Microbiological examination of food

Microbiological analysis of food products is the use of biological, biochemical, molecular or chemical methods for the detection, identification or enumeration of microorganisms in a material. Here some of the common methods have been described.

1. Microbiological Examination of Food

2. Why it’s necessary to examine food qualitatively or quantitatively for microorganisms? The principal objective of microbiological testing is to ensure that - The food meets certain statutory standards. The food meets internal standards set by processing company or external standards required by the purchaser. The food material entering the factory for processing are of required standard and/or meet a standard agreed with supplier. The process control and line sanitation are being maintained .

3. Following are the Microbiological Test Procedures in Usage - TOTAL VIABLE COUNT – Most common test performed on foods. It is also called Standard Plate Count or Aerobic Plate Count. Food dilutions are plated on agar based media containing complex nutrients. (beef extracts, yeast extract, peptone, growth factors etc.) Many plating techniques are employed for enumerating viable cells. They are as follows -

4. The Pour Plate Method A set of petri dish is inoculated with 1 ml aliquots from food dilutions. Incubation is provided as per the required temperature and duration. After incubation plates containing 30-300 colonies should be counted. From this the number of viable cells per gram of food can be calculated.

5. 2) The Spread Plate Method 0.1 ml dilutions are spread evenly over the surface of pre-poured and solidified petri dishes, using sterile L-shaped glass rods. Colonies observed on the surface can be easily observed and picked off.

6. 3) The Drop Plate Method Specially calibrated pipettes delivering 0.02 ml per drop are used and five different drops are delivered onto the surface. The drops are dried before incubation.

7. 4) The Agar Droplet Method Rapid technique used for smaller quantities of materials. Dilutions are prepared in molten agar and colonies develop in the solidified droplets (0.1 ml) during incubation. Counting is facilitated by using a projection viewer which magnifies the droplets.

8. 5) The Spiral Plate Method It is a semi-automatic method. A machine continuously plates a known volume of sample on the surface of a rotating agar plate. The amount of sample deposited decreases as the dispensing stylus is moved from the centre to the perimeter of rotating plate. Counting can be manually done but laser based automatic colony counters have been developed specially for use.

9. COUNTING BY MEASUREMENT OF ATP A bioluminescent technique has been developed based on action involving ATP and the Luciferase enzyme derived from fireflies. ATP + Luciferase → Light The total amount of light produced is directly proportional to the quantity of ATP present and since all bacteria contain roughly the same amount of ATP per cell it should be possible to measure the number of bacterial cells in any sample.

10. COUNTING USING DIRECT EPIFLUORESCENT FILTER TECHNIQUE In this method pre-filtered suspensions of food are subsequently passed through a fine polycarbonate membrane filter. Bacteria retained on the surface of the filter are then stained with fluorescent material and enumerated by means of an epifluorescence microscope.

11. DIRECT MICROSCOPIC COUNT It may be sometimes necessary to obtain total number of microorganisms (viable and non-viable cells) present in food samples. The total number of microbial cells in a given number of microscopic fields is counted and from this the rough calculation of total number of organisms present per gram food can be done.

12. INDICATOR ORGANISMS (presence indicates the possibility of poisoning) eg. Coliforms, Enterococci, Enterobacteriaceae etc. FOOD POISONING ORGANISMS eg. Salmonellas, Staphylococcus aureus, Bacillus aureus, Clostridium botulinum etc. FOOD SPOILAGE ORGANISMS eg. Pseudomonas, Micrococci, Streptococci etc.

13. References – H A Modi (2007) Introductory Food Microbiology Jaipur, India : Aavishkar Publishers

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