Table_2_Frozen Mother’s Own Milk Can Be Used Effectively to Personalize Donor Human Milk.docx

Feeding preterm infants mother’s own milk (MOM) lowers rates of sepsis, decreases necrotizing enterocolitis, and shortens hospital stay. In the absence of freshly expressed MOM, frozen MOM (FMOM) is provided. When MOM is unavailable, preterm infants are often fed pasteurized donor human milk (DHM), rendering it devoid of beneficial bacteria. We have previously reported that when MOM is inoculated into DHM to restore the live microbiota [restored milk (RM)], a similar microbial diversity to MOM can be achieved. Yet, it is unknown if a similar diversity to MOM can be obtained when FMOM is inoculated into DHM. The goal of this study was to determine whether a similar microbial composition to MOM could be obtained when FMOM is used to personalize DHM. To this end, a fresh sample of MOM was obtained and divided into fresh and frozen fractions. MOM and FMOM were inoculated into DHM at different dilutions: MOM/FMOM 10% (RM/FRM10) and MOM/FMOM 30% (RM/FRM30) and incubated at 37°C. At different timepoints, culture-dependent and culture-independent techniques were performed. Similar microbiota expansion and alpha diversity were observed in MOM, RM10, and RM30 whether fresh or frozen milk was used as the inoculum. To evaluate if microbial expansion would result in an abnormal activation on the innate immune system, Caco-2 epithelial cells were exposed to RM/FRM to compare interleukin 8 levels with Caco-2 cells exposed to MOM or DHM. It was found that RM samples did not elicit a significant increase in IL-8 levels when compared to MOM or FMOM. These results suggest that FMOM can be used to inoculate DHM if fresh MOM is unavailable or limited in supply, allowing both fresh MOM and FMOM to be viable options in a microbial restoration strategy.

喂养早产儿母亲自体乳汁（MOM）能够降低败血症的发生率、减少坏死性小肠结肠炎的发病率，并缩短住院时间。在无法获取新鲜挤出MOM的情况下，提供冷冻MOM（FMOM）。当MOM不可用时，早产儿通常被喂养巴氏消毒的供体人乳（DHM），导致其缺乏有益菌群。我们此前曾报道，当MOM接种到DHM中以恢复活性微生物群[恢复乳（RM）]时，可以实现与MOM相似的微生物多样性。然而，当FMOM接种到DHM中时，是否能够获得与MOM相似的多样性尚不清楚。本研究的目标是确定是否可以通过使用FMOM个性化DHM来获得与MOM相似的微生物组成。为此，获取了一份新鲜MOM样本并将其分为新鲜和冷冻部分。MOM和FMOM以不同稀释度接种到DHM中：MOM/FMOM 10%（RM/FRM10）和MOM/FMOM 30%（RM/FRM30），并在37°C下培养。在不同的时间点，进行了依赖于培养和独立于培养的技术。无论使用新鲜或冷冻乳作为接种物，在MOM、RM10和RM30中均观察到相似的微生物群扩张和α多样性。为了评估微生物扩张是否会导致先天免疫系统的异常激活，将Caco-2上皮细胞暴露于RM/FRM中，并与暴露于MOM或DHM的Caco-2细胞比较白细胞介素8水平。发现与MOM或FMOM相比，RM样本并未引起IL-8水平的显著增加。这些结果表明，当新鲜MOM不可用或供应有限时，可以使用FMOM接种DHM，从而使新鲜MOM和FMOM都成为微生物恢复策略中的可行选择。