Slovenia is a small European country with a total of 547 HIV-infected individuals cumulatively reported by the end of 2011. However, the estimated incidence rate of HIV infections increased from 7.0 per million in 2003 to 26.8 per million in 2011. In this study, we assessed the prevalence of transmitted drug resistance (TDR) in the past 6 years (2005-2010) and analyzed the time trend of the proportion of men having sex with men (MSM) and HIV-1 subtype B among newly diagnosed individuals in a 15-year period (1996-2010) in Slovenia. Among 150 patients included in the study, representing 63% of HIV-1 newly diagnosed patients in 2005-2010, TDR was found in seven patients (4.7%). The prevalence of TDR to nucleoside reverse transcriptase inhibitors, nonnucleoside reverse transcriptase inhibitors, and protease inhibitors was 2% (3/150), 2% (3/150), and 0.7% (1/150), respectively. The majority of patients were infected with subtype B (134/150, 89%), while subtype A was detected in 6.0% (9/150), subtype D in 1.3% (2/150), and subtype G and CRF02_AG in 0.7% (one patient each). Three of 150 sequences could not be typed. Infection with subtype B was found to be significantly associated with male gender, Slovenia being reported as the country of the patient's nationality and origin of the virus, CDC class A, mode of transmission with homosexual/bisexual contact, sex with an anonymous person, and a higher CD4(+) count. Among patients carrying the subtype B virus, an MSM transmission route was reported in 87% of patients. Although the prevalence of TDR in Slovenia is still below the European average, active surveillance should be continued, especially among MSM, the most vulnerable population for HIV-1 infection in this part of Europe.