Comparative detection of foot-and-mouth disease virus by reverse transcription loop-mediated isothermal amplification assay and real time polymerase chain reaction in Uganda

 

Hussein Kafeero Mukasa¹*, Frank Norbert Mwiine¹ , David Kalenzi Atuhaire², Sylvester Ochwo¹ and Ann Nanteza¹

 

Research Article | Published March 2016

International Journal of Biotechnology and Food Science, Vol. 4(2), pp. 22-33

 

 

¹College of Veterinary Medicine, Animal resources and Biosecurity, Makerere University, P. O. BOX 7062 Kampala, Uganda.

²Research and development Manager, Centre for Ticks and Tick-Borne Diseases, Private Bag A130, Lilongwe, Malawi.

 

*Corresponding author. E-mail: husseinmukasakafeero@gmail.com.     

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Foot-and-mouth disease (FMD) is a viral disease. FMD diagnosis in the field is based on clinical signs that are shared by other vesicular diseases, hence to confirm FMD a laboratory is needed. Laboratory diagnostic techniques including serology may fail to distinguish between vaccinated and new infection, virus isolation may take up to 4 days to yield results, while molecular techniques including PCR, which are accurate, sensitive, specific and rapid, are costly and require special training of the laboratory staff. These challenges limit laboratory diagnosis yet in Uganda FMD outbreaks are common since the disease is endemic. This work reports the comparative detection of Foot-and-mouth disease virus (FMDV) by reverse transcription-loop mediated isothermal amplification (RT-LAMP) and Real-Time polymerase chain reaction (rRT-PCR) in Uganda based on the 3D polymerase (3D^pol) gene. The rRT-PCR assay is considered as the gold standard. A total of 89 cattle samples that included epithelial tissues (16.9%) and oral swabs (83.1%) were collected from outbreak cases in Eastern Districts of Mbale and Budaka. These were applied to molecular assays of rRT-PCR and RT- LAMP using primers and probes targeting the 3D^pol gene. The diagnostic sensitivity and specificity of RT-LAMP as a screening test and rRT-PCR as the reference test was 94.44% (95% CI = 94.11 to 94.78%) and 98.59% (95% CI = 98.50 to 98.68%), respectively. The kappa value for diagnostic agreement between rRT-PCR and RT-LAMP was 93.0% (95% CI = 83.50 to 100%), showing a perfect agreement. In conclusion, the RT-LAMP assay had a very high sensitivity and specificity when compared to the reference test of rRT-PCR. It was also very rapid since it gave results in 45 to 60 min. Due to its simplicity, sensitivity and specificity, LAMP assay has the potential for use in routine surveillance of FMD in Uganda.

 

Keywords: Foot-and-mouth disease virus, real-time polymerase chain reaction, reverse transcription LAMP, sensitivity, specificity.

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Citation: Mukasa HK, Mwiine FN, Atuhaire DK, Ochwo S, Nanteza A (2016). Comparative detection of foot-and-mouth disease virus by reverse transcription loop-mediated isothermal amplification assay and real time polymerase chain reaction in Uganda. Int. J. Biotechnol. Food Sci. 4(2): 22-33.

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