Adhesion Of Different Clinical Forms Of Sporothrix To Extracellular Matrix Proteins

Sporotrichosis is a common benign subacute or chronic deep mycosisin dermatology which is caused by Sporothrix. This disease not only cancause skin lesions, but also can lead to mucous membranes, bones andeven internal organs and other systemic diseases. The clinicalmanifestations of Sporotrichosis is related to the severity of fungalinfections, it mainly depends on the number of pathogenic fungi, fungalvirulence, the treatment of the wound and the strength of immunity, etc.At present, a large number of studies have shown that the adherence ofSporothrix cell wall composition and several extracellular matrix (ECM)components is one of the induced virulence of pathogenic fungi. TheECM which can be adhered include TypeⅡcollagen, laminin, fibronectin,fibrinogen, thrombin allergenic proteins, etc. The yeast cells and hyphaeof Sporothrix could significantly adhere to type Ⅱ collagen, laminin andfibronectin, there is no significant evidence of adhesion of conidia andfibrinogen as well as thrombin allergenic proteins. The yeast cells can beadhered to the fibrinogen. But the yeast cells can be adhered to thefibrinogen. Lima have confirmed the adhesion of the yeast cells andhyphae of Sporothrix by indirect immunofluorescence and flowcytometry. The Sporothrix cell wall components adhere to the ECM plays a keyrole in the spread of disease, so the exploration of the relation betweenthe adhesion and the Sporothrix pathogenic virulence may be benefit forthe study of pathogenesis of Sporotrichosis.Objective:After culturing the different clinical forms of Sporothrix by thecorresponding culture media at appropriate temperature, respectively combine them with different ECM proteins, through the indirect immunofluorescence we can see the intensity of fluorescence which can reflect the strength of the adhesion of the pathogenic fungi. We can preliminary analyze the relationship between the adh-esion and isolates’s pathogenic virulence by this method.Methods:Culture the60Sporothrix clinical isolates (fixed30, lymphatic type30) on Sabouraud medium at25℃ambient temperature for2weeks,another60isolates (fixed30, lymphatic type30) on brain heart infusionagar medium at37℃for1week, then cultivate them into another brainheart infusion agar medium. Repeat the method above three times. Thenmake them into suspension to combine with fibronectin and laminin. Addthe specific antibodies then stain them by indirect immunofluorescence.The intensity of fluorescence can be observed by immunofluorescencemicroscopy. Make different grades according to the positive rate and the weak positive rate of each group. Count the number of cases of differentlevels in each group and the proportion of total. The results wereanalyzed using statistical software.Results:Experimental strains on Sabouraud medium at25℃and brain heartinfusion agar medium at37℃were growing rapidly. We can see thecolony growth on Sabouraud medium after3-4days. They were brown orwhite moist, smooth, yeast-like colonies. After two weeks we can see thecolony diameter significantly increased, color deepened and have beendark brown, marginal uplift, wrinkles, hair-like colonies; The milky whitecolonies can be seen after one week that grow on BHI, then turned tosmall round plaques, gradual integration into a film after two weeks.Divide this120Sporothrix clinical isolates into eight teams, then makethem into suspension to combine with the corresponding ECM. Makedifferent grades according to the positive rate and the weak positive rateof each group and analyze the results using statistical software. Thedifference was statistically significant（P＜0.05）. The positive rate oflymphatic type of mycelial phase and yeast phase combine withfibronectin and laminin was all higher than the fixed type. The same typebetween the mycelial phase and yeast phase combine with the two kindsof proteins, the positive rate of fluorescence was not significantlydifferent. Conclusion:We try to transforme the mycelial phase of Sporothrix into yeast phase,with conversion prolonged and the times of transfermation increasedfrequency, the number of yeast cells increased significantly. The positiverate of lymphatic type of mycelial phase and yeast phase combine withfibronectin and laminin was all higher than the fixed type. The same typebetween the mycelial phase and yeast phase combine with the two kindsof proteins, the positive rate of fluorescence was not significantlydifferent.