Contemporary ART as Dolutegravir (DTG) has significantly advanced antiretroviral therapy, but relatively few data are available on its impact on the emergence of HIV-1 drug resistance mutations (DRMs). Monitoring the emergence of INSTI-associated DRMs following the introduction of DTG in Suriname will provide general insight and guide national HIV treatment strategies. All people living with HIV (PLHIV) in Suriname, for whom an INSTI drug resistance test was requested between September 2019 and February 2024 ( = 20), were included. HIV-1 integrase gene sequences were determined using Sanger sequencing. INSTI-associated mutations were identified using the Stanford HIV Drug Resistance Database program. The majority of the participants (66.7%) harbored HIV-1 subtype B, and 33.3% were B-recombinant forms. In addition to the INSTI wildtype, a strain was revealed carrying E157EQ and one person harbored a highly INSTI-resistant strain (E138K, G140S, Q148H and N155H). The emergence of a highly INSTI-resistant HIV-1 strain in Suriname, with unusual mutations for ART-experienced PLHIV exposed to DTG as the only INSTI, accentuates the need for continuous monitoring of the emergence of INSTI drug resistance mutations, not only to enable timely interventions and optimized treatment outcomes for PLHIV, but also to steer the decision making for ART protocols, especially for second generation INSTIs.