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  • 1.
    Boiko, Iryna
    et al.
    Ternopil Regional Clinical Dermatovenerologic Dispensary, Clinical Laboratory Department, Ternopil, Ukraine; Department of Functional and Laboratory Diagnostics, I. Horbachevsky Ternopil State Medical University, Ternopil, Ukraine; WHO Collaborating Centre for Gonorrhoea and other STIs, National Reference Laboratory for STIs, Department of Laboratory Medicine, Faculty of Medicine and Health, Örebro University, Örebro, Sweden.
    Golparian, Daniel
    Örebro University, School of Medical Sciences. WHO Collaborating Centre for Gonorrhoea and other STIs, National Reference Laboratory for STIs, Department of Laboratory Medicine.
    Krynsytska, Inna
    Department of Functional and Laboratory Diagnostics, I. Horbachevsky Ternopil State Medical University, Ternopil, Ukraine.
    Unemo, Magnus
    Örebro University, School of Medical Sciences. Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and other STIs, National Reference Laboratory for STIs, Department of Laboratory Medicine.
    High prevalence of Chlamydia trachomatis, Neisseria gonorrhoeae and particularly Trichomonas vaginalis diagnosed using US FDA-approved Aptima molecular tests and evaluation of conventional routine diagnostic tests in Ternopil, Ukraine2019In: Acta Pathologica, Microbiologica et Immunologica Scandinavica (APMIS), ISSN 0903-4641, E-ISSN 1600-0463, Vol. 127, no 9, p. 627-634Article in journal (Refereed)
    Abstract [en]

    Sexually transmitted infections (STIs) remain major public health problems globally. Appropriate laboratory diagnosis of STIs is rare in Ukraine. We investigated the prevalence of Chlamydia trachomatis (CT), Neisseria gonorrhoeae (NG) and Trichomonas vaginalis (TV) using the US FDA-approved Aptima Combo 2 and Aptima TV assays and compared the results with the conventional routine diagnostic tests (CDTs) in Ukraine. Urogenital swabs from consecutive mostly symptomatic females (n = 296) and males (n = 159) were examined. The prevalences were as follows: 10% (n = 47) of TV, 5.3% (n = 24) of CT and 1.5% (n = 7) of NG. The specificity of some CDTs was high, for example, 100% for NG culture, TV IgG ELISA, CT IgM ELISA and CT microscopy, but lower for other CDTs, that is, from 44% to 99.8%. The sensitivity of all CDTs was suboptimal, that is, 71% (n = 5) for NG microscopy, 57% (n = 4) for NG culture, 53% (n = 8) for CT IgG ELISA, 33% (n = 1) for TV IgG ELISA, 28% (n = 13) for TV microscopy, 25% (n = 1) for CT IgA ELISA, 20% (n = 3) for CT IgM ELISA and 0% (n = 0) for CT microscopy. The prevalences of particularly TV and CT were high, but substantial also for NG, in Ternopil, Ukraine. The sensitivities of all CDTs were low, and widespread implementation of validated, quality-assured and cost-effective molecular diagnostic STI tests in Ukraine is imperative.

  • 2.
    Boiko, Iryna
    et al.
    WHO Collaborating Centre for Gonorrhoea and other STIs, National Reference Laboratory for STIs, Department of Laboratory Medicine, Faculty of Medicine and Health, Örebro University, Örebro, Sweden; Clinical Laboratory Department, Ternopil Regional Clinical Dermatovenerologic Dispensary, Ternopil, Ukraine.
    Golparian, Daniel
    Örebro University, School of Medical Sciences. WHO Collaborating Centre for Gonorrhoea and other STIs, National Reference Laboratory for STIs, Department of Laboratory Medicine.
    Krynytska, Inna
    Department of Functional and Laboratory Diagnostics, I. Horbachevsky Ternopil State Medical University, Ternopil, Ukraine.
    Bezkorovaina, Halyna
    Outpatient Department, Ternopil Regional Clinical Dermatovenerologic Dispensary, Ternopil, Ukraine.
    Frankenberg, Arkadii
    Dnipropetrovsk Regional Clinical Dermatovenerologic Dispensary, Dnipro, Ukraine.
    Onuchyna, Margarita
    Clinical Laboratory Department, Dnipropetrovsk Regional Clinical Dermatovenerologic Dispensary, Dnipro, Ukraine.
    Jacobsson, Susanne
    Örebro University, School of Medical Sciences. Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and other STIs, National Reference Laboratory for STIs, Department of Laboratory Medicine.
    Unemo, Magnus
    Örebro University, School of Medical Sciences. Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and other STIs, National Reference Laboratory for STIs, Department of Laboratory Medicine.
    Antimicrobial susceptibility of Neisseria gonorrhoeae isolates and treatment of gonorrhoea patients in Ternopil and Dnipropetrovsk regions of Ukraine, 2013-20182019In: Acta Pathologica, Microbiologica et Immunologica Scandinavica (APMIS), ISSN 0903-4641, E-ISSN 1600-0463, Vol. 127, no 7, p. 503-509Article in journal (Refereed)
    Abstract [en]

    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 have 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 European Committee on Antimicrobial Susceptibility Testing (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.

  • 3.
    Bruni, Mirian Pinheiro
    et al.
    Biology Institute, Federal University of Pelotas (UFPel), Pelotas, Brazil.
    Freitas da Silveira, Mariangela
    School of Medicine, Federal University of Pelotas (UFPel), Pelotas, Brazil.
    Stauffert, Dulce
    School of Medicine, Federal University of Pelotas (UFPel), Pelotas, Brazil.
    Bicca, Guilherme Lucas de Oliveira
    School of Medicine, Federal University of Pelotas (UFPel), Pelotas, Brazil.
    Caetano Dos Santos, Carolina
    Biology Institute, Federal University of Pelotas (UFPel), Pelotas, Brazil.
    da Rosa Farias, Nara Amélia
    Biology Institute, Federal University of Pelotas (UFPel), Pelotas, Brazil.
    Golparian, Daniel
    Örebro University, School of Medical Sciences. WHO Collaborating Centre for Gonorrhoea and other STIs, Department of Laboratory Medicine.
    Unemo, Magnus
    Örebro University, School of Medical Sciences. Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and other STIs, Department of Laboratory Medicine.
    Aptima Trichomonas vaginalis assay elucidates significant underdiagnosis of trichomoniasis among women in Brazil according to an observational study2019In: Sexually Transmitted Infections, ISSN 1368-4973, E-ISSN 1472-3263, Vol. 95, no 2, p. 129-132Article in journal (Refereed)
    Abstract [en]

    OBJECTIVES: Trichomonas vaginalis (TV) infection is the most common non-viral STI globally and can result in adverse pregnancy outcomes and exacerbated HIV acquisition/transmission. Nucleic acid amplification tests (NAATs) are the most sensitive diagnostic tests, with high specificity, but TV NAATs are rarely used in Brazil. We investigated the TV prevalence and compared the performance of the US Food and Drug Association-cleared Aptima TV assay with microscopy (wet mount and Gram-stained) and culture for TV detection in women in Pelotas, Brazil in an observational study.

    METHODS: From August 2015 to December 2016, 499 consecutive asymptomatic and symptomatic sexually active women attending a Gynaecology and Obstetrics Outpatient Clinic were enrolled. Vaginal fluid and swab specimens were collected and wet mount microscopy, Gram-stained microscopy, culture and the Aptima TV assay performed.

    RESULTS: The median age of enrolled women was 36.5 years (range: 15-77). The majority were white, had a steady sexual partner and low levels of education. The TV detection rate was 4.2%, 2.4%, 1.2% and 0% using the Aptima TV assay, culture, wet mount microscopy and Gram-stained microscopy, respectively. The sensitivity of culture and wet mount microscopy was only 57.1% (95% CI 36.5 to 75.5) and 28.6% (95% CI 13.8 to 50.0), respectively.

    CONCLUSIONS: was found among women in Pelotas, Brazil and the routine diagnostic test (wet mount microscopy) and culture had low sensitivities. More sensitive diagnostic tests (NAATs) and enhanced testing of symptomatic and asymptomatic at-risk women are crucial to mitigate the transmission of TV infection, TV-associated sequelae and enhanced HIV acquisition and transmission.

  • 4.
    Donà, Valentina
    et al.
    Institute for Infectious Diseases, University of Bern, Bern, Switzerland.
    Low, Nicola
    Institute of Social and Preventive Medicine, University of Bern, Bern, Switzerland.
    Golparian, Daniel
    Örebro University, School of Medical Sciences. WHO Collaborating Centre for Gonorrhoea, Örebro University Hospital, Örebro, Sweden.
    Unemo, Magnus
    WHO Collaborating Centre for Gonorrhoea, Örebro University Hospital, Örebro, Sweden.
    Recent advances in the development and use of molecular tests to predict antimicrobial resistance in Neisseria gonorrhoeae2017In: Expert Review of Molecular Diagnostics, ISSN 1473-7159, E-ISSN 1744-8352, Vol. 17, no 9, p. 845-859Article in journal (Refereed)
    Abstract [en]

    Introduction: The number of genetic tests, mostly real-time PCRs, to detect antimicrobial resistance (AMR) determinants and predict AMR in Neisseria gonorrhoeae is increasing. Several of these assays are promising, but there are important shortcomings and few assays have been adequately validated and quality assured.

    Areas covered: Recent advances, focusing on publications since 2012, in the development and use of molecular tests to predict gonococcal AMR for surveillance and for clinical use, advantages and disadvantages of these tests and of molecular AMR prediction compared with phenotypic AMR testing, and future perspectives for effective use of molecular AMR tests for different purposes.

    Expert commentary: Several challenges for direct testing of clinical, especially extra-genital, specimens remain. The choice of molecular assay needs to consider the assay target, quality controls, sample types, limitations intrinsic to molecular technologies, and specific to the chosen methodology, and the intended use of the test. Improved molecular- and particularly genome-sequencing-based methods will supplement AMR testing for surveillance purposes, and translate into point-of-care tests that will lead to personalized treatments, while sparing the last available empiric treatment option (ceftriaxone). However, genetic AMR prediction will never completely replace phenotypic AMR testing, which detects also AMR due to unknown AMR determinants.

  • 5.
    Eyre, David W.
    et al.
    Big Data Institute, University of Oxford, Oxford, United Kingdom; Nuffield Department of Medicine, University of Oxford, Oxford, United Kingdom.
    Sanderson, Nicholas D.
    Nuffield Department of Medicine, University of Oxford, Oxford, United Kingdom.
    Lord, Emily
    Oxford University Hospitals NHS Foundation Trust, Oxford, United Kingdom.
    Regisford-Reimmer, Natasha
    Oxford University Hospitals NHS Foundation Trust, Oxford, United Kingdom.
    Chau, Kevin
    Nuffield Department of Medicine, University of Oxford, Oxford, United Kingdom.
    Barker, Leanne
    Nuffield Department of Medicine, University of Oxford, Oxford, United Kingdom.
    Morgan, Markus
    Oxford University Hospitals NHS Foundation Trust, Oxford, United Kingdom.
    Newnham, Robert
    Oxford University Hospitals NHS Foundation Trust, Oxford, United Kingdom.
    Golparian, Daniel
    Örebro University, School of Medical Sciences. WHO Collaborating Centre for Gonorrhoea and Other STIs, National Reference Laboratory for STIs, Örebro University Hospital, Örebro, Sweden.
    Unemo, Magnus
    Örebro University, School of Medical Sciences. Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and Other STIs, National Reference Laboratory for STIs, Örebro University Hospital, Örebro, Sweden.
    Crook, Derrick W.
    Nuffield Department of Medicine, University of Oxford, Oxford, United Kingdom; National Infection Service, Public Health England, Colindale, United Kingdom; National Institute for Health Research Health Protection Research Unit in Healthcare Associated Infections and Antimicrobial Resistance, University of Oxford, Oxford, United Kingdom .
    Peto, Tim E. A.
    Nuffield Department of Medicine, University of Oxford, Oxford, United Kingdom; National Institute for Health Research Health Protection Research Unit in Healthcare Associated Infections and Antimicrobial Resistance, University of Oxford, Oxford, United Kingdom.
    Hughes, Gwenda
    National Infection Service, Public Health England, Colindale, United Kingdom.
    Cole, Michelle J.
    National Infection Service, Public Health England, Colindale, United Kingdom.
    Fifer, Helen
    National Infection Service, Public Health England, Colindale, United Kingdom.
    Edwards, Anne
    Oxford University Hospitals NHS Foundation Trust, Oxford, United Kingdom.
    Andersson, Monique I.
    Oxford University Hospitals NHS Foundation Trust, Oxford, United Kingdom.
    Gonorrhoea treatment failure caused by a Neisseria gonorrhoeae strain with combined ceftriaxone and high-level azithromycin resistance, England, February 20182018In: Eurosurveillance, ISSN 1025-496X, E-ISSN 1560-7917, Vol. 23, no 27, p. 2-7, article id 1800323Article in journal (Refereed)
  • 6.
    Fifer, Helen
    et al.
    Public Health England, London, United Kingdom.
    Natarajan, Usha
    Virgin Care, London, United Kingdom.
    Jones, Lucy
    Virgin Care, London, United Kingdom.
    Alexander, Sarah
    Public Health England, London, United Kingdom.
    Hughes, Gwenda
    Public Health England, London, United Kingdom.
    Golparian, Daniel
    Örebro University, School of Medical Sciences.
    Unemo, Magnus
    Örebro University, School of Health Sciences.
    Failure of Dual Antimicrobial Therapy in Treatment of Gonorrhea2016In: New England Journal of Medicine, ISSN 0028-4793, E-ISSN 1533-4406, Vol. 374, no 25, p. 2504-2506Article in journal (Refereed)
  • 7.
    Foerster, Sunniva
    et al.
    Institute for Infectious Diseases, University of Bern, Bern, Switzerland; Institute of Social and Preventive Medicine, University of Bern, Bern, Switzerland; WHO Collaborating Centre for Gonorrhoea and other STIs, National Reference Laboratory for Pathogenic Neisseria, Örebro University Hospital, Örebro, Sweden.
    Golparian, Daniel
    WHO Collaborating Centre for Gonorrhoea and other STIs, National Reference Laboratory for Pathogenic Neisseria, Örebro University Hospital, Örebro, Sweden.
    Jacobsson, Susanne
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden. WHO Collaborating Centre for Gonorrhoea and other STIs, National Reference Laboratory for Pathogenic Neisseria, Örebro University Hospital, Örebro, Sweden.
    Hathaway, Lucy J.
    Institute for Infectious Diseases, University of Bern, Bern, Switzerland.
    Low, Nicola
    Institute of Social and Preventive Medicine, University of Bern, Bern, Switzerland.
    Shafer, William M.
    Department of Microbiology and Immunology, Emory University School of Medicine, Atlanta GA, USA; Laboratories of Bacterial Pathogenesis, Veterans Affairs Medical Center, Decatur GA, USA.
    Althaus, Christian L.
    Institute of Social and Preventive Medicine, University of Bern, Bern, Switzerland.
    Unemo, Magnus
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden. WHO Collaborating Centre for Gonorrhoea and other STIs, National Reference Laboratory for Pathogenic Neisseria, Örebro University Hospital, Örebro, Sweden.
    Genetic Resistance Determinants, In Vitro Time-Kill Curve Analysis and Pharmacodynamic Functions for the Novel Topoisomerase II Inhibitor ETX0914 (AZD0914) in Neisseria gonorrhoeae2015In: Frontiers in Microbiology, ISSN 1664-302X, E-ISSN 1664-302X, Vol. 6, article id 1377Article in journal (Refereed)
    Abstract [en]

    Resistance in Neisseria gonorrhoeae to all available therapeutic antimicrobials has emerged and new efficacious drugs for treatment of gonorrhea are essential. The topoisomerase II inhibitor ETX0914 (also known as AZD0914) is a new spiropyrimidinetrione antimicrobial that has different mechanisms of action from all previous and current gonorrhea treatment options. In this study, the N. gonorrhoeae resistance determinants for ETX0914 were further described and the effects of ETX0914 on the growth of N. gonorrhoeae (ETX0914 wild type, single step selected resistant mutants, and efflux pump mutants) were examined in a novel in vitro time-kill curve analysis to estimate pharmacodynamic parameters of the new antimicrobial. For comparison, ciprofloxacin, azithromycin, ceftriaxone, and tetracycline were also examined (separately and in combination with ETX0914). ETX0914 was rapidly bactericidal for all wild type strains and had similar pharmacodynamic properties to ciprofloxacin. All selected resistant mutants contained mutations in amino acid codons D429 or K450 of GyrB and inactivation of the MtrCDE efflux pump fully restored the susceptibility to ETX0914. ETX0914 alone and in combination with azithromycin and ceftriaxone was highly effective against N. gonorrhoeae and synergistic interaction with ciprofloxacin, particularly for ETX0914-resistant mutants, was found. ETX0914, monotherapy or in combination with azithromycin (to cover additional sexually transmitted infections), should be considered for phase III clinical trials and future gonorrhea treatment.

