Neisseria gonorrhoeae antimicrobial resistance (AMR) monthly UPDATES - April, 2019

Neisseria gonorrhoeae antimicrobial resistance (AMR) monthly UPDATES

April, 2019

Reviews and Commentary

Deciphering the impact of bystander selection for antibiotic resistance in Neisseria gonorrhoeae.

Olesen SW, Grad YH. (Full Text)

J Infect Dis. 2019 Apr 8. pii: jiz156. doi: 10.1093/infdis/jiz156. [Epub ahead of print]


Epidemiology and Surveillance

Summary and Trends of the Russian Gonococcal Antimicrobial Surveillance Programme, 2005-2016.

Kubanov A, Solomka V, Plakhova X, Chestkov A, Petrova N, Shaskolskiy B, Dementieva E, Leinsoo A, Gryadunov D, Deryabin D. (Full Text)

J Clin Microbiol. 2019 Mar 20. pii: JCM.02024-18. doi: 10.1128/JCM.02024-18. [Epub ahead of print]


The Russian Gonococcal Antimicrobial Surveillance Programme (RU-GASP) was established in 2004 and operated continuously during the years 2005-2016. The aims of this study were to summarize the RU-GASP results over this 12-year period and evaluate the trends in Neisseria gonorrhoeae antimicrobial resistance in Russia. In total, 5,038 verified N. gonorrhoeae isolates from 40 participating regions were tested for susceptibility to six antimicrobials via an agar dilution method. DNA loci involved in antimicrobial resistance were identified via minisequencing or DNA microarray techniques. From 2005 to 2016, increasing susceptibility to penicillin G (from 22.6% to 63.0%), tetracycline (from 34.8% to 53.0%), and ciprofloxacin (from 50.6% to 68.6%) was observed, but resistance to these drugs remained high. The proportions of isolates nonsusceptible to azithromycin and spectinomycin peaked in 2011 and decreased thereafter. Of the isolates, only 6 and 23 were identified as nonsusceptible to ceftriaxone according to the CLSI definitions and EUCAST breakpoint (0.57% of the total population), respectively. Comparison of N. gonorrhoeae antimicrobial resistance genetic determinants in 2005 vs 2016 showed a significant decrease in the number of isolates carrying chromosomal mutations. The proportion of isolates with wild-type genotypes increased from 11.7% in 2005 to 30.3% in 2016. Thus, the RU-GASP can be considered a successful gonorrhea surveillance program, and the current state of N. gonorrhoeae antimicrobial resistance in Russia is less serious than that in other WHO GASP regions.

Detection in the United Kingdom of the Neisseria gonorrhoeae FC428 clone, with ceftriaxone resistance and intermediate resistance to azithromycin, October to December 2018.

Eyre DW, Town K, Street T, Barker L, Sanderson N, Cole MJ, Mohammed H, Pitt R, Gobin M, Irish C, Gardiner D, Sedgwick J, Beck C, Saunders J, Turbitt D, Cook C, Phin N, Nathan B, Horner P, Fifer H. (Full Text)

Euro Surveill. 2019 Mar;24(10). doi: 10.2807/1560-7917.ES.2019.24.10.1900147.


We describe detection in the United Kingdom (UK) of the drug-resistant Neisseria gonorrhoeae FC428 clone, with ceftriaxone resistance and intermediate azithromycin resistance. Two female patients developed infection following contact with UK-resident men from the same sexual network linked to travel to Ibiza, Spain. One case failed treatment with ceftriaxone, and azithromycin and gentamicin, before successful treatment with ertapenem. Both isolates had indistinguishable whole-genome sequences. Urgent action is essential to contain this drug-resistant strain.

First Case of Ceftriaxone-Resistant Multidrug-Resistant Neisseria gonorrhoeae in Singapore.

Ko KKK, Chio MT, Goh SS, Tan AL, Koh TH, Abdul Rahman NB. (Full Text)

Antimicrob Agents Chemother. 2019 Mar 11. pii: AAC.02624-18. doi: 10.1128/AAC.02624-18. [Epub ahead of print]


Amid the global crisis of increasing gonococcal antimicrobial resistance, we report the first case of ceftriaxone-resistant multidrug-resistant Neisseria gonorrhoeae in Singapore...

