Analysis of virulence-associated gene s in Cryptococcus neoformans var.grubii strains of two genotypes with different virulence

Objective To identify non-synonymous single nucleotide polymorphisms （ nsSNPs ） with virulence-associated genes of Cryptococcus neoformans var.grubii strains of two genotypes （ MLMT13 and MLMT36 ） and to investigate their effects on virulence .Methods Specific primers were designed ac-cording to the existing genome database and used to amplify the fragments of virulence -associated genes of Cryptococcus neoformans.The cloned gene fragments were sequenced and aligned with the sequences of Crpy -tococcus neoformans var.grubii strains of MLMT13 and MLMT36 genotypes to analyze SNPs .nsSNPs were screened out to analyze the structure and function of their corresponding proteins .Results Eleven viru-lence-associated genes were sequenced successfully .The alignment analysis showed that besides two identi-cal genes , the other nine genes presented various numbers of SNPs ranging from one to four .nsSNPs were detected in two genes encoding protein HST 302 which was the family member of silent information regular 2 （SIR2） and the UV excision repair protein RAD23, respectively.The phylogenetic analysis demonstrated that most of the RAD23 proteins found in basidiomycetes shared higher homologies with their human orthologs than with those found in other fungal phyla .Conclusion Two virulence-associated genes , Rad23 and Hst302, that determined the virulence difference among Cryptococcus neoformans strains of the two genotypes were detected by using sequencing analysis .The predicated structure and function of their encoded proteins were affected by the existence of nsSNPs to some extent . Key words: Cryptococcus neoformans var.grubii; Multilocus microsatellite genotyping; Viru-lence; Protein