  • 8.
    Gianecini, Ricardo A.
    et al.
    National Reference Laboratory of Sexually Transmitted Diseases, National Institute of Infectious Diseases - ANLIS 'Dr Carlos G. Malbrán', Ciudad Autónoma de Buenos Aires, Argentina.
    Golparian, Daniel
    Örebro University, School of Medical Sciences. WHO Collaborating Centre for Gonorrhoea and Other STIs, Department of Laboratory Medicine, Microbiology.
    Zittermann, Sandra
    Public Health Ontario Laboratories, Toronto, Canada.
    Litvik, Ana
    Rawson Infectious Diseases Hospital, Córdoba, Argentina.
    Gonzalez, Silvia
    Rawson Infectious Diseases Hospital, Córdoba, Argentina.
    Oviedo, Claudia
    National Reference Laboratory of Sexually Transmitted Diseases, National Institute of Infectious Diseases - ANLIS 'Dr Carlos G. Malbrán', Ciudad Autónoma de Buenos Aires, Argentina.
    Melano, Roberto G.
    Public Health Ontario Laboratories, Toronto, Canada.
    Unemo, Magnus
    Örebro University, School of Medical Sciences. Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and Other STIs, Department of Laboratory Medicine, Microbiology.
    Galarza, Patricia
    National Reference Laboratory of Sexually Transmitted Diseases, National Institute of Infectious Diseases - ANLIS 'Dr Carlos G. Malbrán', Ciudad Autónoma de Buenos Aires, Argentina.
    Genome-based epidemiology and antimicrobial resistance determinants of Neisseria gonorrhoeae isolates with decreased susceptibility and resistance to extended-spectrum cephalosporins in Argentina in 2011-162019In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 74, no 6, p. 1551-1559Article in journal (Refereed)
    Abstract [en]

    OBJECTIVES: Our aim was to describe the molecular epidemiology and antimicrobial resistance determinants of isolates of Neisseria gonorrhoeae with decreased susceptibility and resistance to extended-spectrum cephalosporins (ESCs) in Argentina in 2011-16.

    METHODS: Gonococcal isolates (n = 158) with decreased susceptibility and resistance to ESCs collected in 2011-16 across Argentina were subjected to WGS and antimicrobial susceptibility testing for six antimicrobials.

    RESULTS: In total, 50% of the isolates were resistant to cefixime, 1.9% were resistant to ceftriaxone, 37.3% were resistant to azithromycin and 63.9% of the isolates showed an MDR phenotype. Resistance and decreased susceptibility to ESCs was mainly associated with isolates possessing the mosaic penA-34.001, in combination with an mtrR promoter A deletion, and PorB1b amino acid substitutions G120K/A121N. Phylogenetic analysis revealed two main clades of circulating strains, which were associated with the N. gonorrhoeae multiantigen sequence typing (NG-MAST) ST1407 and closely related STs, and characterized by a high prevalence rate, wide geographical distribution and temporal persistence.

    CONCLUSIONS: N. gonorrhoeae isolates with decreased susceptibility and resistance to ESCs in Argentina have emerged and rapidly spread mainly due to two clonal expansions after importation of one or two strains, which are associated with the international MDR NG-MAST ST1407 clone. The identification of the geographical dissemination and characteristics of these predominant clones may help to focus action plans and public health policies to control the spread of ESC resistance in Argentina. Dual antimicrobial therapy (ceftriaxone plus azithromycin) for gonorrhoea needs to be considered in Argentina.

  • 9.
    Golparian, Daniel
    et al.
    WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, Swedish Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Microbiology, Faculty of Medicine and Health, Örebro University Hospital, Örebro, Sweden.
    Boräng, Stina
    Department of Clinical Microbiology, Karolinska University Hospital Huddinge, Stockholm, Sweden .
    Sundqvist, Martin
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden. WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, Swedish Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Unemo, Magnus
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden. WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, Swedish Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Evaluation of the New BD Max GC Real-Time PCR Assay, Analytically and Clinically as a Supplementary Test for the BD ProbeTec GC Qx Amplified DNA Assay, for Molecular Detection of Neisseria gonorrhoeae2015In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 53, no 12, p. 3935-3937Article in journal (Refereed)
    Abstract [en]

    The new BD Max GC real-time PCR assay showed high clinical and analytical sensitivity and specificity. It can be an effective and accurate supplementary test for the BD ProbeTec GC Qx amplified DNA assay, which had suboptimal specificity, and might also be used for initial detection of Neisseria gonorrhoeae.

  • 10.
    Golparian, Daniel
    et al.
    Örebro University, School of Medical Sciences. WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, Department of Laboratory Medicine, Clinical Microbiology.
    Donà, Valentina
    Institute for Infectious Diseases, University of Bern, Bern, Switzerland; Institute of Veterinary Bacteriology, Vetsuisse Faculty, University of Bern, Bern, Switzerland.
    Sánchez-Busó, Leonor
    Pathogen Genomics, The Wellcome Sanger Institute, Wellcome Genome Campus, Hinxton, Cambridgeshire, United Kingdom.
    Foerster, Sunniva
    WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, Department of Laboratory Medicine, Clinical Microbiology, Faculty of Medicine and Health, Örebro University, Örebro, Sweden.
    Harris, Simon
    Pathogen Genomics, The Wellcome Sanger Institute, Wellcome Genome Campus, Hinxton, Cambridgeshire, United Kingdom.
    Endimiani, Andrea
    Institute for Infectious Diseases, University of Bern, Bern, Switzerland.
    Low, Nicola
    Institute of Social and Preventive Medicine, University of Bern, Bern, Switzerland.
    Unemo, Magnus
    Örebro University, School of Medical Sciences. Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, Department of Laboratory Medicine, Clinical Microbiology.
    Antimicrobial resistance prediction and phylogenetic analysis of Neisseria gonorrhoeae isolates using the Oxford Nanopore MinION sequencer2018In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 8, no 1, article id 17596Article in journal (Refereed)
    Abstract [en]

    Antimicrobial resistance (AMR) in Neisseria gonorrhoeae is common, compromising gonorrhoea treatment internationally. Rapid characterisation of AMR strains could ensure appropriate and personalised treatment, and support identification and investigation of gonorrhoea outbreaks in nearly real-time. Whole-genome sequencing is ideal for investigation of emergence and dissemination of AMR determinants, predicting AMR, in the gonococcal population and spread of AMR strains in the human population. The novel, rapid and revolutionary long-read sequencer MinION is a small hand-held device that generates bacterial genomes within one day. However, accuracy of MinION reads has been suboptimal for many objectives and the MinION has not been evaluated for gonococci. In this first MinION study for gonococci, we show that MinION-derived sequences analysed with existing open-access, web-based sequence analysis tools are not sufficiently accurate to identify key gonococcal AMR determinants. Nevertheless, using an in house-developed CLC Genomics Workbench including de novo assembly and optimised BLAST algorithms, we show that 2D ONT-derived sequences can be used for accurate prediction of decreased susceptibility or resistance to recommended antimicrobials in gonococcal isolates. We also show that the 2D ONT-derived sequences are useful for rapid phylogenomic-based molecular epidemiological investigations, and, in hybrid assemblies with Illumina sequences, for producing contiguous assemblies and finished reference genomes.

  • 11.
    Golparian, Daniel
    et al.
    WHO Collaborating Centre for Gonorrhoea and other STIs, National Reference Laboratory for Pathogenic Neisseria, Örebro University Hospital, Örebro, Sverige; Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Fernandes, Prabhavathi
    Cempra Pharmaceuticals, Inc., Chapel Hill NC, United States.
    Ohnishi, Makoto
    National Institute of Infectious Diseases, Tokyo, Japan.
    Jensen, Jörgen S
    Department of Microbiological Surveillance and Research, Statens Serum Institut, Copenhagen, Denmark.
    Unemo, Magnus
    Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and other STIs, National Reference Laboratory for Pathogenic Neisseria, Örebro University Hospital, Örebro, Sverige.
    In vitro activity of the new fluoroketolide solithromycin (CEM-101) against a large collection of clinical Neisseria gonorrhoeae isolates and international reference strains, including those with high-level antimicrobial resistance: potential treatment option for gonorrhea?2012In: Antimicrobial Agents and Chemotherapy, ISSN 0066-4804, E-ISSN 1098-6596, Vol. 56, no 5, p. 2739-2742Article in journal (Refereed)
    Abstract [en]

    Gonorrhea may become untreatable, and new treatment options are essential. We investigated the in vitro activity of the first fluoroketolide, solithromycin. Clinical Neisseria gonorrhoeae isolates and reference strains (n = 246), including the two extensively drug-resistant strains H041 and F89 and additional isolates with clinical cephalosporin resistance and multidrug resistance, were examined. The activity of solithromycin was mainly superior to that of other antimicrobials (n = 10) currently or previously recommended for gonorrhea treatment. Solithromycin might be an effective treatment option for gonorrhea.

  • 12.
    Golparian, Daniel
    et al.
    WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, Swedish Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Faculty of Medicine and Health, Örebro University, Örebro, Sweden.
    Hellmark, Bengt
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden. WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, Swedish Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Faculty of Medicine and Health, Örebro University, Örebro, Sweden.
    Unemo, Magnus
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden. WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, Swedish Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Faculty of Medicine and Health, Örebro University, Örebro, Sweden.
    Analytical specificity and sensitivity of the novel dual-target GeneProof Neisseria gonorrhoeae PCR kit for detection of N-gonorrhoeae2015In: Acta Pathologica, Microbiologica et Immunologica Scandinavica (APMIS), ISSN 0903-4641, E-ISSN 1600-0463, Vol. 123, no 11, p. 955-958Article in journal (Refereed)
    Abstract [en]

    Detection of Neisseria gonorrhoeae relies increasingly on nucleic acid amplification tests (NAATs). The specificity of many gonococcal NAATs has been suboptimal and supplementary testing remains recommended in Europe and several additional countries. The novel dual-target GeneProofNeisseria gonorrhoeae PCR kit, targeting porA pseudogene and 16S rRNA gene, showed a high specificity and sensitivity when isolates of non-gonococcal Neisseria and related species (n=144), and gonococci (n=104) were tested. However, rare gonococcal porA mutants were only detected in the 16S rRNA gene target and two non-gonococcal isolates showed a low-level cross-reactivity in the 16S rRNA gene target. The detection limit for both targets was 1.5 copies per reaction.

  • 13.
    Golparian, Daniel
    et al.
    WHO Collaborating Centre for Gonorrhoea and other STIs, Swedish Reference Laboratory for Pathogenic Neisseria, Örebro University Hospital, Örebro, Sverige; Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Johansson, E.
    WHO Collaborating Centre for Gonorrhoea and other STIs, Swedish Reference Laboratory for Pathogenic Neisseria, Örebro University Hospital, Örebro, Sverige; Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Unemo, Magnus
    Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and other STIs, Swedish Reference Laboratory for Pathogenic Neisseria, Örebro University Hospital, Örebro, Sverige; Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Clinical Neisseria gonorrhoeae isolate with a N. meningitidis porA gene and no prolyliminopeptidase activity, Sweden, 2011-danger of false-negative genetic and culture diagnostic results2012In: Eurosurveillance, ISSN 1025-496X, E-ISSN 1560-7917, Vol. 17, no 9, p. 5-7, article id 20102Article in journal (Refereed)
    Abstract [en]

    We describe a Neisseria gonorrhoeae strain, found in Sweden in 2011, that harbours a N. meningitidis porA gene causing false-negative results in PCRs targeting the gonococcal porA pseudogene. Furthermore, the strain had no prolyliminopeptidase (PIP) activity that many commercial biochemical kits for species verification in culture rely on. Enhanced awareness of the spread of such strains and screening for them can be crucial.

  • 14.
    Golparian, Daniel
    et al.
    World Health Organization Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, Swedish Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Ohlsson, A. K.
    Dept Clin Microbiol, Karolinska Univ Hosp, Stockholm, Sweden.
    Janson, H.
    Dept Clin Microbiol, Cent Hosp Växjö, Växjö, Sweden.
    Lidbrink, P.
    Dept Dermatovenereol, Karolinska Univ Hosp, Stockholm, Sweden.
    Richtner, T.
    Dept Dermatol, Karolinska Inst Södersjukhuset, Stockholm, Sweden.
    Ekelund, O.
    Dept Clin Microbiol, Cent Hosp Växjö, Växjö, Sweden.
    Fredlund, Hans
    Örebro University Hospital. World Health Organization Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, Swedish Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Unemo, Magnus
    Örebro University Hospital. World Health Organization Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, Swedish Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Four treatment failures of pharyngeal gonorrhoea with ceftriaxone (500 mg) or cefotaxime (500 mg), Sweden, 2013 and 20142014In: Eurosurveillance, ISSN 1025-496X, E-ISSN 1560-7917, Vol. 19, no 30, p. 2-5, article id 20862Article in journal (Refereed)
    Abstract [en]

    We describe four cases in Sweden of verified treatment failures of pharyngeal gonorrhoea with ceftriaxone (500 mg; n=3) or cefotaxime (500 mg; n=1) monotherapy. All the ceftriaxone treatment failures were caused by the internationally spreading multidrug-resistant gonococcal NG-MAST genogroup 1407 clone. Increased awareness of treatment failures is crucial particularly when antimicrobial monotherapy is used. Frequent test of cure and appropriate verification/falsification of suspected treatment failures, as well as implementation of recommended dual antimicrobial therapy are imperative.

  • 15.
    Golparian, Daniel
    et al.
    Örebro University, School of Medical Sciences. WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, Swedish Reference Laboratory for Sexually Transmitted Infections, Department of Laboratory Medicine, Clinical Microbiology, Faculty of Medicine and Health, Örebro University, Örebro, Sweden.
    Rose, Lisa
    Department of Clinical Microbiology, University of Dublin, Trinity College, St. James’s Hospital, Dublin, Ireland.
    Lynam, Almida
    Guide Clinic, St. James’s Hospital, Dublin, Ireland.
    Mohamed, Aia
    Department of Clinical Microbiology, University of Dublin, Trinity College, St. James’s Hospital, Dublin, Ireland.
    Bercot, Beatrice
    APHP, St Louis Hospital, Laboratory of Microbiology; French National Reference Center for Bacterial STI, Associated laboratory for gonococci; Paris Diderot University, IAME, Sorbonne Paris Cité, Paris, France.
    Ohnishi, Makoto
    Department of Bacteriology I, National Institute of Infectious Diseases, Tokyo, Japan.
    Crowley, Brendan
    Department of Clinical Microbiology, University of Dublin, Trinity College, St. James’s Hospital, Dublin, Ireland; National Gonococcal Reference Laboratory, St. James’s Hospital, Dublin, Ireland.
    Unemo, Magnus
    Örebro University, School of Medical Sciences. Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, Swedish Reference Laboratory for Sexually Transmitted Infections, Department of Laboratory Medicine, Clinical Microbiology, Faculty of Medicine and Health, Örebro University, Örebro, Sweden.
    Multidrug-resistant Neisseria gonorrhoeae isolate, belonging to the internationally spreading Japanese FC428 clone, with ceftriaxone resistance and intermediate resistance to azithromycin, Ireland, August 20182018In: Eurosurveillance, ISSN 1025-496X, E-ISSN 1560-7917, Vol. 23, no 47, p. 6-9, article id 1800617Article in journal (Refereed)
    Abstract [en]

    We describe a multidrug-resistant Neisseria gonorrhoeae urethritis case with ceftriaxone resistance and azithromycin intermediate resistance in a heterosexual man in Ireland, August 2018. Whole-genome sequencing showed that the isolate IR72 belongs to the internationally spreading multidrug-resistant ceftriaxone-resistant FC428 clade, initially described in Japan in 2015. IR72 was assigned MSLT ST1903, NG-MAST ST17842 and NG-STAR type 1133, including the ceftriaxone resistance-mediating penA-60.001. Global awareness of spreading ceftriaxone-resistant gonococcal strains that threaten recommended dual therapies is essential.

