Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving hydrolase
Patent
1991-11-19
1995-09-12
Wityshyn, Michael G.
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving hydrolase
435 19, 435 24, 435 34, 4352541, 4352551, 435911, 435922, 435944, 530323, 530330, 530331, C12Q 144, C12N 100, C07K 500
Patent
active
054496128
DESCRIPTION:
BRIEF SUMMARY
FIELD OF THE INVENTION
The present invention relates to a novel method of determining, assaying and/or identifying strains of Candida and related strains belonging to the group of the Torulopsis by means of chromogenic substrates of the monoamino acid or peptide type.
PRIOR ART
Candidiases are caused by pathogenic strains of Candida or related strains belonging mainly to the group of the Torulopsis. It is known that, in more than 60% of cases encountered, the strains of Candida responsible for candidiases belong to the species Candida albicans and that, in the other cases, the responsible strains belong to the other species of Candida, for example Candida tropicalis or Candida krusei, or to the group of the related strains of Torulopsis, especially the species Torulopsis glabrata.
It is known that, to mitigate the difficulties (large number of manipulations and long testing times) of the techniques of the prior art [blastesis test (or serum filamentation test), chlamydosporulation test on PCB or RAT media] for the identification of strains of Candida, mainly the strains of the species Candida albicans, and related strains, techniques have been proposed which are known in the biochemical field for detecting enzymes characteristic of yeasts, such as the osidases and aminopeptidases, and which use chromogenic or fluorogenic substrates.
Hitherto, all the techniques for identifying Candida and Torulopsis by the detection of osidases or aminopeptidases have been carried out on strains isolated beforehand.
Tests on oside substrates labeled with the fluorogenic 4-methylumbelliferyl residue (abbreviated to 4MU), i.e. especially 4MU-.beta.-D-glucopyranoside, 4MU-2-acetamido-2-deoxy-.beta.-D-glucopyrano side, 4MU-.beta.-D-galactopyranoside and 4MU-2-acetamido-2-deoxy-.beta.-D-galactopyranoside, for the purpose of detecting .beta.-D-glucosidase, N-acetyl-.beta.-D-glucosaminidase, .beta.-D-galactosidase and, respectively, N-acetyl-.beta.-D-galactosaminidase originating from strains of Candida, are known in particular from D. G. BOBEY et al., J. Clin. Microbiol., 13 (n.degree. 2) pages 393-394 (1981), J. L. PERRY et al., J. Clin. Microbiol., 25(n.degree. 12) pages 2424-2425 (1987), and C. M. SMITKA et al., J. Clin. Microbiol., 27 (n.degree. 1) pages 203-206 (1989). According to these documents, the strains of Candida and Torulopsis are found not to have any .beta.-D-galactosidase, .beta.-L-fucosidase and .beta.-D-glucosidase activity; the strains of Candida albicans and a few strains of Candida tropicalis, on the other hand, have an N-acetyl-.beta.-D-galactosaminidase activity. It is clear that the use of these substrates taken in isolation is not directly applicable to the identification of strains of Candida albicans in a mixture of strains of Candida and, if appropriate, Torulopsis.
Also, the use of oside substrates containing a fluorogenic or chromogenic group such as p-nitrophenyl (abbreviated to PNP), .beta.-naphthyl or 6-bromo-.beta.-naphthyl [obtained by the coupling (with the formation of an ether bridge) of an ose with p-nitrophenol, .beta.-naphthol or, respectively, 6-bromo-.beta.-naphthol] for the determination of isolated strains of Candida albicans is known from M. CASAL et al., Mycopathologia, 81, pages 155-159 (1983), and I. POLACHEK et al., J. Clin. Microbiol., 25, pages 907-910 (1987). According to the teaching of these two documents, said oside substrates react with their target osidase originating from the strains of Candida and related strains. However, it should be pointed out that (i) the substrate .beta.-naphthyl-N-acetyl-.beta.-D-glucosaminide [target enzyme: N-acetyl-.beta.-D-glucosaminidase] gives the same positive reaction with, in particular, the isolated strains of Candida albicans, Candida tropicalis, Candida stellatoidea and Torulopsis glabrata, and (ii) the substrate (6-bromo-.beta.-naphthyl)-.beta.-D-glucopyranoside [target enzyme: .beta.-D-glucosidase] gives the same positive reaction with the isolated strains of Candida albicans, Candida tropicalis, Candida pseudotropicalis, Torul
REFERENCES:
patent: 4874695 (1989-10-01), Pincus
Kalebuia et al., Appl. Microbiol. Biotechnol, vol. 28, pp. 531-536, 1988.
Iida et al., Agric. Biol. Chem., vol. 52, No. 5, pp. 1281-1282, 1988.
Contant epouse Pussard Genevieve
Lepargneur Jean-Pierre
Martinoli Jean-Luc
Quentin Gerard
Mohamed Abdel A.
Serbio
Wityshyn Michael G.
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