The Optimization of an Epifluorescence Microscopy Technique for the Direct Counting of Clumped Bacteria

The enumeration of heterotrophic bacteria is an important measure of water quality in mains'distribution systems. Conventional methods of enumerating bacteria by plate counts do not distinguish between single bacteria and clumps of cells; however, various methods have been reported which use epifluorescence microscopy to give a direct count of bacteria. There appears to be no standard method for the preparation of samples, with the choice of fluorochrome and staining technique varying considerably – even between workers using similar environmental samples. A number of the reported methods were tested, and a technique was developed which enabled a consistent total count of bacterial cells (occurring both singularly and in clumps) to be obtained.