  • 16.
    Golparian, Daniel
    et al.
    National Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Shafer, William M.
    Department of Microbiology and Immunology, School of Medicine, Emory University, Atlanta GA, United States; Laboratories of Bacterial Pathogenesis, Veterans Affairs Medical Center, Decatur GA, United States.
    Ohnishi, Makoto
    National Institute of Infectious Diseases, Tokyo, Japan.
    Unemo, Magnus
    Örebro University Hospital. National Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Importance of Multidrug Efflux Pumps in the Antimicrobial Resistance Property of Clinical Multidrug-Resistant Isolates of Neisseria gonorrhoeae2014In: Antimicrobial Agents and Chemotherapy, ISSN 0066-4804, E-ISSN 1098-6596, Vol. 58, no 6, p. 3556-3559Article in journal (Refereed)
    Abstract [en]

    The contribution of drug efflux pumps in clinical isolates of Neisseria gonorrhoeae that express extensively drug-resistant or multidrug-resistant phenotypes has heretofore not been examined. Accordingly, we assessed the effect on antimicrobial resistance of loss of the three gonococcal efflux pumps associated with a known capacity to export antimicrobials (MtrC-MtrD-MtrE, MacA-MacB, and NorM) in such clinical isolates. We report that the MIC of several antimicrobials, including seven previously and currently recommended for treatment was significantly impacted.

  • 17.
    Golparian, Daniel
    et al.
    Natl Reference Lab Pathogen Neisseria, WHO Collaborating Ctr Gonorrhoea & Other STIs, Örebro University Hospital, Örebro, Sweden.
    Tabrizi, Sepehr N.
    Dept Microbiol & Infect Dis, Royal Womens Hosp, Parkville Vic, Australia.
    Unemo, Magnus
    Örebro University Hospital. Natl Reference Lab Pathogen Neisseria, WHO Collaborating Ctr Gonorrhoea & Other STIs, Örebro University Hospital, Örebro, Sweden; Dept Lab Med, WHO Collaborating Ctr Gonorrhoea & Other STIs, Örebro University Hosp, Örebro, Sweden.
    Analytical Specificity and Sensitivity of the APTIMA Combo 2 and APTIMA GC Assays for Detection of Commensal Neisseria Species and Neisseria gonorrhoeae on the Gen-Probe Panther Instrument2013In: Sexually Transmitted Diseases, ISSN 0148-5717, E-ISSN 1537-4521, Vol. 40, no 2, p. 175-178Article in journal (Refereed)
    Abstract [en]

    Genetic detection of Neisseria gonorrhoeae is replacing culture for increased diagnostic sensitivity. Specificity of several nucleic acid amplification tests is suboptimal. Herein, the Gen-Probe APTIMA Combo 2 and APTIMA GC assays had 100% specificity and 100% sensitivity after confirmatory testing, when testing 298 isolates of non-gonococcal Neisseria and related species and 205 gonococcal isolates.

  • 18.
    Golparian, Daniel
    et al.
    WHO Collaborating Ctr Gonorrhoea & Other Sexually, Natl Reference Lab Pathogen Neisseria, Dept Lab Med,Clin Microbiol, Örebro University Hospital, Örebro, Sweden.
    Unemo, Magnus
    Örebro University Hospital. WHO Collaborating Ctr Gonorrhoea & Other Sexually, Natl Reference Lab Pathogen Neisseria, Dept Lab Med,Clin Microbiol, Örebro University Hospital, Örebro, Sweden.
    Will Genome Analysis Elucidate Evolution, Global Transmission and Virulence of Neisseria Meningitidis Lineages?2015In: EBioMedicine, E-ISSN 2352-3964, Vol. 2, no 3, p. 186-187Article in journal (Refereed)
  • 19.
    Hadad, Ronza
    et al.
    WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, National Reference Laboratory for Sexually Transmitted Infections, Department of Laboratory Medicine, Clinical Microbiology, Örebro University Hospital, Örebro, Sweden.
    Golparian, Daniel
    Örebro University, School of Medical Sciences. WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, National Reference Laboratory for Sexually Transmitted Infections, Department of Laboratory Medicine, Clinical Microbiology, , Örebro University Hospital, Örebro, Sweden.
    Lagos, Amaya C.
    WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, National Reference Laboratory for Sexually Transmitted Infections, Department of Laboratory Medicine, Clinical Microbiology, Örebro University Hospital, Örebro, Sweden.
    Ljungberg, Johan
    Department of Infectious Diseases, Hospital of Halmstad, Halmstad, Sweden.
    Nilsson, Peter
    Department of Clinical Microbiology & Infection Control, Hospital of Halmstad, Halmstad, Sweden.
    Jensen, Jörgen S.
    Department of Microbiology and Infection Control, Sexually Transmitted Infections, Research and Development, Statens Serum Institut, Copenhagen, Denmark.
    Fredlund, Hans
    WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, National Reference Laboratory for Sexually Transmitted Infections, Department of Laboratory Medicine, Clinical Microbiology, Örebro University Hospital, Örebro, Sweden.
    Unemo, Magnus
    WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, National Reference Laboratory for Sexually Transmitted Infections, Department of Laboratory Medicine, Clinical Microbiology, Örebro University Hospital, Örebro, Sweden.
    Macrolide and fluoroquinolone resistance in Mycoplasma genitalium in two Swedish counties, 2011-20152018In: Acta Pathologica, Microbiologica et Immunologica Scandinavica (APMIS), ISSN 0903-4641, E-ISSN 1600-0463, Vol. 126, no 2, p. 123-127Article in journal (Refereed)
    Abstract [en]

    Mycoplasma genitalium, causing non-gonococcal non-chlamydial urethritis and associated with cervicitis, has developed antimicrobial resistance (AMR) to both the macrolide azithromycin (first-line treatment) and the fluoroquinolone moxifloxacin (second-line treatment). Our aim was to estimate the prevalence of resistance, based on genetic AMR determinants, to these antimicrobials in the M. genitalium population in two Swedish counties, Örebro and Halland, 2011-2015. In total, 672 M. genitalium positive urogenital samples were sequenced for 23S rRNA and parC gene mutations associated with macrolide and fluoroquinolone resistance, respectively. Of the samples, 18.6% and 3.2% in Örebro and 15.2% and 2.7% in Halland contained mutations associated with macrolide and fluoroquinolone resistance, respectively. The predominating resistance-associated mutations in the 23S rRNA gene was A2059G (n = 39) in Örebro and A2058G (n = 13) and A2059G (n = 13) in Halland. The most prevalent possible resistance-associated ParC amino acid alterations were S83I (n = 4) in Örebro and S83N (n = 2) in Halland. Resistance-associated mutations to both macrolides and fluoroquinolones were found in 0.7% of samples. Our findings emphasize the need for routine AMR testing, at a minimum for macrolide resistance, of all M. genitalium-positive samples and regular national and international surveillance of AMR in M. genitalium, to ensure effective patient management and rational antimicrobial use.

  • 20.
    Harris, Simon R.
    et al.
    Infection Genomics, Wellcome Sanger Institute, Hinxton, United Kingdom.
    Cole, Michelle J.
    Antimicrobial Resistance and Healthcare Associated Infections Reference Unit, National Infection Service, Public Health England, London, United Kingdom.
    Spiteri, Gianfranco
    European Centre for Disease Prevention and Control, Stockholm, Sweden.
    Sanchez-Buso, Leonor
    Infection Genomics, Wellcome Sanger Institute, Hinxton, United Kingdom.
    Golparian, Daniel
    Örebro University, School of Medical Sciences. WHO Collaborating Centre for Gonorrhoea and Other Sexually Transmitted Infections, Department of Laboratory Medicine, Clinical Microbiology, Faculty of Medicine and Health, Örebro University Hospital, Örebro, Sweden.
    Jacobsson, Susanne
    Örebro University, School of Medical Sciences. Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and Other Sexually Transmitted Infections, Department of Laboratory Medicine, Clinical Microbiology, Faculty of Medicine and Health, Örebro University Hospital, Örebro, Sweden.
    Goater, Richard
    Centre for Genomic Pathogen Surveillance, Wellcome Sanger Institute, Hinxton, United Kingdom.
    Abudahab, Khalil
    Centre for Genomic Pathogen Surveillance, Wellcome Sanger Institute, Hinxton, United Kingdom.
    Yeats, Corin A.
    Centre for Genomic Pathogen Surveillance, Wellcome Sanger Institute, Hinxton, United Kingdom.
    Bercot, Beatrice
    St Louis Hospital, Assistance Publique Hôpitaux de Paris (APHP), Paris, France.
    Borrego, Maria Jose
    National Institute of Health, Lisbon, Portugal.
    Crowley, Brendan
    St James's Hospital, Dublin, Ireland.
    Stefanelli, Paola
    Istituto Superiore di Sanitá, Rome, Italy.
    Tripodo, Francesco
    Antimicrobial Resistance and Healthcare Associated Infections Reference Unit, National Infection Service, Public Health England, London, United Kingdom.
    Abad, Raquel
    Carlos III Health Institute, Madrid, Spain.
    Aanensen, David M.
    Centre for Genomic Pathogen Surveillance, Wellcome Sanger Institute, Hinxton, United Kingdom; Big Data Institute, Li Ka Shing Centre for Health Information and Discovery, Nuffield Department of Medicine, University of Oxford, Oxford, United Kingdom.
    Unemo, Magnus
    Örebro University, School of Medical Sciences. Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and Other Sexually Transmitted Infections, Department of Laboratory Medicine, Clinical Microbiology, Faculty of Medicine and Health, Örebro University Hospital, Örebro, Sweden.
    Public health surveillance of multidrug-resistant clones of Neisseria gonorrhoeae in Europe: a genomic survey2018In: Lancet. Infectious diseases (Print), ISSN 1473-3099, E-ISSN 1474-4457, Vol. 18, no 7, p. 758-768Article in journal (Refereed)
    Abstract [en]

    Background: Traditional methods for molecular epidemiology of Neisseria gonorrhoeae are suboptimal. Whole-genome sequencing (WGS) offers ideal resolution to describe population dynamics and to predict and infer transmission of antimicrobial resistance, and can enhance infection control through linkage with epidemiological data. We used WGS, in conjunction with linked epidemiological and phenotypic data, to describe the gonococcal population in 20 European countries. We aimed to detail changes in phenotypic antimicrobial resistance levels (and the reasons for these changes) and strain distribution (with a focus on antimicrobial resistance strains in risk groups), and to predict antimicrobial resistance from WGS data.

    Methods: We carried out an observational study, in which we sequenced isolates taken from patients with gonorrhoea from the European Gonococcal Antimicrobial Surveillance Programme in 20 countries from September to November, 2013. We also developed a web platform that we used for automated antimicrobial resistance prediction, molecular typing (N gonorrhoeae multi-antigen sequence typing [NG-MAST] and multilocus sequence typing), and phylogenetic clustering in conjunction with epidemiological and phenotypic data.

    Findings: The multidrug-resistant NG-MAST genogroup G1407 was predominant and accounted for the most cephalosporin resistance, but the prevalence of this genogroup decreased from 248 (23%) of 1066 isolates in a previous study from 2009-10 to 174 (17%) of 1054 isolates in this survey in 2013. This genogroup previously showed an association with men who have sex with men, but changed to an association with heterosexual people (odds ratio=4.29). WGS provided substantially improved resolution and accuracy over NG-MAST and multilocus sequence typing, predicted antimicrobial resistance relatively well, and identified discrepant isolates, mixed infections or contaminants, and multidrug-resistant clades linked to risk groups.

    Interpretation: To our knowledge, we provide the first use of joint analysis of WGS and epidemiological data in an international programme for regional surveillance of sexually transmitted infections. WGS provided enhanced understanding of the distribution of antimicrobial resistance clones, including replacement with clones that were more susceptible to antimicrobials, in several risk groups nationally and regionally. We provide a framework for genomic surveillance of gonococci through standardised sampling, use of WGS, and a shared information architecture for interpretation and dissemination by use of open access software.

  • 21.
    Hess, David
    et al.
    Department of Biology, Santa Clara University, Santa Clara CA, United States.
    Wu, Abel
    San Francisco Public Health Laboratory, San Francisco CA, United States.
    Golparian, Daniel
    WHO Collaborating Centre for Gonorrhoea and Other STIs, Örebro University Hospital, Örebro, Sweden.
    Esmaili, Sarah
    Department of Biology, Santa Clara University, Santa Clara CA, United States.
    Pandori, Will
    Department of Biology, Santa Clara University, Santa Clara CA, United States.
    Sena, Emilee
    Department of Biology, Santa Clara University, Santa Clara CA, United States.
    Klausner, Jeffrey D.
    Division of Infectious Diseases, David Geffen School of Medicine, University of California, Los Angeles CA, United States.
    Barry, Pennan
    San Francisco Department of Public Health, San Francisco CA, United States.
    Unemo, Magnus
    WHO Collaborating Centre for Gonorrhoea and Other STIs, Örebro University Hospital, Örebro, Sweden.
    Pandori, Mark
    San Francisco Public Health Laboratory, San Francisco CA, United States.
    Genome Sequencing of a Neisseria gonorrhoeae Isolate of a Successful International Clone with Decreased Susceptibility and Resistance to Extended-Spectrum Cephalosporins2012In: Antimicrobial Agents and Chemotherapy, ISSN 0066-4804, E-ISSN 1098-6596, Vol. 56, no 11, p. 5633-5641Article in journal (Refereed)
  • 22.
    Heymans, Raymond
    et al.
    Public Health Laboratory, Cluster of Infectious Diseases, Health Service of Amsterdam, Amsterdam, The Netherlands.
    Bruisten, Sylvia M.
    Public Health Laboratory, Cluster of Infectious Diseases, Health Service of Amsterdam, Amsterdam, Netherlands; Department of Experimental Virology, University of Amsterdam, Amsterdam, The Netherlands.
    Golparian, Daniel
    WHO Collaborating Centre for Gonorrhoea and Other STIs, Department of Laboratory Medicine, Örebro University Hospital, Örebro, Sweden.
    Unemo, Magnus
    Örebro University Hospital. Örebro University, School of Health and Medical Sciences, Örebro University, Sweden. WHO Collaborating Centre for Gonorrhoea and Other STIs, Department of Laboratory Medicine, Örebro University Hospital, Örebro, Sweden.
    de Vries, Henry J. C.
    STI Outpatient Clinic, Cluster of Infectious Diseases, Health Service of Amsterdam, Amsterdam, Netherlands; Centre for Infectious Disease Control, National Institute for Public Health and the Environment (RIVM), Bilthoven, Netherlands; Department of Dermatology, Academic Medical Center, University of Amsterdam, Amsterdam, Netherlands.
    van Dam, Alje P.
    Public Health Laboratory, Cluster of Infectious Diseases, Health Service of Amsterdam, Amsterdam, Netherlands; Department of Medical Microbiology, Onze Lieve Vrouwe Gasthuis General Hospital, Amsterdam, The Netherlands.
    Clonally Related Neisseria gonorrhoeae Isolates with Decreased Susceptibility to the Extended-Spectrum Cephalosporin Cefotaxime in Amsterdam, the Netherlands2012In: Antimicrobial Agents and Chemotherapy, ISSN 0066-4804, E-ISSN 1098-6596, Vol. 56, no 3, p. 1516-1522Article in journal (Refereed)
    Abstract [en]

    From 2006 to 2008, Neisseria gonorrhoeae isolates were identified with decreased susceptibility to the extended-spectrum cephalosporin (ESC) cefotaxime among visitors of the Amsterdam sexually transmitted infections (STI) clinic, the Netherlands. Spread, clonality, and characteristics of 202 isolates were examined using antibiograms, conventional penA mosaic gene PCR, and N. gonorrhoeae multiple-locus variable-number tandem repeat analysis (NG-MLVA). A strictly defined subset was further characterized by N. gonorrhoeae multiantigen sequence typing (NG-MAST) and sequencing of ESC resistance determinants (penA, mtrR, and porB1b). Seventy-four N. gonorrhoeae isolates with a cefotaxime MIC of >0.125 mu g/ml (group A), 54 with a cefotaxime MIC of 0.125 mu g/ml (group B), and a control group of 74 with a cefotaxime MIC of <0.125 mu g/ml (group C) were included. Fifty-three clonally related penA mosaic-positive isolates (penicillin-binding protein 2 type XXXIV) were identified in group A (n = 47 isolates; 64%) and B (n = 6 isolates; 11%). The 53 penA mosaic-positive isolates were predominantly NG-MAST ST1407 (87%) and contained an mtrR promoter A deletion (98%) and porB1b alterations G101K/A102N. All were assigned to the same NG-MLVA cluster that comprised in total 56 isolates. A correlation was found between decreased cefotaxime susceptibility and ST1407 that was highly prevalent among visitors of the Amsterdam STI clinic. The rapid spread of this strain, which also has been identified in many other countries, might be facilitated by high-risk sexual behavior and should be monitored closely to identify potential treatment failure. Quality-assured surveillance of ESC susceptibility on the national and international levels and exploration of new drugs and/or strategies for treatment of gonorrhea are crucial.

