Similarity in milk microbiota in replicates

In this study, DNA from 44 bovine milk samples were extracted twice and the V3 –V4 region of the 16S rRNA gene was sequenced in two separate runs. The FASTQ files from the two data sets were run through the same bioinformatics pipeline using the same settings and results from the two data sets were compared. Milk samples collected maximally 2 h apart were used as replicate s and permitted comparisons to be made within the same run. Results show a significant difference in species richness between the two sequencing runs although Shannon and Simpson's diversity was the same. Multivariate analyses of all samples demonstrate that the sequencing run was a driver for varia tion. Direct comparison of similarity between samples and sequencing run showed an average similarity of 42%–45% depending on whether binary or abundance-based similarity indices were used. Within-run comparisons of milk samples collected maximally 2 h apart showed an average similarity of 39%– 47% depending on the similarity index used and that similarity differed significantly between runs. We conclude that repeated DNA extraction and sequencing significantly can affect the results of a low microbial biomass microbiota study.
Source: MicrobiologyOpen - Category: Microbiology Authors: Tags: COMMENTARY Source Type: research