Medium for the specific detection of resistant microorganisms

Abstract

A method for distinguishing among a first group of microorganisms belonging to a first taxon of Gram negative bacteria, the first group of bacteria exhibiting a mechanism of resistance to a treatment; a second group of microorganisms belonging to a second taxon of Gram negative bacteria, the second taxon of bacteria being different than said first taxon, and exhibiting a mechanism of resistance to a treatment identical to the mechanism of the first group; and a third group of Gram negative bacteria that is not resistant to the treatment.

What is claimed is: 
     
         1 . A method for distinguishing among at least three groups of microorganisms that may be present in a biological sample, the at least three groups comprising:
 a first group of microorganisms, belonging to a first taxon of bacteria that is vancomycin resistant;   a second group of microorganisms, belonging to a second taxon of vancomycin resistant bacteria that is different than said first taxon; and   a third group of microorganisms that is not resistant or has a natural resistance to vancomycin,   
       the method comprising:
 inoculating a culture medium with the biological sample, the culture medium comprising:
 a first substrate for detecting a first enzymatic activity of said first group of microorganisms; 
 a marker for differentiating the first group of microorganisms and the second group of microorganisms, said marker being a substrate for detecting an enzymatic activity of said second group of microorganisms; and 
 an antimicrobial that is active on the third group of microorganisms, and 
 
 distinguishing among any members of the three groups of microorganisms that are present on the culture medium to determine of which group of microorganisms they are members on the basis of their interactions with the first substrate, the marker, and the antimicrobial. 
 
     
     
         2 . The method of  claim 1 , wherein:
 the first group of microorganisms is enterococcal bacteria that have an acquired resistance to vancomycin;   the second group of microorganisms is enterococcal bacteria that have a natural resistance to vancomycin; and   the third group of microorganisms is enterococcal bacteria that is not resistant to vancomycin.   
     
     
         3 . The method of  claim 1 , wherein:
 the first group of vancomycin-resistant microorganisms comprises  Enterococcus faecalis  and  Enterococcus faecium;      the second group of vancomycin-resistant microorganisms comprises  Enterococcus casseliflavus  and  Enterococcus gallinarum ; and   the third group of microorganisms is not resistant to vancomycin.   
     
     
         4 . The method of  claim 1 , wherein:
 the first group of vancomycin-resistant microorganisms comprises  Enterococcus faecium;      the second group of vancomycin-resistant microorganisms comprises  Enterococcus faecalis ; and   the third group of microorganisms is enterococcal bacteria that is not resistant to vancomycin or that have a natural resistance to vancomycin.   
     
     
         5 . The method of  claim 1 , wherein:
 the first group of vancomycin-resistant microorganisms comprises  Enterococcus faecalis  and  Enterococcus faecium;      the second group of vancomycin-resistant microorganisms comprises  Staphylococcus aureus  that is intermediary resistant or resistant to vancomycin; and   the third group of microorganisms is not resistant to vancomycin.   
     
     
         6 . The method of  claim 1 , wherein:
 the first substrate is an enzymatic substrate for detecting alpha-glucosidase activity;   the marker is a second substrate for detecting a second activity different than alpha-glucosidase activity; and   the antimicrobial is vancomycin.   
     
     
         7 . The method of  claim 6 , wherein the second substrate detects beta-glucosidase or beta-galactosidase activity. 
     
     
         8 . The method of  claim 6 , wherein:
 the first substrate is 5-bromo-4-chloro-3-indolyl-N-methyl-α-D-glucoside or 5-bromo-4-chloro-3-indolyl-α-D-glucoside, and   the second substrate is 6-chloro-3-indolyl-β-D-glucoside, alizarine-β-D-galactoside, 5-bromo-6-chloro-3-indolyl-β-D-glucoside, 5-bromo-6-chloro-3-indolyl-β-D-galactoside or 6-chloro-3-indolyl-β-D-galactoside.   
     
     
         9 . The method of  claim 6 , wherein:
 the first substrate is 5-bromo-4-chloro-3-indolyl-N-methyl-α-D-glucoside or 5-bromo-4-chloro-3-indolyl-α-D-glucoside, and   the second substrate is 6-chloro-3-indolyl-β-D-glucoside or 5-bromo-6-chloro-3-indolyl-β-D-glucoside.   
     
     
         10 . The method of  claim 1 , wherein:
 the first group of vancomycin-resistant microorganisms comprises  Enterococcus faecalis  and  Enterococcus faecium;      the second group of vancomycin-resistant microorganisms comprises  Enterococcus casseliflavus  and  Enterococcus gallinarum;      the third group of microorganisms is not resistant to vancomycin;
 the first substrate is an enzymatic substrate for detecting alpha-glucosidase activity; 
   the marker is a second substrate for detecting a second activity different than alpha-glucosidase activity; and   the antimicrobial is vancomycin.   
     
