Data_Sheet_1_Evaluation of Human Milk Microbiota by 16S rRNA Gene Next-Generation Sequencing (NGS) and Cultivation/MALDI-TOF Mass Spectrometry Identification.xlsx

The aim of the present study was to characterize human milk microbiota (HMM) with 16S rRNA gene amplicon next-generation sequencing and cultivation/matrix-assisted laser desorption/ionization (MALDI)-time of flight (TOF) mass spectrometry (MS) identification approaches. We analyzed 31 human milk samples from healthy Slovenian mothers. To check the accuracy of MALDI-TOF MS identification, several colonies representing most abundant genera and those, which could not be reliably identified by MALDI-TOF, were subjected to Sanger sequencing of their 16S rRNA gene. We showed that cultivation/MALDI-TOF MS was a suitable tool for culture-dependent determination of HMM. With both approaches, Staphylococcus and Streptococcus were found as predominant genera in HMM and the abundance of Staphylococcus was associated with decreased microbial diversity. In addition, we characterized factors that might influence HMM. The use of a breast pump was significantly associated with composition of HMM, higher microbial load, and lower abundance of cultivable staphylococci. Moreover, our study suggests that administration of probiotics to the suckling infant might influence HMM by increased abundance of lactobacilli and the presence of viable probiotic bacteria in human milk. However, since our study was observational with relatively small sample size, more targeted studies are needed to study possible transfer of probiotics to the mammary gland via an external route and the physiological relevance of these events.

本研究旨在运用16S rRNA基因扩增子高通量测序及培养/基质辅助激光解吸电离-飞行时间质谱（MALDI-TOF MS）鉴定方法对人类乳汁微生物组（HMM）进行表征。研究分析了31份来自健康斯洛文尼亚母亲的乳汁样本。为验证MALDI-TOF MS鉴定的准确性，对代表丰度较高的属以及无法通过MALDI-TOF MS可靠鉴定的菌株进行了其16S rRNA基因的Sanger测序。研究结果表明，培养/MALDI-TOF MS是依赖培养确定HMM的适宜工具。两种方法均发现葡萄球菌和链球菌为HMM中的优势属，其中葡萄球菌的丰度与微生物多样性降低相关。此外，我们还对可能影响HMM的因素进行了表征。使用吸奶器与HMM的组成、微生物负荷增加以及可培养葡萄球菌丰度降低显著相关。此外，本研究还表明，对哺乳婴儿补充益生菌可能通过增加乳酸杆菌的丰度和在人类乳汁中存在可存活的益生菌细菌来影响HMM。然而，鉴于本研究为观察性研究且样本量相对较小，仍需进行更针对性的研究，以探讨益生菌通过外部途径转移到乳腺的可能性及其生理学意义。