Standard Method for Testing Microbiological Safety of Drinking Water

Standard Method for Testing Microbiological Safety of Drinking Water


Bacteria originate from faecal matter and pollute streams and groundwater. Most bacteria are only 1 micrometer in diameter, but some bacteria range in size from 0.1 to 10 micrometers. Bacteria of the coliform group are considered the primary indicators of fecal contamination and are often used to assess water quality. Coliform is used to describe a group of Gram-negative, facultative, anaerobic, rod-shaped, non-spore-forming bacteria, capable of growing in bile salts and other surface active agents and able to ferment lactose to produce acid and gas within 48 hours at 35’C. Coliform bacteria, having the same properties at 44’C, are referred to as thermo-tolerant coliforms or E. coli (Escherichia coli).This is nothing but faecal coliform bacteria, found in the intestines of warm-blooded animals, including humans.

Coliform bacteria generated from the waste of warm-blooded animals, may get into the river or groundwater through surface runoff, especially after a heavy rainfall. Water contaminated with faecal matter can seep into bigger bodies of water, which may ultimately affect drinking water. A spring, hand-dug well, buried well, cistern, etc. are all very likely to be contaminated with coliform bacteria. Thus, contamination of water sources by the spread of such bacteria is the leading cause of water quality impairment in rivers. These bacteria may cause a serious health threat to humans, causing diseases like typhoid fever, hepatitis, gastroenteritis, dysentery, etc. Even a very small percentage of faecal coliform bacteria may cause intestinal distress and in more severe cases nausea, vomiting and even death.

It is often difficult to detect specific disease-causing organisms in water that we drink everyday. Therefore, indicator organisms are detected in drinking water, which are of faecal origin. Smoke is an indicator of fire. Similarly, bacteria of faecal origin (coliforms / E. coli) are indicators of pollution from faecal sources, thereby warning that there may be more dangerous organisms like Cholerae, Salmonella, Hepatitis virus etc, present in water. In this exercise, this principle is employed to ensure microbiological safety of our drinking water.


To detect the presence of coliform bacteria in drinking water with the help of multiple tube method.

Materials / Equipment Required

1. Mac Conkey broth double strength 10 ml in 5 tubes.
2. Mac Conkey broth single strength 5 ml in 10 tubes.
3. Dropper to measure water in little quantities.
4. Incubator
5. Water samples
6. Binocular microscope or powerful magnifying lens


  1. First sterilize all the 15 tubes in hot, boiling water.
  2. After cooling, add –
  3. 1. 5 ml of water to 5 tubes of double strength medium
  4. 2. 1 ml of water to 5 tubes of single strength medium
  5. 3. 0.1 ml of water to 5 tubes of single strength medium
  6. Keep all the 15 tubes in the incubator for 48 hours and at a temperature of 35-37’C. If you do not have an incubator a closed chamber with a light bulb and thermometer can be a poor substitute. Periodic manual checking of temperature along with switching off / on of the bulb will be needed.
  7. Now look for some change of colour and gas produced in the tubes.
  8. Plot the number of tubes, which has changed colour and match it with Mc Crady table given below. This table gives you the number of bacteria present in 100 ml of water.
  9. Incubate the tubes showing colour sub culture in fresh medium at a temperature of 44’C.
  10. The positive result is then matched in the same table, which shows thermo tolerant bacteria in 100 ml of water.

How Safe should be our Water? 

  1. Piped and treated (urban)water supply:   No coliforms and E. coli in 100 ml of water
  2. Untreated water entering the system:  E. coli – nil / 100 ml, Coliforms – 3/100 ml
  3. Rural unpiped supply: E. coli – nil / 100 ml, Coliforms 10 / 100 ml



Measurement of the number of coliform bacteria per 100 ml is often used as an important criterion in determining the degree of pollution and the sanitary quality of a simple of water. The allowable number limit per 100 ml of surface water sample is less than 100 organisms. The absence of E. Coli in drinking water is an absolute indispensability.

Further Activity

The presence of faecal coliform bacteria in sources other than drinking water can also pose health hazards. For instance, when irrigating crops with faecal coliform contaminated water (such as wastewater), chances are high that disease may spread after consumption of the grains and vegetables grown with such water. Crops that are eaten uncooked are especially dangerous for the transmission of such diseases. One method for controlling the
spread of such bacterial diseases is by delaying the consumption of the crop, as bacteria dissipates and perishes once exposed to air.


  • Think of some new and innovative solutions to kill faecal coliform bacteria and the disease causing bacteria found in irrigation/municipal water.
  • Is there any quick and easy method of killing the bacteria source in the vegetables you buy before you put it on the table?
  • Do you think disinfectant chemicals such as chloride can effectively kill the bacteria without disturbing the balance of the aquatic environment? Show your teacher with the help of some experiment the possible negative or positive results of using disinfectants to kill bacteria.
  • Imagine yourself the head municipal authority in your village or town. Now think of the measures that you would be taking to stem or control microbiological pollution of water sources in your locality? Make a list of all possible activities / regulations / enforcements that you can think of.

Standard methods for the examination of water and wastewater, 17th edn. American Public Health Association, Washington DC.
