HIV prevalence and knowledge concerning HIV prevention among secondary school students in Tanzania was investigated. Approximately 50% of all secondary school students in Hai district and Moshi town were included in the study. Saliva samples were obtained using the OraQuick® rapid HIV-1/2 antibody assay. Forty-one (1.0%) and 211 (5.5%) students at the rural and urban schools, respectively, tested positive for HIV antibodies in saliva. HIV knowledge and beliefs varied significantly. Noninvasive saliva sample collection for HIV testing was highly acceptable. HIV infection is considerably more widespread among students attending urban rather than rural schools in the population investigated.