Mechanisms of macrolide resistance Ribosomal modification by - - PowerPoint PPT Presentation

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Mar 15, 2023 113 likes •265 views

Mechanisms of macrolide resistance Ribosomal modification by methylase ( erm genes) S. pneumoniae : erm (B) 75-100% of Ery-R strains in Europe S. pyogenes : erm (B) generally <50% of Ery-R strains erm (A) Ribosomal modification by

SLIDE 1

Mechanisms of macrolide resistance

Ribosomal modification by methylase (erm genes)

– S. pneumoniae: erm(B) 75-100% of Ery-R strains in Europe

– S. pyogenes: erm(B) generally <50% of Ery-R strains erm(A)

Ribosomal modification by mutation (rRNA, proteins)

– Occasional in Ery-R S. pneumoniae – Rare in Ery-R S. pyogenes (up to 18% in Eastern Europe) – The only mechanism or highly prevalent in H. pylori, Campylobacter, M. avium

Drug efflux (mef genes)

– <25% in Ery-R S. pneumoniae – >50% in Ery-R S. pyogenes (up to 95%)

SLIDE 2

Genotypes and phenotypes of macrolide resistance in pneumococci

Gene

Phenotype Phenotype acronym 14-15-M 16-M erm(B) MLSB (i) R R(I-S) MLSB (c) R R mef(A) M R S Rarely erm(A) gene

SLIDE 3

MICs of macrolides against S. pneumoniae according to resistance phenotypes/genotypes

MIC (mg/L) MLSB or erm(B) M or mef(A)

range MIC50 MIC90 range MIC50 MIC90

Azithromycin 8->128 >128 >128 2-32 16 16 Clarithromycin 2->128 >128 >128 1-64 8 8 Erythromycin 8->128 >128 >128 1-64 8 8 Roxithromycin 2->128 >128 >128 1-32 8 8 Josamycin 0.5->128 16 >128 0.03-0.025 0.06 0.12 Miocamycin 4->16 2-16 >128 0.25-0.5 - - Spiramycin 16->128 8-128 >128 0.06-1 0.25 0.5

Descheemaker (agar CO2),Fitoussi (agar), Ip (agar), Montanari (NCCLS broth microdil), Nishijima (agar)

SLIDE 4

MICs of macrolides against S. pyogenes with erm genes

Genotype Antimicrobial MIC (mg/L) Range MIC50 MIC90 erm(B) Erythromycin 1->128 >128 >128 Azithromycin 2->128 >128 >128 Clarithromycin 0.5->128 >128 >128 Roxithromycin 2->128 >128 >128 Josamycin 2->128 >128 >128 Miocamycin 16->16 >16 >16 Spiramycin 64->128 >128 >128 erm(A) Erythromycin 1->128 2->128 4->128 Azithromycin 2->128 8->128 32->128 Clarithromycin 0.5->128 1->128 2->128 Roxithromycin 2->128 16->128 16->128 Josamycin 0.06-1 0.03-0.25 0.06- 0.25 Miocamycin 0.06-0.5 0.25 0.25 Spiramycin 0.03-1 0.06-0.25 0.12-0.5

Bingen (NCCLS), Descheemaker (agar), Giovanetti (broth microdil.), Kataja (agar NCCLS)

SLIDE 5

MICs of macrolides against S. pyogenes with mef genes

Genotype Antimicrobial MIC (mg/L) Range MIC50 MIC90 mef(A) Erythromycin 1-32 8 8-16 Azithromycin 2-32 2-8 2-8 Clarithromycin 0.5-16 2-8 4-8 Roxithromycin 2-64 16 16 Josamycin 0.01-1 0.03-0.25 0.12-0.5 Miocamycin 0.06-0.5 0.25 0.25 Spiramycin 0.03-1 0.06-0.25 0.12-0.5

Descheemaker (agar), Giovanetti (broth microdil.), Kataja (agar NCCLS)

SLIDE 6

Ribosomal mutation and macrolide resistance in pneumococci

Azi Cla Ery Rox A2058U/G >32 16->32 >32 >32 A2059G >32 2 8 16 C2610U 0.125 0.03 0.06 0.125 C2611U 0.5 0.03-0.06 0.06-0.12 0.06-0.5 L22 protein 0.06-1 0.12-1 0.25-1 0.5-2 L4 protein 0.5 0.12 0.25 0.5

