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Thermotolerant coliforms

Guideline


Thermotolerant coliforms should not be detected in any 100 mL sample of drinking water. If detected in drinking water, immediate action should be taken including investigation of potential sources of faecal contamination.

General description


Coliforms are gram-negative, non-spore-forming, rod-shaped bacteria that are capable of aerobic and facultative anaerobic growth in the presence of bile salts or other surface active agents with similar growth-inhibiting properties. They are found in large numbers in the faeces of humans and other warm-blooded animals, but many species also occur in the environment.

Thermotolerant coliforms are a sub-group of coliforms that are able to grow at 44.5 ± 0.2°C. E. coli is the most common thermotolerant coliform present in faeces and is regarded as the most specific indicator of recent faecal contamination because generally it is not capable of growth in the environment. In contrast, some other thermotolerant coliforms (including strains of Klebsiella, Citrobacter and Enterobacter) are able to grow in the environment and their presence is not necessarily related to faecal contamination.

Thermotolerant coliforms, including E. coli, can ferment lactose (or mannitol) at 44.5 ± 0.2°C with the production of acid within 24 hours.

Source and occurrence


Thermotolerant coliforms are normal inhabitants of the intestine, generally present in high numbers in human and animal faeces;. However, environmental thermotolerant coliforms, can occur in natural waters. These organisms are of lesser significance.

Method of identification and detection


The presence of thermotolerant coliforms in water samples can be determined using a number of methods. A common method involves membrane filtration (MF) for concentration of the organisms from water, followed by growth in enrichment/selective media or multiple tube dilution (most probable number – MPN) procedures (AS 4276.6 1995, AS 4276.7 1995). Specific secondary tests are used with both MF and MPN procedures to confirm the identification of thermotolerant coliforms.

Indicator value and application in practice


Thermotolerant coliforms can be used as an indicator of faecal contamination but they are not as specific as E. coli, which is the preferred indicator. The group includes types that can grow in the environment and be present in the absence of faecal contamination.

The presence of thermotolerant coliforms may provide evidence of recent faecal contamination. Thermotolerant coliforms can be used to assess:

  • source water quality and potential impacts of human and animal waste;

  • inadequate treatment;

  • post-treatment ingress of human and animal waste into distribution systems;

  • the effectiveness of risk management plans in assuring delivery of safe drinking water at
    consumers’ taps.

Thermotolerant coliforms are not an effective indicator for the presence of enteric protozoa or viruses.

Thermotolerant coliforms should not be present in any 100 mL sample of drinking water. Risk management plans should incorporate corrective actions in the event of the detection of thermotolerant coliforms in drinking water. The presence of these organisms may indicate faecal contamination of the water supply, and if they are detected in drinking water, the cause should always be investigated. Possible causes include inadequate treatment or ingress of contamination. Investigation will generally require further testing.

References


AS 4276.6 (1995). Thermotolerant coliforms and ^ Escherichia coli – Estimation of most probable number (MPN). Standards Association of Australia, Sydney, NSW.

AS 4276.7 (1995). Thermotolerant coliforms and Escherichia coli – Membrane filtration method. Standards Association of Australia, Sydney, NSW.

.

Total coliforms

Guideline


No guideline value has been set for total coliforms in drinking water. If used as an indicator, numbers should be established on a system-specific basis. Increased concentrations should be investigated.

General description


Coliforms are gram-negative, non-spore-forming, rod-shaped bacteria that are capable of aerobic and facultative anaerobic growth in the presence of bile salts or other surface active agents with similar growth-inhibiting properties. They are able to ferment lactose with the production of acid within 48 hours at 35-37°C. Fermentation by these organisms begins with the cleavage of lactose into galactose and glucose by the enzyme ß-galactosidase. Coliforms are oxidase-negative. These characteristics are not taxonomic criteria, but practical working definitions used for water examination purposes.

Coliforms are a diverse group of bacteria including Escherichia coli and other thermotolerant coliforms (see also Fact Sheets on Escherichia coli and Thermotolerant Coliforms, under Microbial Indicators, and Pathogenic Escherichia coli, under Bacteria). Human and animal faeces contains large numbers of coliform bacteria, but there are many species that occur naturally in the environment. For this reason, coliforms can be present and grow in biofilms in drinking-water distribution systems. Coliforms that have been recovered from distribution systems include the non-thermotolerant genera Serratia, Hafnia and Pantoea as well as thermotolerant genera including Klebsiella and Enterobacter. Their presence in water, in the absence of E. coli, does not necessarily indicate faecal contamination.

Source and occurrence


Total coliform bacteria (excluding E. coli) occur in both sewage and natural waters. Some of these bacteria are excreted in the faeces of humans and animals, but many coliforms are heterotrophic and able to multiply in water and soil environments. Total coliforms can also survive and grow in water distribution systems, particularly in the presence of biofilms.

Method of identification and detection


Total coliforms can be quantified in water by a number of techniques. Membrane filtration (MF) can be used for concentration of the organisms from water, followed by growth in enrichment/selective media or multiple tube dilution (most probable number – MPN) procedures (AS 4276.4. 1995, AS 4276.5. 1995). Specific secondary tests are used with both MF and MPN procedures to confirm the identification of coliform organisms.

Alternatively, the presence of coliform bacteria can be detected by testing for the production of the enzyme ß-galactosidase (APHA et al 2005). Enzyme substrate tests incorporate chromogenic substrates such as ortho-nitrophenyl-ß-D-galactopyranoside (ONPG) or chlorophenol red-ß-Dgalactopyranoside (CPRG). When the substrates are hydrolysed, a colour change is produced.

It has been reported that more coliform bacteria may be detected using enzyme substrate-based methodology than with MF-based methodology (Adcock and Saint 1997).

Indicator value and application in practice


Total coliforms (excluding E. coli) are not considered useful as indicators of the presence of faecal contamination and enteric pathogens, as there are many environmental coliforms that are not of faecal origin. The presence of these coliforms may represent release from pipe or sediment biofilms, and may be part of the normal flora of the drinking-water distribution system.

No guideline value has been set for total coliforms in drinking water. If used as an indicator, numbers should be established on a system-specific basis, taking into consideration relevant historical data and an understanding of the characteristics of the system. While coliforms can be used in operational monitoring to indicate inadequate treatment, breakdowns in system integrity, or the presence of biofilms, there are better indicators for these purposes. As a disinfection indicator, the test for total coliforms is far slower and less reliable than direct measurement of disinfectant residual. Heterotrophic Plate Count tests detect a wider range of microorganisms and are generally considered a better indicator of distribution system integrity and cleanliness.

References


Adcock PW, Saint C (1997). Trials of Colilert System. Water, 24(2):22-25.

APHA, AWWA, WEF (American Public Health Association, American Water Works Association, Water Environment Federation) (2005). Standard Methods for the Examination of Water and Wastewater, 21st edition. Method 9223 Enzyme Substrate Coliform Test. American Public Health Association, Washington DC.

AS 4276.4 (1995). Coliforms – Estimation of most probable number (MPN). Standards Association of Australia, Sydney, NSW.

AS 4276.5 (1995). Coliforms – Membrane filtration method. Australian Standards, Standards Association.





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