1st International Workshop on HIV Drug Resistance & Treatment Strategies


25-28 June 1997, St. Petersburg, Florida, USA


GENOTYPING AND PATTERN RECOGNITION OF HIV-1 AND MYCOBACTERIUM USING HIGH DENSITY OLIGONUCLEOTIDE ARRAYS

Antiviral Therapy 1997;2 (Suppl 5):32 (abstract no. 48)

TR Gingeras1, P Small2, M Holodniy3 and J Drenkow1
1Affymetrix, Santa Clara; 2Stanford University, Stanford; and 3VA Medical Center, Palo Alto, California, USA

PURPOSE: Using high density oligonucleotide arrays offers the possibility of examining large quantities of nucleotide sequence with a single hybridization. Applications of these arrays have thus far included: the sequence analysis of viral [1], mitochondrial [2] and human genomic sequences [3], as well as quantitative measurements of multiple murine gene expression [4] and functional mapping of the yeast genome [5]. In this study we have sought to use both the genotyping information and the information provided in the form of the hybridization patterns to speciate clinical Mycobacterium isolates and to provide clade discrimination for HIV-1.

METHODS: A total of 145 Mycobacterium isolates (82 non-tuberculosis and 63 Mycobacterium tuberculosis) including M. tuberculosis and non-tuberculosis species from California and New York as well as isolates and clones of clade B and non-clade B HIV-1 strains were analysed using high-density oligonucleotide genotyping and pattern recognition algorithms were employed to cluster the Mycobacterium isolates into species groups and HIV-1 into clade groups. At the same time, mutations conferring resistance to both anti-M. tuberculosis and HIV-1 drugs were detected.

RESULTS: Of the 63 M. tuberculosis isolates analysed, 44 were found to be phenotypically resistant to rifampicin. In 41 of the 44 of these M. tuberculosis isolates, mutations in rpoB were identified as the resistance conferring lesions. One new mutation (Gln513Glu) was observed. Analysis of the hybridization patterns allowed for the clustering of 121 of the isolates into 10 species groups (see Figure). Identification of HIV-1 isolates from clade B, and non-clade B was also made from the pattern analysis of the hybridizations of the protease and reverse transcriptase genes. From the same hybridization data, mutations conferring resistance to zidovudine, didanosine, zalcitabine and lamivudine in clade B isolates were simultaneously identified.

CONCLUSIONS: The analysis of rpoB gene and the protease/reverse transcriptase genes using genotypic and pattern recognition capabilities of high density oligonucleotide arrays permits the simultaneous detection of drug resistance conferring mutations and speciation/clade identification in Mycobacterium and HIV-1.

  1. Kozal M, Shah N, Naiping S, Yang R, Fucini R, Merigan TC, Richman DD, Morris D, Hubbell E, Chee M & Gingeras TR. Extensive polymorphisms observed in HIV-1 clade B protease gene using high-density oligonucleotide arrays. Nat Med. 1996 Jul;2(7):753-9.

  2. Chee M, Yang R, Hubbell E, Berno A, Huang XC, Stern D, Winkler J, Lockhart DJ, Morris MS & Fodor SPA. Accessing genetic information with high density DNA arrays. Science. 1996 Oct 25;274(5287):610-4.

  3. Hacia JG, Brody LC, Chee MS, Fodor SPA & Collins FS. Detection of heterozygous mutations in BRCA1 using high density oligonucleotide arrays and two-colour fluorescence analysis. Nat Genet. 1996 Dec;14(4):441-7.

  4. Lockhart D, Dong H, Byrne MC, Follettie MT, Gallo MV, Chee MS, Mittmann M, Wang C, Kobayashi M, Horton H & Brown EL. Expression monitoring by hybridization to high-density oligonucleotide arrays. Nat Biotechnol. 1996 Dec;14(13):1675-80.

  5. Shoemaker DD, Lashkari DA, Morris D, Mittmann M & Davis RW. Quantitive phenotypic analysis of yeast deletion mutants using a highly parallel molecular bar-coding strategy. Nat Genet. 1996 Dec;14(4):450-6.

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1997-06-25
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