  • 23.
    Heymans, Raymond
    et al.
    Public Health Laboratory, Cluster of Infectious Diseases, Health Service of Amsterdam, Amsterdam, Netherlands.
    Golparian, Daniel
    Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and other STIs, National Reference Laboratory for Pathogenic Neisseria, Örebro University Hospital, Örebro, Sverige; Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Bruisten, Sylvia M.
    Public Health Laboratory, Cluster of Infectious Diseases, Health Service of Amsterdam, Amsterdam, Netherlands; Department of Experimental Virology, University of Amsterdam, Amsterdam, Netherlands.
    Schouls, Leo M.
    Laboratory for Infectious Diseases and Perinatal Screening, National Institute for Public Health and the Environment (RIVM), Bilthoven, Netherlands.
    Unemo, Magnus
    Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and other STIs, National Reference Laboratory for Pathogenic Neisseria, Örebro University Hospital, Örebro, Sverige; Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Evaluation of Neisseria gonorrhoeae Multiple-Locus Variable-Number Tandem-Repeat Analysis, N. gonorrhoeae Multiantigen Sequence Typing, and Full-Length porB Gene Sequence Analysis for Molecular Epidemiological Typing2012In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 50, no 1, p. 180-183Article in journal (Refereed)
    Abstract [en]

    The performance characteristics of Neisseria gonorrhoeae multilocus variable-number tandem-repeat analysis were evaluated, by comparison with N. gonorrhoeae multiantigen sequence typing and full-length porB sequence typing. Assessment of the relatedness of intra- and interpatient isolates showed that all three genotyping techniques display a high resolution and typeability.

  • 24.
    Ison, Catherine A.
    et al.
    Sexually Transmitted Bacteria Reference Unit, Health Protection Agency, London, UK.
    Golparian, Daniel
    WHO Collaborating Centre for Gonorrhoea and other STIs, National Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Clinical Microbiology, Örebro University Hospital, Örebro, Sweden.
    Saunders, Pamela
    Sexually Transmitted Bacteria Reference Unit, Health Protection Agency, London, UK.
    Chisholm, Stephanie
    Sexually Transmitted Bacteria Reference Unit, Health Protection Agency, London, UK.
    Unemo, Magnus
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden. Örebro University Hospital.
    Evolution of Neisseria gonorrhoeae is a continuing challenge for molecular detection of gonorrhoea: false negative gonococcal porA mutants are spreading internationally2013In: Sexually Transmitted Infections, ISSN 1368-4973, E-ISSN 1472-3263, Vol. 89, no 3, p. 197-201Article in journal (Refereed)
    Abstract [en]

    Objectives: Identification of genetic targets specific to Neisseria gonorrhoeae for use in molecular detection methods has been a challenge. The porA pseudogene in N gonorrhoeae has been commonly used but recently gonococcal isolates giving a negative result in these PCRs have been reported. Here we describe the characterisation of two such gonococcal isolates received by the reference service at the Health Protection Agency, London, England.

    Methods: Phenotypic characterisation was achieved using conventional biochemical and immunological tests, matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF-MS), antimicrobial susceptibility testing, serovar determination and detection of meningococcal PorA using monoclonal antibody 4BG4-E7. Genetic species confirmation was determined using commercial and in house PCRs and 16S rRNA gene sequencing. Molecular typing using the N gonorrhoeae multi-antigen sequence typing (NG-MAST) and multilocus sequence typing (MLST) was performed. The DNA sequence of the full-length gonococcal porA pseudogene was determined and compared with published sequences.

    Results: Both isolates were confirmed, biochemically and immunologically as N gonorrhoeae, but repeatedly gave negative results with two in house real-time PCR assays for the porA pseudogene. Further characterisation of these isolates identified the presence of a meningococcal porA sequence and showed these isolates belong to serovar Bropyst, and to NG-MAST sequence type (ST) 5967 and MLST ST1901.

    Conclusions: Gonococcal isolates that give false negative results with porA pseudogene PCR assays have now been identified in four countries, three of which are in Europe, and do not appear clonal. This report highlights the genetic diversity of N gonorrhoeae, which remains a challenge for the molecular detection methods.

  • 25.
    Jacobsson, Susanne
    et al.
    Örebro University, School of Medical Sciences. Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and Other Sexually Transmitted Infections, National Reference Laboratory for Sexually Transmitted Infections, Department of Laboratory Medicine.
    Boiko, Iryna
    Clinical Laboratory Department, Ternopil Regional Clinical Dermatovenerologic Dispensary, Ternopil, Ukraine.
    Golparian, Daniel
    Örebro University, School of Medical Sciences. WHO Collaborating Centre for Gonorrhoea and Other Sexually Transmitted Infections, National Reference Laboratory for Sexually Transmitted Infections, Department of Laboratory Medicine.
    Blondeel, Karel
    Department of Reproductive Health and Research, World Health Organization (WHO), Geneva, Switzerland.
    Kiarie, James
    Department of Reproductive Health and Research, World Health Organization (WHO), Geneva, Switzerland.
    Toskin, Igor
    Department of Reproductive Health and Research, World Health Organization (WHO), Geneva, Switzerland.
    Peeling, Rosanna W.
    London School of Hygiene and Tropical Medicine (LSHTM), London, UK.
    Unemo, Magnus
    Örebro University, School of Medical Sciences. Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and Other Sexually Transmitted Infections, National Reference Laboratory for Sexually Transmitted Infections, Department of Laboratory Medicine.
    WHO laboratory validation of Xpert((R)) CT/NG and Xpert((R)) TV on the GeneXpert system verifies high performances2018In: Acta Pathologica, Microbiologica et Immunologica Scandinavica (APMIS), ISSN 0903-4641, E-ISSN 1600-0463, Vol. 126, no 12, p. 907-912Article in journal (Refereed)
    Abstract [en]

    Effective tests for diagnosis of sexually transmitted infections (STIs), used point of care to inform treatment and management decisions, are urgently needed. We evaluated the analytical sensitivity and specificity of the Xpert((R)) CT/NG and Xpert((R)) TV tests, examining 339 samples spiked with phenotypically and/or genetically diverse strains of Neisseria gonorrhoeae, Chlamydia trachomatis, and Trichomonas vaginalis, and other related species that may cross-react. The APTIMA Combo 2 test and APTIMA TV test were used as reference tests. The analytical sensitivity for all three agents in the Xpert((R)) CT/NG and Xpert((R)) TV tests was <= 10(2) genome equivalents/reaction. The analytical specificity of both tests was high. False-positive results were identified in the Xpert((R)) TV test when challenging with high concentrations of Trichomonas tenax, Trichomonas gallinae, Trichomonas stableri, and Trichomonas aotus. However, the clinical relevance of these cross-reactions can likely be neglected, because these species have not been identified in urogenital samples from humans. In conclusion, the analytical sensitivity and specificity of the user-friendly Xpert((R)) CT/NG and Xpert((R)) TV tests on the GeneXpert system were high. The results support the use of specimens from also extra-genital sites, for example, pharynx and rectum. However, appropriate clinical validations are additionally required.

  • 26.
    Jacobsson, Susanne
    et al.
    Örebro University Hospital. National Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Örebro University Hospital, Örebro, Sweden.
    Golparian, Daniel
    National Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Örebro University Hospital, Örebro, Sweden.
    Alm, Richard A.
    Infection IMed, AstraZeneca RandD Boston, Waltham MA, United States.
    Huband, Michael
    Infection IMed, AstraZeneca RandD Boston, Waltham MA, United States.
    Mueller, John
    Infection IMed, AstraZeneca RandD Boston, Waltham MA, United States.
    Jensen, Jorgen Skov
    Statens Serum Institut, Copenhagen, Denmark.
    Ohnishi, Makoto
    National Institute of Infectious Diseases, Tokyo, Japan.
    Unemo, Magnus
    Örebro University Hospital. National Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Örebro University Hospital, Örebro, Sweden.
    High In Vitro Activity of the Novel Spiropyrimidinetrione AZD0914, a DNA Gyrase Inhibitor, against Multidrug-Resistant Neisseria gonorrhoeae Isolates Suggests a New Effective Option for Oral Treatment of Gonorrhea2014In: Antimicrobial Agents and Chemotherapy, ISSN 0066-4804, E-ISSN 1098-6596, Vol. 58, no 9, p. 5585-5588Article in journal (Refereed)
    Abstract [en]

    We evaluated the activity of the novel spiropyrimidinetrione AZD0914 (DNA gyrase inhibitor) against clinical gonococcal isolates and international reference strains (n = 250), including strains with diverse multidrug resistance and extensive drug resistance. The AZD0914 MICs were substantially lower than those of most other currently or previously recommended antimicrobials. AZD0914 should be further evaluated, including in vitro selection, in vivo emergence and mechanisms of resistance, pharmacokinetics/pharmacodynamics in humans, optimal dosing, and performance, in appropriate randomized and controlled clinical trials.

  • 27.
    Jacobsson, Susanne
    et al.
    Örebro University, School of Medical Sciences.
    Golparian, Daniel
    Örebro University, School of Medical Sciences.
    Cole, Michelle
    Public Health England, London, UK.
    Spiteri, Gianfranco
    European Centre for Disease Prevention and Control, Stockholm, Sweden.
    Martin, Irene
    Public Health Agency of Canada, Winnipeg, Canada.
    Bergheim, Thea
    Oslo University Hospital Ullevål, Oslo, Norway.
    Borrego, Maria José
    National Institute of Health, Lisbon, Portugal.
    Crowley, Brendan
    St James’s Hospital, Dublin, Ireland.
    Crucitti, Tania
    Institute of Tropical Medicine, Antwerp, Belgium.
    Van Dam, Alje P.
    Public Health Service Amsterdam, Amsterdam, The Netherlands.
    Hoffmann, Steen
    Statens Serum Institut, Copenhagen, Denmark.
    Jeverica, Samo
    Institute of Microbiology and Immunology, University of Ljubljana, Ljubljana, Slovenia.
    Kohl, Peter
    Vivantes Klinikum Neukölln, Berlin, Germany.
    Mlynarczyk-Bonikowska, Beata
    Medical University of Warsaw, Warsaw, Poland.
    Pakarna, Gatis
    Infectology Centre of Latvia, Riga, Latvia.
    Stary, Angelika
    Outpatients’ Centre for Infectious Venereodermatological Diseases, Vienna, Austria.
    Stefanelli, Paola
    Istituto Superiore di Sanitá, Rome, Italy.
    Pavlik, Peter
    HPL Laboratory Ltd, Bratislava, Slovakia.
    Tzelepi, Eva
    Hellenic Pasteur Institute, Athens, Greece.
    Abad, Raquel
    Institute of Health Carlos III, Madrid, Spain.
    Harris, Simon R.
    Pathogen Genomics, Wellcome Trust Genome Campus, The Wellcome Trust Sanger Institute, Hinxton, UK.
    Unemo, Magnus
    Örebro University, School of Health Sciences.
    WGS analysis and molecular resistance mechanisms of azithromycin-resistant (MIC >2 mg/L) Neisseria gonorrhoeae isolates in Europe from 2009 to 20142016In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 71, no 11, p. 3109-3116Article in journal (Refereed)
    Abstract [en]

    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.

  • 28.
    Jacobsson, Susanne
    et al.
    Örebro University, School of Medical Sciences. Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and Other Sexually Transmitted Infections, National Reference Laboratory for Sexually Transmitted Infections, Department of Laboratory Medicine.
    Golparian, Daniel
    Örebro University, School of Medical Sciences. WHO Collaborating Centre for Gonorrhoea and Other Sexually Transmitted Infections, National Reference Laboratory for Sexually Transmitted Infections, Department of Laboratory Medicine.
    Scangarella-Oman, Nicole
    GlaxoSmithKline, Collegeville PA, USA.
    Unemo, Magnus
    Örebro University, School of Medical Sciences. Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and Other Sexually Transmitted Infections, National Reference Laboratory for Sexually Transmitted Infections, Department of Laboratory Medicine.
    In vitro activity of the novel triazaacenaphthylene gepotidacin (GSK2140944) against MDR Neisseria gonorrhoeae2018In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 73, no 8, p. 2072-2077Article in journal (Refereed)
    Abstract [en]

    Objectives: Increased antimicrobial resistance surveillance and new effective antimicrobials are crucial to maintain treatable gonorrhoea. We examined the in vitro activity of gepotidacin, a novel triazaacenaphthylene, and the effect of efflux pump inactivation on clinical Neisseria gonorrhoeae isolates and international reference strains (n = 252) and compared gepotidacin with antimicrobials currently or previously recommended for gonorrhoea treatment.

    Methods: MICs (mg/L) were determined by agar dilution (gepotidacin) or by Etest (seven other antimicrobials). The gyrA and parC genes were sequenced and the impact of inactivation of the MtrCDE, MacAB and NorM efflux pumps on gepotidacin MICs was examined.

    Results: Gepotidacin showed potent in vitro activity against all gonococcal isolates (n = 252; MIC <= 4 mg/L). The modal MIC, MIC50 , MIC90 and MIC range of gepotidacin were 0.5, 0.5, 1 and 0.032-4 mg/L, respectively. Inactivation of the MtrCDE efflux pump, but not MacAB or NorM, decreased the gepotidacin MICs for most strains. No significant cross-resistance between gepotidacin and any other antimicrobials, including the fluoroquinolone ciprofloxacin, was identified. However, the ParC D86N mutation (possibly together with additional antimicrobial resistance mutation), which is associated with fluoroquinolone resistance, was associated with increased gepotidacin MICs.

    Conclusions: Gepotidacin demonstrated high in vitro activity against gonococcal strains, indicating that gepotidacin could potentially be an effective option for gonorrhoea treatment, particularly in a dual antimicrobialtherapy regimen and for patients with resistance or allergy to extended-spectrum cephalosporins. Nevertheless, elucidating in vitro and in vivo resistance emergence and mechanisms in detail, together with further gonorrhoea clinical studies, ideally also including chlamydia and Mycoplasma genitalium are essential.

  • 29.
    Jacobsson, Susanne
    et al.
    Örebro University, School of Medical Sciences. WHO Collaborating Centre for Gonorrhoea and Other Sexually Transmitted Infections, National Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Faculty of Medicine and Health, , Örebro University Hospital, Örebro, Sweden.
    Paukner, Susanne
    Nabriva Therapeutics AG, Vienna, Austria.
    Golparian, Daniel
    Örebro University, School of Medical Sciences. WHO Collaborating Centre for Gonorrhoea and Other Sexually Transmitted Infections, National Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Örebro University Hospital, Örebro, Sweden.
    Jensen, Jörgen S.
    Department of Microbiology and Infection Control, Sexually Transmitted Infections, Research and Development, Statens Serum Institut, Copenhagen, Denmark.
    Unemo, Magnus
    WHO Collaborating Centre for Gonorrhoea and Other Sexually Transmitted Infections, National Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Örebro University Hospital, Örebro, Sweden.
    In Vitro Activity of the Novel Pleuromutilin Lefamulin (BC-3781) and Effect of Efflux Pump Inactivation on Multidrug-Resistant and Extensively Drug-Resistant Neisseria gonorrhoeae2017In: Antimicrobial Agents and Chemotherapy, ISSN 0066-4804, E-ISSN 1098-6596, Vol. 61, no 11, article id e01497-17Article in journal (Refereed)
    Abstract [en]

    We evaluated the activity of the novel semisynthetic pleuromutilin lefamulin, inhibiting protein synthesis and growth, and the effect of efflux pump inactivation on clinical gonococcal isolates and reference strains (n = 251), including numerous multidrug-resistant and extensively drug-resistant isolates. Lefamulin showed potent activity against all gonococcal isolates, and no significant cross-resistance to other antimicrobials was identified. Further studies of lefamulin are warranted, including in vitro selection and mechanisms of resistance, pharmacokinetics/pharmacodynamics, optimal dosing, and performance in randomized controlled trials.

