1. HIV Genotyping: Sharing Experiences and Protocols Justen Manasa PhD Candidate Africa Centre for Health and Population Studies Virology laboratory based at the Nelson R. Mandela School of Medicine University of KwaZulu Natal

2. Introduction Drug resistance genotyping labs to increase –Surveillance –Clinical management

3. Introduction Lack of common standards Limitations in lab infrastructure Cost and complexity of genotypic resistance testing

4. AIM Ensure access to validated, quality assured genotyping services in SATuRN member countries and collaborating institutions

5. Objectives Create laboratory mentorship programmes for Labs setting up genotyping services Create a low cost EQA programme for resource limited settings Create an open access online resource for sharing genotyping protocols for HIV and TB genotyping

6. WHO Accredited Labs

7. Laboratory Protocols @ Bioafrica.net

8. Laboratory Protocols Capillary Sequencing Next Generation Sequencing Allele Specific PCR

9. POLENV nefGAG vpr vifLTR vpu Complete Genome (9719 bases) RTPRIN tat rev Protocols for sequencing the HIV whole genome or specific gene fragment using capillary electrophoresis The HIV genome or its segments have been and are currently being sequencing for many reasons including; Viral evolution Phylogeography Drug resistance Epidemiology The links below provides some of the protocols that have been validated and used for HIV sequencing using capillary electrophoresis. Please click on the segment of your interest to go to the protocol. Page last updated by Justen Manasa, 16 November 2013

10. Paser Ultra-light:Citations ()Comments A Pragmatic Approach to HIV-1 Drug Resistance Determination in Resource-Limited Settings by Use of a Novel Genotyping Assay Targeting the Reverse Transcriptase-Encoding Region Only. AITKEN, S. C., BRONZE, M., WALLIS, C. L., STUYVER, L., STEEGEN, K., BALINDA, S., KITYO, C., STEVENS, W., RINKE DE WIT, T. F. & SCHUURMAN, R. 2013. J Clin Microbiol, 51, 1757-61. Wallis:Citations ()Comments Affordable in-house antiretroviral drug resistance assay with good performance in non-subtype B HIV-1. WALLIS, C. L., PAPATHANASOPOULOS, M. A., LAKHI, S., KARITA, E., KAMALI, A., KALEEBU, P., SANDERS, E., ANZALA, O., BEKKER, L. G., STEVENS, G., DE WIT, T. F. & STEVENS, W. 2010. J Virol Methods, 163, 505-8. CDC:Citations ()Comments Optimization of a low cost and broadly sensitive genotyping assay for HIV-1 drug resistance surveillance and monitoring in resource-limited settings. ZHOU, Z., WAGAR, N., DEVOS, J. R., ROTTINGHAUS, E., DIALLO, K., NGUYEN, D. B., BASSEY, O., UGBENA, R., WADONDA- KABONDO, N., MCCONNELL, M. S., ZULU, I., CHILIMA, B., NKENGASONG, J. & YANG, C. 2011. PLoS One, 6, e28184. SATuRN/Life Technologies:Citations ()Comments An Affordable HIV-1 Drug Resistance Monitoring Method for Resource Limited Settings. MANASA, J., DANAVIAH, S., PILLAY, S., PADAYACHEE, P., MTHIYANE, H., SEEBREGTS, C., RINKE DE WIT, T. F., VILJOEN, J., KATZENSTEIN, D. & DE OLIVEIRA, T. 2013. Jove (in press). Chen:Citations ()Comments In-house human immunodeficiency virus-1 genotype resistance testing to determine highly active antiretroviral therapy resistance mutations in Hong Kong. CHEN, J. H., WONG, K. H., LI, P. C., CHAN, K. K., LEE, M. P., TO, S. W. & YAM, W. C. 2012. Hong Kong Med J, 18, 20-4. NICD:Citations ()Comments Antiretroviral drug resistance surveillance among drug-naive HIV-1-infected individuals in Gauteng Province, South Africa in 2002 and 2004. PILLAY, V., LEDWABA, J., HUNT, G., RAKGOTHO, M., SINGH, B., MAKUBALO, L., BENNETT, D. E., PUREN, A. & MORRIS, L. 2008. Antivir Ther, 13 Suppl 2, 101-7. Protocols for sequencing the HIV whole genome or specific gene fragment using capillary electrophoresis Here we provide detailed full protocols. The protocols are linked to the primary papers in which they were 1 st published. You can also access a summary of publications that have used the same protocol and referenced the primary paper. For each protocol there is also a comment section which should be used by users to discuss issues related to implementation of the protocol Antiretroviral drug resistance surveillance among drug-naive HIV-1-infected individuals in Gauteng Province, South Africa in 2002 and 2004. Antiviral Therapy (2008). Citations (9) Optimization of a low cost and broadly sensitive genotyping assay for HIV-1 drug resistance surveillance and monitoring in resource-limited settings. Plos One (2010). Citations (6) Affordable in-house antiretroviral drug resistance assay with good performance in non-subtype B HIV-1. J. Virol Methods (2010). Citations (5) A Pragmatic Approach to HIV-1 Drug Resistance Determination in Resource- Limited Settings by Use of a Novel Genotyping Assay Targeting the Reverse Transcriptase-Encoding Region Only. J Clin Microbiol (2013). Citations (4) In-house human immunodeficiency virus-1 genotype resistance testing to determine highly active antiretroviral therapy resistance mutations in Hong Kong. Hong Kong Med J (2010). Citations (3) An Affordable HIV-1 Drug Resistance Monitoring Method for Resource Limited Settings. Jove (In Press) Citations (2) Protocols, Primary Publication (orded by publication date)Ranked by number of citations

11. Discussion Registration and comments Blog Moderation of user comments General Trouble shooting guidelines Structure of protocol Videos Any other request?

12. Thank You