Intestinal proteolytic profile changes during larval development of Anticarsia gemmatalis caterpillars

Abstract

Soybean is one of most consumed and produced grains in the world, and Anticarsia gemmatalis is a pest that causes great damage to this crop due to severe defoliation during its larval phase. Plants have mechanisms that lead to the inhibition of proteases in the intestine of these herbivores, hampering their development. Understanding this complex protease inhibitor is important for pest control. The objective of this study was to evaluate the enzymatic profiles of the intestinal proteases of the soybean caterpillar at different instars. For this, the proteolytic profile of the gut in the third, fourth, and fifth instars were analyzed.Irreversible inhibitors of proteases were separately incubated with A. gemmatalis enzyme extracts at the third, fourth, and fifth instar to assess the contribution of these proteases to total proteolytic activity. The enzymatic extracts were also evaluated with specific substrates to confirm changes in the specific activities of trypsin-like, chymotrypsin-like, and cysteine proteases at different instars. The results showed that the protease profile of A.gemmatalis gut changes throughout its larval development.The activity of cysteine proteases was more intense in the first instar. On the contrary, the serine proteases The protease profile of the gut of A. gemmatalis changes throughout larval development. In the third instar, cysteine protease activity is predominant over others. In the fourth and fifth instars, proteolytic activity is mainly due to serine proteases. This difference in activity in the different instars of A. gemmatalis suggests that the expression of the genes coding for proteases also changes throughout the development of the insect. Using serine PIs is a good strategy for controlling insect pests, as these are the main proteolytic enzymes of Lepidoptera. However, the presence of other important protease subclasses in insect digestibility suggests that a combination of serine and cysteine PIs may be promising, aiming to compromise larval development at different instars.

Small peptides inhibit gut trypsin‐like proteases and impairAnticarsia gemmatalis(Lepidoptera:Noctuidae) survival and development

“…A. gemmatalis larvae fed on diets containing the peptides GORE1 and GORE2 showed increased mortality, explained by the fact that they inhibit the main digestive protease (i.e. trypsin‐like) of lepidopteran insects 16,54 . The inhibition of digestive proteases negatively affects protein digestion, causing amino acid deficiency and inefficient protein recycling, which destabilizes larval metabolism and confers insecticidal activity to protease inhibitors 5 .…”

“…2.1 In silico docking of peptides and trypsins of Lepidoptera 2.1.1 Sequences and tridimensional structure prediction The amino acid sequences were obtained by translating trypsin genes identified by zymogram analyses and mass spectrometry, 16 using the Expasy tool. 27 Trypsin protein sequences from other Lepidopteran species (Helicoperva armigera, Manduca sexta, and Spodoptera frugiperda) were used to compare the binding site conservation with the analyzed peptides.…”

“…The total sample volume perchromatography was 2 mL of enzyme extract containing the Table 1. Trypsin-like sequences of Lepidoptera pests obtained from the NCBI database (Anticarsia gemmatalis 1, Anticarsia gemmatalis 2, Helicoperva armigera, Manduca sexta and Spodoptera frugiperda) and identified by Da Silva Junior et al 2019 (Anticarsia gemmatalis 3-6) 16 GenBank Species…”

“…fourth, fifth and sixth) of lepidopteran insects, where 90% of insect feeding occurs. [14][15][16] Exposure of lepidopteran insect larvae to soybean protease inhibitors such as soybean kunitz trypsin inhibitor (SKTI) and soybean Bowman-Birk inhibitor (SBBI) decreases the activity of trypsin-like and chymotrypsin-like enzymes, affecting total proteolytic activity in the midgut. 17,18 However, although protease inhibitors impair insect development, co-evolution with plants has allowed herbivores to develop adaptative mechanisms against these particular molecules.…”

Noncompetitive tight‐binding inhibition ofAnticarsia gemmatalistrypsins byAdenanthera pavoninaprotease inhibitor affects larvae survival

“…The soybean caterpillar Anticarsia gemmatalis Hübner (Lepidoptera: Erebidae) is the main defoliation pest of soybeans that can cause 100% of leaf loss (da Silva‐Júnior et al, 2020; Moscardi et al, 2012). The economic loss in soybean crops caused by the Lepidoptera insect A. gemmatalis has encouraged the search for strategies of pest control that are currently based on synthetic insecticides (Bueno, Carvalho, Santos, Sosa‐Gómez, & Silva, 2017; Krinski, Foerster, & Deschamps, 2018).…”