Characterizing pseudohyphal formation, one of the major virulence factors, in clinical isolates of Saccharomyces cerevisiae in Taiwan
Date Issued
2008
Date
2008
Author(s)
Wu, Yu-Shan
Abstract
Saccharomyces cerevisiae is an emerging opportunistic pathogen. Previous work has shown that American clinical isolates exhibit the ability to grow at 42°C and are capable of pseudohyphal formation at 37°C. S. cerevisiae cells grow in low-ammonium solid media mediating a switch from the yeast to the filamentous form. However, morphology switching is not clear about virulence mechanism. We collected three clinical yeast isolates (YYC1, 2, 3) from National Taiwan University Hospital and found that YYC1 and 3 grow at 42°C and are capable of pseudohyphal differentiation at 37°C, whereas YYC2 could not. We analyzed the genes involved in pseudohyphal formation included TEC1, FLO8 and SSD1. Sequence analysis comparing YYC1, 2, 3 to s288c from Saccharomyces Genome Database showed that YYC1, 2 and 3 have 2 single nucleotide polymorphism(SNPs) in TEC1, at most 37 SNPs in FLO8 gene. And at most 33 SNPs in SSD1 gene. FLO8 in YYC2 are different from those in YYC1 and 3. These three clinical isolates all appeared to be heterozygous in FLO8. We used tetrad dissection to separate the segregants of YYC1 and found that 80 percent of the tetrad analyzed showed 2:2 meiotic segregation for the ability to form pseudohyphae at 37°C. We analyzed six tetrad at nucleotide 814, 2026, 2158 and 2263 sites in FLO8 gene, and found that when the haplotype is A, T, A and T on each site respectively, the results of pseudohyphal formation are positive. The FLO8 alleles in YYC1 segregants were cloned and verified in the strain of s288c background. We developed a simple and rapid method for virulence determination because mouse model is relatively complex. The production of cytokine TNF-α was seen when murine macrophage cell lines were stimulated by the yeast cells. Our result suggested that yeast cells with high virulence indeed stimulated higher cytokine levels in macrophages. YYC1 resembled strains with high virulence, while YYC2 and YYC3 resembled avirulent.
Subjects
pseudohyphae
virulence
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