Background: It is important to identify and manage determinants of virological failure among HIV infected individuals on treatment for achieving viral suppression. This study aimed to identify proportion and factors associated with virological failure among HIV infected individuals receiving first line antiretroviral therapy (ART).Methods: A total of 2670 adult HIV infected individuals attending ART centre at ICMR-National AIDS Research Institute, between January 2005 and June 2019 and having their recent viral load done after implementation of guidelines on routine viral load testing were included. Data were reviewed and analysed.Results: Of the 2670 people living with HIV (PLHIV) on first line antiretroviral therapy, 48% were male and 69% were more than 40 years of age. Mean baseline CD4 count at ART initiation was 252 cells/mm3 (SD:210, IQR 116-313) Overall, 13% (340/2670) of the participants showed virological failure. In multivariate analyses, participants with younger age and males retained significant association. Those with baseline CD4 counts of less than or equal to 500 cells/mm3 at treatment initiation (adjusted OR 1.71; 95% CI 1.08-2.70; p=0.022) and ART adherence ≤95% within last three months of recent viral load determination (adjusted OR 1.55, 95% CI of AOR 1.04-2.32; p=0.031) had higher risk for virological failure as compared to others. PLHIV with ART substitution due to various reasons were almost twice as likely to have virological failure (adjusted OR 1.83, 95% CI 1.44-2.33; p<0.001).Conclusions: It is crucial to focus on factors leading to virological failure among HIV infected individuals attending ART centre. Early linkage to treatment and ART initiation along with adherence counselling at every follow up visit play an important role in mitigating virological failure.