Antimicrobial Agents

1. Antimicrobial Agents Martin Votava Olga Kroftová

2. Overview • If bacteria make it past our immune system and start reproducing inside our bodies, they cause disease. • Certain bacteria produce chemicals that damage or disable parts of our bodies. • Antibiotics work to kill bacteria.Antibiotics are specific to certain bacteria and disrupt their function.

3. What is an Antibiotic? • An antibiotic is a selective poison. • It has been chosen so that it will kill the desired bacteria, but not the cells in your body. Each different type of antibiotic affects different bacteria in different ways. • For example, an antibiotic might inhibit a bacteria's ability to turn glucose into energy, or the bacteria's ability to construct its cell wall. Therefore the bacteria dies instead of reproducing.

4. Antibiotics • Substances produced by various species of microorganisms: bacteria, fungi, actinomycetes- to suppress the growth of other microorganisms and to destroy them. Today the term ATB extends to include synthetic antibacterial agents: sulfonamides and quinolones.

5. History • The German chemist Paul Ehrlich developed the idea of selective toxicity: that certain chemicals that would be toxic to some organisms, e.g., infectious bacteria, would be harmless to other organisms, e.g., humans. • In 1928, Sir Alexander Fleming, a Scottish biologist, observed that Penicillium notatum, a common mold, had destroyed staphylococcus bacteria in culture.

6. Sir Alexander Fleming

7. Fleming’s Petri Dish

8. Zone of Inhibition • Around the fungal colony is a clear zone where no bacteria are growing • Zone of inhibition due to the diffusion of a substance with antibiotic properties from the fungus

10. History • Penicillin was isolated in 1939, and in 1944 Selman Waksman and Albert Schatz, American microbiologists, isolated streptomycin and a number of other antibiotics from Streptomyces griseus.

12. Susceptibility vs. Resistanceof microorganisms to Antimicrobial Agents • Success of therapeutic outcome depends on: • Achieving concentration of ATB at the site of infection that is sufficient to inhibit bacterial growth. • Host defenses maximally effective –MI effect is sufficient – bacteriostatic agents (slow protein synthesis, prevent bacterial division) • Host defenses impaired- bactericidal agents • Complete ATB-mediated killing is necessary

14. Susceptibility vs. Resistance(cont.) • Dose of drug has to be sufficient to produce effect inhibit or kill the microorganism: • However concentration of the drug must remain below those that are toxic to human cells – • If can be achieved – microorganism susceptible to the ATB • If effective concentration is higher than toxic- microorganism is resistant

15. Susceptibility vs. Resistance(cont.) • Limitation of in vitro tests • In vitro sensitivity tests are based on non-toxic plasma concentrations –cut off • Do not reflect concentration at the site of infection • E.g.: G- aer.bacilli like Ps.aeruginosa inhibited by 2 – 4 ug/ml of gentamycin or tobramycin. Susceptible !?

16. Disk Diffusion Test Determination of MIC Str Tet Ery 4 2 1 0 8 Tetracycline (μg/ml) Chl Amp MIC = 2 μg/ml Antibiotic Susceptibility Testing

17. Susceptibility vs. Resistance(cont.) • Plasma concentration above 6-10 ug/ml may result in ototoxicity or nephrotoxicity • Ration of toxic to therapeutic concentration is very low –agents difficult to use. • Concentration in certain compartments – vitreous fluid or cerebrospinal fluid much lower than those in plasma. • Therefore can be only marginally effective or ineffective even those in vitro test states sensitive.

18. Susceptibility vs. Resistance(cont.) • Therefore can be only marginally effective or ineffective even those in vitro test states „sensitive“. • Conversely – concentration of drug in urine may be much higher than in plasma , so „resistant“ agents can be effective in infection limited to urine tract

19. Resistance • To be effective ATB must reach the target and bind to it. • Resistance: • Failure to reach the target • The drug is inactivated • The target is altered

20. Resistance (cont.) • Bacteria produce enzymes at or within the cell surface –inactivate drug • Bacteria possess impermeable cell membrane prevent influx of drug. • Transport mechanism for certain drug is energy dependent- not effective in anaerobic environment. • ATB as organic acids penetration is pH –dependent.

