ColiMinder® Measuring Principle

Simplified Explanations

The question to be answered by testing for E. coli is:

 how high is the level of fecal contamination in a water sample? 

The classical method is utilizing the growth of bacteria (E. coli/Coliforms) and the number of colonies formed, to determine the level of contamination.

The ColiMinder® is using the metabolic activity (specific enzymatic activity)  of target organisms as a measure for how many living E. coli are present per volume of sample to determine the level of contamination.

The measurement technology used in the device is fluorescence spectroscopic detection of E. coli-specific enzymatic activity.

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The classical method to measure bacterial contamination of water is using a fundamental properties of bacteria, their growth and ability to form colonies, as measured variable and measuring method.

This is a self-explanatory way to determine the bacterial contamination of water. This method is a robust and widespread, although it got some disadvantages and limitations, which are accepted due to the fact that they come with the method itself:

  • only culturable cells will form colonies, Viable But Not Culturable (VBNC) cells will not deliver a result, are not detected.
  • If several cells stick together or to a floating particle, these cells will only form one culture and represent one CFU even if they are many.
  • time to result is very long and cannot be made shorter due to the fact that the method is using the growth as measurement parameter which cannot be speeded up.

In order to develop a safe and reliable measurement system for bacterial contamination, it was necessary to identify another fundamental property of living bacteria, which could be measured fast.

The only available genuine property of living bacteria that can be measured directly, is their metabolism which is accomplished by specific enzymes.

The ColiMinder® measurement system is based on the direct measurement of the specific metabolic activity of target organisms within a sample. The metabolic activity (target-specific enzymatic activity) is a direct measure for the activity and thus number of living cells/bacteria within a given sample volume.

Every living organism has a metabolic activity (specific enzymatic activity) to live; if there is no activity present, there are no living organisms.

Measuring the specific enzymatic activity per sample volume, as a direct genuine property of living target organisms, this activity is proportional to the organisms present per sample volume.

In contrary to the classical growth method, which is using the capability of cells forming colonies as a measure to determine the presence of bacteria, the enzymatic method is not differentiating between culturable and VBNC cells. As long as the target organisms have a metabolism they contribute to the measurement signal.

As E. coli is used as an indicator for fecal contamination, the enzymatic method uses, not only  culturable cells, but generates a measurement signal from every living E. coli,  representing a more conservative view on fecal contamination level.

This difference, standard method using only culturable cells, enzymatic method using all living cells to determine contamination, necessarily leads to a correlation that is dependent on the ratio between culturable and VBNC cells.

Within a dilution series of one sample the correlation between both methods is almost 100%. Using standard natural samples the correlation is also strong but not fixed. For that reason it would be not legitimate to claim a direct equalization between MFU/100ml (Modified Fishman Units/100mL) and CFU/100ml or MPN/100ml.The graph below shows results, comparing enzymatic  and IDEXX MPN measurements during a contamination event.

graph

Parallel measurement of enzymatic activity and MPN/100ml during an event at HOAL. (The monitoring network at the Austrian Federal Agency for Water Managements (BAW) Hydrological Open Air Laboratory (HOAL) operated by Vienna University of Technology has been developed for multifactorial water quality analysis.)

The enzymatic activity, for example the E.coli specific ß-Glucuronidase activity, is scientifically exactly defined:

 “MFU (Modified Fishman Unit): One MFU will liberate 1.0 μg of phenolphthalein from phenolphthalein glucuronide per hour at pH 6.8 at 37°C.“

As a direct consequence VWM´s ColiMinder® is calibrated and delivers a result in Modified Fishman Units per 100mL for E.coli contamination, which represents the E.coli -specific enzymatic activity per 100mL  as a direct measure for the number of living organisms per sample volume.

As the measurement parameter of enzymatic activity is scientifically exactly defined it provides the base to adopt respective limits of tolerable contamination expressed in enzymatic activity, for all kinds of applications.

Wolfgang Vogl, CEO VWM GmbH