Cultivation of fungi by slide culture technique

 Aim

To observe morphological characteristics of Fungus

 

Principle

Fungi comprises both molds and yeasts. Molds are filamentous and multi cellular whereas yeasts are usually unicellular.  Fungi possess rigid cell walls containing chitin, mannan and other polysaccharides. They possess true nuclei.  Fungi are important in both beneficial and harmful ways. They act as decomposers, certain fungi are parasites, cause diseases in plants, humans and other animals, they are important in many industrial processes including the making of bread, wine, organic acids, antibiotics, etc. and are important research tools. 

    Morphologically, a mold or filamentous fungi consists of long, branched, threadlike filaments of cells called hyphae that form a mycelium, a tangled mass like aggregation.  

    The isolation, culture, and microscopic examination of molds require the use of suitable selective media and special microscopic slide techniques. The process of transferring hyphae to a slide for staining and observation will break up the structure of hyphae and sporangiophores, so the identification will be very difficult. This problem could be resolved by the slide culture technique where, sporangiophores, hyphae and spores remain intact during staining.

 

Materials required

Cultures - culture of Penicillium and Aspergillus

Reagents - lactophenol cotton blue stain, Ethyl alcohol

Equipments - Bunsen burner, inoculating needle, staining tray, microscope, sterile Petri dish, Filter paper, triangle shaped glass rod, Microscope slides, coverslips, Inoculating needle, forceps, etc.

 

Procedure

1. A triangle shaped glass rod, glass slide, cover slip and a moist piece of cotton was placed in a Petri dish and the set up was sterilized.

2. Using a flame sterilized scalpel, 5 mm square block of Sabouraud’s dextrose medium was cut from the plate of Sabouraud’s agar.

3. The block of agar was aseptically transferred to the slide.

4. The four sides of the agar square was inoculated with the fungus and the coverslip was placed on the upper surface of the inoculated agar block.

5. The set up was covered using the petri dish lid and kept at room temperature for 48 hours.

6. After incubation, the coverslip was carefully removed, the agar block separated from the glass and the coverslip and glass slide were stained separately using lactophenol cotton blue and observed under microscope.

 

Observation

The fungal growth in the first slide culture plate appeared first white and then blackish with a pale colour on the reverse side and colonies in the second slide culture plate appeared greenish blue in colour with a wrinkled appearance on the reverse side. 

 

 

 

Result

On microscopic observation, organisms from the first slide culture plate was identified as Aspergillus species and that from the second slide culture plate was identified as Penicillium species.