Human papillomaviruses from the genus beta (betaPV) are a possible cause of cutaneous squamous cell carcinoma (SCC). We compared the betaPV infections in SCC and in sets of cutaneous tissues collected from a series of individual SCC patients to determine concordance and to assess the adequacy of eyebrow hairs as noninvasive markers of betaPV infection. Biopsies of SCC tumors, perilesional tissue, normal skin from the mirror image of nonfacial SCC and plucked eyebrow hairs were collected from 21 patients with incident SCC living in Queensland, Australia. These were tested for the presence of DNA from 25 different betaPV types. Overall prevalence of betaPV was high in every sample type, ranging from 81% to 95%. The median number of types was significantly higher in the SCC tumour (6), perilesional skin (5) and eyebrow hairs (5) than in normal skin (2). Comparing SCC tissue with other sample types within patients showed 63 overlapping infections with eyebrow hairs (71%; 95% CI: 60-80); 56 with perilesional skin samples (63%; 95% CI: 52-73) and 23 with normal skin samples (26%; 95% CI: 17-36). The sensitivity of eyebrow hair testing for detection of betaPV in the tumor was 82% (95% CI: 57-96) with concordance defined as 50% of betaPV types in common and 29% (95% CI: 10-56) for 100% concordance. These findings support the concept that perilesional skin represents an area of field change involving betaPV preceding SCC development and indicate that eyebrow hairs can serve to some degree as an easily collected marker of tumor betaPV status in epidemiological studies.