In the course of our screening program for active compounds that induce cell morphological changes of Saccharomyces cerevisiae, the culture broth of an entomopathogenic fungus Metarhizium sp. fE61 exhibited a unique morphological phenotype. We conducted an activity-guided isolation from the fermentation broth of Metarhizium sp. fE61 to yield two new macrolide compounds named JBIR-19 (1) and -20 (2) as active substances. Their structures were determined to be 24-membered macrolide analogs containing a 2-aminoethyl phosphate ester on the basis of NMR and other spectroscopic data. Compounds 1 and 2 induced striking elongated morphology of S. cerevisiae at concentrations of 3.1 and 13 microM, but showed weak antiyeast activity at MICs of 200 and >200 microM, respectively.