  • 30.
    Jeverica, Samo
    et al.
    Fac Med, Inst Microbiol & Immunol, Univ Ljubljana, Ljubljana, Slovenia.
    Golparian, Daniel
    WHO Collaborating Centre for Gonorrhoea and Other STIs, National Reference Laboratory for Pathogenic Neisseria, Department of Lab. Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Hanzelka, Brian
    Vertex Pharmaceut Inc, Boston MA, USA.
    Fowlie, Andrew J.
    Vertex Pharmaceut Inc, Boston MA, USA.
    Maticic, Mojca
    Clin Infect Dis & Febrile Illnesses, Univ Med Ctr Ljubljana, Ljubljana, Slovenia..
    Unemo, Magnus
    Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and Other STIs, National Reference Laboratory for Pathogenic Neisseria, Department of Lab. Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    High in vitro activity of a novel dual bacterial topoisomerase inhibitor of the ATPase activities of GyrB and ParE (VT12-008911) against Neisseria gonorrhoeae isolates with various high-level antimicrobial resistance and multidrug resistance2014In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 69, no 7, p. 1866-1872Article in journal (Refereed)
    Abstract [en]

    Clinical resistance to the currently recommended extended-spectrum cephalosporins (ESCs), the last remaining options for empirical antimicrobial monotherapy of gonorrhoea globally, has been reported. New antimicrobials are essential to avoid the emergence of untreatable gonorrhoea. We have investigated the in vitro activity of a novel dual bacterial topoisomerase inhibitor of the ATPase activities of GyrB and ParE (Vertex aminobenzimidazole VT12-008911), compared with antimicrobials currently or previously recommended for gonorrhoea treatment.

    MICs were determined using agar dilution (VT12-008911) or Etest (seven antimicrobials) for international reference strains (naEuroS=aEuroS28) and clinical Neisseria gonorrhoeae isolates (naEuroS=aEuroS220). The latter included three extensively drug-resistant isolates with high-level ceftriaxone resistance, additional isolates with clinical ESC resistance and a high number of isolates with ciprofloxacin resistance and multidrug resistance.

    The MIC50, MIC90 and MIC range of VT12-008911 were 0.064, 0.125 and a parts per thousand currency sign0.002-0.25 mg/L, respectively. One-hundred and seventy (69%) isolates were ciprofloxacin resistant; however, only 54 of those isolates had a VT12-008911 MIC > 0.064 mg/L (47 and 7 with MICaEuroS=aEuroS0.125 mg/L and MICaEuroS=aEuroS0.25 mg/L, respectively). The in vitro activity of VT12-008911 was superior to that of ciprofloxacin and all additional antimicrobials investigated. Time-kill curve analysis showed that VT12-008911 exhibited potent time-dependent bactericidal activity, at or very close to the MIC, against N. gonorrhoeae.

    In vitro results suggest that VT12-008911 might be an effective treatment option for gonorrhoea. However, it will be important to detail the pharmacokinetics/pharmacodynamics, toxicity, selection and mechanisms of VT12-008911 resistance in N. gonorrhoeae and, finally, to perform well-designed in vivo randomized clinical trials.

  • 31.
    Jeverica, Samo
    et al.
    Fac Med, Inst Microbiol & Immunol, Univ Ljubljana, Ljubljana, Slovenia.
    Golparian, Daniel
    WHO Collaborating Centre for Gonorrhoea and other STIs, National Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Maticic, Mojca
    Clin Infect Dis & Febrile Illnesses, Univ Med Ctr Ljubljana, Ljubljana, Slovenia.
    Potocnik, Marko
    Dept Dermatovenereol, Univ Med Ctr Ljubljana, Ljubljana, Slovenia.
    Mlakar, Bostjan
    Surg Ctr Zdrav Splet, Ljubljana, Slovenia.
    Unemo, Magnus
    Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and other STIs, National Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Phenotypic and molecular characterization of Neisseria gonorrhoeae isolates from Slovenia, 2006-12: rise and fall of the multidrug-resistant NG-MAST genogroup 1407 clone?2014In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 69, no 6, p. 1517-1525Article in journal (Refereed)
    Abstract [en]

    To determine the phenotypic and molecular characteristics of Neisseria gonorrhoeae isolates obtained between 2006 and 2012 in Slovenia.

    Gonococcal isolates obtained between 2006 and 2012 in Slovenia (naEuroS=aEuroS194) were investigated with Etest for susceptibility to cefixime, ceftriaxone, penicillin, ciprofloxacin, azithromycin, tetracycline, gentamicin and spectinomycin. All isolates were examined with N. gonorrhoeae multiantigen sequence typing for molecular epidemiology and sequencing of the major extended-spectrum cephalosporin (ESC) resistance determinants (penA, mtrR and penB) was performed.

    The overall prevalence of decreased susceptibility or resistance to cefixime and ceftriaxone (MIC a parts per thousand yen0.125 mg/L) was 11% and 5%, respectively. The decreased susceptibility or resistance showed an epidemic peak in 2011 (33% for cefixime and 11% for ceftriaxone), decreasing to 6% and 4%, respectively, in 2012. ST1407 (9% of isolates), ST21 (6%) and ST225 (6%) were the most common sequence types (STs) during 2006-12. Genogroup G1407 (ST1407 most prevalent ST), an internationally spread clone with decreased susceptibility or resistance to ESCs, was most prevalent (48%) in 2009. However, the G1407 prevalence then declined: in 2010, 30%; in 2011, 28%; and in 2012, 8%. Instead, in 2012 the ESC- and ciprofloxacin-susceptible G21 was the predominant genogroup (26%).

    The prevalence of gonococcal resistance to ESCs in Slovenia has been high, but fluctuating. Fortunately, in 2012 some ESC- and ciprofloxacin-susceptible clones, such as genogroups G21, G1195 and G2992, appeared to have mainly replaced the multidrug-resistant G1407 clone, a replacement also seen in several European countries.

  • 32.
    Jönsson, Agnez
    et al.
    WHO Collaborating Centre for Gonorrhoea and other STIs, Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Foerster, Sunniva
    WHO Collaborating Centre for Gonorrhoea and other STIs, Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden; Institute for Infectious Diseases, University of Bern, Bern, Switzerland; Institute of Social and Preventive Medicine, University of Bern, Bern, Switzerland.
    Golparian, Daniel
    Örebro University, School of Medical Sciences. WHO Collaborating Centre for Gonorrhoea and other STIs, Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Hamasuna, Ryoichi
    Department of Urology, School of Medicine, University of Occupational and Environmental Health, Kitakyushu, Japan.
    Jacobsson, Susanne
    Örebro University, School of Medical Sciences. Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and other STIs, Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Lindberg, Magnus
    Örebro University, School of Medical Sciences. Department of Dermatovenerology, Faculty of Health and Medical Sciences, Örebro University, Örebro, Sweden.
    Jensen, Jörgen Skov
    Department of Microbiology and Infection Control, Sexually Transmitted Infections, Research and Development, Statens Serum Institut, Copenhagen, Denmark.
    Ohnishi, Makoto
    Department of Bacteriology I, National Institute of Infectious Diseases, Tokyo, Japan.
    Unemo, Magnus
    WHO Collaborating Centre for Gonorrhoea and other STIs, Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    In vitro activity and time-kill curve analysis of sitafloxacin against a global panel of antimicrobial-resistant and multidrug-resistant Neisseria gonorrhoeae isolates2018In: Acta Pathologica, Microbiologica et Immunologica Scandinavica (APMIS), ISSN 0903-4641, E-ISSN 1600-0463, Vol. 126, no 1, p. 29-37Article in journal (Refereed)
    Abstract [en]

    Treatment of gonorrhoea is a challenge worldwide because of emergence of resistance in N. gonorrhoeae to all therapeutic antimicrobials available and novel antimicrobials are imperative. The newer-generation fluoroquinolone sitafloxacin, mostly used for respiratory tract infections in Japan, can have a high in vitro activity against gonococci. However, only a limited number of recent antimicrobial-resistant isolates from Japan have been examined. We investigated the sitafloxacin activity against a global gonococcal panel (250 isolates cultured in 1991-2013), including multidrug-resistant geographically, temporally and genetically diverse isolates, and performed time-kill curve analysis for sitafloxacin. The susceptibility to sitafloxacin (agar dilution) and seven additional therapeutic antimicrobials (Etest) was determined. Sitafloxacin was rapidly bactericidal, and the MIC range, MIC50 and MIC90 was ≤0.001-1, 0.125 and 0.25 mg/L, respectively. There was a high correlation between the MICs of sitafloxacin and ciprofloxacin; however, the MIC50 and MIC90 of sitafloxacin were 6-fold and >6-fold lower, respectively. Sitafloxacin might be an option for particularly dual antimicrobial therapy of gonorrhoea and for cases with ceftriaxone resistance or allergy. However, further in vitro and particularly in vivo evaluations of potential resistance, pharmacokinetics/pharmacodynamics and ideal dosing for gonorrhoea, as well as performance of randomized controlled clinical, trials are crucial.

  • 33.
    Lebedzeu, Fiodar
    et al.
    The Republican Research and Practical Center for Epidemiology and Microbiology (RRPCEM), Minsk, Byelarus.
    Golparian, Daniel
    WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, Swedish Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Titov, Leonid
    The Republican Research and Practical Center for Epidemiology and Microbiology (RRPCEM), Minsk, Byelarus.
    Pankratava, Nataliya
    Mogilev Reg Dermato Venerol Dispensary, Mogilyov, Byelarus.
    Glazkova, Slavyana
    The Republican Research and Practical Center for Epidemiology and Microbiology (RRPCEM), Minsk, Byelarus.
    Shimanskaya, Irina
    Mogilev Reg Dermato Venerol Dispensary, Minsk, Byelarus.
    Charniakova, Natallia
    Mogilev Reg Dermato Venerol Dispensary, Vitebsk, Byelarus.
    Lukyanau, Aliaksandr
    The Republican Research and Practical Center for Epidemiology and Microbiology (RRPCEM), Minsk, Byelarus.
    Domeika, Marius
    Dept Prevent & Control Communicable Dis, Uppsala Cty Council, Uppsala, Sweden.
    Unemo, Magnus
    Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, Swedish Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Antimicrobial susceptibility/resistance and NG-MAST characterisation of Neisseria gonorrhoeae in Belarus, Eastern Europe, 2010-20132015In: BMC Infectious Diseases, ISSN 1471-2334, E-ISSN 1471-2334, Vol. 15, article id 29Article in journal (Refereed)
    Abstract [en]

    Background: Gonorrhoea and widely spread antimicrobial resistance (AMR) in its etiological agent Neisseria gonorrhoeae are major public health concerns worldwide. Gonococcal AMR surveillance nationally and internationally, to identify emerging resistance and inform treatment guidelines, is imperative for public health purposes. In 2009, AMR surveillance was initiated in Belarus, Eastern Europe because no gonococcal AMR data had been available for at least two decades. Herein, the prevalence and trends of gonococcal AMR and molecular epidemiological characteristics of N. gonorrhoeae strains from 2010 to 2013 in Belarus, are described. Methods: N. gonorrhoeae isolates (n = 193) obtained in the Mogilev (n = 142), Minsk (n = 36) and Vitebsk (n = 15) regions of Belarus in 2010 (n = 72), 2011 (n = 6), 2012 (n = 75) and 2013 (n = 40) were analyzed in regards to AMR using the Etest method and for molecular epidemiology with N. gonorrhoeae multi-antigen sequence typing (NG-MAST). Results: During 2010-2013, the proportions of resistant N. gonorrhoeae isolates were as follows: tetracycline 36%, ciprofloxacin 28%, penicillin G 9%, azithromycin 5%, and cefixime 0.5%. Only one (0.5%) beta-lactamase producing isolate was detected. No isolates resistant to ceftriaxone and spectinomycin were identified. Overall, the resistance levels to tetracycline, ciprofloxacin and penicillin G were relatively stable. Interestingly, the level of resistance to azithromycin declined from 12% in 2010 to 0% in 2013 (P < 0.05). In total, 70 NG-MAST STs were identified. The predominant STs were ST1993 (n = 53), ST807 (n = 13), ST285 (n = 8) and ST9735 (n = 8). Many novel STs (n = 43, 61%), representing 41% of all isolates, were found. Conclusions: During 2010-2013, the N. gonorrhoeae population in Belarus displayed high and relatively stable resistance levels to tetracycline, ciprofloxacin, and penicillin G, while the resistance to azithromycin declined. One isolate was resistant to cefixime, but no resistance to ceftriaxone or spectinomycin was found. The results of the present surveillance initiated in 2009 were also used to replace penicillin G with ceftriaxone (1 g single dose intramuscularly) as the first-line drug for empiric treatment of gonorrhoea in the national treatment guidelines in Belarus in late 2009. It is essential to further strengthen the surveillance of gonococcal AMR and ideally survey also treatment failures and molecular epidemiological genotypes in Belarus.

  • 34.
    Lewis, David A.
    et al.
    Ctr HIV & Sexually Transmitted Infect, Natl Hlth Lab Serv, Natl Inst Communicable Dis, Johannesburg, South Africa; Dept Internal Med, Fac Hlth Sci, University of Witwatersrand, Johannesburg, South Africa; Sch Med, Div Med Microbiol, Univ Cape Town, Cape Town, South Africa.
    Sriruttan, Charlotte
    Dept Clin Microbiol, Ampath Natl Lab Serv, Centurion, South Africa.
    Mueller, Etienne E.
    Ctr HIV & Sexually Transmitted Infect, Natl Hlth Lab Serv, Natl Inst Communicable Dis, Johannesburg, South Africa.
    Golparian, Daniel
    Dept Lab Med, Swedish Reference Lab Pathogen Neisseria, WHO Collaborating Ctr Gonorrhoea & Other STIs, Örebro University Hospital, Örebro, Sweden.
    Gumede, Lindy
    Ctr HIV & Sexually Transmitted Infect, Natl Hlth Lab Serv, Natl Inst Communicable Dis, Johannesburg, South Africa.
    Fick, Donald
    Meldene Medicross Clin, Melville, South Africa.
    de Wet, Johan
    Springs Medicross Clin, Springs, South Africa.
    Maseko, Venessa
    Ctr HIV & Sexually Transmitted Infect, Natl Hlth Lab Serv, Natl Inst Communicable Dis, Johannesburg, South Africa.
    Coetzee, Jennifer
    Dept Clin Microbiol, Ampath Natl Lab Serv, Centurion, South Africa.
    Unemo, Magnus
    Örebro University Hospital. Dept Lab Med, Swedish Reference Lab Pathogen Neisseria, WHO Collaborating Ctr Gonorrhoea & Other STIs, Örebro University Hospital, Örebro, Sweden.
    Phenotypic and genetic characterization of the first two cases of extended-spectrum-cephalosporin-resistant Neisseria gonorrhoeae infection in South Africa and association with cefixime treatment failure2013In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 68, no 6, p. 1267-1270Article in journal (Refereed)
    Abstract [en]

    To describe the phenotypic and genetic characteristics of the first two cases of extended-spectrum cephalosporin (ESC)-resistant Neisseria gonorrhoeae in South Africa, one of which was associated with verified cefixime treatment failure. Two ESC-resistant N. gonorrhoeae isolates were cultured from the urethral discharge of two men who have sex with men (MSM). One man reported a persistent urethral discharge that had failed to respond to previous therapy with oral cefixime. Agar dilution MICs were determined for eight antibiotics. -Lactam-associated resistance mutations were identified through PCR-based amplification and sequencing for several key genes: penA, mtrR and its promoter, porB1b (penB), ponA and pilQ. For molecular epidemiological characterization, full-length porB gene sequencing, N. gonorrhoeae multiantigen sequence typing (NG-MAST) and multilocus sequence typing (MLST) were performed. Both isolates were resistant to cefixime, ciprofloxacin, penicillin and tetracycline and intermediate/resistant to azithromycin, but susceptible to ceftriaxone, gentamicin and spectinomycin. Both isolates had the type XXXIV penA mosaic allele in addition to previously described resistance mutations in the mtrR promoter (A deletion), porB1b (penB) (G101K and A102N) and ponA1 (L421P). Both isolates had an identical NG-MAST sequence type (ST4822) and MLST sequence type (ST1901). Both isolates were resistant to cefixime and possessed a number of identical mutations in key genes contributing to ESC resistance in N. gonorrhoeae. The two isolates contained the type XXXIV penA mosaic allele and belonged to a successful international MSM-linked multidrug-resistant gonococcal clone (MLST ST1901) associated with several cefixime treatment failures in Europe and North America.

