We produced electrophoretic karyotypes of the reference strain E150 and of seven other isolates from different geographical origins to study the genomic organization of the dimorphic yeast Yarrowia lipolytica. These karyotypes differed in the number and size of the chromosomal bands. The karyotype of the reference stain E150 consisted of five bands of between 2.6 and 4.9 Mb in size. This strain contained at least five rDNA clusters, from 190 to 620 kb in size, which were scattered over most of the chromosomes. The assignment of 43 markers, including rRNA genes and three centromeres, to the E150 bands defined five linkage groups. Hybridization to the karyotypes of other isolates with pools of markers of each linkage group showed that linkage groups I, II, IV and V were conserved in the strains tested whereas group III was not and was split between at least two chromosomes in most strains. Use of a meganuclease I-SceI site targeted to one locus of E150 linkage group III showed that two chromosomes actually comigrated in band III of this strain. Our results are compatible with six chromosomes defining the haploid complement of strains of Y. lipolytica and that, despite an unprecedented chromosome length polymorphism, the overall structure of the genome is conserved in different isolates.

We describe a rapid method developed to identify yeast species commonly found during cheese ripening. It is based on the isolation of species-specific sequences and their hybridization with coarse preparations of genomic DNA. Several strategies were followed for the construction of probes: PCR amplification of sequences available in databases, random cloning of genomic DNA fragments, specific RAPD fragments and PCR-amplified ribosomal DNA fragments. After validation, the probes were applied to the characterization of 400 yeast strains isolated from various French cheeses. Since the strains had been previously identified with classical diagnostic tests, we were able to compare molecular and conventional identification. In addition, the specific probes for one of the species, Debaryomyces hansenii, were used successfully in colony hybridization experiments. The probes developed here proved to be very useful for the screening of large yeast collections and for the assessment of biodiversity within the yeast flora in cheese. yeast / cheese / molecular identification / hybridization / biodiversityResume -Utilisation de sondes ADN specifiques pour la caracterisation rapide de levures isoIees d'ecosystemes complexes. Au cours de ce travail, nous avons developpe une methode a la fois rapide et sensible pour l'identification des levures couramment isolees pendant l'affinage du fromage. Son principe repose sur l'hybridation de sequences specifiques sur les preparations brutes d' ADN genomique des souches a identifier. Pour la construction des sondes, nous avons du suivre plusieurs strategies differentes: amplification de sequences disponibles dans les bases de donnees, clonage au hasard de sequences d' ADN genomique, isolement de sequences specifiques obtenues par amplification RAPD d' ADN genomique ou par amplification d' ADNr. Apres leur validation, les son des ont pu etre mises en ceuvre S354 A.-M. Davila et al.dans differentes applications. Elles ont ete utilisees pour cribler 400 souches de levures isolees de differents from ages fran«ais. Les souches ayant ete prealablement identifiees par des tests classiques, nous avons pu comparer les deux types de methodes d'identification, moleculaire et conventionnelle. La son de specifique de D. hansenii a He testee avec succes dans des essais d'hybridation sur colonie. Au total, ces son des se sont revelees etre de bons outils pour le suivi et l'inventaire de la biodiversite des levures fromageres. levures / fromage / identification moleculaire / hybridation / biodiversite

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