Dataset Information

Conventional and Real-Time PCR Targeting blaOXA Genes as Reliable Methods for a Rapid Detection of Carbapenem-Resistant Acinetobacter baumannii Clinical Strains.

ABSTRACT: Multidrug-resistant Acinetobacter baumannii, particularly those producing carbapenemases, are spread worldwide. A reliable detection of carbapenemases is essential to choose the appropriate antimicrobial therapy and, consequently, prevent the dissemination of carbapenem-resistant strains. The aim of this study is to examine the molecular basis of the carbapenem resistance mechanism and estimation of conventional PCR and real-time PCR usefulness for the detection of oxacillinases when compared to phenotypic carbapenemases detection. The following methods were evaluated: the CarbAcineto NP test, Carbapenem Inactivation Method, CPO panels of semiautomated antimicrobial susceptibility testing method on the BD Phoenix™ M50 system, conventional Polymerase Chain Reaction and real-time PCR. The eazyplex® SuperBug complete A assay was used as the reference method. Among the tested strains, 39 (67.2%) carried the blaOXA-40 gene, while the blaOXA-23 gene was noted amongst 19 (32.8%) isolates. The diagnostic sensitivities of the studied assays were as follows: CarbAcineto NP-65.5%; CIM-100%; CPO-100%; conventional PCR-100%; real-time PCR-100%.

SUBMITTER: Depka D

PROVIDER: S-EPMC9032895 | biostudies-literature | 2022 Mar

REPOSITORIES: biostudies-literature

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Publications

Conventional and Real-Time PCR Targeting blaOXA Genes as Reliable Methods for a Rapid Detection of Carbapenem-Resistant Acinetobacter baumannii Clinical Strains.

Depka Dagmara D Mikucka Agnieszka A Bogiel Tomasz T Rzepka Mateusz M Zawadka Patryk P Gospodarek-Komkowska Eugenia E

Antibiotics (Basel, Switzerland) 20220328 4

Multidrug-resistant Acinetobacter baumannii, particularly those producing carbapenemases, are spread worldwide. A reliable detection of carbapenemases is essential to choose the appropriate antimicrobial therapy and, consequently, prevent the dissemination of carbapenem-resistant strains. The aim of this study is to examine the molecular basis of the carbapenem resistance mechanism and estimation of conventional PCR and real-time PCR usefulness for the detection of oxacillinases when comp ...[more]

PMID: 35453207

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Project description:Nosocomial infections caused by Acinetobacter spp. resistant to carbapenems are increasingly reported worldwide. Carbapenem-resistant Acinetobacter (CRA) is becoming a serious concern with increasing patient morbidity, mortality, and lengths of hospital stay. Therefore, the rapid detection of CRA is essential for epidemiological surveillance. Polymerase chain reaction (PCR) has been extensively used for the rapid identification of most pathogens. In this study, we have developed two multiplex real-time PCR assays to detect and differentiate A. baumannii and non-A. baumannii Acinetobacter spp, and common carbapenemase genes, including blaNDM, blaOXA-23-like, blaOXA-40-like, blaOXA-51-like, and blaOXA-58-like. We demonstrate the potential utility of these assays for the direct detection of blaNDM-, blaOXA-23-like-, blaOXA-40-like-, blaOXA-51-like-, and blaOXA-58-like-positive CRA in clinical specimens. Primers were specifically designed, and two multiplex real-time PCR assays were developed: multiplex real-time PCR assay1 for the detection of Acinetobacter baumannii 16S-23S rRNA internal transcribed spacer sequence, the Acinetobacter recA gene, and class-B-metalloenzyme-encoding gene blaNDM; and multiplex real-time PCR assay2 to detect class-D-oxacillinase-encoding genes (blaOXA-23-like, blaOXA-40-like, blaOXA-51-like,and blaOXA-58-like). The assays were performed on an ABI Prism 7500 FAST Real-Time PCR System. CRA isolates were used to compare the assays with conventional PCR and sequencing. Known amounts of CRA cells were added to sputum and fecal specimens and used to test the multiplex real-time PCR assays. The results for target and nontarget amplification showed that the multiplex real-time PCR assays were specific, the limit of detection for each target was 10 copies per 20 μL reaction volume, the assays were linear over six log dilutions of the target genes (r2 > 0.99), and the Ct values of the coefficients of variation for intra- and interassay reproducibility were less than 5%. The multiplex real-time PCR assays showed 100% concordance with conventional PCR when tested against 400 CRA isolates and their sensitivity for the target DNA in sputum and fecal specimens was 102 CFU/mL. Therefore, these novel multiplex real-time PCR assays allow the sensitive and specific characterization and differentiation of blaNDM-, blaOXA-23-like-, blaOXA-40-like-, blaOXA-51-like-, and blaOXA-58-like-positive CRA, making them potential tools for the direct detection of CRA in clinical specimens and the surveillance of nosocomial infections.

Dataset Information

Conventional and Real-Time PCR Targeting blaOXA Genes as Reliable Methods for a Rapid Detection of Carbapenem-Resistant Acinetobacter baumannii Clinical Strains.

Publications

Conventional and Real-Time PCR Targeting <i>bla</i><sub>OXA</sub> Genes as Reliable Methods for a Rapid Detection of Carbapenem-Resistant <i>Acinetobacter baumannii</i> Clinical Strains.

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