title: Next generation sequencing improves detection of drug resistance mutations in infants after PMTCT failure.
abstract: BACKGROUND
  Next generation sequencing (NGS) allows the detection of minor variant HIV drug resistance mutations (DRMs). However data from new NGS platforms after Prevention-of-Mother-to-Child-Transmission (PMTCT) regimen failure are limited.
  
  
  OBJECTIVE
  To compare major and minor variant HIV DRMs with Illumina MiSeq and Life Technologies Ion Personal Genome Machine (PGM) in infants infected despite a PMTCT regimen.
  
  
  METHODS
  We conducted a cross-sectional study of NGS for detecting DRMs in infants infected despite a zidovudine (AZT) and Nevirapine (NVP) regimen, before initiation of combination antiretroviral therapy. Sequencing was performed on PCR products from plasma samples on PGM and MiSeq platforms. Bioinformatic analyses were undertaken using a codon-aware version of the Smith-Waterman mapping algorithm and a mixture multinomial error filtering statistical model.
  
  
  RESULTS
  Of 15 infants, tested at a median age of 3.4 months after birth, 2 (13%) had non-nucleoside reverse transcriptase inhibitor (NNRTI) DRMs (K103N and Y181C) by bulk sequencing, whereas PGM detected 4 (26%) and MiSeq 5 (30%). NGS enabled the detection of additional minor variant DRMs in the infant with K103N. Coverage and instrument quality scores were higher with MiSeq, increasing the confidence of minor variant calls.
  
  
  CONCLUSIONS
  NGS followed by bioinformatic analyses detected multiple minor variant DRMs in HIV-1 RT among infants where PMTCT failed. The high coverage of MiSeq and high read quality improved the confidence of identified DRMs and may make this platform ideal for minor variant detection.
authors.affiliation: Division of Medical Virology, Department of Pathology, Faculty of Medicine and Health Sciences, Stellenbosch University, Francie van Zilj Drive, Parow 7500, South Africa.
authors.forename: Randall G
authors.initials: RG
authors.lastname: Fisher
authors.affiliation: Department of Medicine, University of California, San Diego, Stein Clinical Research Building #325 (mail code 0679), 9500 Gilman Drive, La Jolla, CA 92093, USA; San Diego Veterans Affairs Healthcare System, (Mail Code 8208), 150 W. Washington Street #100, San Diego, CA 92103, USA.
authors.forename: Davey M
authors.initials: DM
authors.lastname: Smith
authors.affiliation: Department of Medicine, University of California, San Diego, Stein Clinical Research Building #325 (mail code 0679), 9500 Gilman Drive, La Jolla, CA 92093, USA.
authors.forename: Ben
authors.initials: B
authors.lastname: Murrell
authors.affiliation: Department of Genetics, Stellenbosch University, Private Bag X1, 7602 Matieland, South Africa.
authors.forename: Ruhan
authors.initials: R
authors.lastname: Slabbert
authors.affiliation: Institute for Microbial Biotechnology and Metagenomics, New Life Sciences Building, 2nd Floor, Core 2, University of the Western Cape, Modderdam Road, P/Bag X17, Bellville 7530, South Africa.
authors.forename: Bronwyn M
authors.initials: BM
authors.lastname: Kirby
authors.affiliation: Department Paediatrics and Child Health, Stellenbosch University and Tygerberg Children's Hospital, Francie van Zijl Drive, Parow 7500, South Africa.
authors.forename: Clair
authors.initials: C
authors.lastname: Edson
authors.affiliation: Department Paediatrics and Child Health, Stellenbosch University and Tygerberg Children's Hospital, Francie van Zijl Drive, Parow 7500, South Africa; Children's Infectious Diseases Clinical Research Unit (KIDCRU) Ward J8, Tygerberg Hospital, Francie van Zijl Drive, Parow 7500, South Africa.
authors.forename: Mark F
authors.initials: MF
authors.lastname: Cotton
authors.affiliation: University of California, San Diego, AVRC, 220 Dickinson, Suite A, San Diego, CA 92103, USA.
authors.forename: Richard H
authors.initials: RH
authors.lastname: Haubrich
authors.affiliation: Department of Medicine, University of California, San Diego, Stein Clinical Research Building #325 (mail code 0679), 9500 Gilman Drive, La Jolla, CA 92093, USA.
authors.forename: Sergei L
authors.initials: SL
authors.lastname: Kosakovsky Pond
authors.affiliation: Division of Medical Virology, Department of Pathology, Faculty of Medicine and Health Sciences, Stellenbosch University, Francie van Zilj Drive, Parow 7500, South Africa; National Health Laboratory Service, Tygerberg, Francie van Zijl Drive, Parow 7500, South Africa. Electronic address: guvz@sun.ac.za.
authors.forename: Gert U
authors.initials: GU
authors.lastname: Van Zyl
mesh_terms: D019380:Anti-HIV Agents / Q000494:pharmacology* / Q000627:therapeutic use; D023241:Antiretroviral Therapy, Highly Active; D018791:CD4 Lymphocyte Count; D019295:Computational Biology; D024882:Drug Resistance, Viral; D005260:Female; D005838:Genotype; D015658:HIV Infections / Q000175:diagnosis* / Q000188:drug therapy / Q000635:transmission / Q000821:virology*; D015497:HIV-1 / Q000187:drug effects* / Q000235:genetics*; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D007223:Infant; D007231:Infant, Newborn; D018445:Infectious Disease Transmission, Vertical / Q000517:prevention & control; D008297:Male; D008826:Microbial Sensitivity Tests; D009154:Mutation; D059645:Mutation Rate; D012367:RNA, Viral; D012189:Retrospective Studies; D019562:Viral Load
chemical_list: D019380:Anti-HIV Agents; D012367:RNA, Viral
keywords: HIV-1 drug resistance; Minor variant antiretroviral drug resistance mutations; Nevirapine; Next generation sequencing; Non-nucleoside reverse transcriptase inhibitor resistance; Prevention of mother to child transmission