Computational protocol: In-host microevolution of Aspergillus fumigatus: A phenotypic and genotypic analysis

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Protocol publication

[…] Raw reads were quality checked using FastQC (version 0.11.5, Babraham Institute). Reads containing adapter sequences and/or with a Phred score <30 were removed using Trimmomatic (Galaxy version 0.32.3) (, ). Reads were mapped to the Af293 reference genome (release 31, EnsemblFungi) using the very sensitive local align preset mode in Bowtie2 (). Mapping quality was assessed using Qualimap (, ). Single nucleotide polymorphism (SNP) detection was conducted using FreeBayes (). VCFtools vcf-isec was used to assess patterns amongst SNPs and to filter SNPs with a minimum coverage of 5 and a minimum probability of 0.8 (). EnsemblFungi Variant Effect Predictor was used to assess the impact of non-synonymous SNPs (). Both synonymous and non-synonymous SNPs were considered for phylogenetic analysis using the SNPhylo pipeline (), which utilises vcftools (), Phylip (University of Washington, USA) and Muscle () to generate phylogenetic trees by the maximum likelihood method. Integrated Genomics Viewer and Tablet were utilised for visualisation of sequence data (, , ). […]

Pipeline specifications

Software tools FastQC, Trimmomatic, Galaxy, Bowtie2, QualiMap, FreeBayes, VCFtools, PHYLIP, MUSCLE, Tablet
Applications Phylogenetics, Nucleotide sequence alignment
Organisms Aspergillus fumigatus, Homo sapiens, Fungi
Diseases Infection
Chemicals Itraconazole, Voriconazole