Canu et al., 2002

SLIDE 7

Population analysis of erythromycin-susceptible and resistant S. pneumoniae

5 10 15 20 25 30 35 40 45 50

0.03 0.06 0.12 0.25 0.5 1 2 4 8 16 32 64 128

No of strains MIC of erythromycin (mg/L)

S

M MLSB

Phenotype

SLIDE 8

Microbiological breakpoint

Easy to determine for macrolides
May differ by one or two dilutions according to the

methodology used (national committees), but equivalence exists

Clinical significance ?
Two kinds of breakpoints: microbiological and clinical?

! In fact for macrolides, the lower breakpoint is often the microbiological breakpoint

Erythromycin and S. pneumoniae S: BSAC: <=0.5mg/L, CA-SFM: <=1mg/L, NCCLS: <=0.25mg/L

SLIDE 9

Different levels of resistance to macrolides

S. pneumoniae and S. pyogenes with erm(B)

generally highly cross-resistant to all macrolides

S. pneumoniae and S. pyogenes with efflux or erm(A)

generally moderately resistant to 14-15-Md macrolides MIC50 (mg/L)

S. pyogenes
S. pneumoniae

Erythromycin 8 16 Azithromycin 2-8 8 Clarithromycin 2-8 8 Roxithromycin 16 8

SLIDE 10

Various significances of a moderate level of resistance to macrolides

Streptococcus pyogenes with MIC of erythromycin equal

to 8 mg/L may contain

– mef(A) gene: 16-Md macrolides and clindamycin active (« safe » use) – Inducible erm(A) gene: 16-Md macrolides and clindamycin active but risk for selection of constitutive mutants (10-7) – Inducible erm(B) gene: MIC of erythromycin increases after culture in the presence of erythromycin (in vivo?)

! detection of the resistance phenotype ! interpretive reading of the susceptibility test ! answer to the clinician

SLIDE 11

Azithromycin 10 vs 20 mg/kg/d for 3 days in acute GAS pharyngitis (1)

Inclusion

– Children between 2-12 years of age – Acute tonsillopharyngitis – Positive card test for streptococcal antigen confirmed by positive throat culture for GAS

Exclusion

– Antibiotics within 8 days – Hypersensitivity to beta-lactams or macrolides, severe underlying disease, previous inclusion, symptoms suggestive of viral infection

Treatment

– Azithromycin 10 mg/kg/d vs 20 mg/kg/d; 3 days (double-blinded) – Penicillin V 45 mg/kg/d; 10 days (open) – Clinical and bacteriological follow-up: days 14 and 30

Cohen R et al. Pediatric Infectious Diseases Journal, in Press

SLIDE 12

Outcome on day 14 (ITT-assessable patients)

AZM AZM Penicillin V 10 mg/kg/d 20 mg/kg/d Microbiological outcome Total no of patients 158 157 154 No of failures (%) 79 (50) 22 (14) 27 (17.5) Clinical outcome Total no of patients 169 165 167 No of failures (%) 28 (16.6) 14 (8.5) 12 (7.2)

SLIDE 13

Pre- and post-treatment GAS isolates

Isolate MIC of (mg/L) Mechanism Ery Azi Jos of R Pre-treatment 0.064 0.12 0.5 None strains

0.25

Post-treatment Strains 11 V2 0.5 2 1 L4 mutation 124 V2 0.5 1 1 L4 mutation 390 V2 1 2 2 L4 mutation 286 V2 >128 >128 >128 erm(B) 133 V2 8 8 2 unknown 323 V2 32 >128 128 unknown 508 8 8 0.5 mef(A) All pre- and post-treatment strains were related (PFGE and RAPD). Strains 286 and 508 acquired a DNA fragment with erm(B) and mef(A), respectively

SLIDE 14

Resistance by ribosomal mutation in Helicobacter, Campylobacter, M. avium

H. pylori: mutation of 23S rRNA

– MIC of clarithromycin: 8-256 mg/L, depending on the mutation

M. avium: mutation of 23S rRNA

– MIC of clarithromycin > 8 mg/L (generally >128 mg/L) (some strains with MIC=1 mg/L)

Campylobacter: mutation of 23S rRNA and…

– MIC of erythromycin 1->128 mg/L