Nitrite metabolism in Debaryomyces hansenii TOB-Y7, a yeast strain involved in tobacco fermentation.

ABSTRACT

The Italian cigar manufacturing process includes a fermentation step that leads to accumulation of nitrite and tobacco-specific nitrosamines (TSNA), undesirable by-products due to their negative impact on health. In this study, growth and biochemical properties of Debaryomyces hansenii TOB-Y7, a yeast strain that predominates during the early phase of fermentation, have been investigated. With respect to other D. hansenii collection strains (Y7426, J26, and CBS 1796), TOB-Y7 was characterized by the ability to tolerate very high nitrite levels and to utilize nitrite, but not nitrate, as a sole nitrogen source in a chemically defined medium, a property that was enhanced in microaerophilic environment. The ability to assimilate nitrite was associated to the presence of YNI1, the gene encoding the assimilatory NAD(P)Hnitrite reductase (NiR), absent in Y7426, J26, and CBS 1796 by Southern blot data. YNI1 from TOB-Y7 was entirely sequenced, and its expression was analyzed in different media by Northern blot and reverse transcriptase polymerase chain reaction. The evidence that, in D. hansenii TOB-Y7, YNI1 was transcriptional active also in the presence of high ammonia concentration typical of tobacco fermentation, stimulated the development of an improved process that, on a laboratory scale, was proved to be effective in minimizing nitrite and TSNA accumulation.