It is an enveloped, single-stranded RNA virus with a segmented genome of negative and ambisense polarity. RVFV belongs to the Phlebovirus genus from the Phenuiviridae family, Order Bunyavirales. It has been proposed that the virus is able to resist desiccation for long periods within mosquito eggs, which hatch upon flooding starting a new infectious cycle. In Africa, the re-emergence of Rift Valley fever (RVF) is usually associated to heavy rainfall seasons favoring overgrowth of mosquito species capable to transmit the virus trans-ovarially. Rift Valley fever virus (RVFV) is a mosquito-borne pathogen causing severe disease outbreaks affecting humans and livestock in sub-Saharan Africa, Egypt, Yemen, Saudi Arabia and the Indian Ocean islands. Mouse anti-VP2 polyclonal serum was used for VP2 detection and an anti-V5 mAb was used for the detection of NS1Nt (green fluorescence). Detection of VP2 and NS1 Nt transgenes by immunofluorescence of Vero cells infected with serially passaged recombinant virus (p2 to p5). Bars represent the S-segment/VP2 ratio from densitometry values of gel-based DNA fragments of 521bp and 653bp corresponding to the N and VP2 genes respectively. Relative detection of RVFV nucleoprotein N (red) and BTV VP2 (green) genes from ZH548-ΔNSs::VP2 BTV-4 by PCR. Bars represent the L/NS1 ratio from Ct values (right Y axis). Relative detection of L segment RNA (red line) and NS1Nt gene (green) from ZH548-ΔNSs::NS1Nt BTV-4 by RT-qPCR (left Y axis). Vero cell monolayers were infected with of a MOI of 1 with each virus from a first (P1), second (P2), third (P3), or fourth (P4) passage. Schematic of the passage history of the recombinant viruses. S2 Fig: Analysis of the genomic stability of the recombinant viruses.
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