Following heating, discard the pape er form the slide surface and gently wash th
ID: 197886 • Letter: F
Question
Following heating, discard the pape er form the slide surface and gently wash the slide wit 4. running water 5, counter-stain the smear with 0.5% aqueous Safranin for l minute. 6. Tip the dye into the collection bowl. Rinse with water and carefully blot dry. 7. Examine your slide with the 100x oil immersion objective a. Remember to start at 4x and move you way up i n magnification to the 100x objective. Notes: Vegetative cells stain pink/red in this procedure, whereas endospores stain Some endospores may be seen within the former vegetative portion of the cell. **Also prepare Gram stains of the endospore forming and the non-spore forming bacteria f the cultures provided for this exercise. Questions 1) Can everything on your slides be identified by Gram stain? Explain your answer. 2) Why was a basic solution then an acidic solution used for the Acid-fast stain? 3) What environmental conditions might trigger endospore formation? sod spores spear diferent fiom the vegetative cells?Explanation / Answer
Gram stain is used primarily for making a preliminary morphologic identification and confirms the number of bacteria present in the sample. This method cannot be used solely for the identification of bacteria. It can distinguish between gram positive and gram negative bacteria; other bacteria are unable to take dye used in Gram staining and hence can't be seen. In laboratories for clinical microbiology, this method is used in combination with other molecular and traditional techniques in identifying bacteria. Moreover, cannot identify bacteria to the species level. So, Gram staining cannot identify every bacteria placed on the slide. Basic stain are used to stain negatively charged components such as bacterial cell. Examples: methylene blue, safranin, malachite green,basic fuschin, crystal violet. Acid-fast bacteria have mycolic acid in their cell walls which makes them impermeable to many staining procedures including acidic stain. Mycolic acid present in the cell wall gives bacteria negative charge, resistinng the penetration of acidic stain into the cell. Basic stain like carbol fuschin is lipid-soluble and contains phenol, which helps the stain penetrate the cell wall, making the acid-fast cells red in color and non acid fast cell undergoes decolorization and takes counterstain methylene blue and will appear blue. Endospores are a complex, multilayer encapsulated form of the organism consisting of polysaccharides, dipiclonic acid, and high calcium content. The unfavorable environmental coditions trigger endospore formation and helps bacteria to cope adverse conditions. The spores have no water, no ATP production, no macromolecular synthesis, and no active enzymes. Thus, endospores can remain stable in the environment under adverse conditions. Different microorganisms form "spores" or "cysts", but the endospores of low G+C Gram-positive bacteria are much resistant to adverse conditions.