Global challenge of antibiotic-resistant Mycoplasma genitalium Sabine Pereyre USC EA 3671 Mycoplasmal and chlamydial infections in humans INRA - University of Bordeaux -Bordeaux University Hospital National French Reference Center for bacterial IST ECCMID, April 24th 2018 ESCMID eLibrary © by author Disclosures Travel grants from – Diagenode, Belgium – SpeeDx, Australia – Hologic, USA My laboratory has received remuneration for contract work from – bioMérieux, France – Meridian Bioscience, Europe – Diagenode, Belgium – SpeeDx, Australia – Hologic, USA ESCMID eLibrary 2 © by author M. genitalium STI agent responsible for – Non gonococcal urethritis, proctitis – Cervicitis, PID, preterm birth, spontaneous abortion – Carriage frequently asymptomatic Prevalence – Community-based population 1–3% – STI center population (high risk sexual behaviour) 4 – 38% Diagnostic – only NAAT tests – No serology – ≈ no culture antimicrobial susceptibility testing hardly achievable ESCMID eLibrary 3 © by author M. genitalium and antibiotics Intrinsic resistance to ATB targeting the cell wall (-lactams, fosfomycin, glycopeptides) and rifampicin (mutation in rpoB gene) In vitro active antibiotics – Macrolides and related antibiotics (MLSK) – Fluoroquinolones – Tetracyclines ESCMID eLibrary 4 © by author M. genitalium and tetracyclines Relative potency in vitro MIC ranges (µg/ml) Antibiotics M. genitalium M. hominis Ureaplasma spp. Tetracyclines** Doxycycline ≤0.060.01 -–0.31 0.1-2 0.02-1 Minocycline ≤0.01-0.2 0.03-1 0.06-1 MLS group BUT,Eryth rolowmyci neradication rate≤0.01 32->1 000 0.02-16 –RoxiMicrobiologicalthromycin cure of<0 doxycycline.01 : between>16 30 and 40 %0.1-2 – Not a first-line treatment Clarithromycin ≤0.01-0.06 16->256 ≤0.004-2 Azithromycin ≤0.01-0.03 4->64 0.06-4 Pereyre, AntimicrobJosa. mChemotherycin . Vaccines, 3rd edition0.01, 2016;-0.02 Jensen Antimicrob0.05Agnets-2 Chemother 2014;0.03 -Jensen4 J Eur Acad Dermatol Venereol. 2016 ESCMID eLibrary 5 Clindamycin 0.2-1 ≤0.008-2 0.2-64 Pristinamycin ≤0.01-0.02 0.1-0.5 0.1-1 Quinup risti n/ D alfo prist in ©0.05 by 0.03author-2 0.05-0.5 Ketolides Telithromycin ≤0.015 2-32 ≤0.015-0.25 Solithromycin ≤0.000000063- 0.002-0.008 0.002-0.063 0.000125 Fluoroquinolones** Ciprofloxacin 2 0.1-4 0.1-16 Ofloxacin 1-2 0.1-4 0.2-4 Levofloxacin 0.5-1 0.1-2 0.2-2 Moxifloxacin 0.03-0.06 0.06-0.125 0.125-1 Other agents Chloramphenicol 0.5-25 4225 0.4-8 Gentamicin ND 2-16 0.1-13 M. genitalium and MLSK Low MICs Antibiotics M. genitalium M. hominis Ureaplasma spp. MLSK group Erythromycin ≤0.01 32->1 000 0.02-16 Roxithromycin <0.01 >16 0.1-2 Clarithromycin ≤0.01-0.06 16->256 ≤0.004-2 Azithromycin ≤0.01-0.03 4->64 0.06-4 Josamycin 0.01-0.02 0.05-2 0.03-4 Clindamycin 0.2-1 ≤0.008-2 0.2-64 Pristinamycin ≤0.01-0.02 0.1-0.5 0.1-1 Quinupristin/ Dalfopristin 0.05 0.03-2 0.05-0.5 Telithromycin ≤0.015 2-32 ≤0.015-0.25 Solithromycin ≤0.000000063- 0.002-0.008 0.002-0.063 0.000125 ESCMID eLibraryPereyre, Antimicrob. Chemother. Vaccines, 3rd edition , 2016 6 © by author Acquired resistance to macrolides • Point mutations in domain V of 23S rRNA - Single operon encoding 16S and 23S rRNA - A2058G/C/T, A2059G/C/T, A2062G/T (E. coli numbering) high level resistance 2062 23S rRNA Domain V ESCMID eLibraryPeptidyl transferase loop © by author All M. genitalium-positive test should be followed up with an assay capable of detecting macrolide resistance- associated mutations ESCMID eLibrary 8 © by author Detection of macrolide resistance-associated mutations Amplification and sequencing of 23S rRNA – Time-consuming, not adapted to routine Published in-house methods – FRET real-time PCR (Touati et al. J. Clin. Microbiol. 