     
         11 . The method of  claim 1 , wherein:
 the first group of vancomycin-resistant microorganisms comprises  Enterococcus faecium;      the second group of vancomycin-resistant microorganisms comprises  Enterococcus faecalis;      the third group of microorganisms is enterococcal bacteria that is not resistant to vancomycin or that have a natural resistance to vancomycin;
 the first substrate is an enzymatic substrate for detecting alpha-glucosidase activity; 
   the marker is a second substrate for detecting a second activity different than alpha-glucosidase activity; and   the antimicrobial is vancomycin.   
     
     
         12 . The method of  claim 1 , wherein:
 the first group of vancomycin-resistant microorganisms comprises  Enterococcus faecalis  and  Enterococcus faecium;      the second group of vancomycin-resistant microorganisms comprises  Staphylococcus aureus  that is intermediary resistant or resistant to vancomycin;   the third group of microorganisms are not resistant to vancomycin;   the first substrate is an enzymatic substrate for detecting alpha-glucosidase activity;   the marker is a second substrate for detecting a second activity different than alpha-glucosidase activity; and   the antimicrobial is vancomycin.   
     
     
         13 . The method of  claim 1 , wherein:
 the first group of vancomycin-resistant microorganisms comprises  Enterococcus faecalis  and  Enterococcus faecium;      the second group of vancomycin-resistant microorganisms comprises  Enterococcus casseliflavus  and  Enterococcus gallinarum;      the third group of microorganisms is not resistant to vancomycin;   the first substrate is an enzymatic substrate for detecting alpha-glucosidase activity;   the marker is a second substrate for detecting beta-glucosidase or beta-galactosidase activity; and   the antimicrobial is vancomycin.   
     
     
         14 . The method of  claim 1 , wherein:
 the first group of vancomycin-resistant microorganisms comprises  Enterococcus faecium;      the second group of vancomycin-resistant microorganisms comprises  Enterococcus faecalis;      the third group of microorganisms is enterococcal bacteria that is not resistant to vancomycin or that have a natural resistance to vancomycin;   the first substrate is 5-bromo-4-chloro-3-indolyl-N-methyl-α-D-glucoside or 5-bromo-4-chloro-3-indolyl-α-D-glucoside;   the marker is 6-chloro-3-indolyl-β-D-glucoside, alizarine-β-D-galactoside, 5-bromo-6-chloro-3-indolyl-β-D-glucoside, 5-bromo-6-chloro-3-indolyl-β-D-galactoside or 6-chloro-3-indolyl-β-D-galactoside; and   the antimicrobial is vancomycin.   
     
     
         15 . The method of  claim 1 , wherein:
 the first group of vancomycin-resistant microorganisms comprises  Enterococcus faecalis  and  Enterococcus faecium;      the second group of vancomycin-resistant microorganisms comprises  Staphylococcus aureus  that is intermediary resistant or resistant to vancomycin;   the third group of microorganisms is not resistant to vancomycin;   the first substrate is 5-bromo-4-chloro-3-indolyl-N-methyl-α-D-glucoside or 5-bromo-4-chloro-3-indolyl-α-D-glucoside;   the marker is 6-chloro-3-indolyl-β-D-glucoside or 5-bromo-6-chloro-3-indolyl-β-D-glucoside; and   the antimicrobial is vancomycin.   
     
     
         16 . A method for simultaneously distinguishing among at least three groups of microorganisms that are present in a biological sample, the at least three groups comprising:
 a first group of microorganisms belonging to a first taxon of Gram negative bacteria, the first group of bacteria exhibiting an acquired mechanism of resistance to a beta-lactamin and/or cephalosporin;   a second group of microorganisms belonging to a second taxon of Gram negative bacteria, the second taxon of bacteria being different than said first taxon, and exhibiting a mechanism of resistance to the beta-lactamin and/or cephalosporin identical to the mechanism of the first group; and   a third group of Gram negative bacteria that do not exhibit an acquired resistance to the beta-lactamin and/or cephalosporin,   
       the method comprising:
 inoculating a culture medium with the biological sample, wherein the culture medium comprises:
 a first substrate for detecting a first enzymatic or metabolic activity of said first group of microorganisms; 
 a marker for differentiating the first group of microorganisms and the second group of microorganisms, said marker being a substrate for detecting an enzymatic or metabolic activity of said second group of microorganisms; and 
 an antimicrobial that is active on the third group of microorganisms; and 
 
 simultaneously detecting and distinguishing between the first and second groups of microorganisms that are present on the culture medium, and identifying the taxon of the first group of microorganisms and the taxon of the second group of microorganisms on the basis of interactions of the first and second groups of microorganisms with the first substrate, the markers, and the antimicrobial. 
 
