Gonçalves Fátima, Cabanas Joaquim, Costa Inês, Veloso Margarida, Ribeiro Marta, Fernandes Sandra, Diogo Isabel, Sebastião Cruz S, Pingarilho Marta, Pimentel Victor, Abecasis Ana, Gomes Perpétua
Laboratório de Biologia Molecular (LMCBM, SPC, HEM, ULS-LO), Lisboa 1349-019, Portugal.
Global Health and Tropical Medicine (GHTM), Associate Laboratory in Translation and Innovation Towards Global Health (LA-REAL), Instituto de Higiene e Medicina Tropical (IHMT), Universidade NOVA de Lisboa (UNL), Rua da Junqueira 100, Lisboa 1349-008, Portugal; Centro Nacional de Investigação Científica (CNIC), Luanda, Angola; Centro de Investigação em Saúde de Angola (CISA), Caxito, Angola; Instituto Nacional de Investigação em Saúde (INIS), Luanda, Angola.
J Virol Methods. 2025 May;334:115112. doi: 10.1016/j.jviromet.2025.115112. Epub 2025 Feb 7.
HIV-2 affects over 1 million people globally and can lead to AIDS if untreated. Treating people living with HIV-2 (PLHIV-2) is challenging because the virus is inherently resistant to some drugs. Effective treatment monitoring, particularly drug resistance testing, is critical for managing therapeutic failure. Without commercial tests to identify drug resistance mutations (DRM), laboratories have felt the need to develop in-house methods. NGS provides improved sensitivity for detecting minority DRM, which is crucial for effectively treating individuals, especially with limited therapeutic options. This study aimed to evaluate the effectiveness of a hybrid NGS Ion Torrent protocol for the detection of DRM in PLHIV-2 and its use in clinical practice. One hundred samples from PLHIV-2 collected from hospitals across Portugal were analyzed using a hybrid NGS protocol. Of these, 48 samples were also subjected to Sanger sequencing for comparative purposes. NGS successfully amplified 92 % of protease, 91 % of reverse transcriptase, and 49 % of integrase regions. The two sequencing methods agreed on the majority of DRM identified, with the only difference in two samples for the reverse transcriptase, which NGS identified as K70E and M184V, while Sanger did not. Hybrid NGS was able to identify DRM, demonstrating strong statistical agreement. In conclusion, hybrid NGS detected all DRM identified by Sanger, with the added ability to detect minority variants. The implementation of NGS-based protocol can provide clinicians with more comprehensive data, allowing for adjustments to ART regimens, and ultimately improving patient outcomes and quality of care for PLHIV-2.
HIV-2在全球感染超过100万人，若不治疗可导致艾滋病。治疗HIV-2感染者（PLHIV-2）具有挑战性，因为该病毒对某些药物具有内在抗性。有效的治疗监测，尤其是耐药性检测，对于管理治疗失败至关重要。由于缺乏用于鉴定耐药性突变（DRM）的商业检测方法，实验室感到有必要开发内部方法。二代测序（NGS）提高了检测少数DRM的灵敏度，这对于有效治疗个体至关重要，尤其是在治疗选择有限的情况下。本研究旨在评估一种混合NGS Ion Torrent方案检测PLHIV-2中DRM的有效性及其在临床实践中的应用。使用混合NGS方案分析了从葡萄牙各地医院收集的100份PLHIV-2样本。其中48份样本也进行了Sanger测序以作比较。NGS成功扩增了92%的蛋白酶区域、91%的逆转录酶区域和49%的整合酶区域。两种测序方法在鉴定出的大多数DRM上达成一致，仅在两个逆转录酶样本上存在差异，NGS鉴定为K70E和M184V，而Sanger测序未鉴定出。混合NGS能够鉴定DRM，显示出很强的统计学一致性。总之，混合NGS检测出了Sanger测序鉴定出的所有DRM，并且还具有检测少数变异体的能力。基于NGS方案的实施可为临床医生提供更全面的数据，以便调整抗逆转录病毒治疗方案，最终改善PLHIV-2患者的治疗效果和护理质量。
