Loop-Mediated Isothermal Amplification: Rapid And Sensitive Detection Of The Antibiotic Resistance Gene Isaba1-Bla(Oxa-51-Like) In Acinetobacter Baumannii

Carbapenem-resistant Acinetobacter baumannii, which are mainly induced by the production of OXA-type beta-lactamases, are among the leading causes of nosocomial infections worldwide. Among the beta-lactamase genes, the presence of the OXA-51-like gene carrying the upstream insertion sequence, ISAba1, was found to be one of the most prevalent carbapenem resistance mechanisms utilized by these bacteria. Consequently, it is necessary to develop a rapid detection method for ISAba1-bla(OXA-51-like) sequence for the timely and appropriate antibiotic treatment of A. baumannii infection. In this study, a loop-mediated isothermal amplification (LAMP) assay was optimized for ISAba1-bla(OXA-51-like) detection. The LAMP primer set was designed to recognize distinct sequences in the ISAba1-bla(OXA-51-like) gene and could amplify the gene within 25 min at an isothermal temperature of 60 degrees C. This LAMP assay was able to detect the ISAba1-bla(OXA-51-like) gene with high specificity; in addition, no cross reactivity was observed for other types of beta-lactamase producers (OXA-23-like, OXA-40-like, OXA-58-like, and IMP-1), as indicated by the absence of false positive or false negative results. The detection limit for this assay was found to be 10(0) CFU per tube which was 100-fold more sensitive than a polymerase chain reaction assay for ISAba1-bla(OXA-51-like) detection. Furthermore, the LAMP assay provided swift detection of the ISAba1-bla(OXA-51-like) gene, even directly from clinical specimens. In summary, we have described a new, rapid assay for the detection of theISAba1-bla(OXA-51-like) gene from A. baumannii that could be useful in a clinical setting. This method might facilitate epidemiological studies and allow monitoring of the emergence of drug resistant strains. (C) 2015 Elsevier B.V. All rights reserved.