title: Analysis of nevirapine resistance mutations in cloned HIV type 1 variants from HIV-infected Ugandan infants using a single-step amplification-sequencing method (AmpliSeq).
abstract: We analyzed the genetic linkage of nevirapine (NVP) resistance mutations and the genetic complexity of HIV-1 variants in Ugandan infants who were HIV infected despite single dose (SD) prophylaxis. Plasma samples were obtained from six HIV-infected infants who had two or more NVP resistance mutations detected by population sequencing (ViroSeq). ViroSeq PCR products were cloned and transformed, and a single-step amplification-sequencing reaction (AmpliSeq) was used to analyze NVP resistance mutations in cloned HIV-1 variants directly from bacterial colonies. Fifty clones were analyzed for each infant sample. This analysis revealed numerous NVP resistance mutations not detected by population sequencing, genetically linked NVP resistance mutations, and a high degree of genetic complexity at codons that influence NVP susceptibility.
authors.affiliation: Department of Pathology, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.
authors.forename: William Ian
authors.initials: WI
authors.lastname: Towler
authors.forename: Jessica D
authors.initials: JD
authors.lastname: Church
authors.forename: James R
authors.initials: JR
authors.lastname: Eshleman
authors.forename: Mary Glenn
authors.initials: MG
authors.lastname: Fowler
authors.forename: Laura A
authors.initials: LA
authors.lastname: Guay
authors.forename: J Brooks
authors.initials: JB
authors.lastname: Jackson
authors.forename: Susan H
authors.initials: SH
authors.lastname: Eshleman
mesh_terms: D019943:Amino Acid Substitution / Q000235:genetics; D019380:Anti-HIV Agents / Q000494:pharmacology*; D004252:DNA Mutational Analysis; D024882:Drug Resistance, Viral; D015658:HIV Infections / Q000821:virology*; D015497:HIV-1 / Q000187:drug effects* / Q000235:genetics / Q000302:isolation & purification; D006801:Humans; D007223:Infant; D008969:Molecular Sequence Data; D020125:Mutation, Missense; D019829:Nevirapine / Q000494:pharmacology*; D010949:Plasma / Q000821:virology; D012367:RNA, Viral / Q000235:genetics; D017422:Sequence Analysis, DNA / Q000379:methods; D014454:Uganda
chemical_list: D019380:Anti-HIV Agents; D012367:RNA, Viral; D019829:Nevirapine
references.citation: Am J Obstet Gynecol. 2007 Sep;197(3 Suppl):S56-63
references.pmid: 17825651
references.citation: Nucleic Acids Res. 2007;35(13):e91
references.pmid: 17576693
references.citation: Clin Chem. 2005 Jan;51(1):35-9
references.pmid: 15514094
references.citation: J Clin Microbiol. 2004 Aug;42(8):3670-4
references.pmid: 15297515
references.citation: J Clin Microbiol. 2005 Jan;43(1):406-13
references.pmid: 15635002
references.citation: Lancet. 2003 Sep 13;362(9387):859-68
references.pmid: 13678973
references.citation: Biotechniques. 2003 Sep;35(3):614-8, 620, 622
references.pmid: 14513567
references.citation: Antimicrob Agents Chemother. 2006 Jan;50(1):351-4
references.pmid: 16377709
references.citation: Proc Natl Acad Sci U S A. 2006 May 2;103(18):7094-9
references.pmid: 16641095
references.citation: J Infect Dis. 2007 Mar 1;195(5):711-5
references.pmid: 17262714
references.citation: Lancet. 1999 Sep 4;354(9181):795-802
references.pmid: 10485720
references.citation: Nat Methods. 2007 Feb;4(2):123-5
references.pmid: 17206150
references.citation: J Acquir Immune Defic Syndr. 2007 Nov 1;46(3):291-6
references.pmid: 18167645