Ko Daisy, McLaughlin Sherry, Deng Wenjie, Mullins James I, Dragavon Joan, Harb Socorro, Coombs Robert W, Frenkel Lisa M
Center for Global Infectious Disease Research, Seattle Children's Research Institute, Seattle, WA 98109, USA.
Department of Microbiology, University of Washington, Seattle, WA 98195, USA.
Viruses. 2024 Mar 27;16(4):510. doi: 10.3390/v16040510.
HIV-1 typically infects cells via the CD4 receptor and CCR5 or CXCR4 co-receptors. Maraviroc is a CCR5-specific viral entry inhibitor; knowledge of viral co-receptor specificity is important prior to usage. We developed and validated an economical V3- Illumina-based assay to detect and quantify the frequency of viruses utilizing each co-receptor. Plasma from 54 HIV+ participants (subtype B) was tested. The viral template cDNA was generated from plasma RNA with unique molecular identifiers (UMIs). The sequences were aligned and collapsed by the UMIs with a custom bioinformatics pipeline. Co-receptor usage, determined by codon analysis and online phenotype predictors PSSM and Geno2pheno, were compared to existing Trofile data. The cost of V3-UMI was tallied. The sequences interpreted by Geno2pheno using the most conservative cut-off, a 2% false-positive-rate (FPR), predicted CXCR4 usage with the greatest sensitivity (76%) and specificity (100%); PSSM and codon analysis had similar sensitivity and lower specificity. Discordant Trofile and genotypic results were more common when participants had specimens from different dates analyzed by either assay. V3-UMI reagents cost USD$62/specimen. A batch of ≤20 specimens required 5 h of technical time across 1.5 days. V3-UMI predicts HIV tropism at a sensitivity and specificity similar to those of Trofile, is relatively inexpensive, and could be performed by most central laboratories. The adoption of V3-UMI could expand HIV drug therapeutic options in lower-resource settings that currently do not have access to phenotypic HIV tropism testing.
HIV-1通常通过CD4受体以及CCR5或CXCR4共受体感染细胞。马拉维若为一种CCR5特异性病毒进入抑制剂；在使用前了解病毒共受体特异性很重要。我们开发并验证了一种基于V3区Illumina测序的经济检测方法，以检测和定量利用每种共受体的病毒频率。对54名HIV阳性（B亚型）参与者的血浆进行了检测。使用独特分子标识符（UMIs）从血浆RNA中生成病毒模板cDNA。通过定制的生物信息学流程，利用UMIs对序列进行比对和归并。将通过密码子分析以及在线表型预测工具PSSM和Geno2pheno确定的共受体使用情况与现有的Trofile数据进行比较。计算了V3-UMI的成本。Geno2pheno使用最保守的截止值（2%的假阳性率）解释的序列，预测CXCR4使用情况时具有最高的灵敏度（76%）和特异性（100%）；PSSM和密码子分析具有相似的灵敏度但特异性较低。当参与者有不同日期的样本通过任何一种检测方法进行分析时，Trofile和基因分型结果不一致的情况更为常见。V3-UMI试剂成本为62美元/样本。一批≤20个样本需要在1.5天内花费5小时的技术时间。V3-UMI预测HIV嗜性的灵敏度和特异性与Trofile相似，相对便宜，并且大多数中心实验室都可以进行。采用V3-UMI可以在目前无法进行HIV表型嗜性检测的资源匮乏地区扩大HIV药物治疗选择。
