Ten years of external quality assessment (EQA) of Neisseria gonorrhoeae antimicrobial susceptibility testing in Europe elucidate high reliability of data.

BACKGROUND: Confidence in any diagnostic and antimicrobial susceptibility testing data is provided by appropriate and regular quality assurance (QA) procedures. In Europe, the European Gonococcal Antimicrobial Susceptibility Programme (Euro-GASP) has been monitoring the antimicrobial susceptibility in Neisseria gonorrhoeae since 2004. Euro-GASP includes an external quality assessment (EQA) scheme as an essential component for a quality-assured laboratory-based surveillance programme. Participation in the EQA scheme enables any problems with the performed antimicrobial susceptibility testing to be identified and addressed, feeds into the curricula of laboratory training organised by the Euro-GASP network, and assesses the capacity of individual laboratories to detect emerging new, rare and increasing antimicrobial resistance phenotypes. Participant performance in the Euro-GASP EQA scheme over a 10 year period (2007 to 2016, no EQA in 2013) was evaluated. METHODS: Antimicrobial susceptibility category and MIC results from the first 5 years (2007-2011) of the Euro-GASP EQA were compared with the latter 5 years (2012-2016). These time periods were selected to assess the impact of the 2012 European Union case definitions for the reporting of antimicrobial susceptibility. RESULTS: Antimicrobial susceptibility category agreement in each year was ≥91%. Discrepancies in susceptibility categories were generally because the MICs for EQA panel isolates were on or very close to the susceptibility or resistance breakpoints. A high proportion of isolates tested over the 10 years were within one (≥90%) or two (≥97%) MIC log2 dilutions of the modal MIC, respectively. The most common method used was Etest on GC agar base. There was a shift to using breakpoints published by the European Committee on Antimicrobial Susceptibility Testing (EUCAST) in the latter 5 years, however overall impact on the validity of results was limited, as the percentage categorical agreement and MIC concordance changed very little between the two five-year periods. CONCLUSIONS: The high level of comparability of results in this EQA scheme indicates that high quality data are produced by the Euro-GASP participants and gives confidence in susceptibility and resistance data generated by laboratories performing decentralised testing.

WGS analysis and molecular resistance mechanisms of azithromycin-resistant (MIC >2 mg/L) Neisseria gonorrhoeae isolates in Europe from 2009 to 2014.

OBJECTIVES: To elucidate the genome-based epidemiology and phylogenomics of azithromycin-resistant (MIC >2 mg/L) Neisseria gonorrhoeae strains collected in 2009-14 in Europe and clarify the azithromycin resistance mechanisms. METHODS: Seventy-five azithromycin-resistant (MIC 4 to >256 mg/L) N. gonorrhoeae isolates collected in 17 European countries during 2009-14 were examined using antimicrobial susceptibility testing and WGS. RESULTS: Thirty-six N. gonorrhoeae multi-antigen sequence typing STs and five phylogenomic clades, including 4-22 isolates from several countries per clade, were identified. The azithromycin target mutation A2059G (Escherichia coli numbering) was found in all four alleles of the 23S rRNA gene in all isolates with high-level azithromycin resistance (n = 4; MIC ≥256 mg/L). The C2611T mutation was identified in two to four alleles of the 23S rRNA gene in the remaining 71 isolates. Mutations in mtrR and its promoter were identified in 43 isolates, comprising isolates within the whole azithromycin MIC range. No mutations associated with azithromycin resistance were found in the rplD gene or the rplV gene and none of the macrolide resistance-associated genes [mef(A/E), ere(A), ere(B), erm(A), erm(B), erm(C) and erm(F)] were identified in any isolate. CONCLUSIONS: Clonal spread of relatively few N. gonorrhoeae strains accounts for the majority of the azithromycin resistance (MIC >2 mg/L) in Europe. The four isolates with high-level resistance to azithromycin (MIC ≥256 mg/L) were widely separated in the phylogenomic tree and did not belong to any of the main clades. The main azithromycin resistance mechanisms were the A2059G mutation (high-level resistance) and the C2611T mutation (low- and moderate-level resistance) in the 23S rRNA gene.