Whole-genome sequencing as an improved means of investigating Neisseria gonorrhoeae treatment failures.

Background Although rare, Neisseria gonorrhoeae treatment failures associated with ceftriaxone have been reported. The World Health Organization (WHO) recommends standardised protocols to verify these cases. Two cases from Australia were previously investigated using N. gonorrhoeae multiantigen sequence typing (NG-MAST), which has been used extensively to assess treatment failures. Case 1 pharyngeal isolates were indistinguishable, whereas Case 2 pharyngeal isolates were distinguished based on an 18-bp deletion in the major outer membrane porin encoded by the porB gene, questioning the reliability of NG-MAST results. Here we used whole-genome sequencing (WGS) to reinvestigate Cases 1 and 2, with a view to examining WGS to assess treatment failures. METHODS:Pre- and post-treatment isolates for each case underwent Illumina sequencing, and the two post-treatment isolates underwent additional long-read sequencing using Pacific Biosciences. Sequence data were interrogated to identify differences at single nucleotide resolution. RESULTS:WGS identified variation in the pilin subunit encoded by the pilE locus for both cases and the specific 18-bp porB deletion in Case 2 was confirmed, but otherwise the isolates in each case were indistinguishable. CONCLUSIONS:The WHO recommends standardised protocols for verifying N. gonorrhoeae treatment failures. Case 2 highlights the enhanced resolution of WGS over NG-MAST and emphasises the immediate effect that WGS can have in a direct clinical application for N. gonorrhoeae. Assessing the whole genome compared with two highly variable regions also provides a more confident predictor for determining treatment failure. Furthermore, WGS facilitates rapid comparisons of these cases in the future.