Development of a Methodology for the Rapid Detection of Coliform Bacteria

摘要

Work has continued on the development of a rapid method for the enumeration of coliform bacteria, based on the hydrolysis of a fluorogenic substrate by the enzyme beta-D-galactosidase. A validation of the basic manual technique was completed using both laboratory and field samples. A correlation between the percentage of microdroplets and the log of the bacterial concentration was established in both cases. A semiautomated version of the manual technique was rendered feasible through the use of two fluorescent dyes (fluorescein and ethidium bromide) in the detection of microdroplets containing bacteria, as well as those that did not. Extensive studies conducted with E. coli Neotype induced for beta-D-galactosidase activity revealed that fluorochromasia, per se, was beyond the resolution capabilities of the instrument. The use of w/o (water in oil) emulsions was successfully employed in the entrapment of baeteria and the containment of fluorescein. At this point in time, the detection rate of E. coli Neotype emulsions having a concentration about 10 to the 7th power/ml falls within the range of 0.2-0.3%.