Using molecular methods three or five major variants of HTLV-I have been identified; moreover two subtypes of HTLV-II defined as HTLV-IIa and HTLV-IIb with six variants within each of these groups have been described. In the present study we analysed proviral DNA obtained from the peripheral blood mononuclear cells (PBMCs) of a significant group of Italian intravenous drug users (IVDUs), prison inmates and blood donors (BDs) who were HTLV antibody positive. Restriction fragment length polymorphism (RFLP) of amplified LTR region with ApaI, NdeI, DraI, SacI and MaeIII endonucleases was used to define the HTLV-I subtypes, while the different variants of HTLV-IIa and -IIb were defined by RFLP of the LTR region with the AvaII, BglI, SauI, XhoI and BanII endonucleases. The four HTLV-I isolated from BDs were characterized as C type. All the 11 HTLV-II detected in the IVDUs were HTLV-IIb4, while among the prisoners one HTLV-IIb5 and five HTLV-IIb4 were found. Interestingly, in the BDs group two HTLV-IIa0 and one HTLV-IIb4 were detected. It should also be noted that 82% of the IVDUs and 50% of the prisoners were coinfected with HIV, while all the BDs were HIV negative. These data indicate that HTLV-IIb4 is the predominant genotype in Italian IVDUs and prisoners, while the significant variability observed in the BD HTLV-II isolates could be due to the different source of infection among this group.