Why must double-strength lactose broth be used in the MPN method (presomptive test)
Answers
Answer:
Most Probable Number (MPN) is a method used to estimate the concentration of viable microorganisms in a sample by means of replicate liquid broth growth in ten-fold dilutions. It is commonly used in estimating microbial populations in soils, waters, agricultural products and is particularly useful with samples that contain particulate material that interferes with plate count enumeration methods.
MPN is most commonly applied for quality testing of water i.e to ensure whether the water is safe or not in terms of bacteria present in it. A group of bacteria commonly referred to as fecal coliforms act as an indicator of fecal contamination of water. The presence of very few fecal coliform bacteria would indicate that water probably contains no disease‑causing organisms, while the presence of large numbers of fecal coliform bacteria would indicate a very high probability that the water could contain disease‑producing organisms making the water unsafe for consumption.
Contents [hide]
1 Principle
2 Presumptive test
2.1 Requirements
2.2 Preparation of the Medium
2.3 Procedure of MPN test
2.3.1 For untreated (polluted) water
2.3.2 For treated (unpolluted) water
3 Confirmatory Test
4 Completed Test
5 Advantages of MPN
6 Disadvantages of MPN
Principle
Water to be tested is diluted serially and inoculated in lactose broth, coliforms if present in water utilizes the lactose present in the medium to produce acid and gas. The presence of acid is indicated by the color change of the medium and the presence of gas is detected as gas bubbles collected in the inverted Durham tube present in the medium. The number of total coliforms is determined by counting the number of tubes giving positive reaction (i.e both color change and gas production) and comparing the pattern of positive results (the number of tubes showing growth at each dilution) with standard statistical tables.
MPN test is performed in 3 steps
Presumptive test
Confirmatory test
Completed test
Presumptive test
The presumptive test is a screening test to sample water for the presence of coliform organisms.
If the presumptive test is negative, no further testing is performed, and the water source is considered microbiologically safe.
If the presumptive test is negative, no further testing is performed, and the water source is considered microbiologically safe. If, however, any tube in the series shows acid and gas, the water is considered unsafe and the confirmed test is performed on the tube displaying a positive reaction.
The method of the presumptive test varies for treated and untreated water.
Requirements
Medium: Lactose broth or MacConkey broth or Lauryl tryptose (lactose) broth
Glasswares: Test tubes of various capacities (20ml, 10ml, 5ml), Durham tube
Others: Sterile pipettes
Preparation of the Medium
Prepare medium (either MacConkey broth or lactose broth) in single and double strength concentration.
For untreated or polluted water :
Dispense the double strength medium in 10 tubes (10mL in each tube) and single strength medium in 5 tubes (10 mL in each tube)and add a Durham tube in an inverted position.
For treated water:
Dispense the double strength medium in 5 tubes (10mL in each tube) and 50 mL single strength medium in 1 bottle and add a Durham tube in an inverted position.
Examine the tubes to make sure that the inner vial is full of liquid with no air bubbles.
Answer:
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