Analysis of protease activity in Aspergillus flavus and A. parasiticus on peanut seed infection and aflatoxin contamination

Abstract

Aspergillus flavus and A. parasiticus are aflatoxin-producing fungi that can infect peanut seeds in field crops. An association between A. parasiticus proteolytic enzyme activities and peanut fungal infection was examined. For this study, a model of inductive and non-inductive culture media to produce A. parasiticus extracellular protease before infection was used. These A. parasiticus cultures were used to infect peanut seeds of cultivars resistant and susceptible to aflatoxin contamination. Peanut seeds of both cultivars exposed to fungi grown on casein medium (inductive medium) showed higher internal and external infection and a higher fungal protease content than those observed on potato dextrose agar (PDA) and sucrose medium (non-inductive media). A further study showed higher fungal colonisation and aflatoxin contamination in seeds of the resistant cultivar pre-incubated with Aspergillus extracellular proteases than in those incubated without proteases. Moreover, protease activities affected the viability of non-infected resistant cultivar seeds, inhibiting germination and radicle elongation and enhancing seed tissue injury. The results strongly suggest that protease production by A. parasiticus is involved in peanut seed infection and aflatoxin contamination resulting in seed tissue damage, affecting seed viability and facilitating the access of fungi through the testa. The analysis of fungal extracellular proteases formed on peanut seed during infection showed that A. flavus and A. parasiticus produced metallo and serine proteases; however, there were differences in the molecular masses of the enzymes between both species. The greatest activity in both species was by serine protease, that could be classified as subtilase.