Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving viable micro-organism
Patent
1997-09-22
2000-02-01
Saucier, Sandra E.
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving viable micro-organism
435 39, 435 40, 435261, C12Q 104, C12Q 106, C12Q 108, C12N 102
Patent
active
060201509
DESCRIPTION:
BRIEF SUMMARY
FIELD OF THE INVENTION
The present invention relates to a novel method of detecting the presence or absence of microorganisms belonging to the group comprising bacteria and yeasts. This method uses a technique involving separation, culture and development on one and the same gelled system.
It further relates on the one hand to said gelled system as a novel industrial product, and on the other hand to the assay kit for carrying out said method.
Finally, it relates to the use of said method and said gelled system in microbiology (especially in bacteriology or mycology), particularly in the field of the detection of bacterial infections (more particularly the detection of urinary infections and septicemia).
PRIOR ART
When it is desired to assess the presence or absence of microorganisms such as bacteria and yeasts, especially by a colorimetric method, the following results have to be considered:
In practical terms, the following four values (expressed as percentages) have been defined from the TP, FP, TN and FN results which are theoretically possible: measurements.
It is known on the one hand that a colorimetric detection or filtration technique has already been recommended or used for assessing the presence or absence of bacteria in urine, and on the other hand that a gel separation technique has already been recommended or used for revealing erythrocyte agglutinates.
As far as colorimetric detection is concerned, the publication EP-A-0 496 409 has disclosed a method of revealing Gram-negative bacteria. This method comprises the filtration of a biological sample (especially urine) containing bacteria on a porous support (pore diameter 0.75-1.2 .mu.m) in order to retain the bacteria on said support, and then the detection of the Gram-negative bacteria by reaction of their dehydrogenases with a redox color reagent (especially a tetrazolium salt or resazurin), the reduction reactions induced by the Gram-positive bacteria being inhibited by means of a caotropic agent and a non-ionic detergent of the alkylglucoside type.
As far as the colorimetric filtration technique is concerned, tests marketed by VITEK SYSTEMS (Haziewood, Mo.) under the names "Bac-T-Screen.RTM." (semi-automatic test) and "FiltraCheck.RTM.-UTI" (manual test) are known in particular.
These tests are performed by filtering a urine sample on a filter paper impregnated with a developer in order to retain and color the bacteria which may be present in the sample; the filtration is carried out under a pressure difference, so a relatively complicated apparatus is required; the main source of errors (FP and FN results) is due to the possible presence of a large number of erythrocytes, leukocytes and/or epithelial cells in the urine (the presence of erythrocytes and/or leukocytes in the urine is already in itself a sign of dysfunction).
According to the literature--cf. (a) page 272 (Table 3) and page 274 (section "Colorimetric Filtration") of the article by M. PEZZLO, Clin. Microbiol. Rev., 1988, 1 (No. 2), 268-280, and (b) page 86 (paragraph "Systeme coloration filtration" ("Coloration/filtration system") and Tables) of the article by F. W. GOLDSTEIN, Med. Mal. Infect., 1991, 21, 83-88--the Bac-T-Screene.RTM. and FiltraCheck.RTM.-UTI tests are (i) rapid (1-2 minutes), (ii) effective when the bacterial population is .gtoreq.10.sup.5 CFU/ml, but (iii) unsatisfactory when said bacterial population is below 10.sup.5 CFU/ml. Overall, taking these two tests together, the two articles cited above give the following results for urine screenings according to the bacterial populations:
There is therefore a need to improve the reliability of the measurements for bacterial populations below 10.sup.5 CFU/ml.
As far as the gel separation technique for assessing erythrocyte agglutination is concerned, the publications FR-A-2 577 321 and EP-A-0 454 509 are known.
According to FR-A-2 577 321, the formation or presence of erythrocyte agglutinates is revealed by (1) the deposition of a liquid medium containing erythrocytes on a gel which (i) is located in a conical-
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patent: 4717660 (1988-01-01), Schulte
Dorn et al., "Blood Culture Technique Based on Centrifygation : Developmental Phase", Journal of Clinical Microbiology, Mar. 1976, vol. 3, No. 3, pp. 251-257.
Boisard-Beaupere Fran.cedilla.oise
Contant-Pussard Genevieve
Martinoli Jean-Luc
Rousseau Alain
Afremova Vera
Diffusion Bacteriologie Du Var
Saucier Sandra E.
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