To visualize the spread of the virus, Ifnar1 2/2 mice were infected i.p. with 10 4 PFU of rZHDNSs-hRLuc. At 8, 16 and 34 h postinfection, h-coelenterazine was injected i.p. This route of hcoelenterazine administration was preferred to tail-vein injection due to slower kinetics of light production, as previously reported [39] . Mice were observed with real-time in vivo imaging 15 and 20 min after the injection of h-coelenterazine for the whole body and the thorax, respectively. At 8 h post-infection, luminescence was readily detected. Short integration periods (15 s) were sufficient to acquire a significant signal. We observed strong signals between the forelegs in the thoracic cavity, and below the xiphoid cartilage in the abdominal cavity, respectively (Figure 2A) . Imaging of the left profile showed an additional signal in the spleen ( Figure 2B ). In some experiments, animals were euthanized for ex vivo imaging, the organs of the thorax and abdomen were harvested, and the individual organs were imaged ( Figure 2D ). In the thorax, the greatest signal originated from the thymus, whereas the signal from the lungs was only slightly above background. In the abdomen, the pancreas was the most luminescent organ. The spleen and the liver also emitted significant luminescence. On average, a ten-fold higher luminescence signal was observed in the harvested pancreas compared to the liver. This suggests that the liver, the critical target organ of the disease, was not among the main replication sites for RVFV at this early stage of infection. At 16 h post-infection, the whole body luminescence was higher than at 8 h post-infection (Figure 2A) . The signal spread out the abdominal cavity. The high intensity of luminescence in the abdominal cavity precluded detection in other locations unless integration was limited to the thorax ( Figure 2C ). Dissection and ex vivo imaging showed a gradual increase of luminescence in the pancreas and in the liver. Additional sources of luminescence were the intestine mesentery, kidneys, ovaries and uterus in females, the seminal vesicles, preputial glands, epididymis and testis in males ( Figure 2D , and data not shown). The intensity of these signals was quite similar to that measured in the spleen and in the liver (data not shown). At 34 h post-infection, the intensity of the signal led to saturation of the camera using a 0.5 s integration period, thus preventing identification of individual organs (Figure 2A) . Ex vivo imaging revealed that the highest signal was in the liver. Other organs with intense luminescence were the spleen, intestine mesentery and pancreas (data not shown). Quantification of the luciferase expression in living mice during the time course of infection showed that the luminescence signal originated from the thymus remained constant from 8 h postinfection onwards, whereas luminescence profiles were increased in the liver and pancreas, suggesting a progressive increase of viral replication in these organs (Figure 3 ).