     
     
         17 . The method according to  claim 16 , wherein:
 the first group of microorganisms is selected from extended-spectrum β-lactamase producing (ESBL)  E. coli  and high-level chromosome class C cephalosporinases producing (HL Case) bacteria; and   the second group of microorganisms is selected from ESBL  Klebsiella, Enterobacter, Serratia , and  Citrobacter  (KESC) or HL Case bacteria.   
     
     
         18 . The method of  claim 16 , wherein:
 the first group of microorganisms is ESBL  E. coli  bacteria;   the second group of microorganisms is ESBL KESC bacteria; and   the third group of microorganisms is bacteria not resistant to beta-lactamines.   
     
     
         19 . The method of  claim 16 , wherein:
 the first group of microorganisms is HL Case  E. coli  bacteria;   the second group of microorganisms is HL Case KESC bacteria; and   the third group of microorganisms is bacteria not resistant to cephalosporins.   
     
     
         20 . The method of  claim 16 , wherein:
 the first group of microorganisms is ESBL  E. coli  bacteria;   the second group of microorganisms is ESBL KESC bacteria;   the third group of microorganisms is bacteria not resistant to beta-lactamines and/or cephalosporins; and   the biological sample further comprises a fourth group of microorganisms that is ESBL  Proteeae  bacteria.   
     
     
         21 . The method of  claim 16 , wherein:
 the first substrate is a substrate for detecting a beta-glucuronidase or beta-galactosidase enzymatic activity;   the marker is a second substrate for detecting a beta-glucosidase, tryptophanase, or desaminase activity; and   the antimicrobial is ceftazidime.   
     
     
         22 . The method of  claim 16 , wherein:
 the first substrate is a substrate for detecting a beta-glucuronidase or beta-galactosidase enzymatic activity;   the marker is a second substrate for detecting a beta-glucosidase, tryptophanase or desaminase activity; and   the antimicrobial is cefpodoxime and cloxacillin.   
     
     
         23 . The method of  claim 16 , wherein:
 the substrate is a first substrate for detecting a beta-glucuronidase or beta-galactosidase enzymatic activity;   the marker is a second substrate for detecting a beta-glucosidase, desaminase or tryptophanase activity; and   the antimicrobial is ceftriaxone and clavulanic acid.   
     
     
         24 . The method of  claim 16 , wherein:
 the substrate is a first substrate for detecting a beta-glucuronidase or beta-galactosidase enzymatic activity;   the marker is a second substrate for detecting a beta-glucosidase activity;   the antimicrobial is a combination comprising:
 cefpodoxime, 
 cloxacillin, 
 vancomycin, and 
 amphotericin B; and 
   the culture medium further comprises a third substrate for detecting a desaminase activity.   
     
     
         25 . The method of  claim 24 , wherein the antimicrobial further comprises cefsulodine. 
     
     
         26 . The method of  claim 16 , wherein:
 the first group of microorganisms is ESBL  E. coli  bacteria;   the second group of microorganisms is ESBL KESC bacteria;   the third group of microorganisms is bacteria not resistant to beta-lactamines and/or to cephalosporins;   the biological sample further comprises a fourth group of microorganisms that is ESBL  Proteeae  bacteria;   the first substrate is a substrate for detecting a beta-glucuronidase or beta-galactosidase enzymatic activity;   the marker is a second substrate for detecting a beta-glucosidase activity;   the antimicrobial is a combination comprising:
 cefpodoxime, 
 cloxacillin, 
 vancomycin, and 
 amphotericin B; and 
   the culture medium further comprises a third substrate for detecting a desaminase activity.   
     
     
         27 . The method of  claim 16 , wherein:
 the first group of microorganisms is ESBL  E. coli  bacteria;   the second group of microorganisms is ESBL KESC bacteria;   the third group of microorganisms is bacteria not resistant to beta-lactamines and/or to cephalosporins;   the biological sample further comprises a fourth group of microorganisms that is ESBL  Proteeae  bacteria;   the first substrate is a substrate for detecting a beta-glucuronidase or beta-galactosidase enzymatic activity;   the marker is a second substrate for detecting a beta-glucosidase activity;   the antimicrobial is a combination comprising:
 cefpodoxime, 
 cloxacillin, 
 vancomycin, 
 amphotericin B, and 
 cefsulodine; and 
   the culture medium further comprises a third substrate for detecting a desaminase activity.