Despite significant advances in Human Immunodeficiency Virus (HIV-1) treatment, antiretroviral drug resistance remains a critical challenge to the efficacy of Antiretroviral Therapy (ART), as it drives the accumulation of resistance-associated mutations, virological failure, and increased morbidity.1–5 Integrase Strand Transfer Inhibitors (INSTIs) are a class of antiretrovirals with a high genetic barrier to resistance, potent viral suppression, favourable safety profiles, and fewer drug-drug interactions.6 In Brazil, INSTIs are now recommended as the first- and second-line ART regimens following virological failure.7,8 Since 2017, the Brazilian Ministry of Health has adopted the preferred initial regimen of tenofovir disoproxil fumarate, lamivudine (3TC), and Dolutegravir (DTG), a second-generation INSTI.7,8 Recent national guidelines advocate for optimised dual therapy (3TC + DTG), particularly for patients with HIV-associated comorbidities.9 By 2020, approximately 410,000 Brazilian patients were receiving INSTI-based regimens.10 Given Brazil’s continental-scale implementation of these therapies, the widespread use of INSTIs may accelerate the emergence of resistance-associated mutations, potentially compromising their long-term effectiveness.

Tao et al. (2023) identified 99 sequences with INSTI resistance-associated mutations by analysing antiretroviral resistance profiles.11 Scutari et al. (2020) reported an INSTI resistance frequency of 36.4 %, among treatment-experienced individuals and those with long HIV infection duration.12 However, other studies have detected INSTI resistance mutations even in treatment-naïve patients.13,14 The identification of antiretroviral resistance mutations is widely recognised as a cost-effective strategy for optimising antiretrovirals and preventing the initiation or continuation of regimens that are likely to result in virological failure, which is a critical measure for HIV epidemiological control. The World Health Organization recommends routine resistance testing in children at diagnosis and after virological failure.15 According to Joint United Nations Programme on HIV/AIDS (UNAIDS), approximately one-third of children with HIV infection worldwide experience virological failure within 2-years of ART initiation.16–18

In Brazil, Ventosa-Cubillo et al. (2023) analysed HIV-1 sequences from 67 children and adolescents and identified INSTI-associated Drug Resistance Mutations (DRMs) in eight patients.19 Andrade et al. (2017) studied 117 children with HIV infection (mean age = 3.7-years) in Northern Brazil and reported high DRM frequencies, even among those who were treatment-naïve, particularly for non-nucleoside reverse transcriptase inhibitor-associated mutations.20 A retrospective Brazilian study on individuals with virological failure detected INSTI resistance mutations in 22.1 % of participants.10 Notably, a large multicentre study of mother-infant pairs in South Africa, Brazil, and Argentina demonstrated a strong association between maternal DRMs and their transmission to infants.21

Brazilian studies evaluating INSTI resistance have reported high frequencies of polymorphic variants associated with an increased risk of DRMs.22–24 However, data on INSTI resistance profiles, specifically in paediatric populations, remain scarce. Given Brazil's early adoption of INSTI-based regimens as first-line ART, understanding the resistance patterns in children is critical. Surveillance of DRM dissemination represents a fundamental strategy for HIV control, particularly in regions such as Brazil, where INSTIs were rapidly integrated into national treatment guidelines. This study aimed to characterise HIV-1 resistance mutations to INSTIs among children who were vertically infected and received care in the Maranhão and Pernambuco states of Northeast Brazil.

This study analysed 31 paediatric patients diagnosed with HIV-1 infection who experienced virological failure, with a median age of 12-years (IQR: 9–16). Of them, 18 (58 %) were male patients and 13 (42 %) female patients. The median CD4+ T-cell count and viral load at the time of virological failure were 999 cells/mm3 (IQR: 707–1432.5) and 19,235 copies/mL (IQR: 1508.5–270,375), respectively. HIV-1 subtyping of the integrase region revealed subtype B (n = 23, 74.2 %), BF recombinants (n = 5, 16.1 %), and subtype F1 (n = 3, 9.7 %) (Fig. 1). Additionally, six patients (19.4 %) had previously used INSTIs (Table 1). Analysis of resistance to INSTIs (Bictegravir [BIC], Dolutegravir [DTG], Elvitegravir [EVG], Cabotegravir [CAB], and Raltegravir [RAL]) showed that 20 sequences (64.5 %) remained susceptible to BIC and DTG, representing the INSTIs with the lowest resistance levels. In contrast, the highest resistance rate was observed for RAL (n = 21, 67.8 %) (Fig. 2). Twenty-one HIV-1 sequences (67.8 %) exhibited major INSTI resistance mutations. The most frequent resistance mutations were G140S and Q148H (both n = 10, 32.2 %). The Q148K mutation in the integrase gene was observed in 11 patients (35.5 %). Importantly, the Q148H/K mutations conferred resistance to all INSTIs, indicating a high level of resistance in the paediatric population. The N155H mutation was detected in seven patients (22.6 %), of whom two have been receiving antiretroviral therapy that includes INSTIs, and conferred resistance to CAB, EVG, and RAL. Mutations at integrase position 143 (Y143H/R/G) were documented in four patients and conferred resistance to RAL. Among the current ART regimens administered to the patients, the most common were Zidovudine (AZT) + Lamivudine (3TC) + Lopinavir/ritonavir (LPV/r) (n = 10; 32.2 %), AZT + 3TC + Nevirapine (NVP) (n = 6; 19.4 %), and AZT + 3TC + efavirenz (EFV) (n = 5; 16.1 %). The duration of the current ART regimen ranged from 1- to 10-years, with a median of 5.5-years (Table 2).

