Detection and enumeration of coliforms in drinking water:: current methods and emerging approaches

The coliform group has been used extensively as an indicator of water quality and has historically led to the public health protection concept. The aim of this review is to examine methods currently in use or which can be proposed for the monitoring of coliforms in drinking water. Actually, the need for more rapid, sensitive and specific tests is essential in the water industry. Routine and widely accepted techniques are discussed, as are methods which have emerged from recent research developments. Approved traditional methods for coliform detection include the multiple-tube fermentation (MTF) technique and the membrane filter (MF) technique using different specific media and incubation conditions. These methods have limitations, however, such as duration of incubation, antagonistic organism interference, lack of specificity and poor detection of slow-growing or viable but non-culturable (VBNC) microorganisms. Nowadays, the simple and inexpensive membrane filter technique is the most widely used method for routine enumeration of coliforms in drinking water. The detection of coliforms based on specific enzymatic activity has improved the sensitivity of these methods. The enzymes beta-D galactosidase and beta-D glucuronidase are widely used for the detection and enumeration of total coliforms and Escherichia coli, respectively. Many chromogenic and fluorogenic substrates exist for the specific detection of these enzymatic activities, and various commercial tests based on these substrates are available. Numerous comparisons have shown these tests may be a suitable alternative to the classical techniques. They are, however, more expensive, and the incubation time, even though reduced, remains too long for same-day results. More sophisticated analytical tools such as solid phase cytometry can be employed to decrease the time needed for the detection of bacterial enzymatic activities, with a low detection threshold. Detection of coliforms by molecular methods is also proposed, as these methods allow for very specific and rapid detection without the need for a cultivation step. Three molecular-based methods are evaluated here: the immunological, polymerase chain reaction (PCR) and in-situ hybridization (ISH) techniques. In the immunological approach, various antibodies against coliform bacteria have been produced, but the application of this technique often showed low antibody specificity. PCR can be used to detect coliform bacteria by means of signal amplification: DNA sequence coding for the lacZ gene (beta-galactosidase gene) and the uidA gene (beta-D glucuronidase gene) has been used to detect total coliforms and E. coli, respectively. However, quantification with PCR is still lacking in precision and necessitates extensive laboratory work. The FISH technique involves the use of oligonucleotide probes to detect complementary sequences inside specific cells. Oligonucleotide probes designed specifically for regions of the 16S RNA molecules of Enterobacteriaceae can be used for microbiological quality control of drinking water samples. FISH should