|Job Title||Junior Scientist, specialised on Infectious Diseases|
|Date of Scholarship||February-May 2012|
|Host Country||University of Tor Vergata (UTV), Rome, Italy|
|Scholarship||HIV-1 Ultra-Deep Sequencing for the Study of Minority Antiretroviral Drug Resistance Variants|
HIV-1 ULTRA DEEP SEQUENCING FOR THE STUDY OF MINORITY ANTIRETROVIRAL DRUG RESISTANCE VARIANTS
I undertook my training project at the Chair of Virology Laboratory at the Faculty of Medicine and Surgery, University of Rome “Tor Vergata”, Rome, Italy. The training covered the following:
- Theoretical and practical knowledge on HIV-1 ultra-deep sequencing: This training was effective by processing a set of twenty HIV-1 plasma samples using the Roche P454 pyrosequencing technology. Briefly, RNA extraction was performed using QiagenTM, followed by amplicon library construction (library preparation, quantitation and dilution), emulsion amplification (PCR, bead recovery, bead enrichment and bead count), sequencing reaction (preparing the PicoTiter Plate device and start run), and primary data analysis using the AVA software (Amplicon Variant Analyzer). Secondary analysis is underway for the study of HIV-1 minority drug resistant quasi-species, comparison to bulk sequencing, and manuscript write-up.
- Hands-on Bioinformatics for Molecular Phylogeny: Training on phylogenetic tree construction and interpretation, for HIV subtype/clade assignation, was performed using bioinformatics tools, among which BioEdit v.5, MEGA v.5, SplitsTree v.4, Recombinant Detection Program v.3.44, and FigTree
- Analyses and interpretation of HIV drug resistance-associated mutations for clinical management of patients: This skill was acquired using samples processed for HIV-1 drug resistance testing (RNA extraction, RTPCR/Nested PCR, direct sequencing, analysis using Stanford HIV drug resistance database)
- Procedures for sequence submission into the public databank: this hands-on procedure was acquired through the submission of 80 HIV-1 polymerase (protease-reverse transcriptase) gene sequences into the GenBank, using BioEdit v.5 and Sequin softwares.
- Application of these skills into routine research and clinical activities in Cameroon;
- Exposure to other Next Gen Sequencers (Ion TorrentTM) in USA and participation to the New England Biolabs Molecular Biology summer workshop in June 2012;
- One-year Post-Doctorate Research Fellowship on the application of Next Generation Sequencing (P454) in Molecular Virology at the University of Rome, Tor Vergata.
- Collaboration between my home (CIRCB) and host (University of Rome Tor Vergata) institutions has also been strengthened.
I remain very grateful: (1) To the HIV Research Trust for sponsoring this training research project which has exposed me to a wider range of opportunities; (2) To the PRD (Poverty Related Disease) College for my travel and daily subsistence; (3) To the staff of the Virology Laboratory of the University of Rome “Tor Vergata”, especially my Supervisors (Prof Carlo-Federico Perno and Prof Vittorio Colizzi) and training Coordinators (Ass. Prof Francesca Ceccherini-Silberstein and Dr Maria Santoro), for hosting the project, for knowledge and skills transfer, and for offering me a PostDoctoral Research Fellow position; (4) To my Wife (Fokam Rita Mejane) and my Daughter (Giulia Louisiana) for also their constant encouragements and supports.