Red-legged frog, California
Photo©CLCase

Bacteriologic Quality of Water
An environmental project
Christine L. Case

About the MPN test
For further study
Lab Protocol
MPN table
Sample problems

The primary infectious disease threat to our water supply is from sewage wastes. The enteric diseases such as cholera, typhoid fever, enterotoxigenic E. coli, giardiasis, and polio are often the result of sewage–contaminated water which has not been properly treated. Therefore the main emphasis of the drinking water laboratory is to determine the effective-ness of the treatment plant in removing intesti-nal (enteric) microbes.

Tests that determine the bacteriologic quality of water have been developed to prevent transmission of water-borne diseases of fecal origin. However, it is not practical to look for pathogens in water supplies, because pathogens occur in such small numbers that they might be missed by sampling. Moreover, when pathogens are detected, it is usually too late to prevent occurrence of the disease. Rather, the presence of indicator organisms is used to detect fecal contamination of water. An indicator organism must be present in human fe-ces in large numbers and must be easy to detect. The most frequently used indicator organ-isms are the coliform bacteria. Coliforms are aerobic or facultatively anaerobic, gram-negative, nonendospore-forming, rod-shaped bacteria that ferment lactose with acid and gas formation within 48 hours at 35°C. Coliforms are not usually pathogenic, although they can cause opportunistic infections and there are pathogenic strains of E. coli such as O157:H7. Coliforms are not restricted to the human gastrointestinal tract but may be found in other animals and in the soil. Tests that determine the presence of fecal coliforms (of hu-man origin) have been developed.

Established public health standards specify the maximum number of coliforms allowable in each 10 mL of water, depending on the intended use of the water (for example, drinking, water-contact sports, or treated waste water for irrigation or for discharge into a bay or river).

MPN Test

Coliforms can be detected and enumerated in the multiple-tube technique. In this method, dilutions from a water sample are added to lactose fermentation tubes. The lactose broth can be made selective for gram-negative bacteria by the addition of lauryl sulfate or brilliant green and bile. Fermentation of lactose to acid and gas is a positive reaction. The number of coliforms is determined by a statistical estimation called the most probable number (MPN) method. A count of the number of tubes showing acid and gas is then taken, and the figure compared to statistical tables, shown in Table 3. The number is the most probable number of coliforms per 100 ml of water.

For further study

Coliforms include Escherichia, Enterobacter, and Citrobacter. Although the presence of E. coli indicates fecal contamination, the original contamination could have occurred many years earlier and the E. coli being detected are now resident microbiota of the water.

The presence of Enterobacter faecalis or Bacteroides could be better indicators of recent fecal contamination. Design an MPN test for one of these organisms.