Gut protease profiles of different instars of Helicoverpa armigera (Lepidoptera: Noctuidae)

Pest control strategies against Helicoverpa armigera (Hubner) using protease inhibitors have relied on the gut protease profile of the later larval stages of the insect, with serine proteases being considered predominant. Little is known about the gut protease profile of early larval instars. Therefore, the aim of the present study was to detect the levels of gut protease activities in the third-, fourth-, fifth- and late fifth-instar larvae of H. armigera reared on an artificial diet using specific substrates and inhibitors. The analysis of the gut protease profiles of different instars revealed different levels of protease activities at different instar stages. Significant variations were also observed in the specific activities of trypsin, chymotrypsin, cysteine protease, carboxypeptidase-A and amino-peptidase-N across the instars. In general, the activities of the proteases increased from the third to the fifth-instar and then decreased at the onset of pupation in the late fifth instar. Proteolytic activity was optimal at pH 12 for gut extracts from the third-, fourth- and fifth-instar larvae. Bioassays with phenylmethylsulphonyl fluoride (PMSF), a serine-cysteine protease inhibitor, revealed high mortality, and that with sodium ethylenediaminetetraacetic acid (EDTA-Na), a metalloprotease inhibitor, also showed retarded larval growth. The inhibition induced by 0.05% PMSF and 0.05% EDTA-Na in combination was similar to that induced by 0.1% PMSF alone.