Streak Plate Method Procedure and Results (Types & Diagram)

The word “streak” means “a long, thin line.” In the streak plate method procedure, a sample is spread in a petri dish in the shape of a long, thin line over the surface of solid media.

What is Streak Plate Method?

The technique of streak plate is used to isolate organisms (mostly bacteria) from a mixed population into pure cultures. The inoculum is applied to the surface of the agar in a manner that “thins out” the bacteria. Some bacterial cells are well-spaced and separated from one another.

The number of organisms decreases as the original sample is diluted by spotting it across successive quadrants. By the third or fourth quadrant, typically only a few organisms are transferred, resulting in discrete colony forming units (CFUs).

Streak Plate Method Procedure 2

Figure 1: Streak Plate Method

Objectives of Streak Plate Method

  1. From a mixed culture of bacteria, to get a pure culture
  2. To get colonies that are well-separated
  3. To distribute bacteria

Principle of Streak Plate Method

The inoculum is diluted down by smearing it across the agar plate. While streaking in different parts of the plate, the inoculum is diluted until there is only one bacterial cell on the surface of the agar plate every few millimetres. When these single bacteria cells divide and make thousands and thousands of new bacteria cells, an isolated colony is made. Pure cultures can be made by taking colonies that are well-separated and spreading them out again on new agar plates.

Types of Streak Plate Method

Based on the pattern of streaking, the streak plate method can be classified into 6 types, viz.: Quadrant Streaking, T-Streaking, Continuous Streaking, Radiant Streaking, Semi-quantitative Streaking, Zigzag Streaking.

1. Quadrant Streaking

It is the most common and preferred method. Four sections of the agar plate of the same size are streaked. It is also called the “four-quadrant streak” method, the “four sectors” method, and the “four-way streak” method. In this method, each plate is split into four equal parts, and each part is streaked one after the other. The area that gets streaked first is called the “first sector” or “first quadrant,” and it has the most inoculum. Over time, the inoculum in the second, third, and fourth quadrants will get weaker. By the time the fourth quadrant is streaked, the inoculum has been diluted a lot. This means that after the incubation, the colonies will be spread out.

2. T-Streaking

By drawing a “T” on the media, it is split into three sections, and each section is streaked in order. By the time the last section is streaked, the inoculum has been diluted enough that after the incubation, it will grow into separate colonies. Most of the time, streaking is done in a way that’s not continuous. However, if the sample is very dilute, a continuous pattern can also be used.

3. Continuous Streaking

It is another method that is often used. An inoculum is spread evenly from the beginning to the middle of the plate in one continuous motion. During the process, there is no need to split the plate and clean the loop. It is easy and quick, but we can only use it if the inoculum is very diluted or if we need to spread a pure culture instead of isolating one.

4. Radiant Streaking

It is another way to streak. First, the inoculum is spread along one edge, and then it is spread in vertical lines above the edge. Lastly, the lines going up and down are crossed with diagonal lines. This method is suitable for propagating both pure culture and diluted specimens.

5. Semi-quantitative Streaking

This is a variant of continuous streaking. In this method, a certain volume of the liquid specimen is streaked using a calibrated loop. A loopful of the specimen is streaked in a horizontal line in the middle of the Petri plate, and the specimen is spread evenly across the plate with a single back-and-forth motion. This technique permits us to estimate the viable load (in a range, not an exact number) and obtain the pure culture in a single step.

6. Zigzag Streaking

It is another type of continuous streaking in which a loopful of the specimen is streaked in a zigzag pattern across the plate in a single continuous motion. It is common practise to propagate and cultivate large quantities of pure cultures.

Streak Plate Method Requirements

1. Streaking Tool

This sterile instrument is used to streak a specimen across the surface of culture media. Cotton swabs, inoculating loops (both metal and plastic), toothpicks, and wooden, metal, or plastic sticks/wires are used for streaking. The most prevalent is the inoculating loop (nichrome wire loop).

2. Sample culture

Bacteria samples may be in suspension, liquid broth, or colonies on solid media. For streaking, the sample is collected using an inoculating loop and transferred to the surface of fresh culture media.

3. Culture Media

Different types of culture media are used to separate and identify suspected (or known) bacteria. The culture medium is a solid agar that has already hardened before it is used.

4. Bunsen Burner

A Bunsen burner is used to sterilise the loop and make a sterile area around the flame. There are also other chemicals, materials for sterilising, and lab equipment that are needed.

Streak Plate Method Procedure

  1. Set up everything you need, put on your Personal protection equipment, clean the work area, and let all the samples and media that were in the fridge come to room temperature.
  2. If the sample is very strong, diluting it can help get the colonies separated. (But you don’t have to, since the sample will be diluted in the next step.)
  3. Set fire to the inoculating loop and let it cool down. Choose a small part of the colony that is on its own. (Take a loopful of the sample if it is in the suspension.)
Streak Plate Method Procedure

Figure 2: Streak Plate Method Procedure.

