A Heparan-Dependent Herpesvirus Targets the Olfactory Neuroepithelium for Host Entry
Figure 2
MuHV-4 infection localizes to the olfactory neuroepithelium.
a. BALB/c mice were allowed to inhale a 5 µl droplet containing 106 p.f.u. eGFP+ MuHV-4 and 3 days later analysed by immunostaining with a MuHV-4-specific polyclonal rabbit serum (brown) and counterstained with Mayer's Hemalum. The multi-layered main olfactory neuroepithelium and single-layered respiratory epithelium are indicated, as is the neuroepithelium of the vomeronasal organ (VNO). Equivalent staining was seen in >10 mice. Images enlarged ×5 in the right-hand panels show the sharp divide between virus+ neuroepithelium and virus− respiratory epithelium; a representative area of (virus−) squamous epithelium; and background staining of sections from naive mice. b An adjacent section was stained for viral eGFP (brown), which is expressed independently of lytic cycle genes. Again the main olfactory neuroepithelium was virus+ and other sites virus−. c. BALB/c mice were infected with MuHV-4 (104 p.f.u. in 5 µl) and 3 days later analysed for viral tRNA/miRNA expression by in situ hybridization of nose sections with a digoxigenin-labelled riboprobe. The arrows show examples of positive neuroepithelial staining. The respiratory epithelium remained negative. Similar results were obtained in 3 mice. d. BALB/c mice were infected with MuHV-4 (106 p.f.u. in 5 µl) and 4 days later analysed either for viral tRNA/miRNA expression as in c, or for viral lytic antigen expression as in a. Arrows show examples of positive staining. e. BALB/c mice were allowed to inhale a 5 µl droplet containing 106 p.f.u. influenza A/PR/8/34. 1 day later they were analysed by immunostaining with a polyclonal influenza-specific rabbit serum that recognizes predominantly the viral hemagglutinin. Only the respiratory epithelium was virus+. 5 more mice gave similar results.