Mycobiota and fungal secondary metabolites associated with wheat grains harvested in different cultivation areas of Western Australia

A total of 72 wheat grain samples, collected at the end of the 2018 harvest season from 12 different cultivation areas of Western Australia (WA), were analyzed to determine their external appearance (by visual observation), the associated culturable fungal microorganisms (by isolation on Potato Dextrose Agar, PDA) and the concentration of fungal secondary metabolites (by Liquid Chromatography tandem Mass Spectrometry, LC-MS/MS). The majority of grains were healthy in appearance. However, when cultured on PDA a large number of fungal colonies were isolated and, after molecular identification based on the partial sequence of the Internal Transcribed Spacer (ITS) or Translation Elongation Factor 1 alpha (TEF1 alpha) regions, these were shown to belong to seven different fungal genera: Alternaria, Aureobasidium, Cladosporium, Nigrospora, Epicoccum, Fusarium and Pyrenophora. The genus Alternaria was the main component of this fungal community and its distribution was almost homogeneous among the samples derived from different cultivation areas of WA, while its presence was negatively correlated with that of the beneficial genus Aureobasidium. In contrast to Alternaria, fungal colonies belonging to the genus Fusarium showed a very low presence among the samples. Despite the wide range of fungal secondary metabolites assayed, LC-MS/MS analysis detected the presence of only five fungal secondary metabolites: infectopyrone and macrosporin (produced by Alternaria species), and deoxynivalenol (DON), culmorin (CULM) and enniatin B (ENB) (produced by Fusarium species). Alternaria secondary metabolites were present at low concentrations in samples from all cultivation areas (except for two areas that were free of mascroporin). Fusarium secondary metabolites were present with very low levels in samples from only seven cultivation areas (DON and ENB) or three cultivation areas (CULM). These results show that infection by Alternaria species was widespread and that infection by Fusarium species was rare by comparison. The results also show that in the environment studied the comparatively high incidence of infection does not correspond to concerning levels of the most important and known mycotoxins.