.com/2309-608X/7/2/99 |
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