Phylogeny, antimicrobial, antioxidant and enzyme-producing potential of fungal endophytes found in Viola odorata

Abstract

Viola odorata, a medicinal plant, is traditionally used to treat common cold, congestion and cough. Given its medicinal properties and occurrence in the northwestern Himalayas, we isolated and characterized endophytic fungi from this plant morphologically, microscopically and by internal transcribed spacer-based rDNA sequencing. In total, we isolated 27 morphotypes of endophytes belonging to phyla Ascomycota and Basidiomycota. The roots showed the highest diversity of endophyte as well as fungal dominance, followed by leaves and leaf nodes. The fungal extract of VOR16 (Fusarium oxysporum) displayed potent antimicrobial activity against Salmonella typhimurium, Klebsiella pneumoniae and Escherichia coli, with a minimum inhibitory concentration of 0.78, 0.78 and 1.56 μg/mL, respectively, while fungal extract VOLF4 (Aspergillus sp.) exhibited promising antioxidant activity (IC50 of 17.4 μg/mL). To identify the components responsible for various bioactivities, we analyzed the content of penicillin G in the extract of bioactive endophytes. The results suggested that the expression of penicillin G under the fermentation conditions applied was too low to display antimicrobial effects. Thus, the activity may be contributed by a different, novel secondary metabolite. The antioxidant activity of VOLF4 may be attributed to its high content of flavonoids. Of the endophytic fungi assessed, 27% were found to be enzyme producers. The highest zone of clearance was observed in VOLN5 (Colletotrichum siamense) for protease production. Only VOR5 (Fusarium nematophilum) was found to be a producer of cellulase, glutenase, amylase and protease. In summary, this is the first report of the isolation of endophytes, namely Fusarium nematophilum, Colletotrichum trifolii, C. destructivum, C. siamense and Peniophora sp., from V. odorata and their bioactive and enzyme-producing potential.