21. Resistance (cont.) • Acquired by mutation and passed vertically by selection to daughter cells. • More commonly – horizontal transfer of resistance determinant from donor cell, often another bacterial species, by transformation, transduction, or conjugation. • Horizontal transfer can be rapidly disseminated • By clonal spread or resistant strain itself • Or genetic exchange between resistant and further susceptible strains.

22. Resistance (cont.) • Methicilin resistant strains of Staphylococcus aureus clonally derived from few ancestral strains with mecA gene • Encodes low-affinity penicillin-binding protein that confers methicillin resistance. • Staphylococcal beta-lactamase gene, which is plasmid encoded, presumambly transferred on numerous occasions. Because is widely distributed among unrelated strains, identified also in enterococci

23. Selection of the ATB • Requires clinical judgment, detailed knowledge of pharmacological and microbiological factors. • Empirical therapy – initial – infecting organism not identified – single broad spectrum agent • Definitive therapy- microorganism identified – a narrow –spectrum low toxicity regiment to complete the course of treatment

24. Empirical and Definite Therapy • Knowledge of the most likely infecting microorganism and its susceptibility • Gram stain • Pending isolation and identification of the pathogen • Specimen for culture from site of infection should be obtain before initiation of therapy • Definite therapy

25. Penicillins • Penicillins contain a b-lactam ring which inhibits the formation of peptidoglycan crosslinks in bacterial cell walls (especially in Gram-possitive organisms) • Penicillins are bactericidal but can act only on dividing cells • They are not toxic to animal cells which have no cell wall

26. Synthesis of Penicillin • b-Lactams produced by fungi, some ascomycetes, and several actinomycete bacteria • b-Lactams are synthesized from amino acids valine and cysteine

27. b Lactam Basic Structure

28. Penicillins (cont.)Clinical Pharmacokinetics • Penicillins are poorly lipid soluble and do not cross the blood-brain barrier in appreciable concentrations unless it is inflamed (so they are effective in meningitis) • They are actively excreted unchanged by the kidney, but the dose should be reduced in severe renal failure

29. Penicillins (cont.)Resistance • This is the result of production of b-lactamase in the bacteria which destroys the b-lactam ring • It occurs in e.g. Staphylococcus aureus, Haemophilus influenzae and Neisseria gonorrhoea

30. Penicillins (cont.)Examples • There are now a wide variety of penicillins, which may be acid labile (i.e. broken down by the stomach acid and so inactive when given orally) or acid stable, or may be narrow or broad spectrum in action

31. Penicillins (cont.)Examples • Benzylpenicillin (Penicillin G) is acid labile and b-lactamase sensitive and is given only parenterally • It is the most potent penicillin but has a relatively narrow spectrum covering Strepptococcus pyogenes, S. pneumoniae, Neisseria meningitis or N. gonorrhoeae, treponemes, Listeria, Actinomycetes, Clostridia

32. Penicillins (cont.)Examples • Phenoxymethylpenicillin (Penicillin V) is acid stable and is given orally for minor infections • it is otherwise similar to benzylpenicillin

33. Penicillins (cont.)Examples • Ampicillin is less active than benzylpenicillin against Gram-possitive bacteria but has a wider spectrum including (in addition in those above) Strept. faecalis, Haemophilus influenza, and some E. coli, Klebsiella and Proteus strains • It is acid stable, is given orally or parenterally, but is b-laclamase sensitive

34. Penicillins (cont.)Examples • Amoxycillin is similar but better absorbed orally • It is sometimes combined with clavulanic acid, which is a b-lactam with little antibacterial effect but which binds strongly to b-lactamase and blocks the action of b-lactamase in this way • It extends the spectrum of amoxycillin

35. Penicillins (cont.)Examples • Flucloxacillin is acid stable and is given orally or parenterally • It is b-lactamase resistant • It is used as a narrow spectrum drug for Staphylococcus aureus infections

36. Penicillins (cont.)Examples • Azlocillin is acid labile and is only used parenterally • It is b-lactamase sensitive and has a broad spectrum, which includes Pseudomonas aeruginosa and Proteus species • It is used intravenously for life-threatening infections,i.e. in immunocompromised patients together with an aminoglycoside

