Overview on resistance mechanisms in Grampositive. Institute of Medical Microbiology University of Zürich, Switzerland B.

Transcription

1 Overview on resistance mechanisms in Grampositive bacteria Institute of Medical Microbiology University of Zürich, Switzerland B. Berger-Bächi

2 Antibiotic resistance Selective advantage in hospitals: selection by antibiotics: multiresistance accumulation of resistant bacteria spread in nature: competition with other bacteria: fitness community hospital

3 Resistance mutation point mutations IS elements one step/multiple steps acquisition of foreign DNA plasmids, transposons, resistance islands DNA fragments - recombination metabolic status biofilm SCV

4 Mutators: more rapid resistance formation Mobile elements (IS elements) promote resistance formation Resistance mechanisms depend on Gram pos/neg facility and mode of gene exchange physiological/genetic characteristics drug Combination of different resistance mechanisms frequent

5 Resistance strategies drug inactivation target modification prevention of access to target

6 Gram-positive microorganisms no outer membrane = one barrier less against antibiotics

7 Giesbrecht β-lactam resistance Destruction of the drug penicillinases, mobile Modification of target high molecular weight PBPs: different genetic means streptococci: mosaic PBPs staphylococci: additional PBP2a enterococci: PBP5 overproduction, or mutation, plasmid borne PBP3r

8 Mosaic PBPs Streptococcus pneumoniae horizontal gene transfer from commensal streptococci followed by homologous recombination and deletion of intervening sequences of recipient DNA, point mutations PBPs with altered structure and lowered affinity to beta-lactams

9 High penicillin resistance in pneumococci mosaic PBPs require mosaic MurM MurM is involved in the synthesis of the peptidoglycan side chain particular PBP-MurM combinations observed in clinical isolates Trends in Microbiol 2003, 11:547

10 MRSA SCCmec meca coding for an additional low-affinity PBP2a Health care type MRSA (multiresistant) AAC 2006, 50:1001 Emerging community type MRSA (susceptible to most non β-lactam antibiotics)

11 PBP2a transpeptidase with low affinity to penicillinase resistant beta-lactams essential partner: PBP2 in presence of beta-lactams: PBP2a recruits PBP2 to the septal plane

12 Fitness cost: SCCmec reduces the growth rate MSSA MRSA cured MRSA Introduction of SCCmec type I into S. aureus and selection for oxacillin resistance lowers the growth rate. After curing of SCCmec the growth rate is restored to its original value. Doubling time min Oxacillin MIC μg/ml MSSA MRSA cured MRSA AAC 2004, 48:2295

13 Glycopeptides steric hindrance of glycan elongation and peptidoglycan crosslinking

14 vana mediated glycopeptide resistance in enterococci destruction of exisiting peptidoglycan synthesis of a new insensitive target

15 S. aureus: VRSA Transfer of the vana operon to MRSA (Tn1546-like element) MI-VRSA PA-VRSA NY-VRSA VRSA-5 high level resistance low level resistance loss of vana operon, longer lag phase for induction low level resistance low frequency loss of resistance: plasmid or Tn instability

16 VISA: intermediate vancomycin resistant S. aureus Intrinsic chromosomal mutations altered gene expression Multifactorial multiple steps required Strain specific level of resistance attained is strain specific Polyclonal no unique clonal origin

17 Intermediate vancomycin resistance in S. aureus Increased number of false glycopeptide targets - increased cell-wall thickness - decreased cross linking

18 Glycopeptide intermediate resistance affects expression of virulence genes and growth 26 min 38 min doubling time wild type step 1 step 2 step OD 600 spa protein A SA1007 α-hemolysin precursor geh sbi opp1a lipase IgG binding protein Sbi oligopeptide transporter

19 Glycopeptide-r affects methicillin-r teicoplanin MIC MRSA 1 st 2 nd 3 rd step selection for growth on increasing teicoplanin concentrations step mutant methicillin µg/ml

20 Correlation daptomycin-r:visa VISA: thickened cell wall (no correlation with VRSA: altered cell wall composition) Cui et al. AAC 2006, 50:1079

21 Daptomycin, Cubicin 13-member amino acid cyclic lipopeptide compound that contains a water-soluble hydrophilic core with a lipophilic tail binds to bacterial membranes and causes depolarisation of the membrane potential, loss of K+, leads to inhibition of protein, DNA and RNA synthesis. rapid, concentration-dependent bactericidal activity

22 Daptomycin resistance in clinical isolates selection by suboptimal daptomycin therapy no depolarisation of membrane potential membrane binds less daptomycin deletion of an 80 kda membrane protein Kaatz et al. IJAA 2006,28:280

23 Daptomycin resistance formation frequency of spontaneous resistance < growth defects increased membrane potential ΔΨ stepwise decrease in susceptibility by accumulation of mutations MprF: lysyl-phosphatidylglycerol synthetase YycG: sensor histidine kinase of two component sensor transducer (involved in cell permeability to MLS antibiotics) RpoB, RpoC: subunits of RNA polymerase point mutations identified in emerging daptomycin clinical isolates Silverman et al AAC 2001, 45:1799 Friedman et al. AAC 2006, 50:2137

24 MLS B Inhibitors of protein synthesis Macrolide Lincosamide Streptogramin Erythromycin Clindamycin Streptogramin B Ketolide Telithromycin

25 MLS B and ketolide target site bind to 50S subunit of free or initiating ribosomes overlapping binding sites domains V and II of 23S rrna ribosomal proteins L4 and L22

26 MLS Resistance Target site modification Efflux Drug modification

27 Target site modification erm, mono- or di-methylation of the 23S rrna inducible / constitutive (resistant to inducing antibiotic) mutation of the 23S rrna generally in pathogens with few rrn operons: H. pylori, M. avium; S. pneumoniae and other species mutations of ribosomal protein L4/L22 S. pneumoniae/s. aureus, S. pneumoniae

28 Ketolide resistance correlates with degree of dimethylation Streptococcus pneumoniae constitutive ermb resistant to macrolides susceptible to ketolides monomethlyation > dimethylation Ero increases dimethylation ketolide resistance Streptococcus pyogenes constitutive ermb resistant to macrolides and ketolides dimethylation

29 Efflux mef(a) major facilitator resistance to: 14-membered macrolides susceptibility to: lincosamides, streptogramin B associated with conjugative elements: Corynebacteria, Enterococcus, Micrococcus, Streptococcus msr(a) ABC transporter resistance to: 14-membered macrolides, streptogramin B susceptibility to: lincosamides plasmid located: Staphylococcus vga(a) vga(b) ABC transporter (efflux, target protection?) resistance to: streptogramin A susceptibility to: macrolides, lincosamides plasmid located: Staphylococcus

30 Enzymatic modification of the antibiotic vgb: streptogramin B hydrolase Enterococcus, Staphylococcus vat: streptogramin A acetyltransferase Staphylococcus mph: macrolide phosphorylation Staphylococcus, Escherichia lnu: lincosamide nucleotidyl-transferase S. haemolyticus, S. aureus, E. faecium ere: erythromycin esterase Enterobacter, Escherichia, Klebsiella, Citrobacter, Klebsiella, Proteus

31 Combination of multiple resistance mechanisms frequent The efficiency of a resistance mechanism depends on the genetic background of the host strain Antibiotic resistance can modify virulence characteristics of the host strain