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

Cole MJ, Quaye N, Jacobsson S, Day M, Fagan E, Ison C, Pitt R, Seaton S, Woodford N, Stary A, Pleininger S, Crucitti T, Hunjak B, Maikanti P, Hoffmann S, Viktorova J, Buder S, Kohl P, Tzelepi E, Siatravani E, Balla E, Hauksdóttir GS, Rose L, Stefanelli P, Carannante A, Pakarna G, Mifsud F, Cassar RZ, Linde I, Bergheim T, Steinbakk M, Mlynarczyk-Bonikowska B, Borrego MJ, Shepherd J, Pavlik P, Jeverica S, Vazquez J, Abad R, Weiss S, Spiteri G, Unemo M. (Full Text)

BMC Infect Dis. 2019 Mar 25;19(1):281. doi: 10.1186/s12879-019-3900-z.



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.


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.


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.


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.

Antimicrobial susceptibility of Neisseria gonorrhoeae isolates and treatment of gonorrhoea patients in Ternopil and Dnipropetrovsk regions of Ukraine, 2013-2018.

Boiko I, Golparian D, Krynytska I, Bezkorovaina H, Frankenberg A, Onuchyna M, Jacobsson S, Unemo M. (Full Text)

APMIS. 2019 Mar 23. doi: 10.1111/apm.12948. [Epub ahead of print]


Antimicrobial resistance (AMR) in Neisseria gonorrhoeae is a major public health concern globally. However, recent gonococcal AMR data from Eastern Europe are extremely limited and no AMR data for strains spreading in Ukraine has ever been internationally published. We investigated the AMR of N. gonorrhoeae isolates in two regions of Ukraine (Ternopil 2013-2018, Dnipropetrovsk 2013-2014), and, where information was available, the treatment administered to the corresponding gonorrhoea patients. Determination of minimum inhibitory concentration (MIC) of eight antimicrobials was performed using Etest and resistance breakpoints from the EUCAST were applied. Overall, 9.3% of the examined 150 isolates were resistant to ciprofloxacin, 6.0% to tetracycline, 2.0% to azithromycin, and 0.7% to benzylpenicillin. No isolates were resistant to ceftriaxone, cefixime, spectinomycin, or gentamicin. However, one (0.7%) isolate showed a MIC value of 0.125 mg/L for both ceftriaxone and cefixime, i.e. bordering resistance. Eighty-eight (67.2%) of 131 patients were administered dual therapy (ceftriaxone 1 g plus doxycycline/clarithromycin/azithromycin/ofloxacin) and 22 (16.8%) ceftriaxone 1 g monotherapy. Worryingly, 21 (16.0%) patients received monotherapy with clarithromycin/doxycycline/azithromycin/ofloxacin/benzylpenicillin. In conclusion, the antimicrobial susceptibility of gonococcal strains spreading in Ternopil and Dnipropetrovsk, Ukraine during 2013-2018 was high. Low levels of resistance to ciprofloxacin, tetracycline, azithromycin, and benzylpenicillin were found, but no resistance to the internationally recommended ceftriaxone, cefixime, or spectinomycin. Ceftriaxone 1 g should remain as empiric first-line treatment, in dual therapy with azithromycin or doxycycline or in monotherapy. Continued and expanded gonococcal AMR surveillance in Ukraine is essential to monitor the susceptibility to particularly extended-spectrum cephalosporins, azithromycin and doxycycline. This article is protected by copyright. All rights reserved.


Identification of Internationally Disseminated Ceftriaxone-Resistant Neisseria gonorrhoeae Strain FC428, China.

Chen SC, Han Y, Yuan LF, Zhu XY, Yin YP. (Full Text)

Emerg Infect Dis. 2019 Jul 17;25(7). doi: 10.3201/eid2507.190172. [Epub ahead of print]


In 2016, we identified a ceftriaxone-resistant Neisseria gonorrhoeae isolate in China. The strain genotype was identical to the resistant clone FC428 that originated in Japan. Enhanced international collaborative surveillance programs are crucial to track the transmission of the ceftriaxone-resistant clones.

Population-level antimicrobial consumption is associated with decreased antimicrobial susceptibility in Neisseria gonorrhoeae in 24 European countries: an ecological analysis.

Kenyon C, Buyze J, Spiteri G, Cole MJ, Unemo M. (Full Text)

J Infect Dis. 2019 Apr 8. pii: jiz153. doi: 10.1093/infdis/jiz153. [Epub ahead of print]



There are substantial variations in Neisseria gonorrhoeae susceptibility to antimicrobials between different populations, and the reasons for this are largely unexplored. We aimed to assess if the population level consumption of antimicrobials is a contributory factor.


Using antimicrobial susceptibility data from 24 countries in the European Gonococcal Antimicrobial Surveillance Programme and antimicrobial consumption data from the IQVIA MIDAS database, we built mixed effects linear/logistic regression models with country-level cephalosporin, fluoroquinolone and macrolide consumption (standard doses/1000 population/year) as the explanatory variables (from 2009 to 2015) and 1-year lagged ceftriaxone, cefixime, azithromycin and ciprofloxacin geometric mean minimum inhibitory concentrations (MIC) as the outcome variables (2010 to 2016).


