BACKGROUND Human monkeypox, an emerging viral zoonosis first recognized in Africa, has recently emerged in the mid-western US. Initially, it presents with skin eruptions and fevers with diaphoresis and rigors. Clinically, the skin lesions progress from papules to vesiculopustules to resolving eschars. METHODS Three cutaneous biopsy specimens from two patients with polymerase chain reaction (PCR)-proven monkeypox were available for review. The histologic, immunohistochemical and electron-microscopic features were identified. RESULTS The clinical progression of lesions is mirrored histologically with ballooning degeneration of basal keratinocytes and spongiosis of a mildly acanthotic epidermis progressing to full thickness necrosis of a markedly acanthotic epidermis containing few viable keratinocytes. A lichenoid-mixed inflammatory cell infiltrate is present, which exhibits progressive exocytosis with the keratinocyte necrosis. Inflammation of the superficial and deep vascular plexes, eccrine units and follicles is also present. Viral cytopathic effect is manifest by multinucleated syncytial keratinocytes. Immunohistochemically, viral antigen is detected within keratinocytes of the lesional epidermis, follicular and eccrine epithelium and few dermal mononuclear cells. Electron microscopy reveals virions at various stages of assembly within the keratinocyte cytoplasm. CONCLUSIONS The histologic differential diagnosis includes herpes simplex virus, varicella and other pox viruses, such as smallpox. The first one may be differentiated histologically, immunohistochemically and electron microscopically. The last two may be differentiated using PCR assay for the monkeypox extracellular-envelope virus protein gene.