  • 35.
    Mlynarczyk-Bonikowska, Beata
    et al.
    Dept Dermatol & Venereol, Med Univ Warsaw, Warsaw, Poland.
    Serwin, Agnieszka Beata
    Dept Dermatol & Venereol, Med Univ Bialystok, Bialystok, Poland.
    Golparian, Daniel
    WHO Collaborating Centre for Gonorrhoea and other STIs, National Reference Lab. for Pathogenic Neisseria, Dept. of Lab. Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    de Walthoffen, Szymon Walter
    Dept Med Microbiol, Med Univ Warsaw, Warsaw, Poland.
    Majewski, Slawomir
    Dept Dermatol & Venereol, Med Univ Warsaw, Warsaw, Poland.
    Koper, Marta
    Dept Dermatol & Venereol, Med Univ Bialystok, Bialystok, Poland.
    Malejczyk, Magdalena
    Dept Dermatol & Venereol, Med Univ Warsaw, Warsaw, Poland.
    Domeika, Marius
    Dept Prevent & Control Communicable Dis, Uppsala Cty Council, Uppsala, Sweden.
    Unemo, Magnus
    Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and other STIs, National Reference Lab. for Pathogenic Neisseria, Dept. of Lab. Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Antimicrobial susceptibility/resistance and genetic characteristics of Neisseria gonorrhoeae isolates from Poland, 2010-20122014In: BMC Infectious Diseases, ISSN 1471-2334, E-ISSN 1471-2334, Vol. 14, article id 65Article in journal (Refereed)
    Abstract [en]

    Background: In Poland, gonorrhoea has been a mandatorily reported infection since 1948, however, the reported incidences are likely underestimated. No antimicrobial resistance (AMR) data for Neisseria gonorrhoeae has been internationally reported in nearly four decades, and data concerning genetic characteristics of N. gonorrhoeae are totally lacking. The aims of this study were to investigate the AMR to previously and currently recommended gonorrhoea treatment options, the main genetic resistance determinant (penA) for extended-spectrum cephalosporins (ESCs), and genotypic distribution of N. gonorrhoeae isolates in Poland in 2010-2012.

    Methods: N. gonorrhoeae isolates cultured in 2010 (n = 28), 2011 (n = 92) and 2012 (n = 108) in Warsaw and Bialystok, Poland, were examined using antimicrobial susceptibility testing (Etest), pyrosequencing of penA and N. gonorrhoeae multi-antigen sequence typing (NG-MAST).

    Results: The proportions of N. gonorrhoeae isolates showing resistance were as follows: ciprofloxacin 61%, tetracycline 43%, penicillin G 22%, and azithromycin 8.8%. No isolates resistant to ceftriaxone, cefixime or spectinomycin were found. However, the proportion of isolates with an ESC MIC = 0.125 mg/L, i.e. at the resistance breakpoint, increased significantly from none in 2010 to 9.3% and 19% in 2012 for ceftriaxone and cefixime, respectively. Furthermore, 3.1% of the isolates showed multidrug resistance, i.e., resistance to ciprofloxacin, penicillin G, azithromycin, and decreased susceptibility to cefixime (MIC = 0.125 mg/L). Seventy-six isolates (33%) possessed a penA mosaic allele and 14 isolates (6.1%) contained an A501V/T alteration in penicillin-binding protein 2. NG-MAST ST1407 (n = 58, 25% of isolates) was the most prevalent ST, which significantly increased from 2010 (n = 0) to 2012 (n = 46; 43%).

    Conclusions: In Poland, the diversified gonococcal population displayed a high resistance to most antimicrobials internationally previously recommended for gonorrhoea treatment and decreasing susceptibility to the currently recommended ESCs. The decreasing susceptibility to ESCs was mostly due to the introduction of the internationally spread multidrug-resistant NG-MAST ST1407 in 2011. It is essential to promptly revise the gonorrhoea treatment guidelines, improve the gonorrhoea laboratory diagnostics, and implement quality assured surveillance of gonococcal AMR (ideally also treatment failures) in Poland.

  • 36.
    Muhammad, Ibrahim
    et al.
    Department of Laboratory Medicine, WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, National Reference Laboratory for Pathogenic Neisseria, Microbiology, Örebro Univ. Hospital, Örebro, Sweden.
    Golparian, Daniel
    Department of Laboratory Medicine, WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, National Reference Laboratory for Pathogenic Neisseria, Microbiology, Örebro Univ. Hospital, Örebro, Sweden.
    Dillon, Jo-Anne R
    Department of Microbiology and Immunology, College of Medicine, University of Saskatchewan, Saskatoon, Canada.
    Johansson, Åsa
    Department of Clinical Microbiology, Central Hospital, Växjö, Sweden.
    Ohnishi, Makoto
    National Institute of Infectious Diseases, Tokyo, Japan.
    Sethi, Sunil
    Department of Medical Microbiology, Post Graduate Institute of Medical Education and Research (PGIMER), Chandigarh, India.
    Chen, Shao-chun
    National Center for STD Control, Chinese Centers for Disease Control and Prevention, Nanjing, China.
    Nakayama, Shu-ichi
    National Institute of Infectious Diseases, Tokyo, Japan.
    Sundqvist, Martin
    Örebro University Hospital. Department of Laboratory Medicine, WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, National Reference Laboratory for Pathogenic Neisseria, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Bala, Manju
    Apex Regional STD Teaching, Training and Research Centre, VMMC and Safdarjang Hospital, New Delhi, India.
    Unemo, Magnus
    Örebro University Hospital. Department of Laboratory Medicine, WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, National Reference Laboratory for Pathogenic Neisseria, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Characterisation of blaTEM genes and types of β-lactamase plasmids in Neisseria gonorrhoeae - the prevalent and conserved blaTEM-135 has not recently evolved and existed in the Toronto plasmid from the origin2014In: BMC Infectious Diseases, ISSN 1471-2334, E-ISSN 1471-2334, Vol. 14, no 1, article id 454Article in journal (Refereed)
    Abstract [en]

    BACKGROUND: Antimicrobial resistance (AMR) in Neisseria gonorrhoeae is a major concern worldwide. It has been recently feared that the blaTEM-1 gene is, via blaTEM-135, evolving into an extended-spectrum β-lactamase (ESBL), which could degrade all cephalosporins including ceftriaxone. The aims of the present study were to characterize the blaTEM genes, types of β-lactamase plasmids, the degradation of ampicillin by TEM-135 compared to TEM-1, and to perform molecular epidemiological typing of β-lactamase-producing N. gonorrhoeae strains internationally.

    METHODS: β-lactamase producing N. gonorrhoeae isolates (n = 139) cultured from 2000 to 2011 in 15 countries were examined using antibiograms, blaTEM gene sequencing, β-lactamase plasmid typing, and N. gonorrhoeae multiantigen sequence typing (NG-MAST). Furthermore, the blaTEM gene was sequenced in the first described Toronto plasmid (pJD7), one of the first Asian plasmids (pJD4) and African plasmids (pJD5) isolated in Canada. The degradation of ampicillin by TEM-135 compared to TEM-1 was examined using a MALDI-TOF MS hydrolysis assay.

    RESULTS: Six different blaTEM sequences were identified (among isolates with 125 different NG-MAST STs), i.e. blaTEM-1 (in 104 isolates), blaTEM-135 (in 30 isolates), and four novel blaTEM sequences (in 5 isolates). The blaTEM-1 allele was only found in the African and Asian plasmids, while all Rio/Toronto plasmids possessed the blaTEM-135 allele. Most interesting, the first described gonococcal Toronto plasmid (pJD7), identified in 1984, also possessed the highly conserved blaTEM-135 allele. The degradation of ampicillin by TEM-135 compared to TEM-1 was indistinguishable in the MALDI-TOF MS hydrolysis assay.

    CONCLUSIONS: blaTEM-135, encoding TEM-135, is predominantly and originally associated with the Rio/Toronto plasmid and prevalent among the β-lactamase producing gonococcal strains circulating globally. blaTEM-135 does not appear, as previously hypothesized, to have recently evolved due to some evolutionary selective pressure, for example, by the extensive use of extended-spectrum cephalosporins worldwide. On the contrary, the present study shows that blaTEM-135 existed in the Toronto plasmid from its discovery and that blaTEM-135 is highly conserved (not further evolved in the past >30 years). Nevertheless, international studies for monitoring the presence of different blaTEM alleles, the possible evolution of the blaTEM-135 allele, and the types of β-lactamase producing plasmids, remain imperative.

  • 37.
    Olsen, Birgitta
    et al.
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden.
    Thi Lan, Pham
    Department of Dermato-Venerology, Hanoi Medical University, Hanoi, Vietnam; National Hospital of Dermatology and Venerology, Hanoi, Vietnam.
    Golparian, Daniel
    WHO Collaborating Centre for Gonorrhoea and Other STIs, National Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Clinical Microbiology, Örebro University Hospital, Örebro, Sweden.
    Johansson, Emma
    WHO Collaborating Centre for Gonorrhoea and Other STIs, National Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Clinical Microbiology, Örebro University Hospital, Örebro, Sweden.
    Khang, Tran Hau
    Department of Dermato-Venerology, Hanoi Medical University, Hanoi, Vietnam; National Hospital of Dermatology and Venerology, Hanoi, Vietnam.
    Unemo, Magnus
    Örebro University Hospital. Örebro University, School of Medical Sciences. WHO Collaborating Centre for Gonorrhoea and Other STIs, National Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Clinical Microbiology.
    Antimicrobial susceptibility and genetic characteristics of Neisseria gonorrhoeae isolates from Vietnam, 20112013In: BMC Infectious Diseases, ISSN 1471-2334, E-ISSN 1471-2334, Vol. 13, no 1, article id 40Article in journal (Refereed)
    Abstract [en]

    BACKGROUND: Antimicrobial resistance (AMR) in Neisseria gonorrhoeae is a major public health concern worldwide. In Vietnam, knowledge regarding N. gonorrhoeae prevalence and AMR is limited, and data concerning genetic characteristics of N. gonorrhoeae is totally lacking. Herein, we investigated the phenotypic AMR (previous, current and possible future treatment options), genetic resistance determinants for extended-spectrum cephalosporins (ESCs), and genotypic distribution of N. gonorrhoeae isolated in 2011 in Hanoi, Vietnam.

    METHODS: N. gonorrhoeae isolates from Hanoi, Vietnam isolated in 2011 (n = 108) were examined using antibiograms (Etest for 10 antimicrobials), Neisseria gonorrhoeae multi-antigen sequence typing (NG-MAST), and sequencing of ESC resistance determinants (penA, mtrR and penB).

    RESULTS: The levels of in vitro resistance were as follows: ciprofloxacin 98%, tetracycline 82%, penicillin G 48%, azithromycin 11%, ceftriaxone 5%, cefixime 1%, and spectinomycin 0%. The MICs of gentamicin (0.023-6 mg/L), ertapenem (0.002-0.125 mg/L) and solithromycin (<0.016-0.25 mg/L) were relatively low. No penA mosaic alleles were found, however, 78% of the isolates contained an alteration of amino acid A501 (A501V (44%) and A501T (34%)) in the encoded penicillin-binding protein 2. A single nucleotide (A) deletion in the inverted repeat of the promoter region of the mtrR gene and amino acid alterations in MtrR was observed in 91% and 94% of the isolates, respectively. penB resistance determinants were detected in 87% of the isolates. Seventy-five different NG-MAST STs were identified, of which 59 STs have not been previously described.

    CONCLUSIONS: In Vietnam, the highly diversified gonococcal population displayed high in vitro resistance to antimicrobials previously recommended for gonorrhoea treatment (with exception of spectinomycin), but resistance also to the currently recommended ESCs were found. Nevertheless, the MICs of three potential future treatment options were low. It is essential to strengthen the diagnostics, case reporting, and epidemiologic surveillance of gonorrhoea in Vietnam. Furthermore, the surveillance of gonococcal AMR and gonorrhoea treatment failures is imperative to reinforce. Research regarding novel antimicrobial treatment strategies (e.g., combination therapy) and new antimicrobials is crucial for future treatment of gonorrhoea.

  • 38.
    Pellrud, Helena
    et al.
    STD Clinic, Department of Dermatovenereology, Örebro University Hospital, Örebro, Sweden.
    Golparian, Daniel
    WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, Swedish Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Clinical Microbiology, Örebro University Hospital, Örebro, Sweden.
    Nilsson, Christian Steezko
    STD Clinic, Department of Dermatovenereology, Örebro University Hospital, Örebro, Sweden.
    Falk, My
    STD Clinic, Department of Dermatovenereology, Örebro University Hospital, Örebro, Sweden.
    Fredlund, Hans
    Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, Swedish Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Clinical Microbiology, Örebro University Hospital, Örebro, Sweden.
    Unemo, Magnus
    Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, Swedish Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Clinical Microbiology, Örebro University Hospital, Örebro, Sweden.
    Trichomonas vaginalis Infections are Rare Among Young Patients Attending an STI Clinic in Sweden2015In: Acta Dermato-Venereologica, ISSN 0001-5555, E-ISSN 1651-2057, Vol. 95, no 3, p. 343-344Article in journal (Refereed)
  • 39.
    Rumyantseva, Tatiana
    et al.
    Department of Molecular Diagnostics, Central Research Institute for Epidemiology, Moscow, Russian Federation.
    Golparian, Daniel
    WHO Collaborating Centre for Gonorrhoea and Other STIs, National Reference Laboratory for Pathogenic Neisseria, , Örebro University Hospital, Örebro, Sweden; Department of Laboratory Medicine, Microbiology, Faculty of Medicine and Health, Örebro University, Sweden.
    Nilsson, Christian S.
    Department of Dermatovenereology, Örebro University Hospital, Örebro, Sweden.
    Johansson, Emma
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden. WHO Collaborating Ctr Gonorrhoea & Other STIs, Natl Reference Lab Pathogen Neisseria, Dept Lab Med, Microbiol, Örebro University Hospital, Örebro, Sweden.
    Falk, My
    Department of Dermatovenereology, Örebro University Hospital, Örebro, Sweden.
    Fredlund, Hans
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden. WHO Collaborating Ctr Gonorrhoea & Other STIs, Natl Reference Lab Pathogen Neisseria, Dept Lab Med, Microbiol, Örebro University Hospital, Örebro, Sweden.
    Van Dam, Alje
    Public Health Laboratory, Amsterdam Health Centre, Amsterdam, Netherlands; Department of Medical Microbiology, Onze Lieve Vrouwe Gasthuis (OLVG), Amsterdam, Netherlands.
    Guschin, Alexander
    Department of Molecular Diagnostics, Central Research Institute for Epidemiology, Moscow, Russian Federation.
    Unemo, Magnus
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden. WHO Collaborating Ctr Gonorrhoea & Other STIs, Natl Reference Lab Pathogen Neisseria, Dept Lab Med, Microbiol, Örebro University Hospital, Örebro, Sweden.
    Evaluation of the new AmpliSens multiplex real-time PCR assay for simultaneous detection of Neisseriagonorrhoeae, Chlamydiatrachomatis, Mycoplasmagenitalium, and Trichomonasvaginalis2015In: Acta Pathologica, Microbiologica et Immunologica Scandinavica (APMIS), ISSN 0903-4641, E-ISSN 1600-0463, Vol. 123, no 10, p. 879-886Article in journal (Refereed)
    Abstract [en]

    In this study, we performed an evaluation of the new CE-marked multiplex real-time AmpliSens N.gonorrhoeae/C.trachomatis/M.genitalium/T.vaginalis-MULTIPRIME-FRT PCR assay compared to APTIMA tests, i.e., APTIMA COMBO 2assay, APTIMA Trichomonasvaginalis assay (FDA-approved), and two different APTIMA Mycoplasmagenitalium assays (research use only; one of them only used for discrepancy analysis). Vaginal swabs (n=209) and first-void urine (FVU) specimens from females (n=498) and males (n=554), consecutive attendees (n=1261) at a dermatovenerological clinic in Sweden, were examined. The sensitivity of the AmpliSens PCR assay for detection of C.trachomatis (6.3% prevalence), M.genitalium (5.7% prevalence), N.gonorrhoeae (0.3% prevalence), and T.vaginalis (0.08% prevalence) was 97.5% (95% confidence interval (CI): 91.2-99.6%), 81.9% (95% CI: 70.7-89.7%), 100% (95% CI: 40.2-100%) and 100% (95% CI: 16.5-100%), respectively. The specificity of the AmpliSens PCR assay was 100% (95% CI: 99.6-100%) for all agents. The analytical sensitivity and specificity for N.gonorrhoeae detection was excellent, i.e., 55 international gonococcal strains detected and 135 isolates of 13 non-gonococcal Neisseria species were negative. In conclusion, the multiplex real-time AmpliSens N.gonorrhoeae/C.trachomatis/M.genitalium/T.vaginalis-MULTIPRIME-FRT PCR assay demonstrated high sensitivity and excellent specificity for the detection of C.trachomatis, N.gonorrhoeae, and T.vaginalis, and excellent specificity but suboptimal sensitivity for M.genitalium detection.

