Group G-D was inoculated with 107 C6/36 derived RVFV. Group G-E was inoculated with 105 PFU of Vero E6 RVFV stock, and re-inoculated IV with the same inoculum at 1 dpi. Group G-F was inoculated with 105 PFU of C6/36 derived RVFV, and re-inoculated

IV with the same inoculum at 1 dpi. Group G-G was inoculated with 107 C6/36 derived RVFV and re-inoculated SC with the same inoculum at 1 dpi. All goats were kept for four weeks following the inoculation to monitor an antibody development. Serum samples collected at 0, 4, 5, 6, 7, 14, 21 and 28–30 dpi were analyzed for presence of neutralizing antibodies. Differences in susceptibility to RVFV infections were observed between sheep and goats, and also between breeds of sheep. In the first study,

conducted BKM120 in Suffolk-cross sheep, all animals developed viremia at 3 dpi, both by virus isolation and RNA detection when inoculated with 105 PFU of virus produced in Vero cells. However, when the Rideau Arcott cross lambs were inoculated via the same route and the same inoculum, only three out of four animals had detectable RVFV RNA in their blood and only two developed viremia (Fig. 1). Subsequently different inoculation approaches were tested to obtain a more reliable viremia model. Genomic sequences of the inocula were verified prior to the start of the animal inoculations. Concurrently with the infection experiments, characterization on protein level of RVFV 17-AAG generated in Vero E6 cells or the C6/36 was taking place. There was no difference in genome of RVFV generated in Vero E6 cells

compared to virus generated in C6/36 cells, including the stock viruses used in experimental PDK4 inoculations, and the sequences corresponded with sequences published for RVFV ZH501 in Gen Bank. Both viruses had functional NSm and NSs coding genes, as immunoblots of infected cell lysates indicated that all proteins from the M and S segments were expressed. The viruses however differed in protein composition of virions, with the mosquito-cell generated RVFV having an additional large glycoprotein (78 kDa) incorporated into virions [23]. Subcutanous inoculation was used in all primary inoculation. Two doses (105 or 107 PFU/animal) and two different inocula (prepared either in Vero E6 or in C6/36 cells) were tested. The titer of inoculum was confirmed by back-titration at the time of inoculation, and stayed within 0.5 log10 difference from the targeted dose. In specific groups, attempts were made to increase the viremia by re-inoculation, either by the subcutaneous or by the intravenous route at 1 dpi. A summary of the experimental groups is presented in Table 1. Using the same mode of inoculation as for the Suffolk breed (group S-A), the 105 PFU dose of Vero E6 produced RVFV in Rideau Arcott cross lambs (group S-B) lead to development of viremia only in three out of four animals at 2 dpi.