Bacteria and other microbes have particular requirements for growth When they reside in and on our bodies or in the environment, they harvest their

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1 Bacteria and other microbes have particular requirements for growth When they reside in and on our bodies or in the environment, they harvest their food from us or from the environment When we grow bacteria in lab, we are essentially creating a captive environment for bacteria like a bacteria zoo. So we must provide the bacteria we grow in lab with all of the materials that they need to grow

2 In this lab we learn about different types of media that are used to grow bacteria. Some types of media will grow just about any type of bacteria and the others are more selective and only grow specific types of bacteria

3 There are generalized media, like (Nutrient agar)that will grow many different types of microbes. This media is the type most often used to culture bacteria

4 culture medium that allows the growth types of organisms, while inhibiting the growth of other organisms Example: EMB (Eosin Methylene Blue) dyes inhibit Gram (+) bacteria selects for Gram (-) bacteria

5 culture medium that includes ingredients, such as chemical indicators, that produce observable differences between species of bacteria Differentiates between different organisms growing on the same plate Example: Blood agar

6 This media is differential because: Certain bacteria produce enzymes (hemolysins ) that act on the red cells to produce either: Beta hemolysis: Enzymes lyse the blood cells completely, producing a clear area around the colony. Alpha hemolysis: Incomplete hemolysis produces a greenish discoloration around the colony Gamma hemolysis: No effect on the red cells.

7 There are also (selective) and (differential)media

8 MacConkey s is both a selective & differential media. 1. MacConkey s is selective media because it inhibits the growth of some organisms [Gram positive bacteria]. 2. MacConkey s is differential media - lactose fermenters bacteria will grow in red colonies while non-lactose fermenters will be colorless and clear. So if there are colonies of bacteria growing on MacConkey s, it s understood that they are Gram- If those colonies are colorless, they are not lactose fermenters If the colonies have a pinkish appearance, they are lactose fermenters

9

10 MacConkey Agar left: no lactose fermentation right: lactose fermentation

11 FORMS OF CULTURE MEDIA broth liquid medium most common growth media for microorganisms are ( nutrient broths ( liquid nutrient medium

12 Solid media commonly contain 1.5% agar per weight to solidify the liquid media. After sterilization, the media is poured into sterile Petrie plates. Agar is liquefies at 100 C and solidifies at 40 C

13 General Media: Nutrient Broth and Nutrient Agar

14 or After autoclaving the media(in tube) for 20 minutes, the tubes are placed in a slanted position to allow the agar to solidify. These tubes are called slants

15 Slant tubes: are tubes containing a nutrient medium plus a solidifying agent, (agar-agar. The medium has been allowed to solidify at an angle in order to get a flat inoculating surface Agar Plate: are sterile petri plates that are filled with a melted sterile agar medium Microorganisms grow on the surface of agar plates and slants

16 140 Micro Lab 4: Isolation of microbes from different environments

17 Isolation of micro organisms from various environments

18 1. Isolation separating one species from another. 2. Inoculation introduction of a sample into a container of media to produce a culture of observable growth. 3. Incubation conditions that allow growth eg., temperature,humidity etc.. 4. Inspection. 5. Identification.

19 Environmental Sample Normal Flora Samples Surface samples are normally taken using sterile swabs Applying oral sample to surface of agar

20

21 Petri plates with media Soil Rotten fruit Yoghurt Mouth swab etc Incubator

22 Bacteria Fungi

23 Isolation of Bacteria Yoghurt Put a drop of diluted yoghurt on the solid media plate of bacteria. Incubate at 37 C for 1 day. Mouth Take some saliva with the help of a cotton swab. Inoculate it on the media and incubate. Hand Touch the surface of the solid media plate. Incubate 37 C for 1 day..

24 Agar plates are stored upside down to prevent condensation.

25 These plates of bacteria will be incubated at 37 C for 24 hours and then stored at refrigerator until next week when you will observe for results.

26 Isolation of Fungi Air Expose the prepared plate of solid media in air for 5 min Close the lid and incubate at 28 C. After 2-5 days fungus observe for grow Soil Sprinkle a pinch of soil on the solid media plate Close the lid and incubate at 28 C After 2-3 days fungus observe for growth. fruit Clean the rotten part of fruit with alcohol Cut a piece and inoculate it on the media plate Then incubate,after 2-5 days fungus will grow

27 These plates of fungus will be incubated straight at 28 C for 24 hours and then stored at refrigerator until next week when you will observe for results.

28

29 Bacteria Fungi

30 أ.شروق الشهراني

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