Investigation On The Different Morphological Features, Protein Profilings And DRK1 MRNA Level Of Sporothrix Schenckii During Dimorphic Switch

Sporotrichosis is a common cutaneous mycosis which is caused by dimorphic fungus, Sporothrix schenckii(S. schenckii). S. schenckii exhibits a temperature- dependent dimorphic switch. At 25°C it grows as as mycelial phase, whilst at 37°C it forms yeast cells characterized by unicellular conidia. The formation of yeast cells was thought to be a requisite for the pathogenicity of S. schenckii. Objectivity: To investigate the different morphological features, protein profilings and DRK1 mRNA level of S. schenckii during the transition from mould phase to the yeast one.Methods:1, The S. schenckii isolate, ATCC10268, was incubated on Sabouraud’s fluid medium at 25℃for 96H,and then transferred to brain heart infusion liquid medium at 37°C for 24,36 and 48h. The cells were collected and viewed using optical microscope and electron microscope.2, Whole cell proteins from the mould and early stage of yeast in S. schenckii were seperated using 2DE and analyzed with PD Quest Software. 30 selected yeast-specific proteins were identified by peptide mass fingerprinting and interrogated by using the MASCOT search engine 3, cDNA of histidine kinase DRK1 was amplified by RACE PCR technique and sequenced. Bioedit software was used to obtain amino acid sequence and determine the domain structure.Results:1, Cultures of S. schenckii incubated in Sabouraud fluid medium at 25°C and brain heart infusion agar at 37°C exhibited distinct morphological differences.2, Among thousands of protein molecules displayed, more than 300 showed differential expression between mould and early yeast phases. 24 y identified east-phase proteins are related to metabolism, signaling, meiotic mismatch and etc.3 ORF of DRK1 contains 3957bp coding 1319 amino acid. ORFA has the domain structure and sequence of histidine kinase is conserved in dimorphic fungi. ORFA has a histidine-containing H-box, an aspartate-containing D-box, and G-and N-boxes.4,Compared with mould phase of S. schenckii, mRNA level of DRK1 was up-regulated in yeast phase.Conclusions:1, Morphological features of yeast phase appeared when mycelial phase of S. schenckii was transferred to brain heart infusion liquid medium at 37°C for 36H. 2,There were distict differences between protein profilings of S. schenckii’s mould phase and yeast one.3, Histidine kinase DRK1 and other 23 identified proteins were overexpressed in yeast phase of S. schenckii, probably involving the pathogenesis of sporotrichosis.4, ORFA of DRK1 containing 1319 amino acid has the domain structure and sequence of histidine kinase is conserved in dimorphic fungi.