Mononuclear cells play key roles in the pathogenic mechanisms leading to HIV-associated neurocognitive disorders (HANDs). We examined the association between HIV DNA within peripheral blood mononuclear cell (PBMC) subsets and HAND in Nigeria. PBMCs were collected at baseline from 36 antiretroviral naive participants. CD14+ cells and T&B lymphocyte fractions were isolated by, respectively, positive and negative magnetic bead separation. Total HIV DNA within CD14+ and T&B cells were separately quantified using real-time PCR assay targeting HIV LTR-gag and cell input numbers determined by CCR5 copies/sample. Utilizing demographically adjusted T scores obtained from a 7-domain neuropsychological test battery, cognitive status was determined by the global deficit score (GDS) approach, with a GDS of ≥0.5 indicating cognitive impairment. In a linear regression adjusting for plasma HIV RNA, CD4 and lymphocyte count, Beck's depression score, and years of education, there was 0.04 lower log HIV DNA copies within T&B lymphocytes per unit increase in global T score (p = 0.02). Adjusting for the same variables in a logistic regression, the odds of cognitive impairment were 6.2 times greater per log increase in HIV DNA within T&B lymphocytes (p = 0.048). The association between cognitive impairment and HIV DNA within CD14+ monocytes did not reach statistical significance. In this pretreatment cohort with mild cognitive dysfunction, we found a strong association between levels of HIV DNA within the lymphocyte subset and HAND independent of plasma HIV RNA. These findings likely reflect the neurologic impact of a larger HIV reservoir and active viral replication.