Human Effective Microbes, Department of Advanced Convergence, Handong Global University, Pohang 37554, Gyeongbuk, South Korea.
Laboratory of Infection Oncology, Institute of Clinical Molecular Biology, Christian-Albrechts-University Kiel, University Hospital Schleswig Holstein, Schwanenweg 20, 24105 Kiel, Germany.
Folia Microbiol (Praha). 2024 Oct;69(5):1053-1068. doi: 10.1007/s12223-024-01142-7. Epub 2024 Feb 20.
The objective of this study was to isolate, identify, and assess the safety and functionality in vitro of putative probiotic bacterial strains. Isolation procedures were based on standard methods using elective and selective media. The isolates were identified by comparative 16S rRNA sequencing analysis while their safety was determined according to the safety tests recommended by the FAO/WHO such as antibiotic resistance, hemolysin, and biogenic amine production. Most of the isolates did not pass the in vitro safety tests; therefore, only Lactiplantibacillus plantarum (from ant intestine and cheese), Lacticaseibacillus paracasei (from goat milk and kimchi), Enterococcus faecium (from chili doenjang and vegetables with kimchi ingredients), Limosilactobacillus fermentum (from saliva), and Companilactobacillus alimentarius (from kimchi) were identified and selected for further studies. The isolates were further differentiated by rep-PCR and identified to the strain level by genotypic (16S rRNA) and phenotypic (Gen III) approaches. Subsequently, the strain tolerance to acid and bile was evaluated resulting in good viability after simulated gastrointestinal tract passage. Adhesion to mucin in vitro and the presence of mub, mapA, and ef-tu genes confirmed the adhesive potential of the strains and the results of features associated with adhesion such as hydrophobicity and zeta potential extended the insights. This study reflects the importance of fermented and non-fermented food products as a promising source of lactic acid bacteria with potential probiotic properties. Additionally, it aims to highlight the challenges associated with the selection of safe strains, which often fail in the in vitro tests, thus hindering the possibilities of "uncovering" novel and safe probiotic strains.
本研究的目的是分离、鉴定并评估潜在益生菌菌株的安全性和功能。分离程序基于使用选择性和选择性培养基的标准方法。通过比较 16S rRNA 测序分析来鉴定分离株，而其安全性则根据 FAO/WHO 推荐的安全性测试来确定，如抗生素耐药性、溶血素和生物胺产生。大多数分离株未能通过体外安全性测试；因此，仅鉴定和选择了植物乳杆菌（来自蚂蚁肠道和奶酪）、副干酪乳杆菌（来自山羊奶和泡菜）、屎肠球菌（来自辣椒粉味噌和含有泡菜成分的蔬菜）、发酵乳杆菌（来自唾液）和唾液乳杆菌（来自泡菜）进行进一步研究。通过 rep-PCR 进一步区分分离株，并通过基因型（16S rRNA）和表型（Gen III）方法鉴定到菌株水平。随后，评估了菌株对酸和胆汁的耐受性，结果表明在模拟胃肠道通过后具有良好的生存能力。体外黏附到粘蛋白以及 mub、mapA 和 ef-tu 基因的存在证实了菌株的黏附潜力，并且与黏附相关的特征（如疏水性和 ζ 电位）的结果扩展了这些见解。本研究反映了发酵和非发酵食品作为具有潜在益生菌特性的乳酸菌的有前途的来源的重要性。此外，它旨在强调选择安全菌株所面临的挑战，这些菌株通常在体外测试中失败，从而阻碍了“发现”新型安全益生菌菌株的可能性。
