Botrytis cinerea is an important phytopathogenic fungus affecting the fruit production around the world. This fungus is controlled mainly by using synthetic fungicides, but many resistant isolates have been selected by the indiscriminate use of fungicides. Endophytic fungi or secondary metabolites obtained from them become an alternative method of control for this fungus. The aim of this work was to identify endophytic fungi with antifungal activity against the plant pathogenic fungus B. cinerea isolated from plants from Central Andean Precordillera of Chile. Three endophytic fungi (Ac1, Lc1 and Ec1) with antifungal activity against B. cinerea were isolated from native and endemic plants growing in Central Andean Precordillera of Chile. The isolates Lc1 (isolated from Lithraea caustica) and Ac1 (isolated from Acacia caven) were identified as Alternaria spp. and the isolate Ec1 (isolated from Echinopsis chiloensis) was identified as Aureobasidium spp. The isolated endophytic fungi would inhibit B. cinerea through the secretion of diffusible and volatile compounds affecting the mycelial growth, conidia germination and interestingly, it was also shown that the volatile compounds produced by the three isolated endophytic fungi suppressed the sporulation of B. cinerea.

Our research group isolated an endophyte yeast from the endemic plant Echinopsis chiloensis. This yeast, identified as Naganishia sp, produces volatile organic compounds (VOC) with antifungal activity against Botrytis cinerea. Due to the need for alternative control methods for this pathogen, the effect of VOC on B. cinerea was analyzed. On the fourth day of cultivation, in the presence of VOCs, the mycelial growth of B. cinerea stopped. VOCs inhibited 32.8% of the conidia germination and reduced sporulation by 70.6%. These compounds promoted the formation of infection cushions. VOCs caused damage to the cell wall and plasma membrane in B. cinerea. On the other hand, the volatile compounds induced oxidative stress, and led to membrane lipid peroxidation after 16 and 24 h of incubation in the presence of VOCs, because an increased level of malondialdehyde content was observed. These compounds exerted a fungistatic effect on B. cinerea. The volatile compounds emitted by Naganishia sp were identified by gas chromatography coupled with mass spectrometry as 3-methylbutyl acetate, ethylbenzene, 1-ethyl-3-methylbenzene, 2-phenylethanol, 2-heptanone and (5E)-6,10-dimethylundeca-5,9-dien-2-one. The most abundant volatile compound, 2-phenylethanol, inhibited the mycelial growth of B. cinerea with an IC₅₀ value of 0.21 µL mL⁻¹. The effect of this pure compound was also fungistatic. Finally, it was shown that volatile compounds decreased the severity of B. cinerea infection in tomatoes and grapes by 64.7 and 43,1%, respectively. This is the first report identifying volatile compounds emitted by a Naganishia sp. and describing their mode of action against B. cinerea.

Endophytic fungi from the Cactaceae family are an underexplored source of bioactive secondary metabolites with potential applications in sustainable agriculture. This study investigated an endophytic fungus obtained from healthy cladodes of Opuntia ficus-indica growing in the Chilean Andean Precordillera. The influence of culture conditions, specifically pH and nitrogen concentration, on the production of diffusible and volatile antifungal compounds against the phytopathogen Botrytis cinerea was evaluated using dual-culture (confrontation) and sandwiched Petri dish assays. Morphological characteristics and molecular analyses confirmed that the isolate belongs to the genus Alternaria. Antifungal activity increased significantly under acidic conditions and limited nitrogen availability. The strongest inhibition by volatile compounds occurred at pH 4.5 and the lowest concentration of ammonium tartrate. Furthermore, ethyl acetate extracts at 40 mg/L obtained from Alternaria sp. cultures grown at pH 4.5 with 2.3 g/L ammonium tartrate inhibited B. cinerea mycelial growth by 60%. The study provides a framework for improving the yield of antifungal metabolites produced by Alternaria, contributing to the development of biofungicides for gray mold control.