Were identifying an unknown bacteria in lab What I want to k

We\'re identifying an unknown bacteria in lab. What I want to know is the best course of action, the tests to perform to identify the bacteria without doing every single one under the sun. I know that a good starting point is going to be the Gram stain.

Solution

Identification of an unknown bacteria in the lab is based on series of physical, biochemical and molecular reations and observations. Classification is usually based on staining methods, techniques of isolation, bacterial nutritional properties and its biochemical activities as well as its growth characteristics. Listed below are some identification methods;

1. Study of morphology of culture: This includes study of form & elevation, margin of colony, its surface, optical characteristics and also the pigmentation (if any!).

2. Study of cell morphology: Culture morphological features study is followed by study of individual cells of the colony. eg. their shape (coccus, round, bacillus, spiral etc). etc

3. Staining: Different methods of staining bacteria exist. Staining the colourless bacterial cells with methylene blue, carbon fushin or crystal violet (wherein the dying solutions could be acidic or basic) helps in study of morphological characteristics.

Gram stain is used to distinguish between gram positive bacteria (with thickened peptidoglycan layer that holds the primary stain, crystal violet) and gram negative bacteria (thick peptidoglycan layer absent and lose the stain when treated with a decolourizer such as absolute alcohol). Gram-positive cells become purple and gram negative cells become red with secondary stain like iodine.

Acid fast stain differentiates bacterial cells containing lipoidal cell wall. Also, other staining methods are the cell wall stain which stain the cell walls of bacteria comprised of  lipopolysaccharides, phospholipids, and peptidoglycan; spore stain for detection of bacterial species producing spores; capsule stain; flagella stain etc.

4. Respiration test: This determines whether the bacteria is aerobic or anaerobic. Here the bacterial species is made to grow in absence of oxygen (anaerobic conditions) to see whether it can survive or not.

5. Biochemical tests: These tests are the final steps to determining the bacterial species. Here bacterium is tested if it can hydrolyse protein, starch or lipid; can do nitrate reduction or not or have hemolytic propeties or not. These tests are done by varying the medium on the agar plates.

6. Sum up the infomartion obtained from series of observations and tests to identify the unknown bacteria.