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Detection of ciprofloxacin resistance mutations in Campylobacter jejuni gyrA by nonradioisotopic single-strand conformation polymorphism and direct DNA sequencing. 1996

E Charvalos, and E Peteinaki, and I Spyridaki, and S Manetas, and Y Tselentis
Department of Bacteriology, Parasitology, Zoonoses and Geographical Medicine, School of Health Sciences, University of Crete, Heracleion, Greece.

A total of 27 strains of Campylobacter jejuni (24 clinical strains and three laboratory strains) were examined for the presence of point mutations in the quinolone resistance determining region (QRDR) of gyrA gene by nonradioisotopic single-strand conformation polymorphism (non-RI SSCP) analysis with silver stain. Direct DNA sequencing of the polymerase chain reaction (PCR)-amplified DNA fragments confirmed the results obtained by non-RI SSCP analysis and revealed that in clinical strains high-level quinolone resistance [minimal inhibitory concentration (MIC) to ciprofloxacin > or = 16 micrograms/ml] was closely associated with one type of single-point mutation at codon 86 (Thr-Ile). Two strains with MICs of 8 and 1 microgram/ml showed point mutations at codons 86 and 70, respectively. Furthermore, transitions at codon 119 of the gyrA QRDR were identified in 17 strains. Six types of bands were separated in a single electrophoretic step with silver stain within 2 hours after PCR amplification of the gyrA QRDR as follows: type I associated to mutation at codon 70 (Ala-Thr), type II to mutation at codon 90 (Asp-Asn), type III to variant with transition at 119, type IV to wild-type, type V to mutation at codon 86 (Thr-Ile), and type VI to mutation at codon 86 (Thr-Ile) and transition at codon 119. Using four DNA extracts from Cambylobacter coli organisms as templates for amplification of the gyrA QRDR, no PCR products were obtained. Non-RI SSCP was proved to be a simple, rapid, and useful screening method for detecting gyrA mutations associated with ciprofloxacin resistance in C. jejuni.

UI MeSH Term Description Entries
D008969 Molecular Sequence Data Descriptions of specific amino acid, carbohydrate, or nucleotide sequences which have appeared in the published literature and/or are deposited in and maintained by databanks such as GENBANK, European Molecular Biology Laboratory (EMBL), National Biomedical Research Foundation (NBRF), or other sequence repositories. Sequence Data, Molecular,Molecular Sequencing Data,Data, Molecular Sequence,Data, Molecular Sequencing,Sequencing Data, Molecular
D002939 Ciprofloxacin A broad-spectrum antimicrobial carboxyfluoroquinoline. Bay-09867,Ciprinol,Cipro,Ciprofloxacin Hydrochloride,Ciprofloxacin Hydrochloride Anhydrous,Ciprofloxacin Monohydrochloride Monohydrate,Anhydrous, Ciprofloxacin Hydrochloride,Bay 09867,Bay09867,Hydrochloride Anhydrous, Ciprofloxacin,Hydrochloride, Ciprofloxacin,Monohydrate, Ciprofloxacin Monohydrochloride,Monohydrochloride Monohydrate, Ciprofloxacin
D004250 DNA Topoisomerases, Type II DNA TOPOISOMERASES that catalyze ATP-dependent breakage of both strands of DNA, passage of the unbroken strands through the breaks, and rejoining of the broken strands. These enzymes bring about relaxation of the supercoiled DNA and resolution of a knotted circular DNA duplex. DNA Topoisomerase (ATP-Hydrolysing),DNA Topoisomerase II,DNA Topoisomerase II alpha,DNA Topoisomerase II beta,DNA Type 2 Topoisomerase,TOP2A Protein,TOP2B Protein,Topoisomerase II,Topoisomerase II alpha,Topoisomerase II beta,Type II DNA Topoisomerase,alpha, Topoisomerase II,beta, Topoisomerase II
D004352 Drug Resistance, Microbial The ability of microorganisms, especially bacteria, to resist or to become tolerant to chemotherapeutic agents, antimicrobial agents, or antibiotics. This resistance may be acquired through gene mutation or foreign DNA in transmissible plasmids (R FACTORS). Antibiotic Resistance,Antibiotic Resistance, Microbial,Antimicrobial Resistance, Drug,Antimicrobial Drug Resistance,Antimicrobial Drug Resistances,Antimicrobial Resistances, Drug,Drug Antimicrobial Resistance,Drug Antimicrobial Resistances,Drug Resistances, Microbial,Resistance, Antibiotic,Resistance, Drug Antimicrobial,Resistances, Drug Antimicrobial
D004591 Electrophoresis, Polyacrylamide Gel Electrophoresis in which a polyacrylamide gel is used as the diffusion medium. Polyacrylamide Gel Electrophoresis,SDS-PAGE,Sodium Dodecyl Sulfate-PAGE,Gel Electrophoresis, Polyacrylamide,SDS PAGE,Sodium Dodecyl Sulfate PAGE,Sodium Dodecyl Sulfate-PAGEs
D000890 Anti-Infective Agents Substances that prevent infectious agents or organisms from spreading or kill infectious agents in order to prevent the spread of infection. Anti-Infective Agent,Anti-Microbial Agent,Antimicrobial Agent,Microbicide,Microbicides,Anti-Microbial Agents,Antiinfective Agents,Antimicrobial Agents,Agent, Anti-Infective,Agent, Anti-Microbial,Agent, Antimicrobial,Agents, Anti-Infective,Agents, Anti-Microbial,Agents, Antiinfective,Agents, Antimicrobial,Anti Infective Agent,Anti Infective Agents,Anti Microbial Agent,Anti Microbial Agents
D001483 Base Sequence The sequence of PURINES and PYRIMIDINES in nucleic acids and polynucleotides. It is also called nucleotide sequence. DNA Sequence,Nucleotide Sequence,RNA Sequence,DNA Sequences,Base Sequences,Nucleotide Sequences,RNA Sequences,Sequence, Base,Sequence, DNA,Sequence, Nucleotide,Sequence, RNA,Sequences, Base,Sequences, DNA,Sequences, Nucleotide,Sequences, RNA
D016123 Campylobacter jejuni A species of bacteria that resemble small tightly coiled spirals. Its organisms are known to cause abortion in sheep and fever and enteritis in man and may be associated with enteric diseases of calves, lambs, and other animals. Campylobacter fetus subsp. jejuni,Vibrio hepaticus,Vibrio jejuni
D016133 Polymerase Chain Reaction In vitro method for producing large amounts of specific DNA or RNA fragments of defined length and sequence from small amounts of short oligonucleotide flanking sequences (primers). The essential steps include thermal denaturation of the double-stranded target molecules, annealing of the primers to their complementary sequences, and extension of the annealed primers by enzymatic synthesis with DNA polymerase. The reaction is efficient, specific, and extremely sensitive. Uses for the reaction include disease diagnosis, detection of difficult-to-isolate pathogens, mutation analysis, genetic testing, DNA sequencing, and analyzing evolutionary relationships. Anchored PCR,Inverse PCR,Nested PCR,PCR,Anchored Polymerase Chain Reaction,Inverse Polymerase Chain Reaction,Nested Polymerase Chain Reaction,PCR, Anchored,PCR, Inverse,PCR, Nested,Polymerase Chain Reactions,Reaction, Polymerase Chain,Reactions, Polymerase Chain
D017354 Point Mutation A mutation caused by the substitution of one nucleotide for another. This results in the DNA molecule having a change in a single base pair. Mutation, Point,Mutations, Point,Point Mutations

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