Identification of Bifidobacterium longum subsp. infantis ATCC 15697 surface proteins after growth in milk phospholipid-enriched medium

Bifidobacterium infantis is associated with the gut microbiota of breast-fed infants. B. infantis promotes intestinal barrier and immune function through several proposed mechanisms, including interactions between their surface polysaccharides, the host, and other gut microorganisms. Dairy foods and ingredients are some of the most conspicuous food-based niches for this species and may provide benefits for their delivery and efficacy in the gut. Milk phospholipid (MPL)-rich ingredients have been increasingly recognized for their versatile benefits to health, including interactions with the gut microbiota and intestinal cells. Therefore, our objective was to investigate the capacity for MPL to promote survival of B. infantis during simulated digestion and to modulate bacterial polysaccharide production. To achieve these aims, B. infantis was incubated with or without 0.5% MPL in de Man, Rogosa, and Sharpe (MRS) media at 37 °C under anaerobiosis. Survival across the oral, gastric, and intestinal phases using in vitro digestion was measured using plate count, along with adhesion to goblet-like intestinal cells. MPL increased B. infantis survival at the end of the intestinal phase by at least 7% and decreased adhesion to intestinal cells. The bacterial surface characteristics, which may contribute to these effects, were assessed by ζ-potential, changes in surface proteins using comparative proteomics, and production of bound polysaccharides. MPL decreased the surface charge of the bifidobacteria from –17 to –24 mV and increased a 50 kDa protein (3-fold) that appears to be involved in protection from stress. The production of bound polysaccharides was measured using FTIR, HPLC, and TEM imaging. These techniques all suggest an increase in bound polysaccharide production at least 1.7-fold in the presence of MPL. Our results show that MPL treatment is positively correlated with increased survival during simulated digestion, a stress resistance surface protein, and bound polysaccharide production of B. infantis, suggesting its use as a functional ingredient to enhance probiotic and postbiotic effects.

婴儿双歧杆菌（Bifidobacterium infantis）与母乳喂养婴儿的肠道微生物群密切相关。该菌可通过多种潜在机制发挥作用，包括其表面多糖与宿主及其他肠道微生物的相互作用，从而促进肠屏障功能与免疫功能。乳制品及相关配料是该菌最显著的食物来源生态位之一，有助于提升其在肠道中的递送效果与功效。富含牛乳磷脂（Milk phospholipid, MPL）的配料因其对健康的多种有益作用日益受到关注，这类作用包括与肠道微生物群及肠细胞的相互作用。因此，本研究旨在探究牛乳磷脂在模拟消化过程中提升婴儿双歧杆菌存活率、调节细菌多糖生成的能力。为达成上述研究目标，本研究将婴儿双歧杆菌于37℃厌氧条件下，在德-罗-沙（MRS）培养基中进行孵育，实验组添加0.5%的MPL，对照组不添加。采用平板菌落计数法结合体外消化模型，检测该菌在口腔、胃及肠道各消化阶段的存活率，并评估其对杯状样肠细胞的黏附能力。结果显示，MPL可使婴儿双歧杆菌在肠道阶段结束后的存活率提升至少7%，同时降低其对肠细胞的黏附能力。本研究通过ζ电位、比较蛋白质组学分析表面蛋白变化，以及结合型多糖定量检测，探究了可能介导上述效应的细菌表面特性。结果表明，MPL可将双歧杆菌的表面电荷从-17 mV降至-24 mV，并使一种分子量约50 kDa的蛋白表达量上调3倍，该蛋白可能参与细菌的应激保护过程。本研究采用傅里叶变换红外光谱（FTIR）、高效液相色谱（HPLC）及透射电子显微镜（TEM）成像技术检测结合型多糖的生成量，结果均显示，在MPL存在的条件下，结合型多糖的生成量至少提升1.7倍。综上，本研究结果表明，MPL处理与婴儿双歧杆菌的模拟消化存活率提升、抗应激表面蛋白表达上调及结合型多糖生成增加呈正相关，提示MPL可作为一种功能性配料，用于增强益生菌及后生元的应用功效。