1. Antimicrobial Therapy

2. The advent of antimicrobial therapy has dramatically increased the life span and quality of life for humans. More people have died of infection in wartime than have died from swords or bullets. Today, doctors are worried that we are dangerously close to a postantibiotic era where the drugs we have are no longer effective

3. The Origins of Antimicrobial Drugs Naturally occurring antimicrobials – Metabolic products of bacteria and fungi – Microbes produce antibiotics (their weapons) to reduce competition for nutrients and space Derived from: – bacteria in the genera Streptomyces and Bacillus – molds in the genera Penicillium and Cephalosporium Antibiotic= a substance produced by microorganisms that in small amounts inhibits another microorganism. Penicillium

4. Origins of Antimicrobial Therapy Many ancient cultures have used antimicrobials from plants and trees. First systematic attempt to find specific antimicrobials occurred in the early 1900’s.

5. History of Chemotherapy 1910--Paul Erhlich, “father of chemotherapy”, discovered that Salvarsan could treat syphilis 1935—Sulfa drugs were discovered 1928-40—Alexander Fleming discovered antimicrobial action from the mold, Penicillium notatum, however many years passed before penicillin was purified and produced.

6. What is the ideal antimicrobial? Selective toxicity: drug kills the pathogen without damaging the host Solubility in body fluids Toxicity not easily altered by bacteria Non-allergenic Stability: maintenance of a constant, therapeutic concentration in blood and tissue fluids Resistance by microorganisms not easily acquired Long shelf life Reasonable cost

7. The Action of Antimicrobial Drugs

8. Inhibition of Cell Wall Synthesis Penicillin's effect on the Gram negative bacterial cell wall The cell wall is a good, selective target since eukaryotes don’t have peptidoglycan Examples: Penicillin, bacitractin, cephalosporin, vancomycin Penicillins and cephalosporins inhibit the peptide crosslinks that hold the carbohydrate units together. Similar to taking a blow torch and cutting links in a chain link fence.

9. Inhibition of Cell Wall Synthesis: Penicillin Inhibits cell wall synthesis Produced by the mold, Penicillium chrysogenum – A diverse group (1 st, 2 nd, 3 rd generations) Natural (penicillin G and V) Semisynthetic (ampicillin, Augmentin) – Structure Beta-lactam ring Treat streptococci, meningococci, and spirochete infections

10. The R group is responsible for the activity of the drug, and cleavage of the beta-lactam ring will render the drug inactive. Inhibition of Cell Wall Synthesis: Penicillin

11. Injury to the Plasma Membrane Change in permeability of plasma membrane causes loss of important metabolites from cell – Interact with membrane phospholipids – Distorts the cell surface – Leakage of proteins and nitrogen bases

12. Inhibition of Protein Synthesis Exploit difference in ribosomes: Drugs specifically bind to 70S and not 80S because of specific shape ribosomes Erythromycin, Streptomycin, Tetracycline, Chloramphenical Some toxicity since mitochondria have 70S ribosomes

13. Inhibition of Nucleic Acid Synthesis Inhibition of DNA replication Inhibition of transcription of RNA – Modes of action include: Binds and cross-links the double helix Other quinolones – inhibits DNA unwinding enzymes Analogs of purines and pyrimidines that mimic natural bases

14. Inhibition of Folic Acid Synthesis Sulfonamides (sulfa drug) and trimethoprim – Competitive inhibition preventing the metabolism of DNA, RNA, and amino acid

15. Antibiotic Spectrum Broad-spectrum drugs: effective against more than one group of bacteria. Ex. tetracycline antibiotics Narrow-spectrum drugs: target a specific group Ex. polymyxin

16. Biofilm Effects on Antibiotic Treatment Biofilms are unaffected by the same antimicrobials that work against them when they are free living – penetration of the biofilm – different phenotype is expressed by biofilm bacteria, giving them different antibiotic sensitivity Strategies for treating biofilm infections – interrupting quorum sensing signals – adding DNase to antibiotics helps with penetration – impregnating devices with antibiotics

