CCR5 is a G-protein-coupled chemokine receptor that is used as a co-factor by macrophage-tropic (M-tropic) isolates of human immunodeficiency virus-1 (HIV-1) to gain entry into host cells. A 32-bp deletion in the CCR5 gene (CCR5-Delta32) leads to the production of an altered gene product that prevents HIV-1 from entering the host cell. This study was carried out to determine prevalence of CCR5-Delta32 allele frequency in a large Pakistani population sample (n = 821) representing 10 ethnic groups. No individual was homozygous for the mutant allele and the frequency of the CCR5-Delta32 allele ranged from 0.62% to 3.57%. The CCR5-Delta32 allele frequency was generally lower in populations from southern Pakistan. The overall frequency of the CCR5-Delta32 allele in Pakistan was 2.31%, which is much lower than that found in European populations and similar to that in the Middle East. This is consistent with the historical records and genetic data that indicate a close genetic affinity among these populations. This study demonstrates that the Pakistani population is highly susceptible to M-tropic isolates of HIV-1 and public health measures need to be enforced with urgency if Pakistan is to avoid an HIV epidemic.