EXPT NO. 1.

Media Preparation and sterilization

Questions:

1. Why isn’t dry heat used to sterilize nutrient agar?

2. What method of sterilization is best to remove contaminants from culture medium containing
heat sensitive compounds such as vitamins and plant extracts?
3. When do you add heat sensitive components to culture media, before or after sterilization?
Explain your answer.
4. Why is agar not used as food by most bacteria?
5. What temperature and time are necessary to sterilize by dry heat in an oven?
6. Why is dry heat less efficient than moist heat as a means of sterilization?
7. Bacteria require a source of energy and building blocks including minerals. Which component of
nutrient broth supply these needs.

EXPT NO. 2 Culture of Microorganisms

1. Explain why the following steps are essential during subculturing

a. Flaming the inoculating loop before and after each inoculating procedure.
b. Cooling the inoculating loop prior to obtaining culture.
2. Cite the purposes of the subculturing procedure.
3. How are you able to determine the subcultures of the isolates were pure?
4. Why is a straight needle preferable to a loop for obtaining inoculum from streak plate source
5. Can a pure culture be prepared directly from a mixed broth or agar culture?

EXPT NO. 3 Determination of Microbial Load

Questions:

1. What are the advantages and disadvantages of the serial dilution agar plating procedure?
2. In tabular form give the advantage and disadvantage of the following enumeration procedure?
a. Turbidometric Method
b. Serial Dilution and Plating Method
c. MPN Method
3. Why is it necessary to work with diluted cultures for the successful performance of a spread
plate inoculum?
4. Explain how dilution of the culture to be inoculated is accomplished in the streak plate
procedure.
5. Explain why the lines of inoculum in area 3 spread apart?

EXPT NO.4 Staining

 A. Simple Staining
1. What is the purpose of heat fixation
2. Why are basic dyes more effective for bacterial staining than acidic dyes?
3. What is the purpose of simple staining procedure?
B. Negative Staining
1. Can methylene Blue be used in place of the nigrosine for negative staining.
2. What are the practica; advanatages of negative staining.
3. Why do bacteria remain in the negative staining procedure?

C. Gram Staining
1. Distinguish between simple and differentia; staining procedure/
2. Cite the purpose of the following reagents in a differential staining procedure.
a. Primary Stain
b. Counter Stain
c. Decolorizing Agent
d. Mordant
3. Explain the chemical basis of the gram reaction
4. What is the most crucial step in differential staining procedure. Explain
D. Acid Fast Staining
1. Why must heat be used with application of the primary stain during the acid fast staining.
2. Why is the application of heat not required during the application of counterstain
3. Why is acid alcohol and not ethyl alcohol is used as a decolorizing agent?
4. Indicate which group of organisms is best identified by this procedure.
E. Spore Staining
1. Why is heat necessary in spore staining?
2. Explain the function of tap water in spore staining.
3. Explain the properties that differentiate the spore from vegetative cell
4. Distinguish between sporogenesis and germination.
F. Capsular Staining
1. Explain the medical importance of a capsule.
2. Why must het fixation be omitted during the preparation of a smear for capsular staining?
3. Explain the function of copper sulfate in this procedure.

EXPT NO. 5 Biochemical Tests

Questions

1. In Hydrlolysis explain why iodine is used to determine the presence or absence of starch.
2. Cite the enzymes and the end products formed during degradation of starch.
3. Cite the metabolic products of tryptophan degradation.
4. In citrate Untilizationand Hydrogen Sulfide production Tests explain how citrate mayserve as an
energy source for some microorganisms
5. Account for the development of alkalinity in culture capable of using citrate as their carbon
source.

EXPT NO. 6 Nitrate Reduction

Questions

1. Explan the functions of Solution A (Sulfanilic Acid) and Solution B (NEDD)

2. Explain why the development of a red color on addition of Zinc is a negative test
3. Cite the possible products that may be formed as a result of nitrate activity.
4. What is the ecological advantage of nitrate resuction in aquatic system?
5. What is the disadvantage of denitrification in agriculture.

EXPT NO. 7 Anti-Microbial Screening

Questions

1. What metabolites are produced by microorganisms that inhibit associated microorganisms?

2. What are the requirements for an antibiotic to be suitable for medical use?
3. How do antibiotics compare with chemicals in the cure of human diseases?
4. Enumerate the different antibiotics and give the possible modes of action on each
microorganisms.