17. Bioavailability Effects on Antibiotic Treatment The chemical structure of a drug AND the mode of entry of a drug influence: concentration of drug in blood time of drug in blood

18. Actions of Antimicrobial Drugs Treatment of eukaryotic pathogens is more difficult because they are more similar to human cells. Need to target the few differences between cells. – Sterols in cell membrane in fungi Treatment of viral pathogens is also difficult because viruses find protection inside the human cell. Limited drugs available Difficult to maintain selective toxicity Effective drugs – target viral replication cycle – Entry – Nucleic acid synthesis – Assembly/release

19. Drug and Host Interaction Be cautious of toxicity to organs Some drugs can cause allergic reactions – (especially penicillin and sulfa drugs) Many times, drugs will suppress or alter the normal microflora – (good to take extra sources of live cultures(like Lactobacillus acidophilus found in yougurt and milk) to replenish flora Need Effective drugs—be mindful to use the best drug for the job.

20. Testing for Drug Susceptibility Will the treatment kill the pathogen? What concentration of antimicrobial is needed for Minimal Inhibitory Concentration (MIC) Natural selection of bacteria

21. Serial-Dilution Test-Review The minimum inhibitory concentration (MIC) is the concentration required to inhibit growth of a specific isolate in vitro under standardized conditions. It is determined by finding the lowest dilution without visible growth during serial dilution testing. This will vary for individual isolates.

22. Tests to Guide Chemotherapy Disk-Diffusion Method (Kirby-Bauer technique) Same as testing disinfectants Zone of inhibition surrounding the discs is measured and compared with a standard for each drug

23. Tests to Guide Chemotherapy E test uses a strip that contains a gradient of antimicrobial Indicates the concentration of antimicrobial needed to inhibit growth

24. The Effectiveness of Chemotherapeutic Agents Effects of Combinations of Drugs need to be careful of how you take the drug (grapefruit juice effect) and if you are taking any drugs that could interfere with the activity of the prescribed antibiotic (synergism or antagonism). The Future of Chemotherapeutic Agents Many diseases have become resistant to antibiotics. Chemicals produced by plants and animals are providing new antimicrobial agents. Synergy between dalfopristin (right) and quinupristin (left) against a staphylococcal strain resistant to both.

25. The MIC and Therapeutic Index In vitro activity of a drug is not always correlated with the in vivo effect Failure of antimicrobial treatment is due to: – the inability of the drug to diffuse into that body compartment (brain, joints, skin) – resistant microbes in the infection that didn’t make it into the sample collected for testing – an infection caused by more than one pathogen (mixed), some of which are resistant to the drug

26. The MIC and Therapeutic Index Therapeutic index: the ratio of the dose of the drug that is toxic to humans as compared to its minimum effective (therapeutic) dose: – the smaller the ratio, the greater the potential for drug reactions – TI = 1.1 is a risky choice – TI = 10 is a safer choice – the drug with the highest therapeutic index has the widest margin of safety

27. The MIC and Therapeutic Index The physician must take a careful history before prescribing an antibiotic – preexisting conditions that might influence the activity of the drug, Ex. Alcoholism, intestinal damage – history of allergy to a certain class of drugs – underlying liver or kidney disease, our filters! – infants, the elderly, and pregnant women require special precautions – intake of other drugs can result in increased toxicity or failure of one or more drugs. Ex. Birth control! – genetic or metabolic abnormalities – site of infection, route of administration, cost

28. How Does Drug Resistance Develop? Resistance to penicillin developed in some bacteria as early as 1940 In the 1980s and 1990s scientists began to observe treatment failures on a large scale Microbes become newly resistant to a drug after one of the following occurs – spontaneous mutations in critical chromosomal genes – acquisition of entire new genes or sets of genes via horizontal transfer from another species

