1. Today’s talk Dimensions of AB resistance History of AB resistance Biology of AB resistance –Biology of antibiotic action and measurement –Genetics –Biochemistry –Selection Some guiding questions and concepts

2. Dimensions of Antibiotic Resistance Bacterial species –Type of transmission: hospital or community Antibiotic Genetic mechanism of resistance: how does the bug get the genes to be resistant? Biochemical mechanism of resistance: what does the bug do to be resistant? Mechanisms of selection for resistance –Individuals –Populations Study design to assess these mechanisms

3. History of Antimicrobial Resistance

4. Early principles: Paul Ehrlich The therapia sterilisans magna consists in this, that by means of one or at most two injections the body is freed from the parasites…. Here, therefore, the old therapeutic remedy is applicable …. frapper fort et frapper vite A further advantage of combined therapy is, that under the influence of two different medicines the danger of rendering the parasites immune to arsenic, which naturally would be a very great obstacle in connexion with further treatment, is apparently greatly minimized.

5. GC, H. flu ADD DHFR ADENYLATE E.coli ADD ß-LACTAMASE METHYLATE RIBOSOME ACETYLATE EFFLUX PHOSPORYLATE S. pneumo ADD PBP S. aureus ADD ß- LACTAMASE All Gram negs ADD DHTS 1930 1940 1950 1960 1970 1980 1990 2000 GC Mening VRE QMRSA Pseud Kleb E.coli ESBLs AmpCs ? VMRSA Sulfa Penicillin Strept Tetra Chlora Erythro Oxa/amp/ceph Vanco Genta Trimethoprim 3 rd gen cephalosporins Fluroquinolones Carbapenems MRSA ADD PBP MUTATE

6. Resistance comes fast J. Davies 1997

7. P. aeruginosa resistant to imipenem K. pneumoniae non-susceptible to 3rd cephalosporins ICU Patients Non-ICU Patients P. aeruginosa resistant to quinolones Source: NNIS DATA Clinics Chest Med. 20:303-315 Resistance increases quickly

8. Methicillin-resistant S. aureus Methicillin-resistant Coagulase-negative Staph Vancomycin-Resistant enterococci ICU Patients Non-ICU Patients Source: NNIS DATA Clinics Chest Med. 20:303-315 Resistance goes up quickly (2)

9. Still growing: Resistance in Streptococcus pneumoniae in US G Doern et al., Clin Inf Dis. 2005

10. Geographic variation in antibiotic resistance H. Goossens et al. 2005 Lancet

11. Geographic Variation Resistance in 1999 Proportion Penicillin Resistant State

12. The Tragedy of The Commons

13. Resistance goes down slowly, if at all V Enne et al., Lancet 2001 39% R45% R

14. Resistance goes down slowly, if at all Hennessy et al. 2002, CID ~30% decline in prescribing after initial intervention ~25% after expanded

15. But in hospitals, changes can move faster Dunkle et al. Amer J Med 1981

16. Biology of Antimicrobial Resistance

17. How do antibiotics kill? Static v. cidal –Bacteriostatic: prevent cell division (e.g. by preventing protein synthesis) –Bactericidal: kill bacteria directly Cidal drugs often kill only dividing bugs –Cell wall synthesis inhibitors –Others This makes treatment of latent infection especially difficult (TB)

18. Antibiotics and the immune system Immune responses required to kill alongside bactriostatic drugs Also for many bactericidal drugs: phenotypic resistance Wiuff et al. AAC 2005

19. “Normal” Flora S. aureus GNR (E. coli, Klebsiella Enterobacter) Enterococcus faecium S. pneumoniae H. influenzae N. meningitidis

20. Normal flora: Consequences Treatment exerts selection on “innocent bystanders” Most of the harm done by use of a drug may be on species OTHER than the target of treatment –Optimal dosing for treatment ≠ optimal to prevent resistance Most of the exposure of a given species to a given drug may be due to treatment of OTHER infections

