Purpose: To develop a rapid detection method for Rift Valley fever virus (RVFV) diagnosis.
Methods: According to the reference sequences of RVFV published in GenBank, nine overlapping
polymerase chain reaction (PCR) primers and four specific reverse transcription loop-mediated
isothermal amplification (RT-LAMP) primers were designed using DNAStar and LAMP primer design
software, respectively. Based on the synthesis of a conserved part of the RVFV S segment gene
sequence using overlapping PCR, RT-LAMP assay was first established and evaluated after a series of
tests, including, optimization of reaction conditions, and sensitivity and specificity tests.
Result: A target RVFV S segment gene fragment of 288 bp was synthesised. The optimal reaction
conditions for RT-LAMP assay were 63 °C for 45 min: the assay has a specific ladder-like pattern of
amplification bands from about 120 bp. The lowest target gene copy number of RT-LAMP for RVFV
detection was 70 copies. The assay showed good specificity as only the synthesised target RVFV gene
was amplified with no amplification for the detection of Peste des petits ruminants virus, Epidemic
encephalitis B virus,
E. coli
,
Pasteurella multocida
, or
Salmonella
.
Conclusion: This study provides a rapid, sensitive, specific RT-LAMP method for RVFV detection.
