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B. ISOLATING BACTERIA FROM VARIOUS NATURAL SOURCES 1.1 Bacterial Streak Plating (Plate Streak Method)

Introduction: Introduction In nature, whether in the soil, in a pond, on the surface of your skin, or in your gastrointestinal tract, many different kinds of bacteria grow together. To study or identify particular bacteria, you must be able to separate it from the others and grow it in pure culture. The most useful technique for separating bacteria is to streak for individual colonies, and is called the streak plate technique. This method allows the bacteria to be spread out so that a single bacterium can be isolated from all other bacteria.

Objective: Objective To study how to isolate individual bacterial colonies out of various sources, to characterize and identify the bacteria present. To dilute the bacteria to the point where individual bacteria are separate on Petri plate.

Materials and Methods:

Materials and Methods 5 sterile agar plate, water sources from the drain and pipe, sterile water, boiled water and milk, marker pen, wire inoculating loop, and spirit lamp. Use a marker pen to draw a T on the bottom of your Petri plate of nutrient agar as show below.

Set the plate you will be streaking so that its bottom is sitting on the bench top and you can see the T clearly. Rename your Petri plate. Get an inoculating loop and spirit lamp. Light the spirit lamp. Sterilize the loop by holding its tip in the flame until it turns red. Wait a minute until the loop cool before placing it into the bacterial source. Place the inoculating loop into one of the liquid cultures and withdraw the loop. Obtain your Petri plate with agar and lift the lid slightly and remove the lid after complete. Streak the loop containing the bacteria at the top end of the agar plate moving in a zig-zag horizontal pattern. Sterilize the loop again in the flame and let it cool before continuing to spread the bacteria. Rotate the plate about 90 degrees and spread the bacteria from the first streak into a second area using the same zig-zag spread technique. Sterilize the loop again. Rotate the plate about 90 degrees and spread the bacteria from the second streak into the 3rd area in the same pattern. Leave plate in inverted position at room temperature. Record your observation after 48 hours and until 5 days later. Repeat with other water sample


Results and Discussion Petri plate The condition of colonies Colony appearance on agar plate

Day 2 Water from pond Sterile water Milk Mix growth Not present Mix growth

Day 3 Mix growth Not present Mix growth

Day 4 Mix growth Not present Mix growth

Day 5 Mix growth Not present Mix growth Colony : white : tiny yellow : small Clony : white : big yellow : small

Conclusion: Conclusion

Bacteria can exist in any substances. But not all substances containing bacteria like a sterile mater because the sterile was killed the bacteria in water. Bacteria growth when the nutrient is form in the substance. Like milk is rich with nutrient. Question 4 (a) Describe: (i)The colour and whether or not the bacteria colony in opaque, transparent, or translucent Answer: ----------------------------------------(ii) The general bacteria colony appearance, margin, and elevation of the colony (based upon the charts below) Answer: -----------------------------------------

(b) Observe your plates and answer the following questions. (i) What is meant by colony in bacteriology? Answer: Colony is a group of bacteria. Why do you need to isolate the colony? Answer: We need to isolate the colony to individual bacterial (in relative) out of various sources, to characterize and identify the bacteria present.

(iii) Why was the inoculating loop cooled before inserting into sample? Answer: Inoculating loop need to cool before inserting into sample to avoid any organism to be tested not is killed by the hot wire. (iv) Why was the Petri plate inverted during incubation? Answer: Petri plates need to invert during incubation to avoid the water vapour.