This study focused on Nucleoside Reverse Transcriptase drug-resistance profiling and the susceptibility patterns for the plasma samples obtained from HIV-positive naïve patients enrolled at Machakos Level 5 Hospital. The research's specific objectives were to profile resistance to Nucleoside Reverse Transcriptase Inhibitor drugs and then identify the markers for resistance to Nucleoside Reverse Transcriptase Inhibitor. This study used an experimental research design; DNA was extracted from the plasma samples, and PCR was amplified using polymerase-gene specific primers and later Gel electrophoresis. Then finally, cycle sequencing of the polymerase (pol) gen. The amplified products were sequenced, and drug-resistant mutations were determined using Los Alamos HIV DR database. All amplified samples from the PCR had the gel cut/excised and cleaned using the QIA quick gel extraction kit protocol. Sequences with high relatedness were fetched in a FASTA format and aligned using the Mega Evolutionary Genetic Analysis (MEGA) software version 10 using the Neighbour Joining (NJ) algorithm and the 1000 Bootstrap resampling algorithm. The main HIV strain detected in this study was the HIV A1 subtype, the major sub- subtype in Kenya. No other subtypes were noted in the study. Regarding NRTIs, the major mutation noted was D67E which indicated inadequate level, zidovudine resistance, and drug susceptibility to abacavir, emtricitabine, lamivudine, and tenofovir noted with no resistance to NNRTIs. However, there were minor mutations noted. Drug resistance mutations were found in high numbers associated with viral load and treatment time. Importantly, patients with triple and dual-class drug resistance should immediately alter ART regimens to alter the possibility of transmitting multi-drug-resistant HIV-1 strains. This finding emphasizes the importance of targeted resistance monitoring as a tool for addressing the problem.