This study aimed to document the vitamin D status of HIV-infected individuals across a wide latitude range in one country and to examine associated risk factors for low vitamin D. Using data from patients attending four HIV specialist clinics across a wide latitude range in Australia, we constructed logistic regression models to investigate risk factors associated with 25(OH)D <75 nmol L-1. 1788 patients were included; 87% were male, 76% Caucasian and 72% on antiretroviral therapy. The proportion with 25(OH)D <50 nmol L-1 was 27%, and -1 was 54%. Living in Melbourne compared with Cairns (adjusted odds ratio (aOR) 3.30; 95% CI 2.18, 4.99, P <0.001) and non-Caucasian origin (aOR 2.82, 95% CI 2.12, 3.75, P <0.001) was associated with an increased risk, while extreme UV index compared with low UV index was associated with a reduced risk (aOR 0.33; 95% CI 0.20, 0.55, P <0.001) of 25(OH)D <75 nmol L-1. In those with biochemistry available (n = 1117), antiretroviral therapy was associated with 25(OH)D <75 nmol L-1; however, this association was modified by serum cholesterol status. Location and UV index were the strongest factors associated with 25(OH)D <75 nmol L-1. Cholesterol, the product of an alternative steroid pathway with a common precursor steroid, modified the effect of antiretroviral therapy on serum 25(OH)D.