Analysis of microbial composition and sharing in low-biomass human milk samples: a comparison of DNA isolation and sequencing techniques.
Journal
ISME communications
ISSN: 2730-6151
Titre abrégé: ISME Commun
Pays: England
ID NLM: 9918205372406676
Informations de publication
Date de publication:
09 Nov 2023
09 Nov 2023
Historique:
received:
25
05
2023
accepted:
26
10
2023
revised:
18
10
2023
medline:
10
11
2023
pubmed:
10
11
2023
entrez:
9
11
2023
Statut:
epublish
Résumé
Human milk microbiome studies are currently hindered by low milk bacterial/human cell ratios and often rely on 16S rRNA gene sequencing, which limits downstream analyses. Here, we aimed to find a method to study milk bacteria and assess bacterial sharing between maternal and infant microbiota. We tested four DNA isolation methods, two bacterial enrichment methods and three sequencing methods on mock communities, milk samples and negative controls. Of the four DNA isolation kits, the DNeasy PowerSoil Pro (PS) and MagMAX Total Nucleic Acid Isolation (MX) kits provided consistent 16S rRNA gene sequencing results with low contamination. Neither enrichment method substantially decreased the human metagenomic sequencing read-depth. Long-read 16S-ITS-23S rRNA gene sequencing biased the mock community composition but provided consistent results for milk samples, with little contamination. In contrast to 16S rRNA gene sequencing, 16S-ITS-23S rRNA gene sequencing of milk, infant oral, infant faecal and maternal faecal DNA from 14 mother-infant pairs provided sufficient resolution to detect significantly more frequent sharing of bacteria between related pairs compared to unrelated pairs. In conclusion, PS or MX kit-DNA isolation followed by 16S rRNA gene sequencing reliably characterises human milk microbiota, and 16S-ITS-23S rRNA gene sequencing enables studies of bacterial transmission in low-biomass samples.
Identifiants
pubmed: 37945978
doi: 10.1038/s43705-023-00325-6
pii: 10.1038/s43705-023-00325-6
pmc: PMC10636111
doi:
Types de publication
Journal Article
Langues
eng
Pagination
116Informations de copyright
© 2023. The Author(s).
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