  • 40.
    Ryan, L.
    et al.
    Department of Clinical Microbiology, St James’s Hospital, Dublin, Ireland.
    Golparian, Daniel
    Örebro University, School of Medical Sciences. WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, Department of Laboratory Medicine, Clinical Microbiology, Örebro University Hospital, Örebro, Sweden.
    Fennelly, N.
    Department of Clinical Microbiology, St James’s Hospital, Dublin, Ireland.
    Rose, L.
    Department of Clinical Microbiology, St James’s Hospital, Dublin, Ireland.
    Walsh, P.
    Department of Computing, Cork Institute of Technology, Cork, Ireland.
    Lawlor, B.
    Department of Computing, Cork Institute of Technology, Cork, Ireland.
    Mac Aogáin, M.
    Department of Clinical Microbiology, Trinity Translational Medicine Institute, School of Medicine, Trinity College Dublin, Dublin, Ireland.
    Unemo, Magnus
    Örebro University, School of Medical Sciences. Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, Department of Laboratory Medicine, Clinical Microbiology.
    Crowley, B.
    Department of Clinical Microbiology, St James’s Hospital, Dublin, Ireland; Department of Virology, St James’s Hospital, Dublin, Ireland.
    Antimicrobial resistance and molecular epidemiology using whole-genome sequencing of Neisseria gonorrhoeae in Ireland, 2014-2016: focus on extended-spectrum cephalosporins and azithromycin2018In: European Journal of Clinical Microbiology and Infectious Diseases, ISSN 0934-9723, E-ISSN 1435-4373, Vol. 37, no 9, p. 1661-1672Article in journal (Refereed)
    Abstract [en]

    High-level resistance and treatment failures with ceftriaxone and azithromycin, the first-line agents for gonorrhoea treatment are reported and antimicrobial-resistant Neisseria gonorrhoeae is an urgent public health threat. Our aims were to determine antimicrobial resistance rates, resistance determinants and phylogeny of N. gonorrhoeae in Ireland, 2014-2016. Overall, 609 isolates from four University Hospitals were tested for susceptibility to extended-spectrum cephalosporins (ESCs) and azithromycin by the MIC Test Strips. Forty-three isolates were whole-genome sequenced based on elevated MICs. The resistance rate to ceftriaxone, cefixime, cefotaxime and azithromycin was 0, 1, 2.1 and 19%, respectively. Seven high-level azithromycin-resistant (HLAzi-R) isolates were identified, all susceptible to ceftriaxone. Mosaic penA alleles XXXIV, X and non-mosaic XIII, and G120K plus A121N/D/G (PorB1b), H105Y (MtrR) and A deletion (mtrR promoter) mutations, were associated with elevated ESC MICs. A2059G and C2611T mutations in 23S rRNA were associated with HLAzi-R and azithromycin MICs of 4-32 mg/L, respectively. The 43 whole-genome sequenced isolates belonged to 31 NG-MAST STs. All HLAzi-R isolates belonged to MLST ST1580 and some clonal clustering was observed; however, the isolates differed significantly from the published HLAzi-R isolates from the ongoing UK outbreak. There is good correlation between previously described genetic antimicrobial resistance determinants and phenotypic susceptibility categories for ESCs and azithromycin in N. gonorrhoeae. This work highlights the advantages and potential of whole-genome sequencing to be applied at scale in the surveillance of antibiotic resistant strains of N. gonorrhoeae, both locally and internationally.

  • 41.
    Serwin, Agnieszka Beata
    et al.
    Department of Dermatology and Venereology, Medical University of Bialystok, Bialystok, Poland.
    Bulhak-Koziol, Violetta
    Diagnostic and Research Centre for Sexually Transmitted Diseases, Bialystok, Poland.
    Sokolowska, Marianna
    Diagnostic and Research Centre for Sexually Transmitted Diseases, Bialystok, Poland.
    Golparian, Daniel
    Örebro University, School of Medical Sciences. WHO Collaborating Centre for Gonorrhoea and Other STIs, National Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Örebro University Hospital, Örebro, Sweden.
    Unemo, Magnus
    WHO Collaborating Centre for Gonorrhoea and Other STIs, National Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Örebro University Hospital, Örebro, Sweden.
    Trichomonas vaginalis is very rare among women with vaginal discharge in Podlaskie province, Poland2017In: Acta Pathologica, Microbiologica et Immunologica Scandinavica (APMIS), ISSN 0903-4641, E-ISSN 1600-0463, Vol. 125, no 9, p. 840-843Article in journal (Refereed)
    Abstract [en]

    Trichomonas vaginalis is the most common curable sexually transmitted pathogen globally. However, in the European Union (EU), trichomoniasis appears to be a rare condition. The aim of this study was to examine the prevalence of T. vaginalis among females attending an STI centre in Bialystok, Poland, using the highly sensitive and specific APTIMA T. vaginalis assay. Consecutive females, referred by gynaecologists mainly because of abnormal vaginal discharge, were diagnosed using wet mount microscopy, culture and APTIMA T. vaginalis assay. Among 272 women studied, 82% were pre- and 18% postmenopausal. The average age was 36.0 +/- 13.9 (range: 18-86) years. Abnormal discharge (alone or accompanied by itch or vulvovaginal burning) was the most frequent complain in both groups (66.2% and 48.0%). Erythema and discharge were the most frequent abnormal signs (58.6% and 56.0%). Not a single T. vaginalis-positive sample was detected using wet mount microscopy, culture or APTIMA T. vaginalis assay. Despite using the highly sensitive APTIMA T. vaginalis assay for detection, the pathogen could not be identified in females in the studied setting, similar to results from other EU settings. The need for general screening using NAAT for this pathogen while diagnosing vulvovaginal symptoms in females in Poland appears to be low.

  • 42.
    Sethi, Sunil
    et al.
    Dept Med Microbiol, Post Grad Institute of Medicine, Education & Research, Chandigarh, India.
    Golparian, Daniel
    Dept Lab Med, Natl Reference Lab Pathogen Neisseria, WHO Collaborating Ctr Gonorrhoea & Other STIs, Örebro University Hospital, Örebro, Sweden.
    Bala, Manju
    Training & Res Ctr, Apex Reg STD Teaching, WHO GASP SEAR Reg Reference Lab, VMMC & Safdarjang Hosp, New Delhi, India.
    Dorji, Dorji
    JDW NR Hosp, Thimphu, Bhutan.
    Ibrahim, Muhammad
    Dept Lab Med, Natl Reference Lab Pathogen Neisseria, WHO Collaborating Ctr Gonorrhoea & Other STIs, Örebro University Hospital, Örebro, Sweden.
    Jabeen, Kausar
    Aga Khan Univ, Karachi, Pakistan.
    Unemo, Magnus
    Örebro University Hospital. Dept Lab Med, Natl Reference Lab Pathogen Neisseria, WHO Collaborating Ctr Gonorrhoea & Other STIs, Örebro University Hospital, Örebro, Sweden.
    Antimicrobial susceptibility and genetic characteristics of Neisseria gonorrhoeae isolates from India, Pakistan and Bhutan in 2007-20112013In: BMC Infectious Diseases, ISSN 1471-2334, E-ISSN 1471-2334, Vol. 13, article id 35Article in journal (Refereed)
    Abstract [en]

    Background: Knowledge on antimicrobial drug resistance and genetic characteristics of Neisseria gonorrhoeae isolates circulating in India, Pakistan, and Bhutan is sorely lacking. In this paper, we describe the prevalence of antimicrobial resistance and molecular characteristics of N. gonorrhoeae isolates from India, Pakistan, and Bhutan in 2007-2011. Methods: Antimicrobial susceptibility and beta-lactamase production were tested for 65 N. gonorrhoeae isolates from India (n=40), Pakistan (n=18) and Bhutan (n=7) using Etest methodology (eight antimicrobials) and nitrocefin solution, respectively. Resistance determinants, i.e. penA, mtrR, porB1b, gyrA, and parC, were sequenced. N. gonorrhoeae multiantigen sequence typing (NG-MAST) was performed for molecular epidemiology. Results: The highest resistance level was observed for ciprofloxacin (94%), followed by penicillin G (68%), erythromycin (62%), tetracycline (55%), and azithromycin (7.7%). All the isolates were susceptible to ceftriaxone, cefixime, and spectinomycin. Thirty-four (52%) of the isolates were producing beta-lactamase. No penA mosaic alleles or A501-altered alleles of penicillin-binding protein 2 were identified. Forty-nine NG-MAST STs were identified, of which 42 STs have not been previously described worldwide. Conclusions: Based on this study, ceftriaxone, cefixime, and spectinomycin can be used as an empirical first-line therapy for gonorrhoea in India, Pakistan, and Bhutan, whereas ciprofloxacin, penicillin G, tetracycline, erythromycin, and azithromycin should not be. It is imperative to strengthen the laboratory infrastructure in this region, as well as to expand the phenotypic and genetic surveillance of antimicrobial resistance, emergence of new resistance, particularly, to extended-spectrum cephalosporins, and molecular epidemiology.

  • 43.
    Seth-Smith, Helena M. B.
    et al.
    Wellcome Trust Sanger Inst, Hinxton, England.
    Harris, Simon R.
    Wellcome Trust Sanger Inst, Hinxton, England.
    Skilton, Rachel J.
    Fac Med, Mol Microbiol Grp, Southampton Gen Hosp, Univ Southampton, Southampton, England.
    Radebe, Frans M.
    Natl Inst Communicable Dis, Ctr HIV & Sexually Transmitted Infect, National Health Laboratory Service, Johannesburg, South Africa.
    Golparian, Daniel
    Natl Reference Lab Pathogen Neisseria, WHO Collaborating Ctr Gonorrhoea & Other STIs, Örebro University Hospital, Örebro, Sweden.
    Shipitsyna, Elena
    Lab Microbiol, DO Ott Res Inst Obstet & Gynaecol, St Petersburg, Russia.
    Duy, Pham Thanh
    Scott, Paul
    Wellcome Trust Sanger Inst, Hinxton, England.
    Cutcliffe, Lesley T.
    Fac Med, Mol Microbiol Grp, Southampton Gen Hosp, Univ Southampton, Southampton, England.
    O'Neill, Colette
    Fac Med, Mol Microbiol Grp, Southampton Gen Hosp, Univ Southampton, Southampton, England.
    Parmar, Surendra
    Hlth Protect Agcy, Clin Microbiol & Publ Hlth Lab, Addenbrookes Hosp, Cambridge, England.
    Pitt, Rachel
    Sexually Transmitted Bacteria Reference Lab, Health Protection Agency, London, England.
    Baker, Stephen
    Clin Res Unit, Wellcome Trust Major Overseas List, Hosp Trop Dis, Univ Oxford, Ho Chi Minh City, Vietnam.
    Ison, Catherine A.
    Sexually Transmitted Bacteria Reference Lab, Health Protection AgencyLondon NW9 5HT, England..
    Marsh, Peter
    Publ Hlth Lab Southampton, Health Protection Agency, Southampton Gen Hosp, Southampton, England.
    Jalal, Hamid
    Health Protection Agency, Clin Microbiol & Publ Hlth Lab, Addenbrookes Hosp, Cambridge, England.
    Lewis, David A.
    Natl Inst Communicable Dis, Ctr HIV & Sexually Transmitted Infect, National Health Laboratory ServiceJohannesburg, South Africa; Fac Hlth Sci, Dept Internal Med, Univ Witwatersrand, Johannesburg, South Africa.
    Unemo, Magnus
    Örebro University Hospital. Natl Reference Lab Pathogen Neisseria, WHO Collaborating Ctr Gonorrhoea & Other STIs, Örebro University Hospital, Örebro, Sweden.
    Clarke, Ian N.
    Fac Med, Mol Microbiol Grp, Southampton Gen Hosp, Univ Southampton, Southampton, England.
    Parkhill, Julian
    Wellcome Trust Sanger Inst, Hinxton, England.
    Thomson, Nicholas R.
    Wellcome Trust Sanger Inst, Hinxton, England.
    Whole-genome sequences of Chlamydia trachomatis directly from clinical samples without culture2013In: Genome Research, ISSN 1088-9051, E-ISSN 1549-5469, Vol. 23, no 5, p. 855-866Article in journal (Refereed)
    Abstract [en]

    The use of whole-genome sequencing as a tool for the study of infectious bacteria is of growing clinical interest. Chlamydia trachomatis is responsible for sexually transmitted infections and the blinding disease trachoma, which affect hundreds of millions of people worldwide. Recombination is widespread within the genome of C. trachomatis, thus whole-genome sequencing is necessary to understand the evolution, diversity, and epidemiology of this pathogen. Culture of C trachomatis has, until now, been a prerequisite to obtain DNA for whole-genome sequencing; however, as C trachomatis is an obligate intracellular pathogen, this procedure is technically demanding and time consuming. Discarded clinical samples represent a large resource for sequencing the genomes of pathogens, yet clinical swabs frequently contain very low levels of C trachomatis DNA and large amounts of contaminating microbial and human DNA. To determine whether it is possible to obtain whole-genome sequences from bacteria without the need for culture, we have devised an approach that combines immunomagnetic separation (IMS) for targeted bacterial enrichment with multiple displacement amplification (MDA) for whole-genome amplification. Using IMS-MDA ill conjunction with high-throughput multiplexed Illumina sequencing, we have produced the first whole bacterial genome sequences direct from clinical samples. We also show that this method can be used to generate genome data from nonviable archived samples. This method will prove a useful tool in answering questions relating to the biology of many difficult-to-culture or fastidious bacteria of clinical concern.

  • 44.
    Shipitsyna, Elena
    et al.
    Laboratory of Microbiology, Gynaecology and Reproductology, D.O. Ott Research Institute of Obstetrics, St. Petersburg, Russian Federation; WHO Collaborating Centre for Gonorrhoea and Other STIs, Department of Laboratory Medicine, Örebro University Hospital, Örebro, Sweden.
    Rumyantseva, Tatiana
    Laboratory of Molecular Diagnostics, Central Research Institute of Epidemiology, Moscow, Russian Federation.
    Golparian, Daniel
    Örebro University, School of Medical Sciences. WHO Collaborating Centre for Gonorrhoea and Other STIs, Department of Laboratory Medicine, Örebro University Hospital, Örebro, Sweden.
    Khayrullina, Guzel
    Laboratory of Molecular Diagnostics, Central Research Institute of Epidemiology, Moscow, Russian Federation.
    Lagos, Amaya C.
    WHO Collaborating Centre for Gonorrhoea and Other STIs, Department of Laboratory Medicine, Örebro University Hospital, Örebro, Sweden.
    Edelstein, Inna
    Institute of Antimicrobial Chemotherapy, Smolensk State Medical University, Smolensk, Russian Federation.
    Joers, Kai
    United Laboratories, Tartu University Hospital, Tartu, Estonia.
    Jensen, Jorgen S.
    Department of Microbiology and Infection Control, Sexually Transmitted Infections, Research and Development, Statens Serum Institut, Copenhagen, Denmark.
    Savicheva, Alevtina
    Laboratory of Microbiology, D.O. Ott Research Institute of Obstetrics, Gynaecology and Reproductology, St. Petersburg, Russian Federation.
    Rudneva, Natalia
    Tula Regional Clinic of Dermato-Venereology, Ministry of Health of the Tula Region, Tula, Russian Federation.
    Sukhanova, Larisa
    Tula Regional Clinic of Dermato-Venereology, Ministry of Health of the Tula Region, Tula, Russian Federation.
    Kozlov, Roman
    Institute of Antimicrobial Chemotherapy, Smolensk State Medical University, Smolensk, Russian Federation.
    Guschin, Alexander
    Laboratory of Molecular Diagnostics, Central Research Institute of Epidemiology, Moscow, Russian Federation.
    Unemo, Magnus
    Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and Other STIs, Department of Laboratory Medicine, Örebro University Hospital, Örebro, Sweden.
    Prevalence of macrolide and fluoroquinolone resistance-mediating mutations in Mycoplasma genitalium in five cities in Russia and Estonia2017In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 12, no 4, article id e0175763Article in journal (Refereed)
    Abstract [en]

    Background and objective: Resistance in the sexually transmitted bacterium Mycoplasma genitalium to all recommended therapeutic antimicrobials have rapidly emerged. However, to date, internationally reported resistance surveillance data for M. genitalium strains circulating in Eastern Europe are entirely lacking. The aim of this study was to estimate the prevalence of macrolide and fluoroquinolone resistance-associated mutations in M. genitalium in four cities in Russia and one in Estonia, 2013-2016.

