Development of predictive models evaluating the spoilage-delaying effect of a bioprotective culture on different yeast species in yogurt.

Yeast spoilage of fermented dairy products causes challenges for the dairy industry, including economic losses due to wasted product. Food cultures with bioprotective effects are becoming more widely used to help ensure product quality throughout product shelf life. To assist the dairy industry when evaluating product quality throughout shelf life and the effect of bioprotective cultures, we aimed to build stochastic models that provide reliable predictions of yeast spoilage in yogurt with and without bioprotective culture. Growth characterizations of Debaryomyces hansenii, Yarrowia lipolytica, Saccharomyces cerevisiae, and Kluyveromyces marxianus at storage temperatures of 7, 12, and 16°C during a 30-d storage period were conducted in yogurt with and without a bioprotective culture containing Lacticaseibacillus rhamnosus strains. The kinetic growth parameters were calculated using the Buchanan growth model, and these parameters were used as baseline values in Monte Carlo models to translate the yeast growth into spoilage levels. The models were developed using 100,000 simulations and they predicted yeast spoilage levels in yogurt by the 4 yeast types. Each modeled yogurt batch was set to be contaminated with yeast at a concentration drawn from a normal distribution with a mean of 1 log10 cfu/mL and standard deviation of 1 log10 cfu/mL and stored for 30 d at a temperature drawn from a normal distribution with a mean of 6.1°C and a standard deviation of 2.8°C. Considering a spoilage level of 5 log10 cfu/mL, the predicted number of spoiled samples was reduced 3-fold during the first 10 d and by 2-fold at the end of shelf life when a bioprotective culture was added to the yogurt. The models were evaluated by sensitivity analyses, where the main effect factors were maximum yeast population, storage temperature, and yeast strain. The models were validated by comparing the model output to actual observed spoilage data from a European dairy using the bioprotective culture. When the model prediction, based on a mixture of the 4 specific yeast strains, was compared with spoilage data from the European dairy, the observed effect of bioprotective cultures was considerably higher than predicted, potentially influenced by the presence of contaminating strains more sensitive to a bioprotective culture than those characterized here. The developed Monte Carlo models can predict yeast spoilage levels in yogurt at specific production settings and how this may be affected by various parameters and addition of bioprotective cultures.

Identification and characterisation of organisms associated with chocolate pralines and sugar syrups used for their production.

Spoilage of chocolate pralines, due to growth of microorganisms tolerating low water activity, causes problems in the confectionary industry. Therefore, an increased knowledge on which organisms are present in the chocolate fillings and their tolerance towards low aw, pH, ethanol and other preservatives is needed. Using media containing 40-50% glucose (aw 0.872-0.925) bacteria, yeasts and moulds were isolated from chocolate pralines (aw 0.70-0.898) of nine manufactures and sugar syrups (aw 0.854) used as ingredient in chocolate praline production by one of the manufacturers. Isolates were identified by conventional microbiological analyses and by sequencing of their 16S rRNA, 26S rRNA (D1/D2-region) or calmodulin genes. Further, for several species the identity was confirmed by amplification and sequencing of additional genes. In total 677 isolates were identified as belonging to ten different bacteria species, six yeast species and ten mould species with yeast being the most frequently isolated. Bacteria and moulds were found in low numbers, whereas yeast were found in numbers up to 10(7)CFU/g. The most frequently isolated yeast, bacteria and moulds belonged to the species of Zygosaccharomyces rouxii, Bacillus subtilis and Aspergillus terreus, respectively. Fifteen isolates were screened for their ability to grow in presence of low aw (0.65-0.90), low pH (pH=2.0-7.0), ethanol (0-15%), sorbic acid (0-1,500 ppm) and different temperatures (15°C-25°C) relevant for chocolate manufacturing. Z. rouxii was overall the most tolerant organism to the stress factors and grew within the same range of environmental conditions as found in chocolate pralines. It was able to grow at water activities down to 0.70, ethanol concentrations up to 6.0%, pH down to pH2.0, sorbic acid concentrations up to 1,500 ppm and at all temperatures tested. Eurotium amstelodami also showed high tolerance towards all the stress factors except for ethanol. None of the bacteria were able to grow at the conditions tested. However, B. subtilis survived the 60 day incubation period.