US2005009044A1PendingUtilityA1

Oligonucleotide probes for detecting Enterobacteriaceae and quinolone-resistant Enterobacteriaceae

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Assignee: GOVERNMENT OF THE USA AS REPREPriority: Apr 1, 1998Filed: Mar 10, 2004Published: Jan 13, 2005
Est. expiryApr 1, 2018(expired)· nominal 20-yr term from priority
Inventors:Linda WeigelFred C. Tenover
C07H 21/04C12Q 1/6827C12Q 1/689C07H 21/02Y10T436/143333Y02A50/30
40
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Claims

Abstract

Oligonucleotide probes for detecting Enterobacteriaceae species. Unique gyrA coding regions permit the development of probes specific for eight different species: Escherichia coli, Citrobacter freundii, Enterobacter aerogenes, Enterobacter cloacae, Klebsiella oxytoca, Klebsiella pneumoniae, Providencia stuartii and Serratia marcescens. The invention thereby provides methods for the species-specific identification of these Enterobacteriaceae in a sample, and detection and diagnosis of Enterobacteriaceae infection in a subject. Further, nucleic acids are provided for determining quinolone-resistant status of these Enterobacteriaceae.

Claims

exact text as granted — not AI-modified
1 . An isolated nucleic acid probe for identifying an  Enterobacteriaceae  species selected from the group consisting of  Escherichia coli, Citrobacter freundii, Enterobacter aerogenes, Enterobacter cloacae, Klebsiella oxytoca, Klebsiella pneumoniae, Providencia stuartii  and  Serratia marcescens,  wherein the probe selectively hybridizes to a portion of the nucleic acid of SEQ ID NOS: 1-8, or a complementary sequence thereof, respectively.  
     
     
         2 . The isolated nucleic acid probe of  claim 1 , wherein the probe selectively hybridizes to a portion of an  Escherichia coli  nucleic acid of SEQ ID NO:1, or a complementary sequence thereof.  
     
     
         3 . The isolated nucleic acid probe of  claim 1 , wherein the probe selectively hybridizes to a portion of a  Citrobacter freundii  nucleic acid of SEQ ID NO:2, or a complementary sequence thereof.  
     
     
         4 . The isolated nucleic acid probe of  claim 1 , wherein the probe selectively hybridizes to a portion of an  Enterobacter aerogenes  nucleic acid of SEQ ID NO:3, or a complementary sequence thereof.  
     
     
         5 . The isolated nucleic acid probe of  claim 1 , wherein the probe selectively hybridizes to a portion of an  Enterobacter cloacae  nucleic acid of SEQ ID NO:4, or a complementary sequence thereof.  
     
     
         6 . The isolated nucleic acid probe of  claim 1 , wherein the probe selectively hybridizes to a portion of a  Klebsiella oxytoca  nucleic acid of SEQ ID NO:5, or a complementary sequence thereof.  
     
     
         7 . The isolated nucleic acid probe of  claim 1 , wherein the probe selectively hybridizes to a portion of a  Klebsiella pneumoniae  nucleic acid of SEQ ID NO:6, or a complementary sequence thereof.  
     
     
         8 . The isolated nucleic acid probe of  claim 1 , wherein the probe selectively hybridizes to a portion of a  Providencia stuartii  nucleic acid of SEQ ID NO:7, or a complementary sequence thereof.  
     
     
         9 . The isolated nucleic acid probe of  claim 1 , wherein the probe selectively hybridizes to a portion of a  Serratia marcescens  nucleic acid of SEQ ID NO:8, or a complementary sequence thereof.  
     
     
         10 . An isolated nucleic acid probe having a nucleic acid sequence of SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20, SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23, or SEQ ID NO:24, or a complementary sequence thereof.  
     
     
         11 . A method of identifying in a sample an  Enterobacteriaceae  species selected from the group consisting of  Escherichia coli, Citrobacter freundii, Enterobacter aerogenes, Enterobacter cloacae, Klebsiella oxytoca, Klebsiella pneumoniae, Providencia stuartii  and  Serratia marcescens  comprising combining the sample with a nucleic acid probe, wherein the probe selectively hybridizes to a portion of a nucleic acid of SEQ ID NOS: 1-8, or a complementary sequence thereof, respectively, the presence of hybridization with a nucleic acid indicating the identity of the respective species.  
     