    Materials and methods: Consecutive urogenital samples found positive for M. genitalium during diagnostic testing were retrospectively analyzed for resistance-associated mutations in the 23S rRNA and parC genes using pyrosequencing and conventional Sanger sequencing, respectively.

    Results: In total, 867 M. genitalium positive samples from 2013-2016 were analyzed. Macrolide resistance-associated mutations were detected in 4.6% of the samples from Russia (0.7- 6.8% in different cities) and in 10% of the samples from Estonia. The mutations A2059G and A2058G were highly predominating in both Russia and Estonia, accounting together for 90.9% of the cases positive for nucleotide substitutions in the 23S rRNA gene. The rates of possible fluoroquinolone resistance-associated mutations were 6.2% in Russia (2.5-7.6% in different cities) and 5% in Estonia. The mutations S83I and S83N were the most frequent ones in Russia (24.4% each), whereas D87N highly predominated in Estonia (83.3% of all fluoroquinolone resistance-associated mutations). Approximately 1% of the samples in both countries harbored both macrolide and possible fluoroquinolone resistance-associated mutations, with A2058G and S83I being the most frequent combination (37.5%).

    Conclusions: The prevalence of macrolide and fluoroquinolone resistance-associated mutations in M. genitalium was 4.6% and 6.2%, respectively, in Russia, and 10% and 5%, respectively, in Estonia. Despite the relatively low rates of macrolide and fluoroquinolone resistance in these countries, antimicrobial resistance surveillance and testing for resistance-associated mutations in M. genitalium positive cases would be valuable.

  • 45.
    Sánchez-Busó, Leonor
    et al.
    Pathogen Genomics, Wellcome Sanger Institute, Wellcome Genome Campus, Hinxton, UK.
    Golparian, Daniel
    Örebro University, School of Medical Sciences. WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, National Reference Laboratory for Sexually Transmitted Infections, Department of Laboratory Medicine, Clinical Microbiology.
    Corander, Jukka
    Pathogen Genomics, Wellcome Sanger Institute, Wellcome Genome Campus, Hinxton, UK: Helsinki Institute for Information Technology, Department of Mathematics and Statistics, University of Helsinki, Helsinki, Finland; Department of Biostatistics, University of Oslo, Oslo, Norway.
    Grad, Yonatan H.
    Department of Immunology and Infectious Diseases, Harvard TH Chan School of Public Health, Boston, MA, USA; Division of Infectious Diseases, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA.
    Ohnishi, Makoto
    Department of Bacteriology I, National Institute of Infectious Diseases, Tokyo, Japan; Antimicrobial Resistance Research Center, National Institute of Infectious Diseases, Tokyo, Japan.
    Flemming, Rebecca
    Faculty of Classics, University of Cambridge, Cambridge, UK.
    Parkhill, Julian
    Pathogen Genomics, Wellcome Sanger Institute, Wellcome Genome Campus, Hinxton, UK.
    Bentley, Stephen D.
    Pathogen Genomics, Wellcome Sanger Institute, Wellcome Genome Campus, Hinxton, UK.
    Unemo, Magnus
    Örebro University, School of Medical Sciences. Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, National Reference Laboratory for Sexually Transmitted Infections, Department of Laboratory Medicine, Clinical Microbiology.
    Harris, Simon R.
    Pathogen Genomics, Wellcome Sanger Institute, Wellcome Genome Campus, Hinxton, UK.
    The impact of antimicrobials on gonococcal evolution2019In: Nature Microbiology, E-ISSN 2058-5276Article in journal (Refereed)
    Abstract [en]

    The sexually transmitted pathogen Neisseria gonorrhoeae is regarded as being on the way to becoming an untreatable superbug. Despite its clinical importance, little is known about its emergence and evolution, and how this corresponds with the introduction of antimicrobials. We present a genome-based phylogeographical analysis of 419 gonococcal isolates from across the globe. Results indicate that modern gonococci originated in Europe or Africa, possibly as late as the sixteenth century and subsequently disseminated globally. We provide evidence that the modern gonococcal population has been shaped by antimicrobial treatment of sexually transmitted infections as well as other infections, leading to the emergence of two major lineages with different evolutionary strategies. The well-described multidrug-resistant lineage is associated with high rates of homologous recombination and infection in high-risk sexual networks. A second, multisusceptible lineage is more associated with heterosexual networks, with potential implications for infection control.

  • 46.
    Tabrizi, Sepehr N.
    et al.
    Dept Microbiol & Infect Dis, Royal Womens Hosp, Parkville Vic, Australia; Dept Obstet & Gynaecol, Royal Womens Hosp, Univ Melbourne, Parkville Vic, Australia; Murdoch Childrens Res Inst, Parkville Vic, Australia.
    Unemo, Magnus
    Örebro University Hospital. Dept Lab Med, WHO Collaborating Ctr Gonorrhoea & Other STIs, Örebro University Hospital, Örebro, Sweden.
    Golparian, Daniel
    Dept Lab Med, WHO Collaborating Ctr Gonorrhoea & Other STIs, Örebro University Hospital, Örebro, Sweden.
    Twin, Jimmy
    Dept Obstet & Gynaecol, Royal Womens Hosp, Univ Melbourne, Parkville Vic, Australia; Murdoch Childrens Res Inst, Parkville Vic, Australia.
    Limnios, Athena E.
    Dept Microbiol, WHO Collaborating Ctr STD, Prince Wales Hosp, Randwick NSW, Australia.
    Lahra, Monica
    Dept Microbiol, WHO Collaborating Ctr STD, Prince Wales Hosp, Randwick NSW, Australia.
    Guy, Rebecca
    Dept Microbiol, Univ NSW, Sydney NSW, Australia; Kirby Inst, Univ NSW, Sydney NSW, Australia.
    Analytical Evaluation of GeneXpert CT/NG, the First Genetic Point-of-Care Assay for Simultaneous Detection of Neisseria gonorrhoeae and Chlamydia trachomatis2013In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 51, no 6, p. 1945-1947Article in journal (Refereed)
    Abstract [en]

    GeneXpert CT/NG was evaluated with 372 characterized bacterial strains. Sensitivity of 10 genome copies/reaction was obtained for both agents. Four Neisseria mucosa and two Neisseria subflava isolates were positive for one of two gonococcal targets; however, the assay flagged all as negative. The assay was analytically highly sensitive and specific.

  • 47.
    Unemo, Magnus
    et al.
    Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and other STIs, Department of Laboratory Medicine, Örebro University Hospital, Örebro, Sweden.
    Golparian, Daniel
    WHO Collaborating Centre for Gonorrhoea and other STIs, Department of Laboratory Medicine, Örebro University Hospital, Örebro, Sweden.
    Hellmark, Bengt
    Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and other STIs, Department of Laboratory Medicine, Örebro University Hospital, Örebro, Sweden.
    First Three Neisseria gonorrhoeae Isolates with High-Level Resistance to Azithromycin in Sweden: a Threat to Currently Available Dual-Antimicrobial Regimens for Treatment of Gonorrhea?2014In: Antimicrobial Agents and Chemotherapy, ISSN 0066-4804, E-ISSN 1098-6596, Vol. 58, no 1, p. 624-625Article in journal (Refereed)
  • 48.
    Unemo, Magnus
    et al.
    Örebro University Hospital. National Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Golparian, Daniel
    National Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Shafer, William M.
    Department of Microbiology and Immunology, Emory University School of Medicine, Atlanta GA, United States; Laboratories of Bacterial Pathogenesis, Veterans Affairs Medical Center, Decatur GA, United States.
    Challenges with gonorrhea in the era of multi-drug and extensively drug resistance -are we on the right track?2014In: Expert Review of Anti-Infective Therapy, ISSN 1478-7210, E-ISSN 1744-8336, Vol. 12, no 6, p. 653-656Article in journal (Refereed)
    Abstract [en]

    Neisseriagonorrhoeae has retained antimicrobial resistance to drugs previously recommended for first-line empiric treatment of gonorrhea, and resistance to ceftriaxone, the last option for monotherapy, is evolving. Crucial actions to combat this developing situation include implementing response plans; considering use of dual antimicrobial regimens; enhancing surveillance of gonorrhea, gonococcal antimicrobial resistance, treatment failures and antimicrobial use/misuse and improving prevention, early diagnosis, contact tracing and treatment. The ways forward also include an intensified research to identify novel antimicrobial resistance determinants and develop and evaluate appropriate use of molecular antimicrobial resistance testing, ideally point-of-care and with simultaneous detection of gonococci, to supplement culture-based methods and ideally guide tailored treatment. It is crucial with an enhanced understanding of the dynamics of the national and international emergence, transmission and evolution of antimicrobial-resistant gonococcal strains. Genome sequencing combined with epidemiological metadata will detail these issues and might also revolutionize the molecular antimicrobial resistance testing. Ultimately, novel antimicrobials are essential and some antimicrobials in development have shown potent in vitro activity against gonococci. Several of these antimicrobials deserve further attention for potential future treatment of gonorrhea.

  • 49.
    Unemo, Magnus
    et al.
    Örebro University Hospital. WHO Collaborating Ctr Gonorrhoea & Other STIs, Dept Lab Med, Swedish Reference Lab Pathogen Neisseria, Örebro University Hospital, Örebro, Sweden.
    Golparian, Daniel
    WHO Collaborating Ctr Gonorrhoea & Other STIs, Dept Lab Med, Swedish Reference Lab Pathogen Neisseria, Örebro University Hospital, Örebro, Sweden.
    Skogen, Vegard
    Dept Infect Dis, Univ Hosp N Norway, Tromsö, Norway; Inst Clin Med, Univ Tromsö, Tromsö, Norway.
    Olsen, Anne Olaug
    Olafiaklin, Oslo Univ Hosp, Oslo, Norway; Inst Clin Med, Univ Oslo, Oslo, Norway.
    Moi, Harald
    Olafiaklin, Oslo Univ Hosp, Oslo, Norway; Inst Clin Med, Univ Oslo, Oslo, Norway.
    Syversen, Gaute
    Dept Microbiol, Oslo Univ Hosp, Oslo, Norway.
    Hjelmevoll, Stig Ove
    Dept Microbiol & Infect Control, Univ Hosp N Norway, Tromsö, Norway.
    Neisseria gonorrhoeae Strain with High-Level Resistance to Spectinomycin Due to a Novel Resistance Mechanism (Mutated Ribosomal Protein S5) Verified in Norway2013In: Antimicrobial Agents and Chemotherapy, ISSN 0066-4804, E-ISSN 1098-6596, Vol. 57, no 2, p. 1057-1061Article in journal (Refereed)
    Abstract [en]

    Gonorrhea may become untreatable, and new treatment options are essential. Verified resistance to spectinomycin is exceedingly rare. However, we describe a high-level spectinomycin-resistant (MIC, >1,024 mu g/ml) Neisseria gonorrhoeae strain from Norway with a novel resistance mechanism. The resistance determinant was a deletion of codon 27 (valine) and a K28E alteration in the ribosomal protein 5S. The traditional spectinomycin resistance gene (16S rRNA) was wild type. Despite this exceedingly rare finding, spectinomycin available for treatment of ceftriaxone-resistant urogenital gonorrhea would be very valuable.

  • 50.
    Unemo, Magnus
    et al.
    Örebro University, School of Health Sciences. WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, National Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Clinical Microbiology, Örebro University Hospital, Örebro, Sweden.
    Golparian, Daniel
    Örebro University, School of Medical Sciences. WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, National Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Clinical Microbiology, Örebro University Hospital, Örebro, Sweden.
    Sánchez-Busó, Leonor
    Pathogen Genomics, Wellcome Trust Genome Campus, The Wellcome Trust Sanger Institute, Hinxton, UK.
    Grad, Yonatan
    Department of Immunology and Infectious Diseases, Harvard T.H. Chan School of Public Health, Boston, USA; Division of Infectious Diseases Brigham andWomen’s Hospital, Harvard Medical School, Boston, USA.
    Jacobsson, Susanne
    Örebro University, School of Medical Sciences. WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, National Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Clinical Microbiology, Örebro University Hospital, Örebro, Sweden.
    Ohnishi, Makoto
    Department of Bacteriology I, National Institute of Infectious Diseases, Tokyo, Japan.
    Lahra, Monica M.
    WHO Collaborating Centre for Sexually Transmitted Diseases, Department of Microbiology, South Eastern Area Laboratory Services, The Prince of Wales Hospital, Randwick, Sydney, Australia.
    Limnios, Athena
    WHO Collaborating Centre for Sexually Transmitted Diseases, Department of Microbiology, South Eastern Area Laboratory Services, The Prince of Wales Hospital, Randwick, Sydney, Australia.
    Sikora, Aleksandra E.
    Department of Pharmaceutical Sciences, College of Pharmacy, Oregon State University, Corvallis, USA.
    Wi, Teodora
    Department of Reproductive Health and Research, World Health Organization, Geneva, Switzerland.
    Harris, Simon R.
    Pathogen Genomics, The Wellcome Trust Sanger Institute, Wellcome Trust Genome Campus, Hinxton, UK.
    The novel 2016 WHO Neisseria gonorrhoeae reference strains for global quality assurance of laboratory investigations: phenotypic, genetic and reference genome characterization2016In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 71, no 11, p. 3096-3108Article in journal (Refereed)
    Abstract [en]

    Objectives: Gonorrhoea and MDR Neisseria gonorrhoeae remain public health concerns globally. Enhanced, quality-assured, gonococcal antimicrobial resistance (AMR) surveillance is essential worldwide. The WHO global Gonococcal Antimicrobial Surveillance Programme (GASP) was relaunched in 2009. We describe the phenotypic, genetic and reference genome characteristics of the 2016 WHO gonococcal reference strains intended for quality assurance in the WHO global GASP, other GASPs, diagnostics and research worldwide.

    Methods: The 2016 WHO reference strains (n = 14) constitute the eight 2008 WHO reference strains and six novel strains. The novel strains represent low-level to high-level cephalosporin resistance, high-level azithromycin resistance and a porA mutant. All strains were comprehensively characterized for antibiogram (n = 23), serovar, prolyliminopeptidase, plasmid types, molecular AMR determinants, N. gonorrhoeae multiantigen sequence typing STs and MLST STs. Complete reference genomes were produced using single-molecule PacBio sequencing.

    Results: The reference strains represented all available phenotypes, susceptible and resistant, to antimicrobials previously and currently used or considered for future use in gonorrhoea treatment. All corresponding resistance genotypes and molecular epidemiological types were described. Fully characterized, annotated and finished references genomes (n = 14) were presented.

    Conclusions: The 2016 WHO gonococcal reference strains are intended for internal and external quality assurance and quality control in laboratory investigations, particularly in the WHO global GASP and other GASPs, but also in phenotypic (e.g. culture, species determination) and molecular diagnostics, molecular AMR detection, molecular epidemiology and as fully characterized, annotated and finished reference genomes in WGS analysis, transcriptomics, proteomics and other molecular technologies and data analysis.

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