Steven Weger MMIC 7050 Dec. 10 2013

1. Extended spectrum Beta-lactamase (ESBL) and carbapenamase producing E.coli and Klebsiella spp...the next multi-drug resistant catastrophe.  Steven Weger MMIC 7050 Dec. 10 2013

2. β-lactam antibiotics • Natural product application and discovery • Extremely important class of antimicrobial • >50% of all systemically used antimicrobials • High efficacy • Safety profile • Several derivatives

4. Transpeptidaseand D-alanylcarboxypeptidase(Penicillins Binding Proteins, PBP’s) contribute to "cross-linking,” a vital step in completing the cell wall. The Beta-Lactam Ring binds at the active site of the transpeptidase enzyme by mimicking the D-alanyl-D-alanine residues that would normally bind to this site

5. β-lactam Resistance Mechanisms

6. β-lactamases • Hydrolyze amide bond of lactam ring • Plasmid vs. Chromosomal • Inducible vs. constitutive • a.a. sequence • Substrate profile

9. ESBLs • Hydrolyze penicillins, broad- and extended-spectrum cephalosporins, and monobactams • Inhibited by Clavulanic acid • Spectrum of activity extanded by a.a. substitutions w/in enzyme • Plasmid mediated • Most commonly found in Enterobacteriaceae like E.coli and K. pneumoniae

10. ESBLs • 3 main families: • TEM-type -> Penicillin, ampicillin, 1st gen. cephalosporins • Over 200 variants • SHV-type -> Penicillins, ampicillin, 1st gen. cephalosporins • Over150 variants • Other types (CTX-M, OXA, etc.) • CTX-M have increased their spread due to ST131 • Frequently harbored in E. coli

11. ESBL Treatment options • β-lactam/β-lactamase inhibitor combination (e.g. amoxicillin/clavulanate) Clavulanic acid

12. ESBL Treatment options • Carbapenems • Preferred agent • Last line of defense • Resist ESBL hydrolysis

13. AmpCβ-lactamases • Hydrolyze penicillins, broad- and extended-spectrum cephalosporins • NOT inhibited β-lactamase inhibitor’s • Chromosomal or Plasmid encoded • Mutations in promoter can cause hyper-expression of chromosomal ampC • CMY and DHA are most common plasmid-type in E. coli and K. pneumoniae

14. Carbapenemases • Steadily increasing in prevalence • Hydrolyze virtually all β-lactams • CRE = morbidity, mortality • Limited treatment options : aminoglycosides, polymyxins, tigecycline, fosfomycin, and temocillin • Limited by the side effect profiles, nephrotoxicity, administration issues, and efficacy • Highly mobile

15. Molecular epidemiology of extended-spectrum b-lactamase-, AmpC b-lactamase- and carbapenemase-producing Escherichia coli and Klebsiellapneumoniae isolated from Canadian hospitals over a 5 year period: CANWARD 2007–11 Andrew J. Denisuik1, Philippe R. S. Lagace´-Wiens, Johann D. Pitout, Michael R.Mulvey, Patricia J. Simner, Franil Tailor, James A. Karlowsky, Daryl J. Hoban, Heather J. Adam and George G. Zhanelon behalf of the Canadian Antimicrobial Resistance Alliance (CARA)†

16. The Study • Determine the proportion of Escherichia coli and Klebsiella pneumonia collected from Canadian hospitals that produce ESBLs, AmpC β-lactamases, and carbapenemases. • The molecular characteristics of these pathogens as well as the patterns of antibiotic resistance are also described.

17. The study • 5451 E. coli and 1659 K. pneumonia isolates collected from 2007-2011 • AST using broth micro-dilution method • MIC breakpoints using CLSI 2012 guidelines • MDR = Resistance to 3 antimicrobial classes • XDR = Resistance to 5 antimicrobial classes

18. The Study • Putative ESBL-producer: • ceftriaxone and/or ceftazadime MIC of ≥1mg/L • ESBL production confirmed by phenotypic disc test • Putative AmpC-Producer: • ceftriaxone and/or ceftazidime MIC of ≥1 mg/L • cefoxitinMIC ≥32 mg/L that is ESBL negative by the CLSI confirmatory disc test • Putative Carbapenemase-Producer: • ertapenemMIC ≥0.5 mg/L

19. PCR for Resistance Enzyme • ESBL: • blaSHV, blaTEM, blaCTX-M and blaOXA • AmpC: • blaENT, blaDHA, blaFOX, and blaCIT • Promoter sequenced for all PCR Negative for above • Carbapenemase: • blaKPC, blaIMP, blaVIM, blaIMI, blaNDM, blaGES, blaOXA-48 • Genetic relatedness determined by PFGE

22. AmpCE. coli Characterization • Of 115 AmpC E. coli isolates: • 65 contained acquired AmpCβ-lactamase genes • 64 produced CMY-2 • 1 produced FOX-5 • 50 contained mutations in the promoter of the chromosomal ampC gene

23. Genetic Relatedness Among Isolates • Not related (<80% similarity) by PFGE • ST131 found in: • 56% of ESBL-producing E. coli • 29% of AmpC-producing E. coli • Steady  in ST131 over study period • 49%(2007)-72%(2011) • 102 of 153 CTX-M-15 Producers were ST131 • ST131 facilitating spread of CTX-M-15 in Canadian hospitals

24. Antimicrobial Susceptibility/Treatment Options • Greatest activity: amikacin, meropenem, ertapenem, and colistin • ESBL- and AmpCE. coli: highly susceptible (>93%) to piperacillin/tazobactam • ESBL-producing K. pneumonia susceptibility decreased to 67% • Tigecycline: ESBL- and AmpC- producing E. coli susceptibilities of 99.6% and 100% respectively • ESBL- producing K. pneumonia were less susceptible to this antibiotic with 83% susceptibility

25. MDR: • 79% of ESBL-producing E. coli • 69% of ESBL-producing K. pneumoniae • 34% of AmpC-producing E. coli • MDR among the ESBL- producing E. coli increased from 77% in 2007 to 83% in 2011 • 2.6% of ESBL-producing E. coli and 10.4% of ESBL-producing K. pneumonia demonstrated an XDR phenotype • 3 carbapenemase (all KPC) producing isolates: • Resistance to amoxicillin/clavulanate, piperacillin/tazobactam, and ertapenem • susceptible to colistin and tigecycline. Only one of these isolates was resistant to meropenem.

26. Diagnosis • ESBL-producer = inadequate therapy until resistance determined • Mortality doubles in bacteremia • MIC-based screening criteria and phenotypic methods of detection (disc test) -Appropriate therapy determination -Appropriate infection control -> Prevent clonal spread

27. Summary • Proportion of ESBL- and AmpC producing E. coli and K. pneumonia demonstrated significant national increase • ESBL- producing E. coli = dominant group in Canada • Rate of carbapenemase producers in Canadian hospitals will continue to increase • Due to selection pressure • Foreign travel

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