Positive correlations were found between the consumption of cephalosporins and geometric mean MIC of ceftriaxone and cefixime (both P's <0.05). Fluoroquinolone consumption was positively associated with the prevalence of resistance to ciprofloxacin (P<0.05).


Differences in population level consumption of particular antimicrobials may contribute to the variations in the level of antimicrobial resistance in N. gonorrhoeae in different settings. Further interventions to reduce misuse and overuse of antimicrobials in high-consumption populations and core-groups are required.

Systematic review and survey of Neisseria gonorrhoeae ceftriaxone and azithromycin susceptibility data in the Asia Pacific, 2011 to 2016.

George CRR, Enriquez RP, Gatus BJ, Whiley DM, Lo YR, Ishikawa N, Wi T, Lahra MM. (Full Text)

PLoS One. 2019 Apr 3;14(4):e0213312. doi: 10.1371/journal.pone.0213312. eCollection 2019.



Antimicrobial resistance in Neisseria gonorrhoeae is a global concern, with the ongoing emergence of ceftriaxone and azithromycin resistance threatening current treatment paradigms. To monitor the emergence of antimicrobial resistance in N. gonorrhoeae, the World Health Organization (WHO) Gonococcal Antimicrobial Surveillance Programme (GASP) has operated in the Western Pacific and South East Asian regions since 1992. The true burden of antimicrobial resistance remains unknown. In response, the objective of this study was to survey ceftriaxone and azithromycin susceptibility in N. gonorrhoeae across the western Pacific and south-east Asia, and interlink this data with systematically reviewed reports of ceftriaxone and azithromycin resistance.


The WHO Collaborating Centre for Sexually Transmitted Infections and Antimicrobial Resistance, Sydney, coordinated annual surveys of gonococcal susceptibilities with participating laboratories, and additionally undertook a systematic review of reports detailing gonococcal ceftriaxone and azithromycin susceptibility data for locations geographically in the Asia Pacific from 2011 to 2016. It was found that surveillance of gonococcal antimicrobial resistance remains limited in the Asia Pacific, with weaker surveillance of azithromycin versus ceftriaxone. Ninety-three published reports were identified (including national reports) which documented susceptibility data for ceftriaxone and azithromycin. GASP survey data was available for 21 countries, territories or areas, and suggested MICs are increasing for ceftriaxone and azithromycin. Between 2011 and 2016, the percentage of locations reporting >5% of gonococcal isolates with MICs to ceftriaxone meeting WHO's definition of decreased susceptibility (MIC ≥ 0.125 mg/L) increased from 14.3% to 35.3% and the percentage of locations reporting >5% of gonococcal isolates with azithromycin resistance (MIC ≥ 1 mg/L) increased from 14.3% to 38.9%. Published reports were available for several countries that did not provide GASP surveillance responses for ceftriaxone (n = 5) and azithromycin (n = 3) respectively. Over the study period, there was a 183% increase in the number of countries providing surveillance data for GASP for both ceftriaxone and azithromycin, and a 30.6% increase in ceftriaxone MIC testing across the Asia Pacific facilitated by this project.


This study provides the first comprehensive illustration of increasing MICs to ceftriaxone in the Asia Pacific. The survey and literature review additionally detail increasing resistance to azithromycin. Further surveillance system strengthening is required to monitor these trends in order to address and curb gonococcal AMR in the region.

Emergence of Neisseria gonorrhoeae Strains Harboring a Novel Combination of Azithromycin-Attenuating Mutations.

Pham CD, Sharpe S, Schlanger K, St Cyr S, Holderman J, Steece R, Soge OO, Masinde G, Arno J, Schmerer M, Kersh EN; SURRG Working Group. (Full Text)

Antimicrob Agents Chemother. 2019 Mar 27;63(4). pii: e02313-18. doi: 10.1128/AAC.02313-18. Print 2019 Apr.


