An evaluation of reverse transcription loop-mediated isothermal amplification (RTLAMP) for diagnosis of rabies.
Rabies is a neurological disease caused by a lyssavirus belonging to the family rhabdoviridae. The causative agent has a global distribution (except Antarctica) resulting in an estimated 59,000 human deaths worldwide. The true disease burden and public health impact due to rabies remain underestimated due to lack of sensitive laboratory diagnostic methods. This is particularly common in underdeveloped countries. This study was aimed at evaluating Reverse Transcriptase Loop-Mediated Isothermal Amplification (RT-LAMP) as a viable alternative method to RT-PCR and DFAT for diagnosis of rabies in Zambia. RT-LAMP primers were designed based on the most conserved region of N gene sequences covering various rabies virus isolates representative sequences from around Africa. Amplification products of the RTLAMP assay was monitored by visual colour change of fluorescent dye and by agarose gel electrophoresis. To evaluate the RT-LAMP assay’s applicability to rabies, archived samples stored at -80◦C at the University of Zambia repository were used. A total of 84 samples were tested using both RT-LAMP and RT-PCR, with 25 samples coming from Zimbabwe, 40 samples coming from Zambia, and 19 samples coming from Malawi. The results of the RTLAMP were compared with RT-PCR assay. The RT-LAMP assay was successfully optimized with a turnaround time of about 2 hours and diagnostic sensitivity and specificity of 96.1% and 100%, respectively. The limit of detection (LOD) of the RT-LAMP was 2.19x10-5 ηg/μl of rabies sample RNA. In conclusion, RT-LAMP is a promising alternative to RT-PCR and DFAT for the diagnosis of rabies, offering several advantages in terms of cost-effectiveness, simplicity, and sensitivity. While further studies are needed to validate its performance in different settings, RT-LAMP has the potential to become a valuable tool in the global fight against rabies.
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