The power of DNA based methods in probiotic authentication
IntroductionThe global probiotic market is growing rapidly, and strict quality control measures are required to ensure probiotic product efficacy and safety. Quality assurance of probiotic products involve confirming the presence of specific probiotic strains, determining the viable cell counts, and...
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2023-04-01
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| Series: | Frontiers in Microbiology |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fmicb.2023.1158440/full |
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| author | Hanan R. Shehata Hanan R. Shehata Steven G. Newmaster |
| author_facet | Hanan R. Shehata Hanan R. Shehata Steven G. Newmaster |
| author_sort | Hanan R. Shehata |
| collection | DOAJ |
| description | IntroductionThe global probiotic market is growing rapidly, and strict quality control measures are required to ensure probiotic product efficacy and safety. Quality assurance of probiotic products involve confirming the presence of specific probiotic strains, determining the viable cell counts, and confirming the absence of contaminant strains. Third-party evaluation of probiotic quality and label accuracy is recommended for probiotic manufacturers. Following this recommendation, multiple batches of a top selling multi-strain probiotic product were evaluated for label accuracy.MethodsA total of 55 samples (five multi-strain finished products and 50 single-strain raw ingredients) containing a total of 100 probiotic strains were evaluated using a combination of molecular methods including targeted PCR, non-targeted amplicon-based High Throughput Sequencing (HTS), and non-targeted Shotgun Metagenomic Sequencing (SMS).ResultsTargeted testing using species-specific or strain-specific PCR methods confirmed the identity of all strains/species. While 40 strains were identified to strain level, 60 strains were identified to species level only due to lack of strain-specific identification methods. In amplicon based HTS, two variable regions of 16S rRNA gene were targeted. Based on V5–V8 region data, ~99% of total reads per sample corresponded to target species, and no undeclared species were detected. Based on V3–V4 region data, ~95%–97% of total reads per sample corresponded to target species, while ~2%–3% of reads matched undeclared species (Proteus species), however, attempts to culture Proteus confirmed that all batches were free from viable Proteus species. Reads from SMS assembled to the genomes of all 10 target strains in all five batches of the finished product.DiscussionWhile targeted methods enable quick and accurate identification of target taxa in probiotic products, non-targeted methods enable the identification of all species in a product including undeclared species, with the caveats of complexity, high cost, and long time to result. |
| first_indexed | 2024-04-09T17:40:10Z |
| format | Article |
| id | doaj.art-f2bfcabd2b76455ca3864126800d4d6b |
| institution | Directory Open Access Journal |
| issn | 1664-302X |
| language | English |
| last_indexed | 2024-04-09T17:40:10Z |
| publishDate | 2023-04-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Microbiology |
| spelling | doaj.art-f2bfcabd2b76455ca3864126800d4d6b2023-04-17T06:01:23ZengFrontiers Media S.A.Frontiers in Microbiology1664-302X2023-04-011410.3389/fmicb.2023.11584401158440The power of DNA based methods in probiotic authenticationHanan R. Shehata0Hanan R. Shehata1Steven G. Newmaster2Natural Health Product Research Alliance, Department of Integrative Biology, College of Biological Science, University of Guelph, Guelph, ON, CanadaDepartment of Microbiology, Faculty of Pharmacy, Mansoura University, Mansoura, EgyptNatural Health Product Research Alliance, Department of Integrative Biology, College of Biological Science, University of Guelph, Guelph, ON, CanadaIntroductionThe global probiotic market is growing rapidly, and strict quality control measures are required to ensure probiotic product efficacy and safety. Quality assurance of probiotic products involve confirming the presence of specific probiotic strains, determining the viable cell counts, and confirming the absence of contaminant strains. Third-party evaluation of probiotic quality and label accuracy is recommended for probiotic manufacturers. Following this recommendation, multiple batches of a top selling multi-strain probiotic product were evaluated for label accuracy.MethodsA total of 55 samples (five multi-strain finished products and 50 single-strain raw ingredients) containing a total of 100 probiotic strains were evaluated using a combination of molecular methods including targeted PCR, non-targeted amplicon-based High Throughput Sequencing (HTS), and non-targeted Shotgun Metagenomic Sequencing (SMS).ResultsTargeted testing using species-specific or strain-specific PCR methods confirmed the identity of all strains/species. While 40 strains were identified to strain level, 60 strains were identified to species level only due to lack of strain-specific identification methods. In amplicon based HTS, two variable regions of 16S rRNA gene were targeted. Based on V5–V8 region data, ~99% of total reads per sample corresponded to target species, and no undeclared species were detected. Based on V3–V4 region data, ~95%–97% of total reads per sample corresponded to target species, while ~2%–3% of reads matched undeclared species (Proteus species), however, attempts to culture Proteus confirmed that all batches were free from viable Proteus species. Reads from SMS assembled to the genomes of all 10 target strains in all five batches of the finished product.DiscussionWhile targeted methods enable quick and accurate identification of target taxa in probiotic products, non-targeted methods enable the identification of all species in a product including undeclared species, with the caveats of complexity, high cost, and long time to result.https://www.frontiersin.org/articles/10.3389/fmicb.2023.1158440/fullprobioticauthenticationshotgun metagenomic sequencingamplicon-based high throughput sequencingstrain-specificspecies-specific |
| spellingShingle | Hanan R. Shehata Hanan R. Shehata Steven G. Newmaster The power of DNA based methods in probiotic authentication Frontiers in Microbiology probiotic authentication shotgun metagenomic sequencing amplicon-based high throughput sequencing strain-specific species-specific |
| title | The power of DNA based methods in probiotic authentication |
| title_full | The power of DNA based methods in probiotic authentication |
| title_fullStr | The power of DNA based methods in probiotic authentication |
| title_full_unstemmed | The power of DNA based methods in probiotic authentication |
| title_short | The power of DNA based methods in probiotic authentication |
| title_sort | power of dna based methods in probiotic authentication |
| topic | probiotic authentication shotgun metagenomic sequencing amplicon-based high throughput sequencing strain-specific species-specific |
| url | https://www.frontiersin.org/articles/10.3389/fmicb.2023.1158440/full |
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