The Study On Oxidation Resistance And Biological Characteristics Of Trichosporon Asahii After In-vitro Induction

RESEARCH BACKGROUNDThe oxidative stress of fungi is closely related to its infection mechanism,which is also a hot spot of fungal research in recent years.To obtain stable antioxidant strains is a prerequisite for further study on its antioxidant mechanism.Trichosporon asahii（T.asahii）is one of the most important opportunistic pathogens.However,there are few studies on the mechanism of Trichosporon infection.To study whether common oxidants H₂O₂ and Menadione sodium bisulfate（MSB）can induce oxidation resistance of T.asahii in vitro.Recovery-test was performed to observed the stability of strains’ oxidative ability.Changes of physiological characteristics of the strains were also observed.METHODS1.T.asahii clinical standard strain CBS2479 and environmental strain CBS8904 were experim entally serially subcultured in YPD medium containning increasing concentrations of H₂O₂ or MSB until not induced.The resisitant filial generations were subcultured in drug-free medium for 10 times consecutively.The activities of catalase（CAT）and superoxide dismutase（SOD）were determined in different stages of the strains.Enzyme assay data were analyzed by SPSS20.0 software.2.The morphology of different stages of strains was observed using the naked eye and light microscope.3.The MIC of fluconazole,H₂O₂ and MSB for different stages of strains were determined according to the Clinical and Laboratory Standards Institute standard M27-A3.RESULTS1.Determination of antioxidant enzyme activity After induced by H₂O₂,the CAT and SOD values of the CBS2479 resisitant filial strains were increased by about 2.15 and 2.97 times,respectively.The CAT and SOD values of the CBS8904 resisitant filial strains were both increased by about 1.79 times.After induced by MSB,the CAT and SOD values of CBS2479 resisitant filial strains were increased by about 3.36 and 2.68 times respectively.The CAT and SOD values of the CBS8904 resisitant filial strains were increased by about 2.27 and 2.57 times,respectively.After induced by H₂O₂ or MSB,CAT and SOD activity of CBS2479 were increased significantly（P<0.05）.The CAT and SOD after recovery were higher than those before induction（P<0.05）.The CAT of CBS2479-H₂O₂ and CBS2479-MSB strains after recovery-test were lower than that of induction of telophase（P<0.05）,but there were no statistical differences in SOD.After induced by H₂O₂ or MSB,CAT and SOD activity of CBS8904 were increased significantly（P<0.05）.The SOD of the CBS8904-H₂O₂ strains were higher than that before induction（P<0.05）,but there were no significant differences between the CAT before and after treatment（P>0.05）.The CAT and SOD values of CBS8904-MSB strains were higher than those before induction（P<0.05）.The CAT of CBS8904-H₂O₂ and CBS8904-MSB strains after recovery-test and the SOD of CBS8904-MSB after recovery-test were lower than that at the end of induction（P<0.05）,but there were no significant differences of SOD between CBS8904-H₂O₂ after recovery-test and the induction of the final stage（P>0.05）.2.Morphological change After induction,various strains’ morphology of the naked eye changed.Under the light microscope,the mixed state of the pseudo hyphae and spores appeared after induction.3.Determination of drug sensitivity After induction,the MIC values of fluconazole,H₂O₂ and MSB were 2 times as much as that before induction.After recovery-test,the MICH₂O₂ of CBS8904-H₂O₂ and CBS8904-MSB strains were all restored to the level before induction,while the others were the same as the MIC value of the fourth generation.CONCLUSION1.The stable H₂O₂-resistant,MSB-resistant T.asahii strains were successfully induced and obtained by in vitro tests.The antioxidant capacity was relatively stable.2.After induction,various strains’ morphology of the naked eye was changed.The morphology of light microscope also changed:the mixed state of the pseudo hyphae and spores appeared.3.After induction,the tolerance of the strains to fluconazole increased.The tolerance to H₂O₂ and MSB was higher than that before induction.