Assimilation by Yeast Isolates

Carbon and nitrogen assimilation abilities of yeast isolates are among the first properties tested (2-4) in identifying and classifying yeasts. The recommended tests are conventionally done in liquid medium or by an auxanographic method whereby a solid medium containing all the nutrients and growth factors required by a yeast, except a carbon source, is seeded with the test yeast and then different carbon compounds are added at isolated sites on the surface of the hardened agar medium. An enhanced area of yeast growth around a particular carbon source indicates that compound can be assimilated and used for growth by that yeast (5). Previous research done in our laboratory on identification of yeasts from wine musts (K. J. Moore, M. G. Johnson, and J. R. Morris, J. Food Sci., in press) indicates that the liquid medium test methods are too time-consuming and laborintensive, while the conventional solid medium test methods often give equivocal and nonreproducible results. The laborsaving multipoint inoculator method (1) was considered but was discarded because of the cost of fabricating this device as well as the potential for colony crowding and overlap on a 100-mm-diameter plate if 20 or more yeasts are inoculated per plate. In an effort to reduce the time and labor and minimize the expense required to obtain unequivocal results, a study was undertaken to (i) determine whether a disk inoculum-solid medium (DISM) method could be used to test carbon and nitrogen assimilation by yeast isolates from wine musts and (ii) compare the rapidity and reproducibility of this DISM method with conventional static liquid medium methods. Fifty yeast isolates were randomly selected from one lot of yeasts recovered from must fermentations of White Riesling (Vitis vinifera) grapes. Six known pure yeast cultures included as controls in the assimilation assays were Debaryomyces hansenii, Kloeckera apiculata, Kluyveromyces fragilis NRRL Y-2415, Pichia membranaefaciens, Rhodotorula mucilaginosa, and Saccharomyces cerevisiae ATCC 4132. Yeast isolates were maintained on yeast malt agar slants at 4°C. Yeast strains were brought to a state of active growth by loop transferring them from yeas.t malt slants into tubes containing 3 ml of yeast malt broth, followed by incubation at 25°C for 36 to 48 h. Cells were then harvested by centrifugation, using a superspeed centrifuge (Model SS-3; Ivan Sorvall, Inc., Newtown, Conn.) at ca. 2,200 x g for 20 min at room temperature (ca. 25°C). The isolates were washed twice with sterile deionized water before suspending the cells in sterile deionized water to approximate a McFar-