The genome of the insect pathogen Photorhabdus luminescens strain TT01 contains numerous genes predicting toxins and proteases. Within the P. luminescens TT01 genome, the products of two loci, plu 4093-plu 4092 and plu 4437-plu 4436, show oral insecticidal activity against both moth and mosquito larvae. The proteins encoded by these loci, here termed 'Photorhabdus insect related' (Pir) proteins A and B, show similarity both to delta-endotoxins from Bacillus thuringiensis (Bts) and a developmentally regulated protein from a beetle, Leptinotarsa decemlineata. The beetle protein has been inferred to possess juvenile hormone esterase (JHE) activity due to its developmentally regulated pattern of expression and the Photorhabdus proteins PirA and PirB have been proposed to be mimics of insect JHEs that can disrupt insect metamorphosis by metabolizing the insect growth regulator juvenile hormone (JH) [Nat. Biotecbnol. 21 (2003) 1307-1313]. Here we confirm that, when injected together, PirA and PirB from two different Photorhabdus strains have insecticidal activity against caterpillars of the moth Galleria mellonella but show no oral activity against a second moth species Manduca sexta. Direct measurement of JHE activity, however, shows that the Pir proteins are not able to metabolise JH. These data show that the Pir proteins have no JHE activity, as suggested, but leave the mode of action of these interesting proteins uncertain.