2014) – HRM (High Resolution Melting curve analysis) (Twin et al. PloS One 2012) – PCR and pyrosequencing (Salado-Rasmussen et al. Clin. Infect. Dis. 2014) – Taqman PCR using forward primers complementary to 23S mutations (Wold et al. J. Eur. Acas. Dermatol. Venereol. 2015), only if Ct<32. – Taqman PCR using 3 probes and subsequent endpoint fluorescence analysis (Kristiansen et al. J. Clin. Microbiol. 2016) – Single probe PCR and melting curve analysis (Gossé et al. J. Clin. Microbiol. 2016) Commercial kits – ResistancePlusTM MG kit (SpeeDx, Australia) : multiplex real-time PCR (Le Roy, J. Clin. Microbiol. 2017) – S-DiaMGRes kit (Diagenode, Belgium) : multiplex real-time PCR Detection of Mg and 5 mutations – Others expected ESCMID eLibrary 9 © by author Prevalence of macrolide resistance in M. genitalium10 100%(26/26) 56-61% 18% 4.6-6,7% 58% 41% 57% 53% 48% 35% 29% No data 83% in MSM <10% 43-63% 10%-20% 9,8% 20%-40% 40-60% >60% 72-77% Anagrius, PloS one 2013; Tagg, J. Clin. Microbiol. 2013; Pond, Clin. Inf. Dis. 2014; Salado-Rasmussen, Clin. Inf. Dis, 2014; Kikuchi, J. Antimicrob. Chemother. 2014; Hay, Sex. Transm. Dis. 2015; Gushin, BMC Infect. Dis. 2015; Nijhuis, J. Antimicrob. Chemother. 2015; Gesink, Can. Fam. Physician, 2016; Getman, J. Clin. Microbiol. 2016; Gossé, J. Clin. Microbiol. 2016; Shipitsina, Plos One, 2017; Basu, J. Clin. Microbiol. 2017; Tabrizi, J. Clin. Microbiol. 2017; Barbera, Sex. Transm. Dis. 2017; Dumke, Diagn Microbiol infect Dis, 2016; Coorevits, J. Glob. Antimicrob. Resist. 2017; Anderson, J. Clin.ESCMIDMicrobiol. 2017; Unemo, Clin. Microbiol. Infect.2017. eLibrary © by author Treatment studies : azithromycin (AZM) 1g • Meta-analysis on the efficacy of AZM 1g for Mg treatment (Lau et al Clin. Infect. Dis. 2015) 100% 90% 83.5% 80% 67% 70% 60% The efficacy of azithromycin 1 g has 50% 40% decreased 30% 20% 10% 0% Pooled microbial cure rate (%) rate cure microbial Pooled Before 2009 Since 2009 • Selection of resistant mutants during AZM 1g treatment • AZM 1g single dose is no more the 1st line treatment Therapeutic failure if patient infected with a mutated strain Björnelius Sex Transm Infect 2008; Manhart Clin Infect Dis 2013; Mena Clin Infect Dis 2009; Schwebke CID 2011, Sena J Infect Dis 2012; Stamm Sex Transm InfectESCMID 2007; Lau Clin Infect Dis 2015; Horner Clin Infect Dis 2015;eLibrary Jensen BMC Infect Dis 2015 © by author IMPORTANT MESSAGE : Rapid increasing prevalence of macrolide resistance most likely due to widespread use of azithromycin 1g single dose Uncomplicated M. genitalium infection: - In the absence of macrolide resistance-associated mutations Azithromycin 500 mg (day 1), then 250 mg (days 2-5) 85% effective Associated with lower risk of inducing AZM R (conflicting evidence) Patients failing azithromycin 1g single dose cannot be treated successfully with extended 1.5 g AZM 12 Anagrius PLoS One 2013, Bjornelius Sex Transm Infect 2008, Falk J Antimicrob Chemother 2015, Gundevia STI 2015, Read, Clin Inf Dis 2017; Horner Sex ESCMIDTrans Ingfect 2018; Jernberg