title: Optimization of the oligonucleotide ligation assay for the detection of nevirapine resistance mutations in Zimbabwean Human Immunodeficiency Virus type-1 subtype C.
abstract: 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.
authors.affiliation: Biomedical Research & Training Institute (BRTI), Zimbabwe; University of Zimbabwe, College of Health Sciences, Zimbabwe. Electronic address: jmutsvangwa@brti.co.zw.
authors.forename: J
authors.initials: J
authors.lastname: Mutsvangwa
authors.affiliation: Seattle Children's Research Institute, Seattle, WA, USA. Electronic address: ingrid.beck@seattlechildrens.org.
authors.forename: I A
authors.initials: IA
authors.lastname: Beck
authors.affiliation: Biomedical Research & Training Institute (BRTI), Zimbabwe; University of Zimbabwe, College of Health Sciences, Zimbabwe. Electronic address: gwanzura@mweb.co.zw.
authors.forename: L
authors.initials: L
authors.lastname: Gwanzura
authors.affiliation: University of Zimbabwe, College of Health Sciences, Zimbabwe. Electronic address: mnlsmnhnzv2@gmail.com.
authors.forename: M T
authors.initials: MT
authors.lastname: Manhanzva
authors.affiliation: University of Zimbabwe, College of Health Sciences, Zimbabwe. Electronic address: lynda@uz-ucsf.co.zw.
authors.forename: L
authors.initials: L
authors.lastname: Stranix-Chibanda
authors.affiliation: University of Zimbabwe, College of Health Sciences, Zimbabwe. Electronic address: tchipato@zol.co.zw.
authors.forename: T
authors.initials: T
authors.lastname: Chipato
authors.affiliation: Seattle Children's Research Institute, Seattle, WA, USA; University of Washington, Seattle, WA, USA. Electronic address: lfrenkel@u.washington.edu.
authors.forename: L M
authors.initials: LM
authors.lastname: Frenkel
mesh_terms: D019380:Anti-HIV Agents / Q000494:pharmacology*; D024882:Drug Resistance, Viral / Q000235:genetics*; D005838:Genotype; D060005:Genotyping Techniques; D015658:HIV Infections / Q000188:drug therapy / Q000821:virology*; D015497:HIV-1 / Q000235:genetics* / Q000302:isolation & purification; D006801:Humans; D007223:Infant; D007231:Infant, Newborn; D019829:Nevirapine / Q000494:pharmacology*; D015345:Oligonucleotide Probes; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D015030:Zimbabwe
chemical_list: D019380:Anti-HIV Agents; D015345:Oligonucleotide Probes; D019829:Nevirapine
keywords: Genotyping; HIV; HIV drug-resistance; Nevirapine; Oligonucleotide ligation assay; Point mutation