Sampling Methods for Microbiological Examination of Air
Microorganisms in air cause a problem of contamination in food industries, milk processing plants, industrial processes, pathological and research laboratories and mainly in operation theatres.
Therefore, measurement of no. of organisms in air at such places is important. Microorganisms in air may be present in free form or in the form of clumps or chains.
They may adhere to the dust particles or to saliva or to mucus of man. Several methods have been developed for the microbiological examination of air.
Simplest method is exposure of sterile nutrient medium plates to air for different time periods The plates are then incubated during which the settled organisms grow and form colonies.
The number of organisms can be calculated from the number of colonies. But this gives a rough idea about the numbers and kinds of microorganisms in air. Because the volume of air coming in contact with plate is not known.
Therefore, the advanced techniques like solid impaction, liquid impingement, filtration, sedimentation, centrifugation, electrostatic precipitation, thermal precipitation etc. are used. Among these, solid impaction and liquid impingement techniques are most commonly used.
1. Solid Impaction (Sieve Device) :
- Number of sampling devices which work on the principle of impaction of airborne particles.
- In these, an aspirator sucks a known volume of air which collides with nutrient agar in plate.
- This causes the microbes in the air to stick to the surface of an agar medium.
- The plates are then incubated at specific temperature for a specific time to form the colonies.
- These are the most often used methods of detection and enumeration of microbes in air.
- The most widely known device that is based on the solid impaction is the Sieve Device.
- The sieve device is of two types : Single Stage, Multistage Device
a). Single Stage Device :
- It is mechanically very simple.
- It consists of a metal box with a metal cover.
- The cover has a large number of evenly spaced small holes.
- Under this a petriplate containing agar medium can be kept.
- A measured volume of air is drawn through these hole. The particles containing microorganisms are allowed to settle on the agar surface.
- Plates are then incubated for specific time. Then microbial colonies on agar surface are counted.
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| Andersen's Air sampler & Single Stage Sieve Device |
b). Multistage Device :
- It consists of perforated plates with progressively smaller holes at each stage.
- Allows particles to be separated according to size.
- Best known multi-stage samplers is Andersen's air sampler
- In this, air is drawn through a series of six circular plates each perforated with 400 holes.
- The plates in series have progressively smaller holes.
- Therefore, the largest particles being deposited in the first while the smallest in last petri dish.
- Viable particles can be collected on a variety of bacteriological agar media.
- Incubated for counting and identification.
- All the particles collected are sized aerodynamically and can be directly related to human lung deposition.
Advantages of Sieve Device
- The volume of air to be analyzed can be measured.
- An air is drawn with high speed. Therefore, the particles of smaller size that might be escaped can also be settled on agar surface.
- There will be even distribution of air sample on agar surfaces that results in formation of isolated colonies.
- This is helpful to take a colony count.
- The methods are very simple and efficient.
- Therefore, they are most widely used.
2. Liquid Impingement Device :
- In this, a known volume of air is passed in a sterile broth medium, where micro-organisms from air are trapped.
- The aliquots of broth are then used for plating.
- The plates are incubated at room temperature for specific time and the colony count is taken to determine the number of micro-organism.
- There are several devices, among which the most important one is Bead Bubbler Device.
Bead Bubbler Device
- It consists of 250 ml capacity suction flask with side arm linked to suction pump and flow meter.
- The flask is filled with sterile nutrient broth.
- The sterile glass beads of size 5 mm in diameter are also placed in the flask.
- A glass bubbler is fitted in the flask with rubber stopper.
- Diameter of glass bubbler is approximately 14 mm.
- It has many small openings at the base with3 mm diameter.
- A suction flask is added with sterile nutrient broth medium along with sterile glass beads.
- The bubbler is fitted with rubber stopper.
- With help of suction pump and flow meter, known volume of air is allowed to pass through medium.
- Air enters into broth medium in the form of bubbles.
- Because of glass beads bubbles are formed and the clumps or chains of organisms are broken down.
- Aliquots of medium are used to culture on solid agar medium & plates are incubated at room temperature.
- Colony count is taken & number of organisms is determined.
Advantages
- Volume of air to be analyzed can be measured
- Because of glass beads and bubble formed, the micro-organisms from the clumps and chains are separated from each other.
- Thus exact count can be drawn.
