H&E staining and EcPV2 RISH signal distribution in penile lesions. One representative example per lesion (perilesional epithelium #14a, hyperplasia #14, papilloma #12, CIS #6 and SCC #5) is shown. a and b: H&E stained sections photographed using 2x (a) and 20x objectives (b). c and d: RISH stained sections photographed using 2x (c) and 40x (d) objectives. White squares in panel a and c mark the enlarged area shown in panel b and d. Perilesional skin: Squamous epithelium with normal maturation (a and b) and numerous pigmented cells in the basal layer (b). No viral signal is detectable in perilesional skin (c and d). Benign hyperplasia: Thickening of the epithelium and formation of broad rete ridges (a). There is orderly maturation and presence of koilocyte-like cells (white arrows) (b). Strong RISH signal is detectable, consisting of diffuse nuclear staining (DNS) (black arrows), which is detectable also at low magnification (c and d). The nuclear and cytoplasmic granular staining (GS) (small red dots in almost all lesional cells) is just visible at high magnification (d). Papilloma: Finger-like projection of moderately hyperplastic, mildly hyperkeratotic stratified squamous epithelium with normal differentiation and thin central cores of connective tissue (a and b). EcPV2 RISH staining in this case included mild DNS (c) and strong GS (d). CIS: Transepithelial mildly disrupted epithelial maturation can be seen (a and b). The keratinocytes show increased variability of keratinocytes and mitotic figures in the suprabasal region can be noted (white arrows b). There is intra-epithelial keratin pearl formation (white arrow a). EcPV2 RISH GS is just detectable at 40x magnification and no DNS is present (c and d). SCC: Infiltrating islands and trabeculae of moderately differentiated squamous epithelium with keratin pearl formation, (white arrows a). The keratinocytes are highly variable, show numerous mitotic figures (white arrows) and some cells are keratinized (black arrow) (b). The EcPV2 RISH DNS is rarely detectable within the infiltrating islands (c), while strong GS can be noticed (d)