The epidemiologic determinants of seroreactivity to human papillomavirus (HPV) type 16 L1/L2 virus-like particles (VLPs) were assessed separately in HPV-16 DNA-positive and -negative women participating in a nested case-control study of incident cervical neoplasia. Seventy-four women with cervical HPV-16 DNA and 656 cytologically normal HPV-16 DNA-negative subjects were interviewed and tested at two time points for viral DNA and once (at the later time) for VLP seroreactivity. Among subjects who were currently HPV-16 DNA-negative, seroreactivity odds ratios increased from 2.9 for 2-5 male sex partners (vs. 0 or 1) to 5.4 for 6-9 partners and 14.0 for > or = 10. Thus, prior cervical infection may be a major determinant of seroreactivity in HPV-16 DNA-negative women. This trend was not observed in HPV-16 DNA-positive subjects. Seroreactivity was independently associated with oral contraceptive use, particularly in HPV-16 DNA-negative subjects with use for > or = 10 years. Consequently, a possible role for virus-steroid hormone interactions in seroconversion is suggested.