Effect of antifungal proteins (AFPs) on the viability of heat-resistant fungi (HRFs) and the preservation of fruit juices

The control of heat-resistant fungi (HRFs), which cause spoilage of heat-treated fruit products, is considered a challenge for the fruit juice and beverage industry and requires new strategies for the development of antifungal compounds. In this study, four antifungal proteins (AFPs) from Penicillium digitatum (PdAfpB) and Penicillium expansum (PeAfpA, PeAfpB and PeAfpC), were evaluated against conidia from a representative collection of HRFs. A total of 19 strains from 16 different species belonging to the genera Aspergillus, Hamigera, Paecilomyces, Rasamsonia, Sarocladium, Talaromyces and Thermoascus were included in the study. PeAfpA and PdAfpB exhibited potent antifungal activity in synthetic media, completely inhibiting the growth of most of the fungi evaluated in the range of 0.5–32 μg/mL. The efficacy of the four AFPs was also tested in fruit juices against ascospores of five HRFs relevant to the food industry, including P. fulvus, P. niveus, P. variotii, A. fischeri and T. flavus. PdAfpB was the most effective protein in fruit juices, since it completely inhibited the growth of the five species tested in at least one of the fruit juices evaluated. This is the first study to demonstrate the activity of AFPs against fungal ascospores. Finally, a challenge test study showed that PdAfpB, at a concentration of 32 μg/mL, protected apple fruit juice artificially inoculated with ascospores of P. variotii for 17 days, highlighting the potential of the protein as a preservative in the fruit juice industry.