Mould spoilage of foods and beverages: Using the right methodology.

Fungal spoilage of products manufactured by the food and beverage industry imposes significant annual global revenue losses. Mould spoilage can also be a food safety issue due to the production of mycotoxins by these moulds. To prevent mould spoilage, it is essential that the associated mycobiota be adequately isolated and accurately identified. The main fungal groups associated with spoilage are the xerophilic, heat-resistant, preservative-resistant, anaerobic and psychrophilic fungi. To assess mould spoilage, the appropriate methodology and media must be used. While classic mycological detection methods can detect a broad range of fungi using well validated protocols, they are time consuming and results can take days or even weeks. New molecular detection methods are faster but require good DNA isolation techniques, expensive equipment and may detect viable and non-viable fungi that probably will not spoil a specific product. Although there is no complete and easy method for the detection of fungi in food it is important to be aware of the limitation of the methodology. More research is needed on the development of methods of detection and identification that are both faster and highly sensitive.

Intermediate Thermoresistance in Black Yeast Asexual Cells Variably Increases with Culture Age, Promoting Survival and Spoilage in Thermally Processed Shelf-Stable Foods.

ABSTRACT: Black yeasts are a functional group that has caused spoilage in cold-filled and hot-filled beverages, as well as other water activity-controlled food products. We established quantitative thermoresistance parameters for the inactivation of 12 Aureobasidium and Exophiala isolates through isothermal experiments and a challenge study. Culture age (2 versus 28 days) variably affected the thermoresisitance among the black yeast strains. Variation in thermoresistance exists within each genus, but the two most resistant strains were the Exophiala isolates. The two most heat-resistant isolates were Exophiala phaeomuriformis FSL-E2-0572, with a D60-value of 7.69 ± 0.63 min in 28-day culture and Exophiala dermatitidis YB-734, with a D60-value of 16.32 ± 2.13 min in 28-day culture. Although these thermoresistance levels were, in some cases, greater than those for conidia and vegetative cells from other common food spoilage fungi, they were much more sensitive than the ascospores of heat-resistant molds most associated with spoilage of hot-filled products. However, given that black yeasts have caused spoilage in hot-filled products, we hypothesized that this intermediate degree of thermoresistance may support survival following introduction during active cooling before package seals have formed. A challenge study was performed in an acidic (apple cider) and water activity-controlled (maple syrup) product to evaluate survival. When apple cider was hot filled at 82°C, black yeast counts were reduced by 4.1 log CFU/mL 24 h after the heat treatment, but the survivors increased up to 6.7 log CFU/mL after 2 weeks. In comparison, the counts were below the detection limit after both 24 h and 14 days of shelf life in both products when filled at the boiling points. This suggests that ensuring water microbial quality in cooling tunnels and nozzle sanitation may be essential in mitigating the introduction of these fungi.

Sexual reproduction as the cause of heat resistance in the food spoilage fungus Byssochlamys spectabilis (anamorph Paecilomyces variotii).

Paecilomyces variotii is a common cosmopolitan species that is able to spoil various food- and feedstuffs and is frequently encountered in heat-treated products. However, isolates from heat-treated products rarely form ascospores. In this study we examined by using molecular techniques and mating tests whether this species can undergo a sexual cycle and form ascospores. The population structure of this species was examined by analyzing the nuclear ribosomal internal transcribed spacer 1 (ITS1) and ITS2 and the 5.8S rRNA gene, as well as partial beta-tubulin, actin, and calmodulin gene sequences. Phylogenetic analyses revealed that P. variotii is a highly variable species. Partition homogeneity tests revealed that P. variotii has a recombining population structure. In addition to sequence analyses, mating experiments indicated that P. variotii is able to form ascomata and ascospores in culture in a heterothallic manner. The distribution of MAT1-1 and MAT1-2 genes showed a 1:1 ratio in the progeny of the mating experiments. From the sequence analyses and mating data we conclude that P. variotii is the anamorph of Talaromyces spectabilis and that it has a biallelic heterothallic mating system. Since Paecilomyces sensu stricto anamorphs group within Byssochlamys, a new combination Byssochlamys spectabilis is proposed.

Inactivation of stress-resistant ascospores of Eurotiales by industrial sanitizers.

Different fungi, including the genera Aspergillus (Neosartorya), Paecilomyces (Byssochlamys) and Talaromyces, produce (asco)spores that survive pasteurization treatments and are regarded as the most stress-resistant eukaryotic cells. The sensitivity of the ascospores to treatments with industrial sanitizers containing chlorine dioxide and iodine (iodophors) has never been assessed before. Ascospores of 4 species of Eurotiales were tested and showed clear variations in sensitivity. The most resilient species, T. macrosporus and Pae. variotii (=B. spectabilis) survive 75, but not 200 ppm chlorine dioxide solution treatments. These species were able to survive 75 ppm iodine solution treatments, but relatively low amounts of ascospores (100-1000 spores) could be inactivated after 16 h of treatment. Inactivated spores did not show any sign of germination after 7 days following treatment on growth medium. As judged by microscopy, iodine inactivation resulted in visibly distorted ascospores. For the interpretation of results, the state of dormancy or activation of ascospores is highly important.

The Antimicrobial Activity of Phenolic Compounds Against Listeria monocytogenes and Their Effectiveness in a Model Milk System.

The minimum inhibitory concentration (MIC) of several phenolic compounds against eight strains of Listeria monocytogenes in tryptose phosphate agar (TPA) was determined. Based upon concentration, the most effective compound was the phenolic antioxidant tertiary butylhydroquinone (TBHQ) which had a MIC of 64 µg/ml. Among the FDA approved food antimicrobials, the most effective was propyl paraben with a MIC of 512 µg/ml. Propyl paraben and TBHQ were then compared to potassium sorbate, a commonly used food antimicrobial, in a model milk system containing 10% nonfat milk solids. In this study, only one strain of the test microorganisms, Scott A, was used and two levels of inoculum, 10 and 1000 CFU/ml, were tested. As expected with the basic pH of the model system, both phenolic compounds were significantly more effective than potassium sorbate against L. monocytogenes at 35°C. Both compounds caused a noticeable increase in lag phase of this microorganism. There was about a three log difference in viable cell counts between propyl paraben and TBHQ and the control. The TBHQ was inconsistent in its activity. The inhibitory action of propyl paraben was not affected by the level of inoculum and had consistent activity throughout testing.