Next Generation Sequencing in Defining the Composition of Bacterial Microbiota of Donated Human Milk

Abstract

Purpose: Human milk microbiota can significantly influence children’s health. This study aimed to investigate the composition and diversity of the bacterial microbiota in donated human milk (DHM) and analyze the possible influencing factors. Methods: Archival samples from single-donor pools of raw DHM expressed from 88 different donors within the first six months postpartum were included. Donor-, infant-, and milk-related data were collected and analyzed using descriptive and comparative statistical methods. The hypervariable region V1–V3 of the bacterial 16S ribosomal ribonucleic acid gene was sequenced using the Illumina MiSeq platform to characterize the microbial profile of the donated milk. Results: Staphylococcus was the most abundant genus in all analyzed samples. The genus Lactobacillus was also present in all samples, with relative abundances ranging from 0.06– 62%. Bacteria of the genus Bifidobacterium were present in 62 (70% n=62) of the samples. A statistically significant difference in Lactobacillus abundance was observed based on the storage container type before pasteurization (p=0.015, median 0.804 in bottles vs. 0.289 in bags). Alpha diversity was higher in milk samples from exclusively breastfed infants, and in the samples that were expressed using manual techniques. Conclusion: This study provides evidence of the potential influence of donor infantrelated factors and milk characteristics on the composition and alpha diversity of bacterial microbiota in DHM. Notably, previous studies utilizing high-throughput sequencing technology to examine the human milk microbiota have not focused on pooled DHM samples.