High Levels of Dual-Class Drug Resistance in HIV-Infected Children Failing First-Line Antiretroviral Therapy in Southern Ethiopia

Abstract

Clinical monitoring of pediatric HIV treatment remains a major challenge in settings where drug resistance genotyping is not routinely available. As a result, our understanding of drug resistance, and its impact on subsequent therapeutic regimens available in these settings, remains limited. We investigate the prevalence and correlates of HIV-1 drug resistance among 94 participants of the Ethiopia Pediatric HIV Cohort failing first-line combination antiretroviral therapy (cART) using dried blood spot-based genotyping. Overall, 81% (73/90) of successfully genotyped participants harbored resistance mutations, including 69% (62/90) who harbored resistance to both Nucleoside Reverse Transcriptase Inhibitors (NRTIs) and Non-nucleoside Reverse Transcriptase Inhibitors (NNRTIs). Strikingly, 42% of resistant participants harbored resistance to all four NRTIs recommended for second-line use in this setting, meaning that there are effectively no remaining cART options for these children. Longer cART duration and prior regimen changes were significantly associated with detection of drug resistance mutations. Replicate genotyping increased the breadth of drug resistance detected in 34% of cases, and thus is recommended for consideration when typing from blood spots. Implementation of timely drug resistance testing and access to newer antiretrovirals and drug classes are urgently needed to guide clinical decision-making and improve outcomes for HIV-infected children on first-line cART in Ethiopia.

Keywords: Ethiopia; HIV; children; dried blood spots; drug resistance; first-line combination antiretroviral therapy (cART); genotyping; pediatrics; treatment failure.

Figure 1

Prevalence of HIV-1 drug resistance among Ethiopian children experiencing virologic failure of first-line cART. Maximum-likelihood phylogeny inferred from the inclusive consensus sequences of 90 participants for whom genotyping was successful. Drug resistance codons were removed from the alignment prior to phylogenetic inference [48]. Scale indicates expected substitutions per nucleotide site. Colours indicate resistance genotype. Reference strains HXB2 (subtype B, green), MJ4 (subtype C-Botswana, orange) and KU319528 (subtype C-Ethiopia, pink) are included. (Inset) Distribution of HIV-1 resistance genotypes, stratified by drug class.

Figure 2

Prevalence of NRTI and NNRTI resistance mutations in participants with resistant genotypes: (A) NRTI resistance mutation (red) frequencies among 64 participants with at least one NRTI resistance mutation; and (B) NNRTI resistance mutation (blue) frequencies among 72 participants with at least one NNRTI resistance mutation. All mutations observed in >5% of resistant participants are shown.

Figure 3

Implications of resistant genotypes on recommended first- and second-line regimens: (A) prevalence of resistance to recommended first-line NNRTIs (blue) and NRTIs (red) among participants harboring NNRTI (N = 72) and NRTI (N = 64) resistance, respectively; and (B) burden of multi-NRTI resistance among participants harboring NRTI resistance (N = 64). Resistance to individual drugs was defined using the Stanford Drug Resistance Database, where genotypes exhibiting any level of reduced susceptibility to a given drug were considered “resistant” [51,52].

Figure 4

Concordance between resistance genotypes amplified with and without an initial Reverse Transcriptase (RT) step: (A) Resistance genotype concordance for 68 participants for whom paired PCR and RT-PCR genotypes were available (one participant per row). Color denotes resistance at the level of drug class. “O” symbols indicate that the genotype harbors at least one NRTI resistance mutation that is not observed in its associated pair, while “X” symbols indicate that the genotype harbors at least one NNRTI resistance mutation that is not observed in its associated pair; (B) Total number of resistance mutations observed in genotypes obtained with (right) and without (left) an initial RT step, where connecting lines indicate linked pairs.

Figure 5

Impact of replicate genotyping on resistance interpretation: (A) Proportion of individuals for whom all replicate resistance genotypes were concordant at the individual drug level (grey) versus those where interpretation based on a single genotype would have underestimated the degree of resistance (green). Analysis was based on 71 participants for whom at least two replicate genotypes, derived from any amplification type, were available; (B) Log₁₀ baseline plasma viral load of participants for whom all replicate resistance genotypes were concordant (grey) versus discordant (green) at the individual drug level; (C) Same as (B); but for log₁₀ plasma viral load at failure (D) same as (B), but for CD4+ T-cell count at baseline.