We used the findings from two, cross-sectional studies of HIV serostatus and risk behaviours to assess the effects of knowledge of HIV serostatus and risk behaviours (relating to sex and injection drug use) among injecting drug users (IDUs). Respondent-driven sampling was used simultaneously at two sites in Estonia (the capital Tallinn, and the second-largest city of Ida-Virumaa County, Kohtla-Jarve). The research tool was an interviewer-administered survey. Biological samples were collected for HIV testing. Participants were categorised into three groups based on HIV testing results and self-report on HIV serostatus: HIV-negative (n=133); HIV-positive unaware of their serostatus (n=75); and HIV-positive aware of their serostatus (n=168). In total, 65% of the participants tested positive for HIV. Of those 69% were aware of their positive serostatus. HIV-positive IDUs aware of their serostatus exhibited more risk behaviours than their HIV-positive counterparts unaware of their serostatus or HIV-negative IDUs. Effective prevention of HIV among IDUs should therefore, include programmes to reduce high-risk sexual and drug use behaviours at the public health scale and enhanced prevention efforts focusing on HIV-infected individuals.