Background: Persistent low-level viraemia (PLLV) is a risk factor for virologic failure among people receiving antiretroviral therapy (ART). Objectives: We assessed the prevalence and predictors of PLLV among individuals receiving Dolutegravir-based ART in southern Nigeria. Design: This retrospective cohort study used routine program data from electronic medical records of persons receiving Dolutegravir-based first-line ART in 154 PEPFAR/USAID-supported health facilities in Akwa Ibom and Cross Rivers states, Nigeria. Methods: Clients on first-line Dolutegravir-based ART ⩾6 months, who had a viral load result in the 12 months preceding October 2021 (baseline), and a second viral load result by September 2022 were included. Persons with low-level viraemia (LLV) (viral load 51–999 copies/ml) received additional adherence support. The outcome analysed was PLLV (two consecutive LLV results). Indices were summarized using descriptive statistics, and predictors of PLLV were determined using multivariate logistic regression. Results: In total, 141,208 persons on ART were included, of which 63.3% ( n = 89,944) were females. The median age was 36 [29–44] years, median ART duration was 19 [11–42] months. At the end of the study, 10.5% (14,759/141,208) had initial LLV, 90.1% (13,304/14,759) of which attained undetectable viral load (⩽50 copies/ml), and 1.1% (163/14,759) transitioned to virologic failure (⩾1000 copies/ml) by the end of the study. PLLV prevalence was 0.9% (1292/141,208). Increasing ART duration [adjusted odds ratio (aOR) = 1.0; 95% confidence interval (CI): 1.005–1.008; p < 0.001] and viral suppression (<1000 copies/ml) before initial LLV (aOR = 1.7; 95% CI: 1.50–2.00; p < 0.001) were positively associated with PLLV, while receipt of tuberculosis preventive therapy reduced the likelihood of PLLV (aOR = 0.3; 95% CI: 0.10–0.94; p = 0.039). Conclusion: PLLV was uncommon among individuals receiving dolutegravir-based ART and was associated with longer ART duration, prior viral suppression, and non-receipt of tuberculosis preventive therapy. This strengthens recommendations for continuous adherence support and comprehensive health services with ART, to prevent treatment failure.