Practical No. 2 Culture Media and Laboratory Equipments

1. Practical No. 2 Culture Media and Laboratory Equipments Department of Microbiology College of Medicine

2. Knowing the microscopic morphology and staining characteristics of a microorganism present in a clinical specimen, the microbiologist can make appropriate decisions as to how it should be cultivated and what biologic properties must be demonstrated to identify it fully.

3. A suitable culture medium must be provided, and it must contain the nutrients essential for the growth of the microorganism to be studied. Most media designed for the initial growth and isolation of microorganisms are rich in protein components derived from animal meats. Many bacteria are unable to break down proteins to a utilizable form and must be provided with extracted or partially degraded protein material (peptides, proteoses, peptones, amino acids). Meat extracts or partially cooked meats are the basic nutrients of many culture media. Some carbohydrate and mineral salts are usually added as well. Such basal media may then be supplemented, or enriched, with blood, serum, vitamins, other carbohydrates and mineral salts, or particular amino acids as needed or indicated.

4. In this exercise, we will prepare a basic nutrient broth medium and also a nutrient agar from commercially available dehydrated stock mixtures containing all necessary ingredients except water. The term ''nutrient broth'' (or agar) refers specifically to basal media prepared from meat extracts, with a few other basic ingredients, but lacking special enrichment. We will also see how liquid and agar media are appropriately dispensed in flasks, bottles, or tubes for sterilization prior to use, and how a sterile agar medium is then poured aseptically into Petri dishes.

5. Materials: • Dehydrated nutrient agar • Dehydrated nutrient broth • A balance, and weighing filter papers • A 1-liter Erlenmeyer flask, cotton plugged • A 1-liter glass beaker • A 1-liter graduated cylinder • Glass stirring rode (at least 10 cm long) • 10-ml pipettes (cotton plugged) • Test tubes (screw capped or cotton plugged) • Petri dishes

6. Requirements for bacterial growth • Nutrients H2O, C-source, N-source,proteins ,sugars, Inorganic salts, • Temperature • pH • humidity culturemedium  Nutrients, pH incubator :37c,24hr

7. Culture medium • Is the mixture of various nutrients that is suitable for the growth of microorganisms. • Types of Culture Media based on the function and chemical components based on the physical state

8. Based on the chemical components: Synthetic medium:prepared manually , known concentration ,used for physiological studies. Semi Sythetic medium:prepared manually with known concentration and supplemented ,used for isolation and physiological studies. Natural Medium: yeast extract ,meat extract their concentration s are not known, used for growth.

9. Based on function: 1- Isolation medium :All nutrients used for growth of unknown bacteria.e.g. nutrients agar 2-Enriched Medium Additional or special nutrients(e.g., serum, growth factors, trace elements) are added to support some fastidious bacterial growth. e.g. blood agar.

10. 3-Selective Medium the medium that can prevent the certain bacterial growth while permitting others. e.g. SS agar

11. 4-Differential Medium Some special substrates and indicators are added into the media in order to produce a visual differentiation when several bacteria grow on the same kind of medium. e.g. EMB agar (Eosin-methylene blue agar).

12. 5-Biochemical test:different compounde. e.g.fermentation test, mediaTriple sugar iron slant

13. 6- maintenance media :preservation of bacteria for long period of time. 7-Antibiotic sensitivity test medium: Allows the growth of most bacteria .  e.g: Müeller-Hinton agar is most frequently used in this antibiotic susceptibility test. 

14. According to physical properties, media can be classified three types: 1. solid medium ( 2% of agar) agar plate /slant medium 2. semi-solid medium (1% of agar) 3. liquid medium 2 1 3 1