Reverse Transcriptase-Loop Mediated Isothermal Amplification (RT-LAMP): a rapid detection method for Sac brood viral disease infecting Apis cerana indica Fabricius

Abstract

Sac brood virus (SBV) also called AcSBV that infects Apis cerana indica, the Indian honey bee, is an important and serious virus disease in honey bee colonies causing losses upto 80% in South India. The methods currently employed to diagnose the bee virus, require sophisticated equipments, have low specificity and sensitivity and are also costly. Hence, there is a need to develop a simple and cheap method for the detection of SBV. In this research work, Reverse Transcriptase loop-mediated isothermal amplification (RT-LAMP) assay which is a newer method that is relatively simpler and cheaper than the conventional RT-PCR was tested. The RT-PCR and RT-LAMP methods were compared for their efficiencies to diagnose the AcSBV. The cDNA isolated from 11 virus infected honey bee samples were used for the study. The detection rate of RT-PCR and RT-LAMP was 10/11(91%) and 3/11(27%) for the field samples, respectively. The RT-LAMP assay was more specific than RT-PCR for the detection of SBV and more primer sets from conserved regions of the genome have to be designed and used in detection to reduce the chances of missing positive samples. The results demonstrate clearly that this LAMP-based assay is quick, cheap, rapid and easier to detect the target gene, within 2 hrs and just a standard laboratory water bath or heat block is a useful tool for the rapid and sensitive diagnosis of SBV infection of bees.