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Standardization of Reverse Transcription Loop-Mediated Isothermal Amplification (Rt-Lamp) Diagnostic Test for Rapid Detection of Foot and Mouth Disease Virus

Standardization of Reverse Transcription Loop-Mediated Isothermal Amplification (Rt-Lamp) Diagnostic Test for Rapid Detection of Foot and Mouth Disease Virus

Tahira Kamal1*, Saeed-Ul-Hassan Khan2, Amir Bin Zahoor3, Khalid Naeem3, Muhammad Naeem Riaz1, Siddra Tayyab Akhtar4, Ghulam Muhammad Ali1*

1National Institute of Genomics and Advanced Biotechnology, NARC, Islamabad; 2Quaid-I-Azam University, Islamabad; 3Animal Health, NARC, Islamabad; 4Punjab University, Lahore, Pakistan.

Email: tahirakhurram3@gmail.com; drgmail@yahoo.ca

 

ABSTRACT

Foot and Mouth Disease Virus (FMD) is a RNA virus, member of Picornaviridae. It has seven serotypes with no cross protection among these. FMD is responsible for heavy economic losses in cattle and buffalo. So rapid and accurate diagnosis of FMD is uttermost need of this time. Several lab.techniques are developed for rapid detection of FMD but these are expensive and time consuming. Elisa, cell culture, Reverse Transcriptase (RT-PCR) and real time PCR need specific high quality equipments in lab. Whereas Lamp PCR is simple and accurate test for the rapid diagnosis of FMD. This study has been done initially with FMD known serotypes (O, A, Asia1) already in use at animal health.Viral RNAs were extracted using R Neasy Minikit (Qiagen) according to instruction manual. After extraction RNAs were eluted in 60 ul elution buffer and stored at -70C. Each strain of FMD virus was serially diluted to -5dilution in order to check the efficacy of test. Reaction was completed in 1 hr and 10 min at temperatures 600C. It was also tested in hot water bath on same temperature. Results were same in both techniques. Results showed that LAMP was more sensitive for FMD strain O (positive reaction upto -4 dilution) than others strains A and Asia -1 (positive reaction upto -3 dilutions). Further studies will be conducted to develop LAMP PCR as field test that does not require RNA extraction.
 

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