Background: Recognized as a resistance mechanism responsible for the emergence and prevalence of antimicrobial resistance, integron is widely distributed and spread among clinical microorganisms and play a key role in the dissemination of such antimicrobial resistance, which may eventually contribute to the unleashing of "Super Bugs" In this study, detection assays based on loop-mediated isothermal amplification (LAMP) methodologies targeting on class 1 to class 3 integrase genes was developed and evaluated.
Methods: LAMP methodology was employed to develop novel detection assays on class 1, 2 and 3 integrons. Firstly, this protocol was specifically designed to detect such integrons by targeting integrase genes intI1, intI2 and intI3. Development, evaluation and optimization of such LAMP assays was studied, including the reaction temperature, volumn, time, sensitivity and specificity of both primers and targets. A total of 1082 strains, including 397 integron positive and 685 integron negative microorganisms, were included for the application verification of the established LAMP assays.
Results: The indispensability of each primer was confirmed, and the optimal amplification was obtained under 63 °C for 45 min, with 25 μl reaction found to be the most cost-efficient volume. As application was concerned, all of the 397 integron-positive isolates yielded positive amplicons and other 685 integron-negative bacteria were negative for the integron-LAMP assays, revealing totaling 100% detection rate and specificity.
Conclusions: The established integron-LAMP assays was demonstrated to be a valid and rapid detection method for integrons screening, which may aid in both the laboratory and clinical integron screening for microorganisms.
Keywords: Bacterial integrons; Class 1 integron; Class 2 integron; Class 3 integron; Loop-mediated isothermal amplification (LAMP); Virulome.
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