Report on the Swiss-Colombian workshop: “Assembly, annotation and comparison of bacterial genomes”
Falquet et al. (2016) EMBnet.journal 22, e872 http://dx.doi.org/10.14806/ej.22.0.872
Received 29 August 2016; Published 9 November 2016
This workshop was organised as part of the SwissColombian project, A pilot integrative knowledgebase for the characterization and tracking of multi resistant Acinetobacter baumannii in Colombian Hospitals, sponsored by the Leading House Cooperation and Development Centre of the École Polytechnique Fédérale Lausanne (EPFL).
The aim of this project is to develop a prototype centralised knowledgebase. Initially, we selected complete genome sequences obtained from a collection of Acinetobacter baumannii strains collected from the Antimicrobial-Resistant Healthcare-Associated Infections Surveillance Program, during 2012–2015, by the Colombian National Health Institute (NHI) and the Biotechnology Institute of the National University of Colombia (IBUN-UNAL). In addition, complete Acinetobacter baumannii genome sequences were added from public databases. The prototype will consist of fully assembled and annotated genomes associated with geographical, temporal and clinical data, allowing tracking of a variety of infection outbreaks. The resulting knowledgebase will serve as a reference to help clinicians to track rapid dissemination of highly pathogenic and resistant strains.
The workshop was held at the Bioinformatics centre of the National University of Bogotá, 23-27 May 2016, and gathered 18 participants from diverse institutions in Colombia
Each half-day was split into theoretical lectures (60-90 minutes each), followed by hands-on practicals (150 minutes each). The programme is shown in Table 1.
|Day 1||Introduction to UNIX and computer clusters Introduction to sequencing techniques, QC and data cleaning (adapter removal, trimming, filtering, etc.)|
|Day 2||De novo assembly
Assembly by re-mapping
|Day 3||SNP and small indel calling: how to detect variants?
Annotation and profi ling of resistance and virulence factors
|Day 4||Comparative genomics (core/pan genomes, structural variants, phylogeny distribution)|
|Day 5||Presentation of individual research projects of participants|
Figure 1. Participants and trainers.
Organisation of the work
Given the lack of access to a high-performance cluster, the participants were divided into nine groups of two, each being responsible for the analysis of a set of pairedend 100 bp reads from Illumina sequencing of a strain of Acinetobacter baumannii from the Sequence Read Archive (SRA). To distribute the workload, we divided the work across fi ve computing nodes (16 cores, 64 Gb RAM). After a brief reminder of computing and UNIX operating system basics, participants had the opportunity to refresh their knowledge of the command line. The genome analysis comprised data quality control with FastQC2, cleaning both the adapter content with CutAdapt (Martin, 2011) and low quality sequences with sickle3. The cleaned sequences were assembled with SOAPdenovo (Luo et al., 2012), using various kmers, and SPAdes (Bankevich et al., 2012). The draft genomes were compared using summary statistics, QUAST (Gurevich et al., 2013) and MAUVE (Darling et al., 2010). The best draft genome was annotated using Prokka (Seemann, 2014) and a set of HMMs built from the Virulence Factor database (Chen et al., 2016) and downloaded from the ResFam database (Gibson et al., 2015). The gff fi les of ten genomes (nine, plus reference) were compared, looking for core and pan genomes using Roary (Page et al., 2015) and Phandango4. The reads were also re-mapped to the reference genome, and SNPs and indels called with BWA (Li and Durbin, 2010), SAMtools and BCFtools (Li, 2011). The SNP vcf fi les were annotated with snpEff , fi ltered with SnpSift (Cingolani et al., 2012) and finally visualised with IGV (the orvaldsdottir et al., 2013). After conversion to multi-fasta format, a tree was constructed with FastTree (Price et al., 2010) and visualised using the Newick viewer (Boc et al., 2012).
Finally, participants had the opportunity to present their own current research work and to receive feedback from other course participants and trainers, promoting an enriching exchange of valuable research experiences in the area of genomics.
Figure 1. Participants and trainers.
Evaluation of the course
Participants from diff erent research institutions expressed satisfaction with the high academic level of the course in general. They gave high value to the knowledge shown by trainers, and to the materials used in the lectures and practical exercises. Some respondents said that knowledge acquired during the course had allowed them to solve their own data-analysis problems. They also made recommendations regarding the inclusion of additional practicals, and the possibility of additional access to the servers used for the hands-on sessions, in order to become more familiar with Linux. Course servers will be available to them for a few months more.
Figure 2. Workshop hand-on session.
According to the attendees’ course evaluation and the organisers comments, this workshop was very useful both for biologists working on assembly and annotation of bacterial genomes, and researchers of the Colombian NHI, interested in tracking resistance and virulence factors in clinical isolates.
This work was supported by the “Leading House Cooperation and Development Centre”. We thank Hermes Perez Cardona and Dra. Maria Teresa Reguero, for their help with local organisation of the workshop.
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