37. Penicillins (cont.)Adverse effects • Allergy (in 0.7% to 1.0% patients). Patient should be always asked about a history of previous exposure and adverse effects • Superinfections(e.g.caused by Candida ) • Diarrhoea : especially with ampicillin, less common with amoxycillin • Rare: haemolysis, nephritis

38. Penicillins (cont.)Drug interactions • The use of ampicillin (or other broad-spectrum antibiotics) may decrease the effectiveness of oral conraceptives by diminishing enterohepatic circulation

39. Antistaphylococcus penicillins • Oxacillin, cloxacillin • Resistant against staphylococcus penicillinasis

40. Cephalosporins • They also owe their activity to b-lactam ring and are bactericidal. • Good alternatives to penicillins when a broad -spectrum drug is required • should not be used as first choice unless the organism is known to be sensitive

41. Cephalosporins • BACTERICIDAL- modify cell wall synthesis • CLASSIFICATION- first generation are early compounds • Second generation- resistant to β-lactamases • Third generation- resistant to β-lactamases & increased spectrum of activity • Fourth generation- increased spectrum of activity

42. Cephalosporins • FIRST GENERATION- eg cefadroxil, cefalexin, Cefadrine - most active vs gram +ve cocci. An alternative to penicillins for staph and strep infections; useful in UTIs • SECOND GENERATION- eg cefaclor and cefuroxime. Active vs enerobacteriaceae eg E. coli, Klebsiellaspp,proteus spp. May be active vs H influenzae and N meningtidis

43. Cephalosporins • THIRD GENERATION- eg cefixime and other I.V.s cefotaxime,ceftriaxone,ceftazidine. Very broad spectrum of activity inc gram -ve rods, less activity vs gram +ve organisms. • FOURTH GENERATION- cefpirome better vs gram +ve than 3rd generation. Also better vs gram -ve esp enterobacteriaceae & pseudomonas aerugenosa. I.V. route only

45. Cephalosporins (cont.)Adverse effects • Allergy (10-20% of patients wit penicillin allergy are also allergic to cephalosporins) • Nephritis and acute renal failure • Superinfections • Gastrointestinal upsets when given orally

46. Aminoglycosides (bactericidal)streptomycin, kanamycin, gentamicin, tobramycin, amikacin, netilmicin, neomycin (topical) • Mode of action - The aminoglycosides irreversibly bind to the 16S ribosomal RNA and freeze the 30S initiation complex (30S-mRNA-tRNA) so that no further initiation can occur. They also slow down protein synthesis that has already initiated and induce misreading of the mRNA. By binding to the 16 S r-RNA the aminoglycosides increase the affinity of the A site for t-RNA regardless of the anticodon specificity. May also destabilize bacterial membranes. • Spectrum of Activity -Many gram-negative and some gram-positive bacteria • Resistance - Common • Synergy - The aminoglycosides synergize with β-lactam antibiotics. The β-lactams inhibit cell wall synthesis and thereby increase the permeability of the aminoglycosides.

47. AminoglycosidesClinical pharmacokinetics • These are poorly lipid soluble and, therefore, not absorbed orally • Parenteral administration is required for systemic effect. • They do not enter the CNS even when the meninges are inflamed. • They are not metabolized.

48. Aminoglycosides (cont.)Clinical pharmacokinetics • They are excreted unchanged by the kidney (where high concentration may occur, perhaps causing toxic tubular demage) by glomerular filtration (no active secretion). • Their clearance is markedly reduced in renal impairment and toxic concentrations are more likely.

49. Aminoglycosides (cont.)Resistance • Resistance results from bacterial enzymes which break down aminoglycosides or to their decreased transport into the cells.

50. Aminoglycosides (cont.)Examples • Gentamicin is the most commonly used, covering Gram-negative aerobes, e.g. Enteric organisms (E.coli, Klebsiella, S. faecalis, Pseudomonas and Proteus spp.) • It is also used in antibiotic combination against Staphylococcus aureus. • It is not active against aerobic Streptococci.