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IDEM approves Coliscan ^® MF for NPDES permittees use

As you may know, the Indiana Department of Environmental Management (IDEM) has announced that all NPDES permittees that must conduct E. coli testing may now use the Coliscan^® MF method for the coming season. This decision was made following extensive comparative work done at the City of Elkhart Water and Wastewater Laboratory under the guidance of Lynn A. Newvine, laboratory director, who recommended the method to IDEM for consideration as an improvement over the m-TEC method which has been the primary approach in the past. A portion of the comparative work has been published in the April, 1999 issue of Water Environment & Technology, pp. 57-59, authored by A. K. Umble, L. E. Kolo, L. A. Newvine, G. N. Roth and J. N. Roth.

The IDEM decision was made following consultations with USEPA officials who indicated that the state of Indiana (and any other state that has primacy) has the authority to specify the method(s) to be used. More information can be obtained from Barbara McDowell of IDEM.

Email: , Phone: 317-233-6464

Anyone who has performed the m-TEC method is aware of its inherent potential problems and drawbacks. Following is a comparison of the Coliscan^® MF and m-TEC methods.

Advantages of the Coliscan ^® MF method are numerous.

1. There is no initial 2 hour resuscitation period requiring operator attention and a change of incubators, as well as the placement of the plates in waterproof bags for the water bath incubator.

2. Temperature is not critical. 24 hr incubation at 35ºC ±0.5 is the only requirement. Less time is used in checking and calibrating incubators and no switching from incubator to incubator is needed. (Also, much less expensive incubators can be used.

3. There is no need to do the additional urease test required by the m-TEC method. This saves time and expense. Also, some bacterial types (eg. Pseudomonas spp. ) can result in gross false positive results with the m-TEC method.

4. Coliscan^® MF broth is easier to use and plates are easier to prepare than m-TEC agar medium, which must be mixed, sterilized and dispensed into plates, and then stored until use. The Coliscan^® MF broth is kept frozen (with a 6 month shelf life) until needed and then the thawed broth is dispensed aseptically into plates containing sterile absorbent pads and fresh medium is assured.

5. Coliscan^® MF broth is provided in safe-to-use plastic bottles (20 mL for 10 tests/bottle) which are recyclable. There are no dangerous-to-handle glass vials to break open.

6. Coliscan^® MF medium is based upon well proven technology which incorporates chromogenic enzyme substrates. It is superior to the modified m-TEC medium developed by the EPA laboratories which uses only one chromogenic substrate (for the enzyme glucuronidase) and requires the resuscitation period followed by elevated incubation temperature.

With the modified m-TEC medium, it is stated that E. coli is indicated by magenta colored colonies. This is generally true, but it is possible for some temperature tolerant, glucuronidase positive organisms to exist, which could result in false positive readings. Also, if other (non-magenta) colonies grow on the modified m-TEC medium, they should be classified as "fecal" coliforms. However, there is no surety that these non-colored colonies are in fact true coliforms as they may not be galactosidase positive, and the modified m-TEC medium does not test for galactosidase.

The Coliscan^® MF medium avoids the potential problems described above by the incorporation of two different chromogenic substrates, one for the enzyme glucuronidase (X-gluc which colors positive colonies a teal green color), and one for the enzyme galactosidase (Red-gal^® which colors positive colonies a pink color). Since E. coli is unique among the coliforms as a producer of glucuronidase, it utilizes the X-gluc substrate, causing the expression of the teal green color. However, since E. coli is a true coliform, it also produces galactosidase and so utilizes the second substrate also and the combination of the teal green and the pink colors results in E. coli colonies having a dark blue/purple color. If there are any non-E. coli coliform organisms in the sample, they appear as pink galactosidase positive colonies, while non-coliform, galactosidase negative organisms are colorless.

7. It should be pointed out that if an operator wishes to determine both E. coli and "fecal coliforms", the Coliscan^® MF medium is well prepared to accomplish this task. All that is needed is to initiate the same procedure for incubation as that called for with the modified m-TEC method. The 44.5ºC incubation temperature will eliminate non-temperature tolerant organisms so that the E. coli colonies will be blue/purple, but other temperature tolerant coliforms will be pink, and any temperature tolerant non-coliforms will be colorless. This versatility allows the operator to distinguish temperature tolerant "fecal coliforms" which sometimes occur (commonly members of the genus Klebsiella from the more important E. coli.

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