The surviving hamsters from this experiment were re-challenged with PICV to assess the ability of DEF201 to enhance longer-term protection via acquired immunity. With the exception of 4 animals in the 24 h DEF201 pretreatment group, and a single animal in the rAd EV 48 h group, all animals that were challenged with PICV on day 0 of the experiment survived a second challenge Figure 3 . DEF201 extended pre-exposure prophylaxis protects hamsters from lethal PICV challenge. Animals were treated i.n. with a single dose 10 8 pfu of DEF201, the rAd EV control virus, or PBS placebo 7 or 14 days prior to PICV infection. Animal weights were measured two weeks prior to, and at the time of, PICV challenge. The effect of 7-day and 14-day pretreatments on A) weight change over the two-week period prior to PICV challenge and the extended PICV prophylaxis efficacy data for the B) 7-day and C) 14-day pretreatments are shown. ***P,0.001 compared to respective placebo-treated animals. b P,0.01, c P,0.001 compared to respective rAd EV-treated animals. doi:10.1371/journal.pone.0026072.g003 on day 28 ( Figure 5 ). All six naïve animals that were initially shaminfected succumbed as expected.