The PhenePlate system for studies of the diversity of enterococcal populations from the food chain and the environment.

ABSTRACT

The growing interest in ecological investigations, such as studies of the "flow" of bacteria through the food chain, has resulted in a great need for simple typing techniques for bacteria that can be used when a large number of isolates need to be analysed. The present study describes a simple method for biochemical fingerprinting of enterococci, the PhenePlate RF (PhP-RF) system. The system is based on a 96-well microplate containing eight sets of 11 dehydrated reagents, selected to have a high discriminatory power among enterococcal isolates. The PhP plates are inoculated easily by picking single colonies directly from the primary agar culture and suspending them in the first well of each row in the microplate. The kinetics of each reaction is evaluated by measuring the absorbance value of each well three times during 64 h, whereupon a biochemical fingerprint is calculated as the mean value for each reagent over the three readings. The PhP-RF method was shown to be highly reproducible, even when results were compared between different laboratories. The discriminatory power, measured as Simpson's diversity index (Di), was as high as 0.96 for all enterococci. The PhP-RF method could also be used as a preliminary species identification method, by comparing the biochemical fingerprints of unknown strains to those of a set of reference strains of known species. Most strains of Enterococcus faecalis, Enterococcus faecium and Enterococcus hirae were correctly identified using this method. We conclude that the PhenePlate RF system is useful for rapid typing of enterococcal populations, and it is especially useful as a first screening method for ecological studies, when many isolates per sample need to be analysed.