The nimbleness of Neisseria gonorrhoeae to evade the effect of antibiotics has perpetuated the fight against antibiotic-resistant gonorrhea for more than 80 years. The ability to develop resistance to antibiotics is attributable to its indiscriminate nature in accepting and integrating exogenous DNA into its genome. Here, we provide data demonstrating a novel combination of the 23S rRNA A2059G mutation with a mosaic-multiple transferable resistance (mosaic-mtr) locus haplotype in 14 N. gonorrhoeae isolates with high-level azithromycin MICs (≥256 μg/ml), a combination that may confer more fitness than in previously identified isolates with high-level azithromycin resistance. To our knowledge, this is the first description of N. gonorrhoeae strains harboring this novel combination of resistance determinants. These strains were isolated at two independent jurisdictions participating in the Gonococcal Isolate Surveillance Project (GISP) and in the Strengthening the U.S. Response to Resistant Gonorrhea (SURRG) project. The data suggest that the genome of N. gonorrhoeae continues to shuffle its genetic material. These findings further illuminate the genomic plasticity of N. gonorrhoeae, which allows this pathogen to develop mutations to escape the inhibitory effects of antibiotics.


Novel detection strategies and diagnostics

Evaluation of the ResistancePlus GC (beta) assay: a commercial diagnostic test for the direct detection of ciprofloxacin susceptibility or resistance in Neisseria gonorrhoeae.

Ebeyan S, Windsor M, Bordin A, Mhango L, Erskine S, Trembizki E, Mokany E, Tan LY, Whiley D, GRAND2 Study Investigators. (Full Text)

J Antimicrob Chemother. 2019 Mar 20. pii: dkz108. doi: 10.1093/jac/dkz108. [Epub ahead of print]



To evaluate the performance of the ResistancePlus GC (beta) assay for the simultaneous detection of Neisseria gonorrhoeae and gyrA S91 markers of resistance (S91F) and susceptibility (WT) to ciprofloxacin, from both clinical specimens and isolates.


Performance was assessed on several sample banks, including N. gonorrhoeae isolates (n = 822), non-gonococcal isolates (n = 110), N. gonorrhoeae-positive clinical specimens (n = 402) and N. gonorrhoeae-negative specimens (n = 290). Results were compared with previous testing data, including S91 genotyping and phenotypic resistance profiles.


Overall, the assay demonstrated 100% sensitivity for N. gonorrhoeae detection in clinical isolates. For gyrA S91 mutation detection in clinical isolates, the assay showed 100% sensitivity/specificity compared with the genotype, and >99%/>97% sensitivity/specificity when compared with phenotype. For positive clinical specimens, the assay demonstrated >96% sensitivity for N. gonorrhoeae detection and 100% sensitivity/specificity for gyrA S91 mutation detection. The assay demonstrated >99% specificity for N. gonorrhoeae detection against non-gonococcal isolates and 100% specificity for negative clinical specimens.


The ResistancePlus GC (beta) assay is suitable for the detection of N. gonorrhoeae and gyrA markers associated with resistance/susceptibility to ciprofloxacin directly in clinical samples. This assay could be implemented for the individualized treatment of gonorrhoea infections as well as to enhance current antimicrobial resistance surveillance methods.


Novel antibiotics, mechanisms, vaccine, and other advances

Targeting Lipooligosaccharide (LOS) for a Gonococcal Vaccine.

Gulati S, Shaughnessy J, Ram S, Rice PA. (Full Text)

Front Immunol. 2019 Feb 27;10:321. doi: 10.3389/fimmu.2019.00321. eCollection 2019.


The increasing incidence of gonorrhea worldwide and the global spread of multidrug-resistant strains of Neisseria gonorrhoeae, constitute a public health emergency. With dwindling antibiotic treatment options, there is an urgent need to develop safe and effective vaccines. Gonococcal lipooligosaccharides (LOSs) are potential vaccine candidates because they are densely represented on the bacterial surface and are readily accessible as targets of adaptive immunity. Less well-understood is whether LOSs evoke protective immune responses. Although gonococcal LOS-derived oligosaccharides (OSs) are major immune targets, often they undergo phase variation, a feature that seemingly makes LOS less desirable as a vaccine candidate. However, the identification of a gonococcal LOS-derived OS epitope, called 2C7, that is: (i) a broadly expressed gonococcal antigenic target in human infection; (ii) a virulence determinant, that is maintained by the gonococcus and (iii) a critical requirement for gonococcal colonization in the experimental setting, circumvents its limitation as a potential vaccine candidate imposed by phase variation. Difficulties in purifying structurally intact OSs from LOSs led to "conversion" of the 2C7 epitope into a peptide mimic that elicited cross-reactive IgG anti-OS antibodies that also possess complement-dependent bactericidal activity against gonococci. Mice immunized with the 2C7 peptide mimic clear vaginal colonization more rapidly and reduce gonococcal burdens. 2C7 vaccine satisfies criteria that are desirable in a gonococcal vaccine candidate: broad representation of the antigenic target, service as a virulence determinant that is also critical for organism survival in vivo and elicitation of broadly cross-reactive IgG bactericidal antibodies when used as an immunogen.

