Objective: To evaluate if levels of pregnancy-associated plasma protein-A (PAPP-A), free β-hCG and nuchal translucency (NT) used in first trimester aneuploidy screening and α-fetoprotein (AFP), unconjugated oestriol (UE3) and free β-hCG in the second trimester are altered in pregnancies with human immunodeficiency virus (HIV) infection. Methods: Median MoM values of biochemical markers and δ-NT in 92 women with HIV infection in the first trimester were compared with those from 912 HIV-seronegative controls. In the second trimester, biochemical marker levels were compared in 52 women with HIV infection and 378 HIV-seronegative controls. Results: First trimester free β-hCG median MoM levels in HIV-infected pregnancies were not different to controls (0.978 vs. 0.981, p = 1.000), as were PAPP-A levels (1.190 vs. 1.102, p = 0.099) and δ-NT (0.1374 vs. 0.0445, p = 0.0631). Second trimester levels of free β-hCG were not significantly different (1.0575 vs. 0.9619, p = 0.1827), as was AFP (0.9734 vs. 0.9350, p = 0.6576), although UE3 was significantly lower (0.970 vs. 1.110, p = 0.0005). Conclusions: In the first trimester, marker levels are not affected by the presence of HIV infection, and risks for aneuploidy are likely to be accurate in this group. Further studies are required to evaluate if UE3 levels continue to be low in HIV-infected women since this may have an impact on screening in the second trimester.