Sequence analysis of the penA gene, encoding penicillin-binding protein 2 (PBP2), in 30 penicillin-intermediate (PenI) Neisseria meningitidis strains showed altered gene sequences due to the translocation of exogenous DNA blocks derived from commensal neisseriae, which are known to have PBP2 proteins with decreased affinity for the antibiotic. In order to obtain a rapid and reproducible method for predicting the PenI phenotype, a real-time PCR assay was set up with primers and probes designed on the basis of the penA gene. The A→G mutation at codon 566, in the transpeptidase domain of the penA gene (which is present in the whole sample of 30 PenI strains and in all the 41 sequences of PenI meningococci isolated worldwide and has been deposited in the sequence databank), was chosen as a marker of penA translocations. Two hybridization probes were designed to distinguish the wild-type penA gene in penicillin-susceptible (PenS) meningococci from the mutated penA gene at codon 566 in PenI strains. Thermal analysis of probe hybridization revealed a melting temperature difference of at least 6°C between PenI and PenS strains. This real-time PCR protocol characterizes the penicillin phenotype of N. meningitidis in a few hours without DNA sequencing and is useful for rapid screening of the penicillin-intermediate genotype among meningococcal isolates.

Prediction of decreased susceptibility to penicillin of Neisseria meningitidis strains by real-time PCR / Stefanelli, P; Carattoli, A; Neri, A; Fazio, C; Mastrantonio, P.. - In: JOURNAL OF CLINICAL MICROBIOLOGY. - ISSN 1098-660X. - 41:10(2003), pp. 4666-4670. [10.1128/JCM.41.10.4666-4670.2003]

Prediction of decreased susceptibility to penicillin of Neisseria meningitidis strains by real-time PCR

Carattoli A;
2003

Abstract

Sequence analysis of the penA gene, encoding penicillin-binding protein 2 (PBP2), in 30 penicillin-intermediate (PenI) Neisseria meningitidis strains showed altered gene sequences due to the translocation of exogenous DNA blocks derived from commensal neisseriae, which are known to have PBP2 proteins with decreased affinity for the antibiotic. In order to obtain a rapid and reproducible method for predicting the PenI phenotype, a real-time PCR assay was set up with primers and probes designed on the basis of the penA gene. The A→G mutation at codon 566, in the transpeptidase domain of the penA gene (which is present in the whole sample of 30 PenI strains and in all the 41 sequences of PenI meningococci isolated worldwide and has been deposited in the sequence databank), was chosen as a marker of penA translocations. Two hybridization probes were designed to distinguish the wild-type penA gene in penicillin-susceptible (PenS) meningococci from the mutated penA gene at codon 566 in PenI strains. Thermal analysis of probe hybridization revealed a melting temperature difference of at least 6°C between PenI and PenS strains. This real-time PCR protocol characterizes the penicillin phenotype of N. meningitidis in a few hours without DNA sequencing and is useful for rapid screening of the penicillin-intermediate genotype among meningococcal isolates.
2003
DNA; gamma glutamyltransferase; penicillin binding protein; penicillin binding protein 2; penicillin derivative; unclassified drug, antibiotic sensitivity; article; bacterial gene; codon; DNA sequence; drug protein binding; gene mutation; gene sequence; hybridization; melting point; Neisseria meningitidis; nonhuman; nucleotide sequence; pena gene; penI gene; penS gene; phenotype; polymerase chain reaction; prediction; priority journal; sequence analysis; thermal analysis, Base Sequence; Humans; Microbial Sensitivity Tests; Molecular Sequence Data; Mutation; Neisseria meningitidis; Penicillin Resistance; Penicillins; Polymerase Chain Reaction; Predictive Value of Tests; Reproducibility of Results; Ribonucleoproteins; Saccharomyces cerevisiae Proteins; Sequence Analysis, DNA; Temperature; Time Factors, Bacteria (microorganisms); Negibacteria; Neisseria; Neisseria meningitidis
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Prediction of decreased susceptibility to penicillin of Neisseria meningitidis strains by real-time PCR / Stefanelli, P; Carattoli, A; Neri, A; Fazio, C; Mastrantonio, P.. - In: JOURNAL OF CLINICAL MICROBIOLOGY. - ISSN 1098-660X. - 41:10(2003), pp. 4666-4670. [10.1128/JCM.41.10.4666-4670.2003]
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11573/1284916
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