Mutsvangwa J, Beck I A, Gwanzura L, Manhanzva M T, Stranix-Chibanda L, Chipato T, Frenkel L M
Biomedical Research & Training Institute (BRTI), Zimbabwe; University of Zimbabwe, College of Health Sciences, Zimbabwe.
Seattle Children's Research Institute, Seattle, WA, USA.
J Virol Methods. 2014 Dec 15;210:36-9. doi: 10.1016/j.jviromet.2014.09.005. Epub 2014 Sep 17.
An oligonucleotide ligation assay (OLA) designed to detect Human Immunodeficiency Virus type-1 (HIV)-drug-resistance to the nevirapine (NVP) selected mutations K103N, Y181C, V106M and G190A was used to evaluate 200 archived dried blood spots (DBS) from infected infants participating in the Zimbabwean Early Infant Diagnosis (EID) Program. Consensus sequencing of specimens with indeterminate OLA results was performed to identify genetic sequence polymorphisms that appeared to compromise performance of the OLA. When consistent patterns of polymorphisms were observed the probes were redesigned, and DBS specimens with indeterminate OLA results were retested with the new Zimbabwe-specific (ZW) probes. OLA results obtained in Zimbabwe were compared to repeat testing in a US reference laboratory. 188/200 (94%) DBS yielded polymerase chain reaction (PCR) amplification of HIV pol. ZW probes reduced indeterminate OLA results from 5.2% to 2.8% of codons evaluated (p=0.02), with 98.2% concordance between results obtained in the Zimbabwean and US laboratories. Optimization of OLA probes to accommodate polymorphisms in regional HIV variants improved OLA performance, and comparison to the USA results showed successful implementation of the OLA in Zimbabwe for detection of NVP resistance mutations in DBS specimens.
一种旨在检测人类免疫缺陷病毒1型（HIV-1）对奈韦拉平（NVP）耐药相关选择突变K103N、Y181C、V106M和G190A的寡核苷酸连接测定（OLA），用于评估参与津巴布韦早期婴儿诊断（EID）项目的感染婴儿的200份存档干血斑（DBS）样本。对OLA结果不确定的样本进行一致性测序，以鉴定似乎影响OLA性能的基因序列多态性。当观察到一致的多态性模式时，重新设计探针，并用新的津巴布韦特异性（ZW）探针重新检测OLA结果不确定的DBS样本。将在津巴布韦获得的OLA结果与美国参考实验室的重复检测结果进行比较。200份DBS样本中有188份（94%）实现了HIV pol的聚合酶链反应（PCR）扩增。ZW探针将不确定的OLA结果从所评估密码子的5.2%降至2.8%（p=0.02），津巴布韦和美国实验室获得的结果之间的一致性为98.2%。优化OLA探针以适应区域HIV变体中的多态性可提高OLA性能，与美国结果的比较表明，OLA在津巴布韦成功用于检测DBS样本中的NVP耐药突变。
