ABSTRACT
We describe an indirect detection method for bacterial identification and differentiation, based on selective adsorption of several fluorescent dyes. The lipid, protein, and nucleic acid components of fixed whole cells are stained with two mixtures, each containing two fluorochromes. The unadsorbed dyes were measured simultaneously with a video fluorometer [Clin Chem 22: 1483, 1976]. A dye-absorption matrix of the response can be generated, and we did so for each of nine bacteria. These responses were compared to a control or "complete" response matrix, and the response ratios of each bacterial species for each of the four dyes were calculated and plotted to obtain a characteristic pattern. From the response-ratio plots, plus simple pattern-recognition techniques, we could differentiate among all the bacteria. This rapid, sensitive technique is potentially applicable to a wide variety of bacteria.
Subject(s)
Coloring Agents , Enterobacteriaceae/analysis , Fluorometry/methods , Staphylococcus/analysis , Streptococcus/analysis , Acridine Orange , Fluorescein-5-isothiocyanate , Fluoresceins , Lipids/analysis , Nucleic Acids/analysis , Proteins/analysis , ThiocyanatesABSTRACT
A recent study has shown that an emission-excitation matrix could be used to provide a unique fingerprint for the selective identification and characterization of certain species of Pseudomonas. This paper describes the results of a systematic study of the variables that contribute to the uniqueness of the fingerprint. Additional insight concerning the nature of the fluorescent pigments of these species is gained by analysis with a high-performance liquid chromatograph/video fluorometer combination. While providing information about the analytical variables contributing to the analysis, this combination also added the variable of chromatographic retention time for unambiguous fingerprinting.
Subject(s)
Pigments, Biological/analysis , Pseudomonas/analysis , Autoanalysis , Chromatography, High Pressure Liquid , Computers , Fluoresceins , History, Modern 1601- , Pseudomonas aeruginosa/analysis , Pseudomonas fluorescens/analysis , Species Specificity , Spectrometry, Fluorescence/methodsABSTRACT
Fluorescent pseudomonads may be identified through examination of the fluorescence "profiles" of diffusible pigments released into the growth medium. The profiles are obtained in the form of an emission-excitation matrix, the elements of which correspond to fluorescence intensity as a function of multiple exciting and emitting wavelengths. Use of a rapid scanning fluorometer to acquire these data provides a relatively fast method for identification of the microorganisms after incubation. This procedure can potentially be expanded for the identification of other microorganisms.
