Background:  Primary ART resistance may hinder the efficacy of mother-to-child-transmission (MTCT) prophylaxis and limit future ART options for both mother and child. Allele-specific polymerase chain reaction (AS-PCR) could increase the sensitivity of detecting primary drug resistance in pregnant HIV-1-infected women relative to standard genotyping methods.

Methods:  We determined the prevalence of primary lamivudine (3TC) and nelfinavir (NFV) resistance in ART-naïve HIV-1-infected pregnant women enrolled in the Women and Infants Study Group (WITS) cohort by standard dideoxinucleotide chain termination sequencing (SQ) and AS-PCR. Blinded plasma specimens collected between 1998 and 2003 were analyzed in 2006. Nested PCR products including the protease (PR) and the first half of the reverse transcriptase (RT) -coding region of pol were analyzed by SQ. The presence of HIV-1 variants with the D30N and M184V mutations in pol was investigated in all specimens using AS-PCR, with a sensitivity cut-off of 0.1% and 0.4%, respectively. Phylogenetic analyses were performed to rule out specimen contamination.

Results:  Of the 61 women who fulfilled the inclusion criteria, the average age was 28.1 (range 17.9 to 42.7) years. The CDC AIDS category was A in 47 women, B in 13, and C in 1. Two women reported previous intravenous drug use. Median CD4⁺ count at the time of specimen collection was 446 (IQR 312 to 607) cells/mm³, median HIV-1 RNA levels were 11851 (IQR 1604 to 20,097) copies/mL. Resistance data were available from 45 women (74%). The prevalence of M184V mutation was 8.8% and 13.3% by SQ and AS-PCR, respectively. The prevalence of D30N mutation was 2.2% and 6.7% by SQ and AS-PCR, respectively. Whereas D30N was usually detected as a minor variant, M184V was more likely found as a predominant variant. The frequency of natural polymorphisms in the PR-coding region by SQ was:  L10V (11.1%), I13V (13.3%), K20R/M (2.2%), L33I/V (2.2%), M36I (15.6%), D60E (8.9%), I62V (15.6%), L63P (44.4%), A71V (6.7%), V77I (15.6%), and I93L (11.1%). Only L214F (91.1%) polymorphism was found in the RT-coding region.

Conclusions:  This study suggests a high prevalence of primary 3TC and NFV resistance in HIV-1-infected pregnant women in the United States. AS-PCR improves the sensitivity for M184V and D30N detection 1.5-fold and 3-fold, respectively, relative to SQ. These results support routine genotypic resistance testing before initiating MTCT prophylaxis in the United States.