Identification And Drug Sensitive Test Of Trichosporon Spp. Isolated From Giant Pandas

Giant pandas are rare and endangered mammals unique to China and are China's "national treasures." Studies had shown that trichosporium is a dominant genus of giant panda epidermis fungi that can cause local and deep infections.This research intended to study the interspecies identification,genotyping,susceptibility testing,morphological development and pathogenicity of Trichosporon spp.isolated from giant panda surface,so as to provided a reference for the diagnosis and treatment of giant panda trichosporosis.(1)This study was conducted through the PCR amplification,sequencing,and phylogenetic analysis of the ITS,D1/D2 and IGS1 locus of 29 Trichosporon species for interspecies identification and genetyping;The morphological development process was observed by the micro culture method;Twenty-nine isolates of Trichosporon were identified as 11 species by molecular biology methods,including Trichosporon laibachii(4 strains),Trichosporon gracile(1 strain),Trichosporon brassicae(1 strain),Trichosporon domesticum(1 strain),Trichosporon guehoae(3 strains),Trichosporon asteroides(5 strains),Trichosporon jirovecii(5 strains),Trichosporon cutaneum(1 strain),Trichosporon shinodae(1 strain),Trichosporon middelhovenii(3 strains)and Trichosporon monilliforme(4 strains).T.jirovecii and T.asteroides were dominant species(Account for 17% of all isolates),besides T.middelhovenii and T.shinodae were isolated from surface of animals for the first time.Four strains of T.laibachii(JYZ 3252,JYZ 921,JYZ321,and JYZ 912)and one strain of T.monilliforme(JYZ 372)were found to be novel genotypes,and T.jirovecii was from genotype1.T.asteroides was the same genotype as pathogen of a disseminated trichosporosis.Different species of Trichosporon had distinct morphological developmental processes and most of Trichosporon had typical features,such as Trichosporon gracile mycelium septation and short thick mycelium during the development of Trichosporon shinodae etc.These differences could be used as an important basis for interspecies identification.(2)Mice were artificially infected by skin rubbing,intraperitoneal injection and intradermal injection.The liver and skin were taken for pathological analysis.Pathogenicity studies showed that T.cutaneum,T.laibachii,T.asteroides,T.brassicae,T.guehoae,T.middelhovenii and T.shinodae all damaged liver and skin of mice,and the pathogenicity was strong.T.asteroides had the strongest pathogenicity and might provoke invasive infection.The pathological characteristics of different species of Trichosporon infection of the liver and skin were basically similar.Trichosporon mostly caused hepatocyte necrosis,hepatocyte space enlargement or hepatocyte swelling,skin lymphocyte infiltration,thickening of stratum corneum,and infected tissue had a strong tendency to hemorrhage.(3)Drug sensitive test of 29 isolates of Trichosporon used 9 kinds of clinical commonly antifungal medicine(Voriconazole,Ketoconazole,Clotrimazole,Miconazole,Fluconazole,Amphotericin B,Terbinafine,Flucytosine and Itraconazole)as qualitative drugs for the experiment by the Danish Rosco disc diffusion method;The drug sensitivity results showed that Voriconazole was the most effective for Trichosporon,Terbinafine was useless,Flucytosine had no indication.Disparity of drug resistance between strains was existing.T.shinodae was a strain that was resistant to terbinafine and flucytosine and moderately sensitive to fluconazole.The drug resistance of the other Trichosporon was weak.Conclusion:Twenty-nine isolates of Trichosporon were identified as 11 species.Four strains of T.laibachii and one of T.monilliforme(JYZ372)were found to be novel genotypes.Different species of Trichosporon had distinct morphological developmental processes which could be used as an important basis for interspecies identification.Pathogenicity studies showed that T.cutaneum,T.laibachii,T.asteroides,T.brassicae,T.guehoae,T.middelhovenii and T.shinodae all had stronger pathogenicity.The pathological characteristics of different species of Trichosporon were basically similar,and infected tissue had a strong tendency to hemorrhage.The drug sensitivity results showed that Voriconazole was the most effective for Trichosporon.Disparity of drug resistance between strains was existed.T.shinodae was more resistant,and the rest of Trichosporon spp.isolated from the giant pandas had weaker drug resistance.