Brettanomyces is the major microbial cause for wine spoilage worldwide and causes significant economic losses. The reasons are the production of ethylphenols that lead to an unpleasant taint described as 'phenolic odour'. Despite its economic importance, Brettanomyces has remained poorly studied at the metabolic level. The origin of the ethylphenol results from the conversion of vinylphenols in ethylphenol by Brettanomyces hydroxycinnamate decarboxylase. However, no information is available on the vinylphenol reductase responsible for the conversion of vinylphenols in ethylphenols. In this study, a vinylphenol reductase was partially purified from Brettanomyces bruxellensis that was active towards 4-vinylguaiacol and 4-vinylphenol only among the substrates tested. First, a vinylphenol reductase activity assay was designed that allowed us to show that the enzyme was NADH dependent. The vinylphenol reductase was purified 152-fold with a recovery yield of 1.77%. The apparent K(m) and V(max) values for the hydrolysis of 4-vinylguaiacol were, respectively, 0.14 mM and 1900 U mg(-1). The optimal pH and temperature for vinylphenol reductase were pH 5-6 and 30 degrees C, respectively. The molecular weight of the enzyme was 26 kDa. Trypsic digest of the protein was performed and the peptides were sequenced, which allowed us to identify in Brettanomyces genome an ORF coding for a 210 amino acid protein.

Aims: Spore germination requires microscopic observation whereas fungal growth results in a macroscopic examination. This paper aims at establishing a relationship between the percentage of germinated spores and parameters easily available from visible development. Methods and Results: About 225 spores of Mucor racemosus were inoculated on PDA medium and incubated at 15°and 25°C. Germination kinetics were modelled by a logistic function. Fungal development provided two parameters, a growth rate, l, and a lag period, k, defined as the slope of the straight line of the graph radius (mm) vs time (h) and the intercept of this line with the X-axis, respectively. Conclusions: It was found that the lag period coincided with the completion of the germination process, although the number of spores inoculated should be controlled carefully. Significance and Impact of the Study: Providing that this result can be generalized, this procedure would constitute a significant breakthrough for predicting food spoilage by moulds.

The influence of ethanol vapor on germination of Penicillium chrysogenum was determined on yeast nitrogen base plus glucose agar medium at 25 degrees C. Ethanol vapors were generated by 0 to 6% (wt/wt) ethanol solutions at the bottom of hermetically closed petri dishes. The logistic equation was used to describe the data as the percentage of germination versus time and to estimate the germination time. The effect of ethanol concentration on germination time was described by a new reparameterized equation, resulting in an estimated limiting ethanol concentration of 4.3%. Up to 3% ethanol, all spores germinated, and the germination time increased with increasing ethanol concentration. At 3.5 and 4%, some spores formed abnormal germ tubes and others were inhibited at the swelling stage. The inhibiting effect of ethanol was reversible under these experimental conditions.