title: Multiplex real-time PCR probe-based for identification of strains producing: OXA48, VIM, KPC and NDM.
abstract: The spread of multi-resistant enterobacteria, particularly carbapenem-resistant Enterobacteriaceae (CRE), in both community and hospital settings is a global problem. The phenotypic identification of CRE is complex, occasionally inconclusive and time consuming. However, commercially available molecular assays are very expensive, and many do not allow the simultaneous identification of all genetic markers of resistance that have been recognised in CRE (bla KPC, bla OXA-48, bla VIM and bla NDM). The aim of the present study is to describe a new test: a multiplex real time PCR probe-based assay designed for the simultaneous detection of KPC, OXA-48, VIM and NDM in a short time (no longer than 90 min from the extraction of DNA to detection). Our assay correctly identified 63 CRE isolates and all standard reference strains tested, in agreement with and extending the results of phenotypic identification tests; additionally, a KPC-VIM co-expressing Enterobacter aerogenes isolate was identified using the new assay, whereas traditional methods failed to detect it. The assay was also able to correctly detect 28 CRE-producers from 50 positive blood cultures, again detecting, in four specimens, the presence of CRE co-expressing KPC and VIM, which were only partially identified by traditional methods. Finally, when used directly on rectal swabs, the assay enabled the identification of CRE-carrier patients, for whom isolation is mandatory in a hospital setting.
authors.affiliation: Department of Experimental Medicine and Surgery, "Tor Vergata" University of Rome, Via Montpellier 1, 00133, Rome, Italy.
authors.forename: Marco
authors.initials: M
authors.lastname: Favaro
authors.forename: Mario
authors.initials: M
authors.lastname: Sarti
authors.forename: Carla
authors.initials: C
authors.lastname: Fontana
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D004755:Enterobacteriaceae / Q000145:classification / Q000201:enzymology* / Q000235:genetics / Q000302:isolation & purification*; D004756:Enterobacteriaceae Infections / Q000175:diagnosis* / Q000382:microbiology; D060885:Multiplex Polymerase Chain Reaction / Q000379:methods*; D015345:Oligonucleotide Probes; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D013997:Time Factors; D001618:beta-Lactamases / Q000235:genetics / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D015345:Oligonucleotide Probes; D001618:beta-Lactamases; C063912:carbapenemase