FIG 1 .
Plaque morphologies in HSV-1 stocks, before and after plaque purification. Lab-passaged stocks of classical HSV-1 KOS and F strains contain multiple plaque morphologies. (A) With multiple rounds of limiting dilution, distinct plaque morphologies can be separated into populations that breed true and exhibit greatly reduced diversity (see Table 1 for details). (B) HSV-1 KOS can be separated into large, syncytial, and hypersyncytial variants. (C) HSV-1 F can be separated into large, syncytial, and small variants. (D) We used a previously described BAC-cloned HSV-1 F genome that has been modified to encode an mRFP fusion to the viral capsid (38). Recovery of this cloned genome into mammalian cells regenerates the plaque-morphology diversity observed in HSV-1 F stocks. Phase and fluorescence images reveal that in addition to diversity in plaque morphology, fluorescently tagged viral stocks exhibit diversity in fluorescence (additional images can be found in Fig. S1 in the supplemental material). (E) The low-passage-number clinical isolates HSV-1 H166 and H166Syncytial are distinct strains isolated from the cerebrospinal fluid of the same patient during an episode of viral meningitis. The size and appearance of cytopathic and syncytial plaques mirror those seen in the lab-isolated variants of HSV-1 KOS and F. Viruses were plated at limiting dilutions on monolayers of Vero cells and then fixed and stained with methylene blue at 72 hpi. Images were exported from Nikon NIS-Elements software. Contrast was inverted using Adobe Photoshop to show plaques more clearly. Bars: 1 mm (A), 5 mm (B, C, and E), and 2 mm (D). Further quantifications of plaque size distribution and frequency are found in Fig. 2 and Tables 1 and 2.