开菲尔谷物由细菌和酵母的复杂共生混合物组成,据报道,牛奶和水开菲尔饮料具有许多增进健康的特性。这项工作的目的是研究开菲尔谷物中的微生物群落,并探索从中获得有用益生菌菌株的可能性。共158个微生物菌株,代表六种真菌和17种细菌,是从一家新加坡家庭酿酒师收集的牛奶和水开菲尔谷物中分离出来的。基于16SrRNA测序,分离的属包括乳酸菌,Liquorilactacillus,乳酸菌,慢乳酸杆菌,明串珠菌,乳球菌,醋杆菌,葡糖酸杆菌,肠球菌,梭菌属,发酵单胞菌,酵母菌,克鲁维酵母,Pichia,Lachancea,念珠菌,和Brettanomyces.为了表征这些分离株,漏斗法,涉及许多表型和基因组筛选试验,用于鉴定具有最高益生菌潜力的酸乳来源的微生物菌株。特别重点放在检查开菲尔分离株对肠道病原体的病原体抑制特性上,这些病原体构成了相当大的全球健康负担。测试的肠道病原体包括芽孢杆菌属,沙门氏菌,弧菌,梭菌属,克雷伯菌属,埃希氏菌,和葡萄球菌。进行了良好的扩散测定,以确定开菲尔分离株抑制肠道病原体生长的倾向,并使用竞争性粘附/排除测定法来确定开菲尔分离株竞争或排除肠病原体对Caco-2细胞的粘附的能力。七株扁乳杆菌,副干酪乳杆菌,萨umensisLiquorilactobacillus,瑞士乳杆菌,和酸奶慢乳杆菌,最终被鉴定为潜在的益生菌,并结合形成一个\"开菲尔益生菌混合物。\"理想的益生菌特性,包括在酸和胆汁环境中的良好生存,胆汁盐水解酶活性,抗氧化活性,非细胞毒性和对Caco-2细胞的高粘附性,缺乏毒力或抗菌素抗性基因。此外,维生素和γ-氨基丁酸(GABA)合成基因,在这些开菲尔分离株中被鉴定出来。总的来说,本研究中衍生的益生菌候选物是特征明确的菌株,具有良好的安全性,可以作为对抗肠道疾病的新型药物。这些源自开菲尔的益生菌也为现有的益生菌菌株增加了多样性,并可能为消费者提供替代产品形式,以获得开菲尔的健康益处。
Kefir grains consist of complex symbiotic mixtures of bacteria and yeasts, and are reported to impart numerous health-boosting properties to milk and water kefir beverages. The objective of this work was to investigate the microbial communities in kefir grains, and explore the possibility of deriving useful probiotic strains from them. A total of 158 microbial strains, representing six fungal and 17 bacterial species, were isolated from milk and water kefir grains collected from a Singapore-based homebrewer. Based on 16S rRNA sequencing, isolated genera included Lactobacillus, Liquorilactobacillus, Lacticaseibacillus, Lentilactobacillus, Leuconostoc, Lactococcus, Acetobacter, Gluconobacter, Oenococcus, Clostridium, Zymomonas, Saccharomyces, Kluyveromyces, Pichia, Lachancea, Candida, and Brettanomyces. To characterize these isolates, a funnel approach, involving numerous phenotypic and genomic screening assays, was applied to identify kefir-derived microbial strains with the highest probiotic potential. Particular focus was placed on examining the pathogen inhibitory properties of kefir isolates toward enteric pathogens which pose a considerable global health burden. Enteric pathogens tested include species of Bacillus, Salmonella, Vibrio, Clostridium, Klebsiella, Escherichia, and Staphylococcus. Well diffusion assays were conducted to determine the propensity of kefir isolates to inhibit growth of enteric pathogens, and a
competitive adhesion/exclusion assay was used to determine the ability of kefir isolates to out-compete or exclude attachment of enteric pathogens to Caco-2 cells. Seven bacterial strains of Lentilactobacillus hilgardii, Lacticaseibacillus paracasei, Liquorilactobacillus satsumensis, Lactobacillus helveticus, and Lentilactobacillus kefiri, were ultimately identified as potential probiotics, and combined to form a \"kefir probiotics blend.\" Desirable probiotic characteristics, including good survival in acid and bile environments, bile salt hydrolase activity, antioxidant activity, non-cytotoxicity and high adhesion to Caco-2 cells, and a lack of virulence or antimicrobial resistance genes. In addition, vitamin and γ-aminobutyric acid (GABA) synthesis genes, were identified in these kefir isolates. Overall, probiotic candidates derived in this study are well-characterized strains with a good safety profile which can serve as novel agents to combat enteric diseases. These kefir-derived probiotics also add diversity to the existing repertoire of probiotic strains, and may provide consumers with alternative product formats to attain the health benefits of kefir.