Dynamics and characterization of yeasts during natural fermentation of Italian sausages

A multiphasic approach was used to investigate the yeast ecology in Italian fermented sausages. Culture-dependent and -independent methods were applied to identify the yeast species during the maturation process and to characterize the numerically dominant species. Plating analysis and subsequent molecular identification of the isolates highlighted the dominance of Debaryomyces hansenii, but at least other three yeast species -Candida zeylanoides, Pichia triangularis and Metschnikowia pulcherrima - contributed to the fermentation as well. Direct denaturing gradient gel electrophoresis analysis confirmed that D. hansenii was the main yeast species present and its activity was also demonstrated. No other yeasts species were detected on the direct denaturing gradient gel electrophoresis gels, whereas DNA of Penicillium farinosum, Penicillium viridicatum and Mucor racemosus were present. Molecular characterization by RAPD-PCR analysis of the D. hansenii isolates demonstrated a shift in its population from the beginning to the end of the maturation of the sausages. Strains present during the early stages of the fermentation were grouped in clusters that differed from those isolated in the final phases of the maturation, underlining the genetic differences between these two populations of D. hansenii. However, all the isolates were able to grow in the presence of 3.5% sodium chloride and at 10 degrees C, evidence that these parameters did not select the species present at the end of the maturation period.