Background: Medically important pathogens are responsible for the death of millions every year. For many of these pathogens, there are limited options for therapy and resistance to commonly used drugs is fast emerging. The availability of genome sequences of many eukaryotic microbes is providing critical biological information for understanding parasite biology and identifying new drug and vaccine targets. Methods: We developed automated search strategies in the Eukaryotic Pathogen Database Resources (EuPathDB) to construct a protein list and retrieve protein sequences of folate transporters encoded in the genomes of 200 eukaryotic microbes. The folate transporters were categorized according to features including mitochondrial localization, number of transmembrane helix, and protein sequence relatedness. Results: We identified 234 folate transporter proteins associated with 63 eukaryotic microbes including 48 protozoa, 13 fungi the others being algae and bacteria. Phylogenetic analysis placed 219 proteins into a major clade and 15 proteins into a minor clade. All the folate transporter sequences from the malaria parasite, Plasmodium, belonged to the major clade. The identified folate transporters include folate-binding protein YgfZ, folate/pteridine transporter, folate/biopterin transporter, reduced folate carrier family protein and folate/methotrexate transporter FT1. About 60% of the identified proteins are reported for the first time. Phylogeny computation shows the similarity of the proteins identified. Conclusion: These findings offer new possibilities for potential drug development targeting folate-salvage proteins in eukaryotic pathogens.