Inter- and intra-specific variation in destruxin production was detected in Metarhizium and may be important in determining virulence and/or specificity against insects. Strains of M. anisopliae var. anisopliae produced different amounts of destruxins A, B and E, but strain V220 did not produce any destruxins. M. anisopliae var. majus, M. flavoviride and M. album which are reported to be specific towards Coleoptera, Orthoptera and Hemiptera, respectively, had different destruxin profiles with destruxin A predominant. In time course studies on M. anisopliae var. anisopliae V245, destruxin E levels declined with time while destruxin A levels increased. The most virulent strains, Ma23 and V245, produced large quantities of destruxins but some low toxin producers were also virulent, suggesting that destruxins are not the only pathogenicity determinants. Some weakly to moderately pathogenic strains were highly pathogenic when injected into Galleria mellonella larvae, demonstrating the importance of the cuticle as a barrier to fungal infection. Only trace amounts of destruxin A or a combination of A and B could be detected in Galleria larvae infected with M. anisopoliae var. anisopliae or M. anisopliae var. majus. No destruxins were detected in larvae infected with M. flavoviride. Destruxin production may be influenced by the nutrients in insects or culture media.