STREAK PLATE TECHNIQUE

STREAK PLATE TECHNIQUE

AIM

      To isolate the pure strains of bacteria by Streak plate technique (Quadrant streaking method).

PRINCIPLE   

          The Streak plate technique is a rapid qualitative isolation method used for the isolation of a pure culture of the bacteria from the mixed population.  Streaking method was first developed by Loeffler and Gaffky in Robert Koch’s laboratory, which involves the dilution of bacteria by systematically streaking them over the exterior of the agar in a Petridish to obtain isolated colonies which will then grow into the number of cells or isolated colonies. The inoculum is streaked over the agar surface in such a way that it “thins out” the bacteria. Some individual bacterial cells are separated and well-spaced from each other. Aseptic techniques are used to maintain microbiological cultures and to prevent contamination of the growth medium.

        Many type of procedures are performed for the Streak plate technique, but the four ways or Quadrant streak is mostly done. As the original sample is diluted by streaking it over successive quadrants, the number of organisms decreases. Usually, by the third or fourth quadrant, only a few organisms are transferred which will give discrete colony forming units (cfu). When the bacteria are streaked and isolated, the causative agent of a bacterial disease can be identified. Apart from the Quadrant streaking method, some other Streaking methods also used in the Microbiology Laboratory. They are Continuous streaking, T streaking, Radiant streaking and Spiral streaking.

            Streak plating method that has two major disadvantages. They are

a)    Firstly, users will not be able to grow obligate anaerobes using this method.

b)    Secondly, only organisms that were viable in the original sample are able to be grown.  

MATERIALS REQUIRED

• Bacterial culture
• Inoculation Loop
• Bunsen burner
• Nutrient agar plates
• Incubator

PROCEDURE

a)  Sterilize the inoculating loop in the Bunsen burner by putting the loop into the flame until it is red hot. Allow it to cool.

b)  Pick an isolated colony from the agar plate culture and spread it over the first quadrant (approximately 1/4 of the plate) using close parallel streaks.

c)  Turn the Nutrient agar plate at 90° angle and lightly sweep the loop 1-2 times through the inoculated area, then streak into the next quadrant without overlapping the previous streaks (Step - 1).

d)   Flame the loop and allow the loop to cool.

e)  Turn the Nutrient agar plate at 90° angle and lightly sweep the loop 1-2 times through the inoculated area, then streak into the next quadrant without overlapping the previous streaks (Step - 2).

f)    Flame the loop and allow the loop to cool.

g)   Turn the Nutrient agar plate at 90° angle and lightly sweep the loop 1-2 times through the inoculated area, then streak into the next quadrant without overlapping the previous streaks (Step - 3).

h)   Flame the loop and allow the loop to cool.

i)  Invert the streaked Nutrient agar plate and incubate at 37 °C for 24 hours in an Incubator.

 

OBSERVATION AND RESULTS

       The streaked Nutrient agar plate is incubated at 37 °C for 24 hours. After incubation, examine the bacterial colonies grown in the plate carefully. All colonies should have the same general appearance. If there is more than one type of colony, each type should be streaked again on a separate plate to obtain a pure culture.

Figure – 1: Serial dilution technique