The biocontrol potential of antagonistic bacteria and yeasts isolated from tomato phyllosphere and fructoplane was evaluated against fruit rot of tomato caused by Alternaria alternata. A total of 48 bacterial and 24 yeast isolates were obtained from field-grown tomato leaves/fruits collected from Barka, Oman. The antagonistic activities of these isolates were evaluated against A. alternata using in vitro dual culture assay. Among the 72 bacterial and yeast isolates tested, one yeast (Meyerozyma guilliermondii) and two bacteria (Enterobacter roggenkampii and Pseudomonas aeruginosa) showed inhibitory activity against A. alternata. Examination of hyphal morphology of A. alternata at the edge of inhibition zone using scanning electron microscopy revealed abnormalities such as twisting, shriveling, distortion and collapsing of hyphae. Postharvest treatment of tomato fruits with E. roggenkampii, P. aeruginosa and M. guilliermondii reduced the fruit rot lesion size by 60%, 57% and 50%, respectively, compared to untreated control. The antifungal metabolites produced by the antagonistic yeast M. guilliermondii were fractionated using thin-layer chromatography. Characterization of the bioactive fractions using gas chromatography-mass spectrometry revealed the presence of different types of alkanes, predominantly tricosane and pentacosane, suggesting that volatile compounds, especially alkanes, produced by M. guilliermondii might also be involved in the suppression of A. alternata.