A new method for detection of spoilage yeasts and molds in dairy products

Introduction. Fungi (yeasts and moulds) are recognized as one of the main contaminants of dairy products including yogurt and sour milk. These microorganisms can also cause spoilage in a wide range of processed, preserved and refrigerated food products. During the past years, several molecular methods based on immunological and genotypic techniques have been developed for revealing the presence of undesirable microorganisms, including fungi, in different food matrices. However, no commercial kit are already available to detect viable yeasts and moulds in dairy products. Materials and methods. Five antibodies against yeasts and molds were selected from commercially available antibodies and used to produce functionalized magnetic beads to be used to capture and separate microrganisms associated to dairy products. Four yeast type species (Debaryomyces hansenii, Kluyveromyces marxianus, Geotrichum candidum and Pichia anomala) and four mold species (Alternaria alternata, Aspergillus niger, Penicillium italicum and Rhizopus stolonifer) were used. Milk, yogurt and soft cheese were tested as matrices. A RT-PCR protocol was developed for detection of yeast and molds mRNA extracted from contaminated foods. Results. A new method for yeast and molds enrichment from different food dairy products (milk, yogurt and soft cheese) based on the use of antibody coated magnetic beads was developed. A new RT-PCR assay based on a nested amplification was optimized for the detection of yeast and molds in artificially contaminated dairy products. Discussion. The correlation between the amplification signal and the microbial count will allow to use this method for viable contaminants quantification in dairy products. This method can avoid the labor expensive food matrices treatments, often cause of loss of sensitivity. This approach will be transferred also in other food matrices. This procedure can be implemented by the use of automated enrichment systems already available for pathogen microorganisms.