Fungistatics

Transcription

Slide Culture Technique for Fungi Slide culture method is a rapid method of preparing fungal colonies for examination and identification. This method permits the fungi to be studied virtually insitu with as little disturbance as possible. Fungi are identified mostly by close examination of its morphology and the characteristics it possess. In slide cultures, we are growing the fungi directly on the slide on a thin film of agar. Materials required: 7-10 day old fungal culture Sterile sabourauds agar plate Sterile Petri dish in which a sterile filter paper of 9cm diameter is placed U-shaped glass rod placed in the petriplate Microscope slides and coverslips Scalpel and Inoculating loop 95% ethanol Lactophenol cotton blue stain Sterile distilled water Procedure for Slide Culture Preparation Arrange all the materials in the laminar chamber (DSCO6119) Aseptically, with a pair of forceps, place a sheet of sterile filter paper in a Petri dish Place a sterile U-shaped glass rod on the filter paper Pour sterile water on filter paper to completely moisten it. Gently flame a scalpel to sterilize and cut a 5 mm square block of the medium from the plate containing sterile Sabouraud’s agar Pick up the block of agar by inserting the scalpel and carefully transfer this block aseptically to the centre of the slide. Aseptically, place a sterile cover slip on the upper surface of the agar cube. Flame the inoculating loop until red hot Take the plate containing fungal culture to be examined Remove a small amount of fungal culture from the plate Inoculate four sides of the agar square with spores or mycelial fragments of the fungus to be examined Cover the Petri dish and incubate at room temperature for 48 hours. After 48 hours, examine the slide .Mycelial growth and spore producion is observed through the four corners the agar cube spreading over to the cover slip. Procedure for Application of Stain Place a drop of lactophenol cotton blue stain on a clean microscope slide. Remove the cover glass from the slide culture and discard the block of agar place the cover glass with mold grown side down, on the drop of lactophenol cotton blue stain on the slide. Examine the slide under low power and then high power objective of the microscope. Morphological features of the Fungi was observed. Slide culture method is more advantageous as there is no need to remove a portion of the fungus from a culture plate and transfer it to the slide. Thus there is less chance for the features that are key to identification, notably the spore-bearing structures, to be damaged. • By doing this, there is no need to remove a portion of the fungus from a culture plate and transfer it to the slide. So there is less chance for the features that are key to identification, notably the spore-bearing structures, to be damaged.