Sex Transm Infect 2008, JenseneLibrary Clin Infect 2009 © by author Uncomplicated M. genitalium infection: - Macrolide-resistant M. genitalium infection - Second line treatment for persistent M. genitalium infection Moxifloxacin 400 mg/d - 7 to 10 days Complicated M. genitalium infection (PID, epididymitis) Moxifloxacin: 400 mg - 14 days ESCMID eLibrary 13 © by author M. genitalium and fluoroquinolones Only moxifloxacin and sitafloxacin have low MICs Antibiotics M. genitalium M. hominis Ureaplasma spp. Fluoroquinolones Ciprofloxacin 2 0.1-4 0.1-16 Ofloxacin 1-2 0.1-4 0.2-4 Levofloxacin 0.5-1 0.1-2 0.2-2 Moxifloxacin 0.03-0.06 0.06-0.125 0.125-1 Sitafloxacin (Japan) 0.125 Deguchi, J Infect Chemother 2014; Pereyre, Antimicrob. Chemother. Vaccines, 3rd edition, 2016 ESCMID eLibrary 14 © by author Fluoroquinolone resistance in M. genitalium • Mutations in the bacterial target genes of fluoroquinolones - Most frequent mutations in parC (Topoisomerase IV) Primarily Ser83 and Asp87 - Rare mutations in gyrA (DNA gyrase) • Many mutations have not been evaluated by in vitro MIC determination • Molecular detection only: amplification and sequencing of target genes ESCMID eLibrary © by author Prevalence of fluoroquinolone resistance-associated mutations16 10.2% 6.2% 4.1% 20% 4.5% 10% 6% 13.6% 8% 31- 47% No data <10% 12-19% 10%-20% 20%-40% 40-60% >60% 23% Bissessor Clin Infect Dis 2015; Deguchi, Clin Infect Dis 2016; Dumke, DMID 2016; Kikuchi J Antimicrob Chemother 2014; Le Roy Emerg Infect Dis 2016; Pond Clin Infect Dis 2014; Shipitsina PLoS one 2017; Couldwell Int J STD and AIDS 2013; Gesink Can family Physian 2016; Tagg J Clin Microbiol 2013; Murray Emerg Infec Dis 2017; Barbera ESCMIDSex Transm infect 2017; Anderson, J Clin Microbiol 2017, UnemoeLibrary, Clin Microbiol Infect 2017. © by author M. genitalium fluoroquinolone treatment studies Meta-analysis on the efficacy of moxifloxacin for M. genitalium treatment – 17 studies, 252 patients 100 % 100 89 % 90 80 70 60 50 40 30 20 10 0 Before 2010 Since 2010 Pooled microbial cure rate (%) rate cure microbial Pooled ESCMID eLibraryYi et al Int J STD AIDS 2017 © by author Dual class resistance Prevalence – Japan : 17-30% – Australia : 8.6 - 9.8% – Denmark : 4.2% Mainly due to successive treatment failures of macrolides then fluoroquinolones Enormous clinical implications – No other highly effective antibiotics available What can we use as third line therapy? Murray Emerg Infect Dis 2017, Kikuchi J Antimicrobiol Chemother 2014; Le Roy Emerg Infect Dis 2016; Degushi Clin Infect Dis 2016; Kikuchi J Antimicrobiol ESCMIDChemother 2014 eLibrary 18 © by author • Third-line treatment for persistent MG infection after AZM and MXF Doxycycline 100 mg x2 daily for 14 days Pristinamycin 1 g x4 daily for 10 days ESCMID eLibrary © by author Pristinamycin MICs – AZM-resistant strains have higher MIC90 (0.5 mg/l) than AZM-susceptible strains (MIC90=0.125 mg/l) – but MICs close to the breakpoint : 1 mg/l (French society for microbiology, not dedicated to mycoplasmas) Maximum dose of 1g 4 times/day for 10 days was retained Evaluation in patients after azithromycin failure (2012-2016, Australia) – 114 persons : infection cured in 75% Option during pregnancy and in other situations where fluoroquinolones have failed or are contraindicated Not registered in many countries Renaudin et al.