1. General Procedure

  1. There is a label on the bottom of the Petri dish. Typical labels include the organism’s name and the date.
  2. The nichrome wire loop should be sterilised using the Bunsen burner flame.
  3. With the aid of a sterile nichrome wire loop, remove a sample of bacterial culture from the culture tube.
  4. Apply a bacterial culture to the nutrient agar plate. The agar plate’s lid must be removed between the Bunsen burner and the labelled quadrants must be streaked out.
  5. In a laminar airflow cabinet, the entire process is performed under strict aseptic conditions.

2. Quadrant Streaking

  1. Nichrome wire loop must be sterilised using a Bunsen burner flame.
  2. Cool the loop of wire between the burners.
  3. Label petri dish with a marker and indicate the four quadrants.
  4. Heat the test tube containing the bacterial culture.
  5. Insert the wire loop made of nichrome into the culture tube.
  6. Spread bacterial suspension between the four-quadrant of the plate.
  7. Plates are incubated at 37°C for 24 hours.

3. T-Streaking

  1. The nichrome wire loop must be sterilised using a Bunsen burner flame.
  2. The wire loop between the Bunsen burner must be cooled.
  3. The wire loop is dipped into the broth culture containing the bacterial mixture.
  4. Insert the nichrome wire loop onto the nutrient agar plate, then streak the bacterial suspension in a zigzag pattern to create a T-shaped pattern.
  5. After 24 hours of incubation, you will observe isolated colonies in the third sector. There will be less expansion in the second sector and the greatest expansion in the first.

4. Continuous Streaking

  1. Hold the Petri plate at an angle of 60° with your left hand. (If you write with your left hand, hold the plate with your right).
  2. Put the loop at one end of the plate and start streaking the inoculum in a straight line toward the middle of the plate.
  3. Turn the plate by 180°, and then follow step (ii) to streak the other half of the plate without sterilising the loop.

5. Radiant Streaking

  1. Hold the Petri plate at an angle of 60° with your left hand. (If you write with your left hand, hold the plate with your right).
  2. Use a gentle zigzag motion to spread the inoculum over the near edge of the agar plate.
  3. Clean the loop and let it cool down. Then, the streak from the starting point moved outward in a circle until it reached the far edge of the Petri dish.
  4. Relight the loop and let it cool down. Then, draw lines that go across the radial lines.

6. Semi-quantitative

  1. Hold the Petri plate at an angle of 60° with your left hand. Take a loop (urine) of the sample using a calibrated loop.
  2. Draw the sample into a vertical or horizontal line in the middle of the plate. This line is called the “primary streak.”
  3. Using the same loop, the inoculum was spread by moving it back and forth (zigzagging) over the primary streak.

Result of Streak Plate Method

The streaked plate is incubated at 37°C for 24 hours. Examine the colonies grown on 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.

Tips for Streak Plate Method

  1. Use only a small amount of inoculum.
  2. Streak lightly so that you do not gouge the agar.
  3. Flame the loop after you streak each quadrant.
  4. Make sure the surface of the plate is free of droplets of condensed moisture.

Applications of Streak Plate Method

  1. Used to separate a pure culture from a mixed culture so that morphological, biochemical, and molecular tests and other applications can be conducted.
  2. Used to classify specimens as pure or hybrid species.
  3. Used for studying bacterial colony characteristics.
  4. Utilized to create a population of genetically identical individuals.
  5. Used for inoculating clinical specimens in diagnostic laboratories to cultivate isolated pathogen colonies.
  6. Used in urine culture to isolate pathogens and semi-quantify uropathogens to determine infection severity. This is achieved using a calibrated loop.

Advantages of Streak Plate Method

  1. Inoculation using this method is straightforward, dependable, uncomplicated, and straightforward to carry out.
  2. It makes well-separated colonies of people who are genetically the same, so we can do more tests and applications on the isolates. So, it is used to make a clinical diagnosis.
  3. Since dilution is part of the process of inoculation (or “streaking”), there is no need to dilute the sample separately.
  4. Permit the user to manually control the sample, as well as the sample size and the inoculating area in a petri dish.
  5. The cultivation of aerobic organisms can be accomplished in a suitable manner that requires less time.
  6. Different patterns of streaking make it possible to choose the right method based on sample size, the number of Petri dishes available, and other factors.
  7. We can use both a sample from a broth or suspension and a colony from a solid medium.

Limitations of Streak Plate Method

  1. Only aerobic or facultative aerobic bacterial isolates could be grown. The primary suspension should contain the viable (living) bacterium.
  2. There is a chance of tearing the agar surface during streaking if one is not skilled enough, and the media is freshly prepared.
  3. It is unsuitable if the sample size is large and has a very high viable count. If we take heavy inoculum there may not be isolated colonies following the incubation.