Genetic identification of cycloheximide-resistant yeasts isolated from traditional dairy products

Traditional dairy products harbor a complex microbiota composed by several microbial groups، including lactic acid bacteria (LAB) and yeasts. The common method to detect LAB is to culture the products on media containing cycloheximide (CHX) to prevent yeast growth by interfering with protein synthesis. However، some yeast species and strains show natural or acquired resistance to this antibiotic and thus can be specifically selected in media with CHX. The aim of this research was to identify such CHX-resistant yeasts. To this purpose، 25 samples of home-made dairy products were plated on MRS medium with 0. 01% CHX; after incubation (48 h، 30˚C)، 32 colonies of presumptive yeasts were picked up. Catalase test and morphological investigation confirmed that 19 were yeasts. After DNA isolation، randomly amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) with primer M13 was applied to gain information on genomic relatedness among isolates. Visual comparison of RAPD-PCR pattern allowed selecting five representative isolates for further analysis. Primers ITS1 and ITS4 were used for specific amplification، and the PCR-products were sequenced after purification. The results of sequencing revealed that all isolates belong to Kluyveromyces marxianus، a species with a high technological potential.