Development of a new genotyping tool for molecular monitoring of HIV drug resistance

Anti-HIV drug resistance poses a threat to effective use of the available HIV drugs. Despite the introduction of HIV combination therapy, antiviral drug resistance was found to emerge with increasing duration of therapy. Rapidly accumulating mutations in the HIV genome are responsible for drug resistance. For adequate therapeutic decisions on individual as well as on population level, it is essential to identify the prevalence of specific mutations that are associated with antiviral drug resistance. In particular in resource poor settings, where only a small number of cheap HIV drugs are available, monitoring of mutation frequencies in both treatment naïve and patients receiving ART are of great importance to optimize the use of the few affordable drugs and to maintain treatment options. With the initiation of a HIV cohort and implementation of ART at the Ifakara Health Research and Development Center, Tanzania, great advances were made in rolling back HIV morbidity in the local population. To maintain this progress, the availability of diagnostic tests and monitoring tools needs to be secured at Ifakara Center.

The state-of-the-art monitoring tool for HIV drug resistance is direct sequencing of PCR products amplified from HIV cDNA. The mutational pattern detected in the sequence data is interpreted with the help of publicly available algorithms that predict antiretroviral response to a given set of mutations. Historically laboratory methods as well as computational analyses were based on HIV-1 subtype B, the most common strain in North America as well as in Switzerland.

Cheap and robust tools for monitoring drug resistance must replace the current methodology. The Molecular Diagnostics group at STI collaborates with T. Klimkait (Institute of Medical Microbiology, University of Basel) for developing a highly parallel micro array-based genotyping technique. The technological platform had been originally developed at STI for monitoring malaria drug resistance (Crameri et al. 2007) and has already been successfully applied in a number of field studies the past (Mugittu et al. 2006). The dedicated equipment necessary for micro array-based genotyping is already available at Ifakara.

Our prototype HIV micro array currently provides genotyping data on 18 single nucleotide polymorphisms (SNPs) within the reverse transcriptase gene that are associated with antiviral drug resistance. It is planned to extend the number of SNPs to include molecular markers from other genes as well. We target specifically the non-B HIV-1 subtypes A, C, and D, presumably predominant in Tanzania. In a prospective analysis genotypes at baseline will be analyzed with respect to treatment failures during the longitudinal cohort surveys. We hope to implement locally low cost surveillance of transmitted ART resistance. The availability of monitoring tools in Ifakara will foster evidence-based public health decisions on ART in the Kilombero District of Tanzania, where a roll out of ART is on its way.

Dr. Ingrid Felger, Molecular Diagnostics, Department of Medical Parasitology and Infection Biology, Swiss Tropical Institute, Basel
Janet Gare
Dr. Thomas Klimkait

References
Crameri A, Marfurt J, Mugittu K, Maire N, Regös A, Coppee JY, Sismeiro O, Burki R, Huber E, Laubscher D, Puijalon O, Genton B, Felger I, Beck HP.Rapid microarray-based method for monitoring of all currently known single-nucleotide polymorphisms associated with parasite resistance to antimalaria drugs.J Clin Microbiol. 2007 Nov;45(11):3685-91.

Mugittu K, Genton B, Mshinda H, Beck HP.Molecular monitoring of Plasmodium falciparum resistance to artemisinin in Tanzania. Malar J. 2006 Dec 19;5:126.

 

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