Atividade larvicida e inibição in vitro de proteases intestinais de Aedes aegypti por extratos de sementes de plantas da caatinga

Abstract

Arboviruses are epidemiologically relevant viruses transmitted by arthropods, affecting especially tropical countries, such as Brazil. Currently, the fight against arboviruses is carried out mainly through population control of Aedes aegypti, main urban vector mosquito. Recently, the search for eco friendly alternatives for vector control has been encouraged, such as the development of botanical insecticides. In this scenario, the Caatinga, a center of biodiversity, represents an important source of plant species for prospecting insecticidal molecules. The present research investigated the potential activity of seed extracts from 3 plant species from the Caatinga, Amburana cearensis, Anadenanthera colubrina and Erythrina velutina, against A. aegypti larvae, and sought to evaluate, in vitro, the ability to inhibit intestinal proteases of the mosquito larvae. Thus, bioassays were performed exposing the larvae to different extract concentrations, and the percentage of larval survival at 24 and 48 hours was analyzed in order to estimate the lethal concentrations 50% (LC50). For the in vitro tests, homogenates were produced from the larvae midguts, in order to analyze the inhibition by exposure to extracts quantitatively, through an enzyme activity assay, and qualitatively, through zymography. The aqueous extracts of A. cearensis (CE), A. colubrina (AE) and E. velutina (ME) compromised larval survival, presenting LC50 equal to 0.67 mg/mL, 0.28 mg/mL and 1, 85 mg/mL, after 24h of exposure, and 0.24 mg/mL, 0.05 mg/mL and 0.91 mg/mL, after 48h of exposure, respectively. The initial characterization of the extracts suggests an efficiency in the extraction conditions, with dry weight yield between 14.53 mg and 34.50 mg, total yield between 7.56% and 54.16%, and soluble protein contents between 1,16 mg/ml and 2,30 mg/ml. The extracts soluble proteins possibly contributed for the enzyme activity impairment of A. aegypti larvae intestinal homogenate proteases, with specific inhibitory activity in CE (662.65 IU/mg), AE (226.19 IU/mg) and ME (496.11 IU/mg). The zymography analysis revealed the presence of proteolytic bands of high relative molecular weight (above 66 kDa), medium (between 45 kDa and 30 kDa) and low (below 30 kDa), and that the extracts affected distinct proteases groups in the larval homogenate. The results indicate the inhibition of intestinal proteases may comprise one of the modes of action for the larvicidal activity of the extracts, especially for the CE extract. Complementary tests are underway to confirm this relationship.