Objective: To describe the prevalence of the L76V protease inhibitors resistance-associated mutation (PI-RAM) in relation with patients’ characteristics and protease genotypic background in HIV-1 B- and “non-B”-infected patients. Methods: Frequency of the L76V mutation between 1998 and 2010 was surveyed in the laboratory database of 3 clinical centers. Major PI-RAMs were identified according to the IAS-USA list. Fisher’s and Wilcoxon tests were used to compare variables. Results: Among the overall 29,643 sequences analyzed, the prevalence of L76V was 1.50%, while was 5.42% in PI-resistant viruses. Since 2008 the prevalence of L76V was higher in “non-B”-infected than in B-infected patients each year. Median time since diagnosis of HIV-1 infection and median time under antiretroviral-based regimen were both shorter in “non-B”- than in B-infected patients (8 vs 11 years, P<0.0001; and 7 vs 8 years, P = 0.004). In addition, “non-B”-infected patients had been pre-exposed to a lower number of PI (2 vs 3, P = 0.016). The L76V was also associated with a lower number of major PI-RAMs in “non-B” vs B samples (3 vs 4, P = 0.0001), and thus it was more frequent found as single major PI-RAM in “non-B” vs B subtype (10% vs 2%, P = 0.014). Conclusions: We showed an impact of viral subtype on the selection of the L76V major PI-RAM with a higher prevalence in “non-B” subtypes observed since 2008. In addition, in “non-B”-infected patients this mutation appeared more rapidly and was associated with less PI-RAM.