     
         12 . The method of identifying an  Enterobacteriaceae  species of  claim 11 , comprising combining the sample with a nucleic acid probe, wherein the probe selectively hybridizes to a portion of a nucleic acid of SEQ ID NO: 1, or a complementary sequence thereof, the presence of hybridization indicating  Escherichia coli  in the sample.  
     
     
         13 . The method of identifying an  Enterobacteriaceae  species of  claim 11 , comprising combining the sample with a nucleic acid probe, wherein the probe selectively hybridizes to a portion of a nucleic acid of SEQ ID NO:2, or a complementary sequence thereof, the presence of hybridization indicating  Citrobacter freundii  in the sample.  
     
     
         14 . The method of identifying an  Enterobacteriaceae  species of  claim 11 , comprising combining the sample with a nucleic acid probe, wherein the probe selectively hybridizes to a portion of a nucleic acid of SEQ ID NO:3, or a complementary sequence thereof, the presence of hybridization indicating  Enterobacter aerogenes  in the sample.  
     
     
         15 . The method of identifying an  Enterobacteriaceae  species of  claim 11 , comprising combining the sample with a nucleic acid probe, wherein the probe selectively hybridizes to a portion of a nucleic acid of SEQ ID NO:4, or a complementary sequence thereof, the presence of hybridization indicating  Enterobacter cloacae  in the sample.  
     
     
         16 . The method of-identifying an  Enterobacteriaceae  species of  claim 11 , comprising combining the sample with a nucleic acid probe, wherein the probe selectively hybridizes to a portion of a nucleic acid of SEQ ID NO:5, or a complementary sequence thereof, the presence of hybridization indicating  Klebsiella oxytoca  in the sample.  
     
     
         17 . The method of identifying an  Enterobacteriaceae  species of  claim 11 , comprising combining the sample with a nucleic acid probe, wherein the probe selectively hybridizes to a portion of a nucleic acid of SEQ ID NO:6, or a complementary sequence thereof, the presence of hybridization indicating  Klebsiella pneumoniae  in the sample.  
     
     
         18 . The method of identifying an  Enterobacteriaceae  species of  claim 11 , comprising combining the sample with a nucleic acid probe, wherein the probe selectively hybridizes to a portion of a nucleic acid of SEQ ID NO:7, or a complementary sequence thereof, the presence of hybridization indicating  Providencia stuartii  in the sample.  
     
     
         19 . The method of identifying an  Enterobacteriaceae  species of  claim 11 , comprising combining the sample with a nucleic acid probe, wherein the probe selectively hybridizes to a portion of a nucleic acid of SEQ ID NO:8, or a complementary sequence thereof, the presence of hybridization indicating  Serratia marcescens  in the sample.  
     
     
         20 . An isolated nucleic acid probe capable of determining the quinolone resistance status of an  Enterobacteriaceae  species selected from the group consisting of  Escherichia coli, Citrobacter freundii, Enterobacter aerogenes, Enterobacter cloacae, Klebsiella oxytoca, Klebsiella pneumoniae, Providencia stuartii  and  Serratia marcescens,  wherein the probe selectively hybridizes to a portion of a nucleic acid of SEQ ID NOS:1-8, or a complementary sequence thereof, respectively.  
     
     
         21 . The isolated nucleic acid probe of  claim 20 , wherein the probe selectively hybridizes to a portion of an  Escherichia coli  nucleic acid of SEQ ID NO:1, or a complementary sequence thereof.  
     
     
         22 . The isolated nucleic acid probe of  claim 20 , wherein the probe selectively hybridizes to a portion of a  Citrobacter freundii  nucleic acid of SEQ ID NO:2, or a complementary sequence thereof.  
     
     
         23 . The isolated nucleic acid probe of  claim 20 , wherein the probe selectively hybridizes to a portion of an  Enterobacter aerogenes  nucleic acid of SEQ ID NO:3, or a complementary sequence thereof.  
     
     
         24 . The isolated nucleic acid probe of  claim 20 , wherein the probe selectively hybridizes to a portion of an  Enterobacter cloacae  nucleic acid of SEQ ID NO:4, or a complementary sequence thereof.  
     
     
         25 . The isolated nucleic acid probe of  claim 20 , wherein the probe selectively hybridizes to a portion of a  Klebsiella oxytoca  nucleic acid of SEQ ID NO:5, or a complementary sequence thereof.  
     
     
         26 . The isolated nucleic acid probe of  claim 20 , wherein the probe selectively hybridizes to a portion of a  Klebsiella pneumoniae  nucleic acid of SEQ ID NO:6, or a complementary sequence thereof.  
     
