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BackgroundThe role of human papillomavirus (HPV) in keratoacanthoma (KA) remains unclear.ObjectivesTo identify possible differences in HPV DNA, detected by next-generation sequencing (NGS), between KAs and cutaneous squamous cell carcinomas (cSCCs), which may suggest different pathogenesis.Materials & MethodsWe extracted DNA from formalin-fixed and paraffin-embedded (FFPE) tissue blocks from samples of 151 patients (105 with cSCCs and 46 with KAs). HPV DNA was detected using the NGS-based Ezplex® kit. HPV detection rates and other clinical characteristics were compared.ResultsHPV was detected in 6.7% (7/105) of cSCC and 10.9% (5/46) of KA samples. Eight alpha-HPV genotypes (16, 57, 81, 31, 33, 45, 53, and 58) were detected, with HPV 16 being the most common. Only one type (57) is commonly classified as cutaneous type, and the rest are all mucosal types. HPV detection rate did not significantly differ between the KA and cSCC groups.ConclusionHPV detection was relatively low in KA and cSCC samples. HPV might be related to the pathogenesis of only selected KA and cSCC cases.