Plant defence mechanisms can reduce the digestive enzyme activity of insect pests. The aim of this study was to determine the relationship between the production of proteinase inhibitors, lipoxygenase and polyphenol oxidase activity in Coffea arabica (Catuai IAC 15) plants, and the digestive enzyme activity in the pest Leucoptera coffeella (Lepidoptera: Lyonetiidae) after feeding on the plant. The production of proteinase inhibitors was evaluated with L‐BApNA as a substrate. We studied lipoxygenase activity with linoleic acid and polyphenol oxidase activity with catechol substrates, in coffee plants damaged (T1) and not damaged (T2) by L. coffeella. L. coffeella digestive enzyme activity was verified by trypsin‐like (substrate l‐BApNA and l‐TAME), chymotrypsin‐like (BTpNA and ATEE), cysteine proteases (l‐BApNA) and total protease (azocasein). Proteinase inhibitor production and lipoxygenase and polyphenol oxidase activity in C. arabica increases (P ≤ 0.05) with L. coffeella damage. Our results provide important information that these enzymatic activities may play a role in plant defence processes in C. arabica. Trypsin‐like activity increases, whereas chymotrypsin‐like and cysteine protease activity decrease in the midgut of L. coffeella, which acts as a defence mechanism.

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The recognition of protease inhibitors with insecticidal activity is important as a basis for the development of mimetic peptides with potential use as biorational insecticides. We sprayed benzamidine on soybean plants and assessed whether this potent synthetic trypsin-inhibitor has protease inhibitory, insecticidal and deterrent effects on the velvetbean caterpillar Anticarsia gemmatalis Hübner (Lepidoptera: Erebidae). Activity of trypsin inhibition in soybean leaves was increased and total proteolytic activity in the midgut extract from larvae fed on these leaves was reduced by benzamidine. Different concentrations of benzamidine sprayed on the plant caused approximately 50 % of larval mortality, and larval choice and moth preference and oviposition were all negatively affected. Low concentrations of benzamidine increased mortality and hindered insect choice and oviposition as well as higher doses. Since many synthetic protease inhibitors are usually expensive, small doses of benzamidine may be effective to protect soybean against A. gemmatalis attack. Our results highlight the potential of synthetic protease inhibitors for insecticidal and deterrent purposes in insect pest management.

Os besouros serradores são conhecidos por causarem danos em várias espécies arbóreas. Estes besouros roletam galhos e os utilizam para ovipositar e completar o desenvolvimento de sua prole. O objetivo deste trabalho foi descrever a ocorrência de um caso de O. dejeanii Thomson, 1868 (Coleoptera: Cerambycidae) em Pyrus pyrifolia (Burm F.) Nakai. (Rosaceae) (pereira), visando caracterizar suas injúrias e verificar a possibilidade de desenvolvimento neste vegetal. A pereira se encontrava em um pomar, no Município de Viçosa, MG. Foram recolhidos três galhos roletados de uma pereira, sendo que em dois destes foi coletada uma fêmea em cada. O diâmetro e comprimento médio dos galhos foram de 3,47±0,15 cm e 3,31±0,17 m, respectivamente. O terço inferior do galho roletado apresentou 52% das incisões de posturas. Foi observada uma fêmea efetuando duas posturas ao longo do galho e registrado o tempo de 10 min e 13 min, respectivamente. O. dejeanii utiliza P. pyrifolia como hospedeiro alternativo, apresentando potencial para se tornar praga e reduzir a produção.

Endogenous mechanisms of plant resistance to herbivorous insects could be adapted to enable the development of alternative strategies for pest control. Activation of the lipoxygenase (LOX) pathway by jasmonic acid, the hormone responsible for activating genes encoding protease inhibitors (PI), is one of the main direct defenses of plants against insects. In this study, soybean cultivars at the V3 stage that were either susceptible or resistant to herbivorous insects were used to investigate changes in the activity of LOX and concentration of PIs in response to 24-or 48-h feeding by fourth and fifth instar Anticarsia gemmatalis (Lepidoptera: Noctuidae) larvae. LOX activity (0.0-0.007 nm s -1 /mg) and the PI concentration (60 to 125 mg of trypsin inhibitor / mg of protein and 90 to 120 mg of trypsin inhibitor / mg protein after 24 and 48 h after attack, respectively) both increased in response to damage caused by A. gemmatalis. Proteolytic activity decreased by approximately 50% in cultivars with different degrees of resistance. Raised enzyme profiles in the midgut of A. gemmatalis may be related to tryptic enzymes that increased, compensating for the inhibition of proteases by entomotoxic substances of the soybean cultivars.Key words: plant defense, lipoxygenases, velvet caterpillar. RESUMEN

Anticarsia gemmatalis represents a relevant factor for lowering soybean and other legume crop productivities.Protease inhibitors affect protein degradation and reduce the availability of amino acids, impairing the development and survival of insect pests. To evaluate the possible use of proteinaceous protease inhibitors in the management of this pest, the activities of midgut proteases and the growth and development of A. gemmatalis larvae exposed to soybean Bowman-Birk trypsin-chymotrypsin inhibitor (SBBI) and soybean Kunitz trypsin inhibitor (SKTI) were determined. The survival curves obtained using Kaplan-Meier estimators indicated that SKTI and SBBI stimulated larval survival. However, the development of A.gemmatalis was delayed, and prepupal weight decreased in the presence of both inhibitors. The results showed that SKTI and SBBI inhibited the trypsin-like and total proteolytic activities of larvae on the 12th day after eclosion. On the 15th day after eclosion, larvae exposed to SKTI increased the activities of trypsin and total proteases.Although SKTI and SBBI did not affect the survival of the insect, they had effects on midgut proteases in a stage wherein A. gemmatalis fed voraciously, increased the larval cycle, and decreased prepupal weight. These findings provide baseline information about the potential of proteinaceous protease inhibitors to manage the velvetbean caterpillar, avoiding chemical pesticides.K E Y W O R D S biotic stress, insect attack, insect performance, protease inhibitory

Anticarsia gemmatalis Hübner, 1818 (Lepidoptera) is a major pest of soybean in the Brazil. It is known that the reduction of proteolytic activity by the ingestion of protease inhibitors reduces digestion and larval development of the insects. Control via inhibition of the digestive enzymes necessitates deeper knowledge of the enzyme kinetics and the characterization of the inhibition kinetics of these proteases, for better understanding of the active centers and action mechanisms of this enzyme. Trypsin-like proteases found in the gut of Anticarsia gemmatalis were purifi ed in a p-aminobenzamidine agarose column. Kinetic characterization showed K M 0.503 mM for the L-BApNA substrate; V max = 46.650 nM s -1 ; V max /[E]= 9.256 nM s -1 mg L -1 and V max /[E]/K M = 18.402 nM s -1 mg L -1 mM. The K i values for the inhibitors benzamidine, berenil, SKTI and SBBI were 11.2 µM, 32.4 µM, 0.25 nM and 1.4 nM, respectively, and all revealed linear competitive inhibition. The SKTI showed the greatest inhibition, which makes it a promising subject for future research to manufacture peptide mimetic inhibitors.

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