Folate, together with other B-vitamins, plays a crucial role in epigenetic regulation. Folate-mediated one-carbon metabolism refers to a complex network of interconnected metabolic pathways, which ultimately result in a supply of methyl groups for DNA, RNA or protein methylation. These epigenetic marks, particularly DNA methylation, have been proposed as plausible mechanisms underlying associations between folate and various disease outcomes, such as neural tube defects (NTDs), asthma, cardiometabolic disorders and cancer. However, evidence for the impact of folate on most of these disease outcomes, except for NTDs, is hampered by inconsistencies, with studies investigating the relationship between folate, DNA methylation and disease being subject to further complexities (i.e. identification of specific genomic loci affected, directions of differential methylation, and timing of epigenetic changes throughout the life course). Folate is closely related to other key role players in one-carbon metabolism (i.e. vitamin B12, homocysteine, methionine and choline) and therefore compensatory changes in interrelated metabolic pathways may account for inconsistent reports. Moreover, estimated folate intake does not necessarily correspond to circulating levels (correlations ranging from r=0.05–0.54), pointing towards methodological issues or other genetic, physiological or environmental factors determining folate levels. One particular aspect that remains unexplored is the contribution of bacterial folate biosynthesis to folate status and DNA methylation patterns.