Gibberella zeae incites Fusarium head blight (FHB), a devastating disease that causes extensive yield and quality losses to wheat and barley. Of over 700 microbial strains obtained from wheat anthers, 54 were able to utilize tartaric acid as a carbon source when the compound was supplied as choline bitartrate in liquid culture. Four tartaric acid-utilizing and three nonutilizing strains reduced FHB in initial tests and were selected for further assays. Antagonists were effective against three different isolates of G. zeae when single wheat florets were inoculated with pathogen and antagonist inoculum. All seven antagonists increased 100-kernel weight when applied simultaneously with G. zeae isolate Z3639 (P ≤ 0.05). Bacillus strains AS 43.3 and AS 43.4 and Cryptococcus strain OH 182.9 reduced disease severity by >77, 93, and 56%, respectively. Five antagonists increased 100-kernel weight of plants inoculated with G. zeae isolate DAOM 180378. All antagonists except one increased 100-kernel weight, and four of seven antagonists reduced disease severity (P ≤ 0.05) when tested against G. zeae isolate Fg-9-96. In spray-inoculation experiments, Bacillus strains AS 43.3 and AS 43.4 and Cryptococcus strains OH 71.4 and OH 182.9 reduced disease severity, regardless of the sequence, timing, and concentration of inoculum application (P ≤ 0.05), though 100-kernel weight did not always increase when antagonists were applied 4 h after inoculum of G. zeae. Overall, 4 of 54 isolates that utilized tartaric acid in vitro were effective against G. zeae versus only 3 of 170 isolates tested that did not utilize tartaric acid (P ≤ 0.05, χ-square test of goodness of fit), demonstrating the potential benefit of prescreening candidate antagonists of FHB for their ability to utilize tartaric acid. Biological control shows promise as part of an integrated pest management program for managing FHB.