In vitro assessment of gentamicin and azithromycin-based combination therapy against Neisseria gonorrhoeae isolates in India.

Sood S, Agarwal SK, Singh R, Gupta S, Sharma VK. (Full Text)

J Med Microbiol. 2019 Mar 14. doi: 10.1099/jmm.0.000953. [Epub ahead of print]



The public health burden of infections caused by Neisseria gonorrhoeae is magnified due to high rates of resistance to traditional antimicrobials. The aim of this study was to evaluate the in vitro efficacy of an alternative dual therapy comprising gentamicin and azithromycin.


The E-test method was used to determine the minimum inhibitory concentrations (MICs) of gentamicin and azithromycin individually prior to testing in combination using the cross or 90o angle formation method. A total of 70 clinical isolates of N.gonorrhoeae displaying varying ceftriaxone MICs along with 2 reference strains (WHO K and P) and 1 ceftriaxone-resistant QA isolate were examined. The fractional inhibitory concentration index (FICI) was calculated and the results were interpreted using the following criteria: synergy, FICI ≤0.5; indifference or additive, FICI >0.5 to ≤4.0; and antagonism, FICI >4.0.


A total of 54 (77.1 %) isolates displayed indifference, while 16 (22.9 %) demonstrated synergy. When azithromycin was tested alone, the MICs ranged from 0.016 to 2 µg ml-1 . However, in combination with gentamicin, the mean MIC value of all isolates decreased from 0.275 µg ml-1 to 0.090 µg ml-1 (P=0.05).When gentamicin was tested alone, the MICs ranged from 0.25 to 8 µg ml-1, with a mean MIC of 4.342 µg ml-1, whereas in combination with azithromycin it decreased significantly to 2.042 µg ml-1 (P=0.04).


No antagonism was observed in this combination, suggesting that it could be a future treatment option as we prepare for a post-cephalosporin era. However, comprehensive in vivo evaluations are warranted and recommendations should be made based on clinical trials.

Development of flow cytometry based adherence assay for Neisseria gonorrhoeae using 5'-carboxyfluorosceinsuccidyl ester.

Thakur SD, Obradovic M, Dillon JR, Ng SH, Wilson HL. (Full Text)

BMC Microbiol. 2019 Mar 25;19(1):67. doi: 10.1186/s12866-019-1438-2.



Neisseria gonorrhoeae is an obligate human pathogen and its adherence to host cells is essential for its pathogenesis. Gonococcal adherence assays are based on the enumeration of bacteria attached to human cells on solid media. Because conventional adherence assays are based on bacterial counts, they are often time consuming to perform and prone to observer bias. A flow cytometry based method, using the cell-permeable fluorescent dye 5'-carboxyfluoroscein succidyl ester (CFSE), was developed to dramatically increase the number of adherent N. gonorrhoeae quantified per assay while improving repeatability and removing observer bias. Piliated N. gonorrhoeae F62 were stained with CFSE then the staining reaction was quenched with foetal bovine serum. Human cervical ME-180 cells were infected with CFSE-stained N. gonorrhoeae (multiplicity of the infection 100:1) for 2 h. Infected cells were washed to remove loosely adhered bacteria. Flow cytometry was used to quantify the percentage of ME-180 cells associated with CFSE-stained N. gonorrhoeae and a minimum of 30,000 events were recorded. Real time-PCR analysis targeting opa gene (encoding N. gonorrhoeae opacity associated gonococcal outer membrane protein) was performed on infected ME-180 cells to confirm the flow cytometric adherence assay results. A rabbit was immunized with heat-killed N. gonorrhoeae F62 to generate hyperimmune serum. The functional compatibility of the assay was confirmed by studying the effect of N. gonorrhoeae F62 antiserum on blocking adherence/invasion of CFSE-stained bacteria to ME-180 cells.


We observed that 20.3% (+/- 1.0) ME-180 cells were associated with CFSE-stained N. gonorrhoeae. Heat-inactivated hyperimmune serum, at 1:10 to 1:80 dilutions, significantly inhibited gonococcal adherence by 6 and 3 fold, respectively. Real time-PCR analysis targeting opa gene confirmed that hyperimmune serum blocked adherence/invasion of N. gonorrhoeae to the ME-180 cells in a dilution-dependent manner.


Flow cytometric analysis was amenable to quick, easy and high-throughput quantification of the association of N. gonorrhoeae with ME-180 cells and was functionally confirmed using PCR analysis. These approaches may be adapted for in vitro and in vivo adherence studies related to gonococcal pathogenesis.