     
         27 . The isolated nucleic acid probe of  claim 20 , wherein the probe selectively hybridizes to a portion of a  Providencia stuartii  nucleic acid of SEQ ID NO:7, or a complementary sequence thereof.  
     
     
         28 . The isolated nucleic acid probe of  claim 20 , wherein the probe selectively hybridizes to a portion of a  Serratia marcescens  nucleic acid of SEQ ID NO:8, or a complementary sequence thereof.  
     
     
         29 . An isolated nucleic acid probe having a nucleic acid sequence of SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:27, SEQ ID NO:28, SEQ ID NO:29, SEQ ID NO:30, SEQ ID NO:31, SEQ ID NO:32, or SEQ ID NO:33, or a complementary sequence thereof.  
     
     
         30 . A method of determining the quinolone resistance of an  Enterobacteriaceae  species selected from the group consisting of  Escherichia coli, Citrobacter freundii, Enterobacter aerogenes, Enterobacter cloacae, Klebsiella oxytoca, Klebsiella pneumoniae, Providencia stuartii  and  Serratia marcescens  in a sample, comprising combining the sample with a nucleic acid probe, wherein the probe selectively hybridizes to a nucleic acid of SEQ ID NOS:9-16, or a complementary sequence thereof, respectively, the presence of hybridization with a nucleic acid indicating the quinolone resistance of the respective species.  
     
     
         31 . The method of determining the quinolone resistance status of an  Enterobacteriaceae  species of  claim 30 , comprising combining the sample with a nucleic acid probe, wherein the probe selectively hybridizes to a nucleic acid of SEQ ID NO:9, or a complementary sequence thereof, the presence of hybridization indicating quinolone resistance of the  Escherichia coli  in the sample.  
     
     
         32 . The method of determining the quinolone resistance status of an  Enterobacteriaceae  species of  claim 30 , comprising combining the sample with a nucleic acid probe, wherein the probe selectively hybridizes to a nucleic acid of SEQ ID NO:10, or a complementary sequence thereof, the presence of hybridization indicating quinolone resistance of the  Citrobacter freundii  in the sample.  
     
     
         33 . The method of determining the quinolone resistance status of an  Enterobacteriaceae  species of  claim 30 , comprising combining the sample with a nucleic acid probe, wherein the probe selectively hybridizes to a nucleic acid of SEQ ID NO:11, or a complementary sequence thereof, the presence of hybridization indicating quinolone resistance of the  Enterobacter aerogenes  in the sample.  
     
     
         34 . The method of determining the quinolone resistance status of an  Enterobacteriaceae  species of  claim 30 , comprising combining the sample with a nucleic acid probe, wherein the probe selectively hybridizes to a nucleic acid of SEQ ID NO:12, or a complementary sequence thereof, the presence of hybridization indicating quinolone resistance of the  Enterobacter cloacae  in the sample.  
     
     
         35 . The method of determining the quinolone resistance status of an  Enterobacteriaceae  species of  claim 30 , comprising combining the sample with a nucleic acid probe, wherein the probe selectively hybridizes to a nucleic acid of SEQ ID NO:13, or a complementary sequence thereof, the presence of hybridization indicating quinolone resistance of the  Klebsiella oxytoca  in the sample.  
     
     
         36 . The method of determining the quinolone resistance status of an  Enterobacteriaceae  species of  claim 30 , comprising combining the sample with a nucleic acid probe, wherein the probe selectively hybridizes to a nucleic acid of SEQ ID NO:14, or a complementary sequence thereof, the presence of hybridization indicating quinolone resistance of the  Klebsiella pneumoniae  in the sample.  
     
     
         37 . The method of determining the quinolone resistance status of an  Enterobacteriaceae  species of  claim 30 , comprising combining the sample with a nucleic acid probe, wherein the probe selectively hybridizes to a nucleic acid of SEQ ID NO:15, or a complementary sequence thereof, the presence of hybridization indicating quinolone resistance of the  Providencia stuartii  in the sample.  
     
     
         38 . The method of determining the quinolone resistance status of an  Enterobacteriaceae  species of  claim 30 , comprising combining the sample with a nucleic acid probe, wherein the probe selectively hybridizes to a nucleic acid of SEQ ID NO:16, or a complementary sequence thereof, the presence of hybridization indicating quinolone resistance of the  Serratia marcescens  in the sample.

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