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Coliform bacteria and E. coli in drinking water. Comparison of EU reference method with alternative methods
2 Danish equivalency study 2000/2001 on MPN (DS 2255) against Colilert
In 2000/2001 a Danish equivalency study was performed between the existing national reference method (DS 2255:1983) and Colilert. Both methods are MPN-methods but with two different detection principles. DS 2255 defines coliform bacteria as bacteria able to ferment lactose to acid and gas at 37°C in MacConkey broth and E. coli as coliform bacteria furthermore able to ferment lactose at 44°C in MacConkey broth and producing indole from tryptophane at 44°C. In Colilert coliforms are defined by the activity of ß-galactosidase and E. coli by the activity of ß-glucuronidase.
Fricker, Niemela & Lee (2000) states that it is clear the many coliform bacteria are unable to ferment lactose within 48 hours whereas they will be able to demonstrate the activity of ß-galactosidase. Therefore differences between the two methods may be expected as fully agreeable results may not be achieved by two different detection principles.
The different principles however does not discriminate per se one method for another as there are more different definitions of the group of coliform bacteria and coliform bacteria and E. coli detected with either of the methods are unwanted in drinking water.
2.1 Materials and methods
A total of 64 samples of drinking water were analysed in two laboratories using both the Danish national MPN method and Colilert for coliform bacteria and E. coli. The results from both methods were then compared.
2.2 Results
Of the 64 samples examined most (51) were negative using both methods. The results of the remaining 13 tests are given in Table 1.
Table 1 Results of coliform and E. coli analyses from 13 samples of water using the Danish national method (DS 2255:1983) and Colilert
| DS 2255:1983 |
Colilert |
| Coliform bacteria |
E. coli |
Coliform bacteria |
E. coli |
| <1 |
<1 |
2 |
<1 |
| 1 |
1 |
10 |
<1 |
| <1 |
<1 |
16 |
<1 |
| >161 |
<1 |
>201 |
<1 |
| <1 |
<1 |
1 |
<1 |
| <1 |
<1 |
4 |
<1 |
| 1 |
<1 |
<1 |
<1 |
| <1 |
<1 |
1 |
<1 |
| 1 |
<1 |
1 |
<1 |
| 1 |
<1 |
1 |
<1 |
| 49 |
49 |
53 |
14 |
| 70 |
46 |
47 |
8 |
| 23 |
23 |
64 |
13 |
2.3 Conclusions
There were no significant differences between the number of coliform bacteria found using the two different methods when comparing (Students t-test) paired results for coliform bacteria when detected by both methods. However Colilert detected coliform bacteria in 12 samples and the Danish reference method in only eight. This indicates that the use of Colilert will increase the number of samples found to contain coliform bacteria compared to the use of the Danish MPN-method. It is likely, based on data generated during the European study that also the actual number of coliform bacteria detected with Colilert will be higher in some samples than that detected by traditional methods.
For E. coli there was complete agreement between the two methods regarding which samples were positive (excluding one sample with the finding of 1 E. coli/100 ml with the Danish reference method and <1 /100 ml with Colilert, which is found to be within the statistical uncertainty). However, the total numbers of E. coli were higher using the Danish reference method (three samples; n = 13) although non-significant (Students t-test). The most likely explanation for the higher counts is the possible misidentification of E. coli using traditional detection principles. Traditional methodologies identify E. coli based on its ability to grow at 44°C, ferment lactose and produce indole from tryptophan. Some other coliform species (notably strains of Klebsiella oxytoca) also possess these characteristics and may lead to false positive results for E. coli.
In conclusion, the use of Colilert is likely to give rise to an increase in the number of samples found to contain coliform bacteria. Furthermore Colilert may lead to an increase in the actual number of coliform bacteria detected in a given sample over that detected using the Danish standard method. For E. coli, the number of samples found to be positive by Colilert is likely to be similar to the findings with DS 2255:1983, but the actual number of E. coli in the samples may be slightly lower using Colilert.
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Version 1.0 February 2007, © Danish Environmental Protection Agency
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