Complete genome assembly of clinical multidrug resistant Bacteroides fragilis isolates enables comprehensive identification of antimicrobial resistance genes and plasmids

constitutes a significant part of the normal human gut microbiota and can also act as an opportunistic pathogen. Antimicrobial resistance and the prevalence of antimicrobial resistance genes are increasing, and prediction of antimicrobial susceptibility based on sequence information could support targeted antimicrobial therapy in a clinical setting. Complete identification of insertion sequence (IS) elements carrying promoter sequences upstream of resistance genes is necessary for prediction of antimicrobial resistance. However, assemblies from short reads alone are often fractured due to repeat regions and the presence multiple copies of identical IS elements. Identification of plasmids in clinical isolates can aid in the surveillance of the dissemination of antimicrobial resistance and comprehensive sequence databases support microbiome and metagenomic studies. Here we test several short-read, hybrid and long-lead assembly pipelines by assembling the type strain CCUG4856T (=ATCC25285=NCTC9343) with Illumina short reads and long reads generated by Oxford Nanopore Technologies (ONT) MinION sequencing. Hybrid assembly with Unicycler, using quality filtered Illumina reads and Filtlong filtered and Canu corrected ONT reads produced the assembly of highest quality. This approach was then applied to six clinical multidrug resistant isolates and, with minimal manual finishing of chromosomal assemblies of three isolates, complete, circular assemblies of all isolates were produced. Eleven circular, putative plasmids were identified in the six assemblies of which only three corresponded to a known cultured plasmid. Complete IS elements could be identified upstream of antimicrobial resistance genes, however there was not complete correlation between the absence of IS elements and antimicrobial susceptibility. As our knowledge on factors that increase expression of resistance genes in the absence of IS elements is limited, further research is needed prior to implementing antimicrobial resistance prediction for from whole genome sequencing.