Fig. 1.

Phylogenetic characterisation of HIV-1 pol sequences obtained from children and adolescents with virological failure in Northeast Brazil. Red: Subtype B; Green: Subtype F1; Blue: BF recombinants.

Fig. 2.

Incidence of antiretroviral resistance to integrase inhibitors in paediatric patients with virological failure in Northeast Brazil. S, Susceptible; I, Intermediate; R, Resistant; BIC, Bictegravir; DTG, Dolutegravir; EVG, Elvitegravir; CAB, Cabotegravir; RAL, Raltegravir.

The current study aimed to retrospectively analyse the antiretroviral resistance profiles of INSTIs in virologically failing patients under 16-years of age who were followed by Brazil's Unified Health System. Given the scarcity of research on ART resistance in paediatric populations, our study population selection is a key strength. We analysed 31 HIV-1 integrase region sequences from virologically failing children and adolescents in Northeast Brazil for antiretroviral resistance mutations. Six patients (19.4 %) received INSTIs before genotypic testing for virological failure. The highest resistance frequency was observed for RAL (n = 21; 67.8 %), indicating a high-level resistance to at least one INSTI.

The predominant HIV-1 subtype was B (74.2 %), which is consistent with phylogenetic data from previous Brazilian studies that identified this as the most common subtype in both the Northeast and other Brazilian regions, except for Southern Brazil.29–31 Among the non-B subtypes, BF showed the highest incidence (19 %), aligning with the findings of a previous study conducted in the Maranhão State, Northeast Brazil. However, 2017 data identified HIV-F as the most common subtype in the Pernambuco State, Northeast Brazil,32 which contrasts with our findings. The predominance of subtype B may reflect its status as the first subtype to be isolated in the Western world, which facilitated its subsequent spread.

BIC and DTG showed the lowest resistance rates, with 64.5 % of sequences remaining susceptible. Notably, DTG, included in Brazil's first-line adult ART regimen as per the Ministry of Health guidelines, has recently been formulated as dispersible tablets for HIV treatment and prevention in children aged 2-months to 6-years,33 significantly improving paediatric treatment feasibility.

Regarding the most prevalent INSTI resistance mutations, in descending order of frequency, we identified G140S, Q148H, N155H, Q148K, Y143H, Y143HR, and Y143R. The high frequency of G140S and Q148H mutations (both n = 10; 32.2 %) raises significant concerns, as they confer resistance to all INSTIs. Mutations at positions 140, 143, 148, and 155 of the HIV-1 integrase gene induce resistance to multiple INSTIs.33 For instance, N155H (22.6 %, n = 7/31) conferred resistance to most INSTIs except DTG. These findings in the paediatric population may suggest consequences associated with prior INSTI exposure, either through the individual’s treatment history or potential maternal transmission. Mutations most associated with INSTI resistance (G140S, Q148H, and N155H) were observed in various combinations among patients infected with subtypes B and F1. Considering the population's median age of 12-years, approximately 35 % showed DTG resistance (only introduced in Brazil's public health system in 2016), and only 19.4 % (n = 6/31) had prior INSTI exposure. We hypothesised that INSTI resistance may be emerging rapidly, with treatment adherence likely playing a crucial preventive role.34 Mutation distribution analysis revealed that G140S and Q148H frequently co-occurred in patients infected with subtype B virus, whereas N155H appeared in patients infected with subtype B or F1 virus. Patients infected with recombinant BF virus showed INSTI-sensitive profiles, with only one patient exhibiting resistance mutations.

Despite the inclusion criterion adopted in the study, selecting only patients with virological failure who had genotyping results available may have introduced a selection bias towards individuals already suspected of having HIV drug resistance, continuous genotypic monitoring of these patients is essential for therapeutic adjustments to improve clinical outcomes, particularly in paediatric populations. This study highlights the critical need for the surveillance of INSTI resistance mutations, as emerging resistance may compromise treatment efficacy and safety. Systematic resistance monitoring ensures that INSTIs remain a viable therapeutic option, while advancing HIV research to develop more potent treatments and tailored prevention strategies for vulnerable populations. Through these efforts, healthcare systems can implement preventive measures, where early detection of viral resistance may curb the transmission of drug-resistant strains and guide policymakers in optimising antiretroviral selection.

This study was approved by the Research Ethics Committee of Federal University of Pernambuco, under approval number 5.180.229 (December 20, 2021), in accordance with the guidelines of the National Health Council of Brazil (Resolution 466/12).

The ethics committee accepted the exemption from the patient consent statement, as only HIV-1 medical records and genomic data were obtained.

Permission to reproduce material from other sources was obtained where necessary. All reproduced content is appropriately cited and used with authorization from the original copyright holders.

None.

This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.

The data that support the findings of this study are available from the corresponding author upon reasonable request.