A Novel Close-tube Loop Mediated Isothermal Amplification (LAMP) Assay for Rapid Detection of Brucella

Abstract

Background: Brucellosis is one of the important diseases affecting both human as well as livestock. Rapid diagnosis of the pathogen is highly essential for undertaking effective therapeutic measures. Loop mediated isothermal amplification (LAMP) assay, one of the robust nucleic acid detection platforms, is used especially for rapid disease diagnosis. However, persistent contamination has been the major bottleneck in LAMP which can be eliminated with adoption of appropriate closed-tube formats.Methods: In the present report, two sets of LAMP primers targeting omp2b gene of Brucella were designed and standardized for detection of all the major Brucella spp. Result: There was absence of amplification in any of the non-Brucella species in contrast to detection of white precipitation in unaided eye in daylight as well as greenish fluorescence under ultraviolet light in all of the Brucella species. It was found to be more sensitive than conventional PCR as relative sensitivity was found to be 0.34pg for first set of primers and 34fg for second set of primers, as compared to 3.4pg through bcsp31 PCR.