Comparison of MiSeq, MinION, and hybrid genome sequencing for analysis of Campylobacter jejuni

Abstract

Abstract The sequencing, assembly, and analysis of bacterial genomes is central to tracking and characterizing foodborne pathogens. The bulk of bacterial genome sequencing at the US Food and Drug Administration is performed using short-read Illumina MiSeq technology, resulting in highly accurate but fragmented genomic sequences. The MinION sequencer from Oxford Nanopore is an evolving technology that produces long-read sequencing data with low equipment cost. The goal of this study was to compare Campylobacter genome assemblies generated from MiSeq and MinION data independently, as well as hybrid genome assemblies combining both data types. Two reference strains and two field isolates of C. jejuni were sequenced using MiSeq and MinION, and the sequence data were assembled using the software programs SPAdes and Canu, respectively. Hybrid genome assembly was performed using the program Unicycler. Comparison of the C. jejuni 81-176 and RM1221 genome assemblies to the PacBio reference genomes revealed that the SPAdes assemblies had the most accurate nucleotide identity, while the hybrid assemblies were the most contiguous. Assemblies generated only from MinION data using Canu were the least accurate, containing many indels and substitutions that affected downstream analyses. The hybrid sequencing approach was the most useful for detecting plasmids, large genome rearrangements, and repetitive elements such as rRNA and tRNA genes. The full genomes of both C. jejuni field isolates were completed and circularized using hybrid sequencing, and a plasmid was detected in one isolate. Continued development of nanopore sequencing technologies will likely enhance the accuracy of hybrid genome assemblies and enable public health laboratories to routinely generate complete circularized bacterial genome sequences.