Screening Botrytis cinerea for proteases suitable for removing haze-forming proteins in winemaking

Protein haze in wine is caused by plant pathogenesis related (PR0 proteins that precipitate when heated, cause the wine to look cloudy and render it unsaleable. Advancements have been made to reduce the economic, environmental and wine quality impacts that come with traditional bentonite stabilisation. Enzymatic treatment with recombinant proteins has shown some promise, however genetically modified organisms cannot be used in Australian commercial winemaking. In order to find a non-genetically modified alternative for protein haze removal, 99 isolates of Botrytis cinerea were initially screened for strong growth and sporulation, with 55 of these isolates selected and screened for protease production by skim milk plate clearing assay. These results coupled with guaiacol plate assays to assess laccase production from these isolates, which would oxidize the finished wine, resulted in 6 B. cinerea isolates that were assessed for protease secretion in liquid culture. Analyses of the protease and laccase production of these isolates by fluorescence microplate and syringaldazine assays identified 6 B. cinerea isolates that secrete high levels of protease, to be used in continuing research.