29. Development of Drug Resistance Chromosomal drug resistance – usually results from spontaneous random mutation – can be the result of a phenotypic, rather than a genotypic change; slowing or stopping of metabolism so that the microbe can’t be harmed by the antibiotic

30. Development of Drug Resistance Resistance through horizontal transfer – Resistance (R) factors: plasmids containing antibiotic resistance genes (some have 6-7 resistance genes!) – Can be transferred through conjugation, transformation, or transduction – Plasmids encoded with drug resistance are naturally present in microbes before they have been exposed to an antibiotic – Transposons that have duplicated and inserted genes for drug resistance into plasmids

31. Resistance plasmids can carry many resistance genes YIKES! We now have a “super bug”!

32. Resistant Strains Rare x x Resistant Strains Dominant Antimicrobial Exposure x x x x x x x x x x Antibiotic Resistant Strain Propagation Antimicrobial exposure causes pressure to select resistant strains. Without antibiotics, selective pressure decreases and antibiotic resistance genes may be lost.

33. Intermicrobial transfer of plasmids containing resistance genes

34. S. aureus Penicillin 1950s Penicillin-resistant S. aureus Clinical Implications in the Development of Drug Resistant Staphylococcus aureus How do we treat S. aureus infections when they become completely resistant to our last line antibiotic, Vancomycin? Methicillin 1970s Methicillin- resistant S. aureus (MRSA) Vancomycin-resistant enterococci (VRE) Vancomycin 1990s 1997 Vancomycin intermediate- resistant S. aureus (VISA) Vancomycin Resistant S. aureus 1997

35. Mechanisms Associated with Drug Resistance 1.Drug inactivation 2.Decreased permeability 3.Activation of drug pumps 4.Change in drug binding site 5.Use of alternate metabolic pathways

36. Beta-lactamase breaks the beta-lactam ring and renders penicillin inactive Penicillins -Original penicillin was narrow-spectrum and susceptible to microbial counterattacks -Molecule has been altered and improved upon over the years -Later penicillins overcome the limitations of the original molecule Synthetic Advances in Antibiotics

37. The Future of Chemotherapeutic Agents Some drugs are actually a combination of 2 drugs. This nifty example shows an ingenious way to start with one antibiotic and then if it is being degraded a second one can be released.

38. New Approaches to Antimicrobial Therapy Using bacteriophages – Eastern European countries use mixtures of bacteriophages as medicine, but these drugs have never been approved for use in the West – Biophage-PA used to treat ear infections caused by Pseudomonas aeruginosa biofilms – Other researchers are incorporating bacteriophages into wound dressings – Advantage to bacteriophage is their narrow specificity; only infect one species of bacterium

39. Methicillin-resistant Staphylococcus aureus(MRSA) Vancomycin-resistant Enterococci (VRE) Non-Intensive Care Unit Patients Intensive Care Unit Patients Antibiotic Resistance is Prevalent and Rising Source: National Nosocomial Infections Surveillance (NNIS) System

40. Preventing Antibiotic Resistance Prevent infection Stay healthy Diagnose and treat infection effectively Don’t treat viral infections with antibiotics Use effective antibiotics Use antimicrobials wisely Take full course of antibiotics Stop feeding antibiotics to livestock Prevent transmission Wash your hands Isolate infections

41. Take the full course of antibiotics to decrease resistance Preventing Antibiotic Resistance

42. Antibiotics are sold world wide without a prescription

43. Development of Drug Resistance A wide variety of soil bacteria can not only survive in the presence of many antibiotics, but can use the antibiotics as fuel

44. Development of Drug Resistance A large population of natural environmental bacteria exists with antibiotic resistance capabilities that might be transferred to disease-causing bacteria Non-disease-causing flora of humans and animals can also harbor antibiotic resistance genes that can jump into pathogenic bacteria with which they share space Yep, it’s a cow peeing