1887

Abstract

The phylogenetic position of has previously been established on the basis of the sequence of rRNA genes. In order to confirm the relationship between and other yeast species, particularly , using non-rRNA gene sequences the gene was chosen for analysis. Three overlapping fragments that together span the entire gene () were amplified from a recombinant phage isolated from a genomic DNA λ library using PCR. These were cloned and used to determine the contiguous sequence of the gene. Analysis of the sequence data revealed the presence of a 1131 bp ORF interrupted by a single 632 bp intron at the 5′ extremity of the gene. Comparison of the sequence with the homologue () revealed that although the exons are 97.9% identical the introns are only 83.4% identical. Phylogenetic trees generated using exon and intron sequences from a range of yeast species unequivocally confirmed the phylogenetic position of as a unique taxon within the genus Analysis of the -associated intron sequences from 10 epidemiologically unrelated isolates from disparate geographical locations showed a very low level of intraspecies sequence variation. In order to develop an accurate and rapid method to identify from primary isolation plates the significant divergence between the and intron sequences was exploited by designing -specific PCR primers. Using a rapid boiling method to produce template DNA directly from colonies from primary isolation plates in 10 min, these primers were used in a blind test with 122 isolates of , 53 isolates of , 10 isolates of and representative isolates of other clinically relevant and other yeast species. Only the isolates yielded the -specific 288 bp amplimer. Use of this technique on colonies suspected to be allows their correct identification as in as little as 4 h.

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/content/journal/micro/10.1099/13500872-145-8-1871
1999-08-01
2019-09-17
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http://instance.metastore.ingenta.com/content/journal/micro/10.1099/13500872-145-8-1871
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