Fungal Staining

      Aim

To perform fungal staining and to observe fungi 

       Principle

The lactophenol cotton blue (LPCB) wet mount is most widely used method for the staining and observation of fungi.  The LPCB stain has following three components, Phenol to kill any live organism, Lactic acid to preserve fungal structures and Cotton blue to stain the chitin and cellulose of the fungal cell wall in blue colour. 

       Materials required

1. Cultures – Cultures of Penicillium, Aspergillus

2. Reagents – lactophenol cotton blue

3. Equipments – Glass slides, cover slips, Bunsen burner, inoculating needle, staining tray, microscope

 

           Procedure

1. A drop of lacto phenol cotton blue was put on a clean microscopic glass slide
2. A portion of mycelial mat from the fungal culture was transferred in the drop of lactophenol cotton blue using a flamed and cooled inoculation needle.
3. The fungal mycelia were gently spread and mixed with the stain using the inoculation needle. 
4. A cover slip was kept on the surface of slide and observed under microscope.              

Observation 

The slide was observed under low and high power objectives of the microscope and type of spores and arrangement of hyphae noted.

Result

Aspergillus spp. is  recognized by its conidiophores terminating in an apical vesicle and phialides are attached to the vesicle and bear conidia in chains. 

Penicillium spp. is identified by conidiophores with secondary branches that give it a brush-like appearance. Spores (conidia) are produced in chains from the tips of the phialides.

                                                                      Aspergillus

                                                                             Penicillium