J Vet Sci.  2014 Sep;15(3):423-426. 10.4142/jvs.2014.15.3.423.

A reverse transcription loop-mediated isothermal amplification assay to rapidly diagnose foot-and-mouth disease virus C

• Ding YZ
• Zhou JH
• Ma LN
• Qi YN
• Wei G
• Zhang J
• Zhang YG

• KMID: 2155626
• DOI: http://doi.org/10.4142/jvs.2014.15.3.423

Abstract

A reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed to rapidly detect foot-and-mouth disease virus serotype C (FMDV C). By testing 10-fold serial dilutions of FMDV C samples, sensitivity of the FMDV C RT-LAMP was found to be 10 times higher than that of conventional reverse transcription-PCR (RT-PCR). No cross-reactivity with A, Asia 1, or O FMDV or swine vesicular disease virus (SVDV) indicated that FMDV C RT-LAMP may be an exciting novel method for detecting FMDV C.

Keyword

foot-and-mouth disease; foot-and-mouth disease virus serotype C; reverse transcription loop-mediated isothermal amplification; reverse transcription-PCR, sensitivity

MeSH Terms

Animals
Foot-and-Mouth Disease/*diagnosis
Foot-and-Mouth Disease Virus/genetics
Nucleic Acid Amplification Techniques/*methods/veterinary
Reverse Transcriptase Polymerase Chain Reaction/veterinary
Reverse Transcription/genetics
Sensitivity and Specificity

Figure

• 

  Fig. 1 Comparing the sensitivity of RT-LAMP and RT-PCR for the detection of C-type FMDV using agarose gel electrophoresis. (A) Lane M1, DNA marker DL-100 (1500, 1000, 900, 800, 700, 600, 500, 400, 300, 200 and 100 bp; Takara Bio,Japan). Lanes 1~7, different dilutions (3.25 × 105 ng/mL to 3.25 × 10-1 ng/mL) of FMDV C subjected to the RT-LAMP assay. (B) Lane M2, DNA marker DL-2000 (2000, 1000, 750, 500, 250 and 100 bp; Takara Bio). Lanes 1~5, different dilutions (3.25 × 104 ng/mL to 3.25 × 100 ng/mL) of FMDV C subjected to RT-PCR. RT-PCR products corresponded to specific amplification of the FMDV C VP1 gene with a detection limit of 3.25 × 100 ng/mL. In contrast, the detection limit for RT-LAMP was 3.25 × 10-1 ng/mL.

• 

  Fig. 2 Analysis of RT-LAMP specificity for FMDV C. The ability of the assay to distinguish FMDV C from FMDV A, O, and Asia 1 as well as SVDV was evaluated. Lane M, DNA marker DL-100 (1500, 1000, 900, 800, 700, 600, 500, 400, 300, 200 and 100 bp; Takara Bio); Lane 1, FMDV C; Lane 2, FMDV A; Lane 3, FMDV Asia 1; Lane 4, FMDV O; and Lane 5, SVDV.

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