Rapid Nanopore Whole-Genome Sequencing for Anthrax Emergency Preparedness

Abstract

Human anthrax cases necessitate rapid response. We completed Bacillus anthracis nanopore whole-genome sequencing in our high-containment laboratory from a human anthrax isolate hours after receipt. The de novo assembled genome showed no evidence of known antimicrobial resistance genes or introduced plasmid(s). Same-day genomic characterization enhances public health emergency response.

Keywords: Bacillus anthracis; anthrax; bacteria; bioterrorism and preparedness; emergency preparedness; nanopore; whole-genome sequencing; zoonoses.

Figure 1

Time required to detect antimicrobial resistance markers in Bacillus anthracis strain Ba0914 by using WGS and summary of assembly results. A) Comparison of time to complete rapid nanopore (MinION) and short-read (iSeq) sequencing laboratory workflows. Workflows include DNA extraction, library preparation, WGS, and bioinformatics analysis. B) Comparison of nanopore-based and short-read sequencing–based data used to assemble the B. anthracis chromosome and plasmid sequences and to detect known AMR mutations and genes. Mutations associated with fluoroquinolone resistance in B. anthracis are located within the quinolone resistance–determining regions of gyrA, gyrB, parC, and parE genes. AMR genes contained in the Resfinder database (https://cge.cbs.dtu.dk) were queried against the assemblies. The rsiP mutation associated with penicillin resistance was not included. The nanopore assembly was generated by using the first 120,000 basecalled reads. AMR, antimicrobial resistance; WGS, whole-genome sequencing.

Figure 2

Circular maps of the whole-genome–sequenced Bacillus anthracis Ba0914 chromosome and 2 plasmids, pXO1 and pXO2, assembled by using rapid nanopore sequencing and short-read sequencing. (Maps are not to scale.)