21. Measuring resistance: Minimal Inhibitory Concentration (MIC) Broth microdilutionEtest

22. MIC is a simplification Regoes et al. AAC 2004

23. Limitations of MIC Subpopulations Depends on in vitro conditions: pH, etc – not necessarily same as in vivo One parameter summary of the curve Ignores physiologic variation Regoes et al. AAC 2004

24. Biological Aspects of Resistance Genetics: how is drug resistance coded? –And how can it move from one bug to another? –Think of: floppy disk, memory stick, punch card Biochemistry/mechanism: what does a bug do to become resistant? –Think of: iTunes, RealPlayer, Microsoft Media Player How resistance is selected: how do we increase the frequency of resistant bugs? –Block that metaphor!

25. Intrinsic resistance All members of a species are resistant, and have been since before clinical use –Tuberculosis and penicillin: naturally encodes beta-lactamase –Vancomycin-producing species and vancomycin: alters its cell wall to be insensitive (same as resistance in targets!) –Don’t activate prodrug Isoniazid or pyrazinamide and non-mycobacteria: not chemically altered to become active

26. Transformation Plasmid transfer Genetic Mechanisms of Resistance Acquisition Mutation

27. Implications Mutation: easy to get a resistant strain in almost any patient: mutation frequencies ~10 -7 – 10 -10 –Unless multiple mutations are required to confer resistance! –Examples: Tuberculosis, HIV Other mechanisms –Very complex mechanisms of resistance can evolve, because they can move as a block from one bug to the next –Can even transfer from one species to another –Emergence of a resistant bug in a single host is unlikely, unless a mix of resistant bugs and sensitive bugs is present

28. Courtesy Tom O’Brien, BWH Epidemic Plasmids

29. Integrons System for combining resistance (and other) genes Can take up new genes via integrase and add them to the “package” Often on transposons or plasmids www.mmb.usyd.edu.au/coleman/

30. Reduced permeability Efflux Degradation Detoxification Target alteration: enzyme Target alteration: mutation Target amplification Inactivate the activator of the prodrug Biochemical mechanisms

31. Mechanisms: their consequences Most mechanisms are quite specific to one drug or class of drugs –Enzymes to alter drug or target –Target changes or amplification A few mechanisms confer resistance to more than one class of drug –Efflux pump: “MDR transporter”

32. Mechanisms High-level resistance: completely resistant to any achievable concentration Partial resistance: small change in MIC

33. Mechanisms of selection

34. How Antimicrobial Use Increases Resistance: Mechanisms Rx “Acquired” Resistance: selection within host – Patient infected with a susceptible organism –Treatment selects a resistant variant Emerg Inf Dis 2002 8:347

35. How Antimicrobial Use Increases Resistance: Mechanisms “Primary” Resistance: Selection in host population –Patient infected with a resistant organism –Competitive mechanism: Treatment selects by reducing transmission of susceptible infections Rx

36. How Antimicrobial Use Increases Resistance: Mechanisms Increasing susceptibility to colonization –Patient carries a “normal flora” –Treatment increases susceptibility to colonization by opening ecologic niche Rx

37. Summary of Mechanisms of Selection of Resistance by Abx Rx

38. Is antibiotic use harmful to individuals?

39. Co-selection Dental fillings installed and removed from experimental monkeys Summers 1993 AAC

40. Concepts and questions Antibiotic resistance is interesting: how bad is it? How can we measure the costs? Compared to what?

41. What can we do to/for an individual patient to prevent resistant infections?

42. Why have some kinds of drug resistance increased fast, others slowly or not at all?

43. What can we do to the population as a whole to reduce the risk of reistant infections?

44. In what circumstances does doing what is best for the patient Increase the burden of resistance in the community? Reduce the burden of resistance in the community? Both but on different time scales?

45. What would happen if we stopped using antibiotics tomorrow? To disease? To resistance?