Researchers from the BMGF project took part in capacity building on next-generation metagenomic sequencing (mNGS) and the use of the CZID platform for bioinformatics analysis in San Francisco.
Training on next-generation metagenomic sequencing (mNGS) and the use of the CZID platform for bioinformatics analysis was held at the Chan Zuckenberg Biohub in San Francisco, California, from 17 June to 03 July 2023. Participants from various countries attended, including Dr Amoikon Tiemélé Laurent-Simon, research assistant at the Centre Suisse de Recherches Scientifiques en Côte d'Ivoire (CSRS), and Dr Diplo Tchepe Flore Bernadette, associate researcher at the Institut Pasteur de Côte d'Ivoire (IPCI).
The first week of training was devoted to laboratory activities. These activities were marked by three theory classes. The first dealt with how to progress in a molecular biology laboratory, and the usefulness of SPRI (Solid Phase Reversible Immobilisation) beads in mNGS manipulations. The second covered the design of a study plan for metagenomic analysis, from sample collection to library preparation, with particular emphasis on the removal of human nucleic acids from total nucleic acid samples extracted from cerebrospinal fluid (CSF). And the last on the basic principles of Illumina technology.
The practical part of the training took place in the laboratory. It consisted of (i) removing human nucleic acids from CSF nucleic acid extracts, (ii) cleaning RNA samples with SPRI beads, (iii) making the RNA library, (iv) cleaning the RNA library of primer dimers, (v) combining the library into a pool when library quantities were low, (vi) selecting nucleic acids according to their size from SPRI beads, (vii) preparing a solution of SPRI beads from SPRI bead powder at the desired concentration, (viii) performing Kappa amplification, (ix) loading the library onto an illumina Miseq sequencer and performing quality control (QC) of the libraries using the Tapestation and fragment analyzers, agarose gel electrophoresis and Qubit. Analysis of the results obtained was carried out at the end of each practical activity.
The week of 26-30 June was devoted to bioinformatics analyses. These activities were marked by (i) an introduction to the use of the CZID platform https://czid.org , (ii) the preparation of metadata, (iii) the demultiplexing of sequences from the Illumina platform using BaseSpace https://basespace.illumina.com, (iv) the loading of sequences onto CZID, (v) the interpretation of the results obtained and the search for RAMs (antibiotic resistance genes). Finally, an exchange on ONT (Oxford Nanopore technology) took place between the trainer and the face-to-face participants and CSRS researchers online via Zoom.
In addition to these training days, the Côte d'Ivoire team, warmly welcomed by CZ Biohub, was officially presented to the various scientists and students present, and met the President of CZ Biohub, Professor Joe DeRisi, Dr Paul Lebel and his team of bioengineers, Dr Abigail Glascock, bioinformatics researcher and Professor John Pak, head of the "Protein Sciences" group. During his presentation to all the participants, Dr Amoikon also had the opportunity to talk about the CSRS and its activities, the research team led by Dr Diallo Kanny, principal investigator of the BMGF project, and the BMGF project for which the training course was organised.
"The training objectives were achieved. The trainers reaffirmed their availability if we had any misunderstandings or concerns. We met scientists with whom we could collaborate. The remaining CSF samples will be sequenced and the sequences will be shared with the team with a view to continuing the training on bioinformatics analysis, but this time online", says Dr Amoikon Tiemele Simon.
