Background; Effective antiretroviral therapy (ART) is crucial towards the success of both treatment and prevention of HIV-1 infection. Development of drug resistance against potent and widely used drugs such as Tenofovir (TDF) poses a challenge towards achievement of the 2030 targets. Furthermore, East Africa and Tanzania specifically has diverse HIV-1 subtypes, which might affects response to treatment and selection of particular mutations that will compromise the subsequent treatment regimens. For many years, a clinicoimmunological criterion has been used to switch ART regimens without viral load measurements resulting in delays in switching and/or unnecessary switching. Drug resistance testing is not done before initiation of ART or during switching therefore people may be kept on regimens that are not effective resulting in treatment failure. Objective; To determine the prevalence of antiretroviral drug resistance mutations and molecular diversity of HIV-1 among adults on 1st line ART containing TDF with virological failure at KCMC Moshi, Tanzania. Methods A well-structured questionnaire will be administered to the study participants who gave verbal informed consent by the nurse/doctor; physical examination and blood collection will be done. Other information which will be collected includes age, gender, date of HIV infection diagnosis, baseline CD4 cells count, the current viral loads (not more than six months old), date started ARV, 1st line ART regimes, previous ARV exposure (PMTCT, PEP), adherence to ARV, co-morbid conditions, blood pressure, weight, height, and pregnancy status. Absolute CD4+ count (cells/μl) will be determined using xii FACSCountTM SW Version 1.4 BD, control reagents will be run for verification of the test. Viral load will be determined using NucliSENS EasyQ HIV-1 V 2.0 according to manufacturer guidelines, negative controls will be run concurrently. Genetic sequencing will be done using a 16 capillaries 3130 XL ABI Prism Genetic Analyzer sequencer (Applied Biosystems, Foster City, Canada.