Figure 2. Cerebrospinal fluid sample of a 39-year-old man in Montpellier, France, infected with USUV who had an atypical neurologic presentation. The sample was amplified for 6 days on C6/36 cells, and the supernatant was used to infect Vero cells or primary human astrocytes. A) Cytopathic effect (presence of adherent dead cells and absence of heaps; all dead cells were scattered) was observed at day 5 postinfection of a Vero cell culture. Scale bar indicates 100 μm. B) Mock or infected primary human astrocytes were fixed at day 4 postinfection and labeled with pan-flavivirus antibody (MAB10216, clone D1–4G2) by indirect immunofluorescence (green). Strong labeling was observed in some cells (arrows). Nuclei are labeled with DAPI (4′,6-diamidino-2-phenylindole) (blue). NI, not infected; USUV, Usutu virus. Scale bar indicates 10 μm.