The human immunodeficiency virus (HIV) is associated with a significant burden of disease, including medical and non-medical costs. Therefore, it is considered to be a priority for all health authorities. The aim of this study is to determine healthcare and treatment costs of caring for PLHIV at one of the tertiary care university hospitals in Riyadh, Saudi Arabia. This was a micro-costing, retrospective, observational study from a tertiary care university hospital and included all confirmed HIV-infected patients who visited infectious disease clinics in the period from 1 January 2015 to 31 December 2018. A total of 42 PLHIV were included in this study. The mean age of the study participants was 38.76 ± 11.47 years with a mean disease duration of 5.27 ± 4.81 years. The majority of patients were male (85.7%) and Saudi (88.1%). More than half of included patients (59.5%) had a CD4 count of more than 500. During the study period, 26 patients (61.9%) were initiated on a single-tablet regimen. Overall, the main cost-driver was antiretroviral medications, which cumulatively represented more than 64% of the total cost. Patients who developed opportunistic infections had a statistically significant (p = 0.033) higher financial impact, both as a total and on a patient level, than those presented without opportunistic infections. On a patient level, the mean and median costs were higher and statistically significant for those with co-morbidities than those without co-morbidities (p = 0.002). The majority of the economic burden of PLHIV is attributable to antiretroviral therapy use. The healthcare costs of PLHIV can vary greatly, depending on the presenting illness, clinical stage, developed opportunistic infection, co-morbidity, and pharmacological therapy.