Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving viable micro-organism
Patent
1996-01-26
1999-10-05
Dees, Jose G.
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving viable micro-organism
435 29, C12Q 104
Patent
active
059622512
DESCRIPTION:
BRIEF SUMMARY
The present invention relates to a method for demonstrating the presence or absence of a particular strain of microorganism in a culture medium.
The detection of microorganisms is very important, in particular in the food industry, in relation to water monitoring or in medicine, in view of the fact that these microorganisms may not only prove to be pathogenic agents, but can also consist of agents that reveal some types of contamination.
Various methods enable the presence of microorganisms in a medium of some kind to be demonstrated, consisting in taking a sample of the medium in question and then in promoting the growth of the microorganisms present by culture on or in a suitable medium.
In order to simplify the demonstration of the microorganisms present, the use has been proposed, in the detection medium, of colored compounds whose presence is characteristic of a given microorganism.
The coloration often reveals an enzyme activity associated with the microorganism in question, and the outcome of this activity may result in a modification of the pH of the medium, revealed by a colored indicator (EP-A-0 395 532), or alternatively in the liberation of a chromophoric or fluorophoric compound (FR-A-2-684,110).
Chromophores or fluorophores are compounds generally obtained by enzymatic hydrolysis of corresponding chromogenic or fluorogenic compounds present in the culture medium.
Fluorophores emit a characteristic radiation by fluorescence.
Chromophoric compounds are characterized by a color with a dominant wavelength.
Among known chromophoric compounds, indoxyl derivatives, hydroquinoline or alternatively naphthoic derivatives, or naphthyl and phenyl derivatives, may be noted in particular.
In order to differentiate two different genera of microorganisms in a culture medium, the proposal has even been made to introduce two chromogenic agents each liberating a chromophoric compound with a color characteristic of the presence of a particular microorganism (U.S. Pat. No. 5,210,022).
Although all of these media are efficacious in 10 detecting microorganisms of a specific genus, such as, for example, Salmonella, Candida or E. coli, and distinguishing them from other species, they do not, however, permit the detection of a large number of microorganisms of different genera on the same culture medium, or the differentiation of pathogenic species from others among microorganisms of the same genus.
Such a distinction appears to be all the more important for certain species of yeasts, such as Candida albicans which is responsible for more than 50% of pathologies associated with yeasts.
In point of fact, it was unexpectedly demonstrated that at least four different colorations enabling the strains to be characterized could be observed by introducing into the culture medium at least two chromogens which are substrates for enzymes of a particular strain, the two chromogens being chosen so that, in the culture medium, the presence of at least one strain is revealed by an accessory color, namely:
In many cases, the accessory color does not correspond to the mixture of the colors of the corresponding two chromophores, and is a color whose dominant wavelength does not correspond to the dominant wavelength of the mixture df chromophores liberated by the chromogens present in the culture medium, taken separately.
The dominant wavelength of the chromophores and the accessory colors may be calculated by reference to daylight, as defined by the CIE (International Commission on Energy as illuminant D.sub.65, using any standard method for measuring the color of an object, especially with a spectrocolorimeter.
Hence the present invention relates to a method for demonstrating the presence or absence of a particular strain of microorganism in a culture medium, for which at least two chromogens which are enzyme substrates for said strain are introduced, said chromogens being chosen so that, in the culture medium, the presence of said strain is revealed by an accessory color.
The chromogens are, in particular, substrates for the followi
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Dees Jos,e G.
Jones Dameron
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