{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,20]],"date-time":"2026-08-20T15:24:34Z","timestamp":1787239474998,"version":"build-2736575974"},"reference-count":34,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2026,8,20]],"date-time":"2026-08-20T00:00:00Z","timestamp":1787184000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["frontiersin.org"],"crossmark-restriction":true},"short-container-title":["Front. Nutr."],"abstract":"\n Background<\/jats:title>\n Folate deficiency is a global nutritional problem associated with multiple adverse health outcomes, including impaired one-carbon metabolism and elevated homocysteine levels (hyperhomocysteinemia). Gut microbiota-mediated folate biosynthesis has emerged as a promising strategy for improving the host\u2019s folate status. This study aimed to isolate folate-producing probiotic strains, clarify their folate synthesis mechanisms, and evaluate their regulatory effects on folate metabolism and gut microbiota.<\/jats:p>\n <\/jats:sec>\n \n Methods<\/jats:title>\n High-throughput cultivation and screening were performed to isolate folate-producing candidate probiotics. Whole-genome sequencing analysis, pathway reconstruction, and metabolite profiling in fermented milk were performed to explore folate biosynthesis pathways and microbial cross-feeding interactions. A folate-deficient mouse model was established to evaluate the effects of a candidate probiotic cocktail on serum folate, homocysteine (Hcy) levels, and gut microbiota composition using quantitative PCR (qPCR) and 16S rRNA gene sequencing.<\/jats:p>\n <\/jats:sec>\n \n Results<\/jats:title>\n \n High-throughput screening identified 8 high-folate-producing candidate probiotic strains, including\n Lactiplantibacillus plantarum<\/jats:italic>\n and\n Heyndrickxia coagulans<\/jats:italic>\n , from over 1,000 isolates. Genomic analysis revealed that most commonly used probiotics lacked para-aminobenzoic acid (pABA) biosynthesis genes but retained downstream modules, suggesting a reliance on cross-feeding with pABA-producing gut commensals such as Bacteroides. Metabolite profiling of fermented milk demonstrated that selected strains significantly increased bioactive 5-methyltetrahydrofolate (5-MeTHF) and tetrahydrofolate levels.\n In vivo<\/jats:italic>\n , only a high-dose candidate probiotic cocktail significantly elevated serum folate (\n p<\/jats:italic>\n \u202f&lt;\u202f0.05) and reduced homocysteine levels (\n p<\/jats:italic>\n \u202f&lt;\u202f0.05) in deficient mice. Fecal qPCR confirmed dose-dependent transient persistence of the administered bacterial species. Consistent with the qPCR data, 16S rRNA gene sequences demonstrated significant enrichment of these administered species observed in the high-dose group. Furthermore, beta-diversity analysis found that high-dose candidate probiotic supplementation promoted a shift in the gut microbiota composition toward a normal profile, partially mitigating the dysbiosis induced by the folate-deficient diet. This effect was accompanied by a significant enrichment of potential short-chain fatty acid producers (e.g.,\n Lachnospiraceae<\/jats:italic>\n and\n Oscillospiraceae<\/jats:italic>\n ) and the depletion of potential opportunistic pathogens.\n <\/jats:p>\n <\/jats:sec>\n \n Conclusion<\/jats:title>\n This study screened high-folate-producing candidate probiotic strains and demonstrated their ability to synthesize the active form of 5-MeTHF. Moreover, folate-producing candidate probiotic cocktail treatment significantly improved folate status and Hcy metabolism and modulated the gut microbiota by enriching potential beneficial bacterial taxa. These findings suggested that folate-producing probiotics may serve as a promising microbiota-based strategy to improve folate availability and homocysteine metabolism.<\/jats:p>\n <\/jats:sec>","DOI":"10.3389\/fnut.2026.1845105","type":"journal-article","created":{"date-parts":[[2026,8,20]],"date-time":"2026-08-20T14:32:13Z","timestamp":1787236333000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":0,"title":["Isolation of folate-producing probiotic candidates and their effects on homocysteine metabolism and gut microbiota composition"],"prefix":"10.3389","volume":"13","author":[{"given":"Mingfang","family":"Pan","sequence":"first","affiliation":[{"name":"Institute of Biopharmaceutical and Health Engineering, Shenzhen International Graduate School, Tsinghua University","place":["Shenzhen, China"]},{"name":"Shenzhen Evergreen Medical Institute","place":["Shenzhen, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Changming","family":"Ye","sequence":"additional","affiliation":[{"name":"Shenzhen Evergreen Medical Institute","place":["Shenzhen, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yun","family":"Song","sequence":"additional","affiliation":[{"name":"Shenzhen Evergreen Medical Institute","place":["Shenzhen, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Minqing","family":"Tian","sequence":"additional","affiliation":[{"name":"Shenzhen Evergreen Medical Institute","place":["Shenzhen, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Runming","family":"Wang","sequence":"additional","affiliation":[{"name":"Institute of Biopharmaceutical and Health Engineering, Shenzhen International Graduate School, Tsinghua University","place":["Shenzhen, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ping","family":"Chen","sequence":"additional","affiliation":[{"name":"Inspection and Testing Center, Key Laboratory of Cancer FSMP for State Market Regulation","place":["Shenzhen, China"]},{"name":"AUSA Research Institute, Shenzhen AUSA Pharmed Co., Ltd.","place":["Shenzhen, China"]},{"name":"Precision Nutrition Innovation & Transformation Public Service Platform","place":["Shenzhen, China"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1965","published-online":{"date-parts":[[2026,8,20]]},"reference":[{"key":"ref1","doi-asserted-by":"publisher","first-page":"2867","DOI":"10.3390\/nu12092867","article-title":"B vitamins and one-carbon metabolism: implications in human health and disease","volume":"12","author":"Lyon","year":"2020","journal-title":"Nutrients"},{"key":"ref2","doi-asserted-by":"publisher","first-page":"66","DOI":"10.2174\/1573403X19666230209111854","article-title":"Homocysteine, 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