Controlling Microbial Growth in the Body: Antimicrobial Drugs
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1 PowerPoint Lecture Presentations prepared by Mindy Miller-Kittrell, North Carolina State University C H A P T E R 10 Controlling Microbial Growth in the Body: Antimicrobial Drugs
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3 bacteria fungi HOW? WHAT? virus protozoa
4 Antimicrobials Antibiotics (antimicrobial agents produced naturally by microbes) and semisynthetic and synthetic chemicals Typically used for treatment of disease and not for environmental control (like earlier chemicals) Some used for antimicrobial control outside the body Nisin (kill bacteria in cheese) Natamycin (kill fungi in cheese) is Lysozyme an antibiotic?
5 Antibiotic s targets on microbe Mechanisms of Antimicrobial Action Inhibition of Cell Wall Synthesis Disruption of Cytoplasmic Membranes Inhibition of Protein Synthesis Inhibition of Metabolic Pathways/Nucleic Acid Synthesis Prevention of Virus Attachment
6 Figure 10.2 Mechanisms of action of microbial drugs. Inhibition of pathogen's attachment to, or recognition of, host Arildone Pleconaril Human cell membrane Inhibition of DNA or RNA synthesis Actinomycin Nucleotide analogs Quinolones Rifampin Inhibition of cell wall synthesis Penicillins Cephalosporins Vancomycin Bacitracin Isoniazid Ethambutol Echinocandins (antifungal) Bacteria/fungi Inhibition of protein synthesis Aminoglycosides Tetracyclines Chloramphenicol Macrolides Antisense nucleic acids Disruption of cytoplasmic membrane Polymyxins Polyenes (antifungal) Inhibition of general metabolic pathway Sulfonamides Trimethoprim Dapsone
7 Figure 10.1 Antibiotic effect of the mold Penicillium chrysogenum. Staphylococcus aureus (bacterium) Penicillium chrysogenum (fungus) Zone where bacterial growth is inhibited
8 Mechanisms of Antimicrobial Action Antibiotics Antimicrobial agents produced naturally by organisms Common usage: antibiotic = antibacterial
9 The History of Antimicrobial Agents Natural Semisynthetics Chemically altered antibiotics that are more effective, longer lasting, or easier to administer than naturally occurring ones Synthetics Antimicrobials that are completely synthesized in a lab
10 Mechanisms of Antimicrobial Action Inhibition of Cell Wall Synthesis in Bacteria Beta-lactams are most prominent in this group Functional groups are beta-lactam rings Beta-lactams bind to enzymes that cross-link NAM subunits Bacteria have weakened cell walls and eventually lyse
11 How cell walls are made and maintained NAG-NAM polylinker NAM-NAM crosslink NAG NAM NAG-NAM chain Crossbridge between NAM and NAM Growth Divide New NAG and NAM subunits New NAG and NAM subunits are inserted into the wall by enzymes, allowing the cell to grow. Other enzymes link new NAM subunits to old NAM subunits with peptide cross-links. A bacterial cell wall is made of peptidoglycan, which is made of NAG-NAM chains that are cross-linked by peptide bridges between the NAM subunits.
12 Figure 10.3c-e Bacterial cell wall synthesis and the inhibitory effects of beta-lactams on it. Beta-lactams Functional groups are beta-lactam rings inhibit new NAM-NAM crosslinks Penicillin G (natural) Methicillin (semisynthetic) Penicillins Cephalothin (natural) Cephalosporins New cross-links inhibited by beta-lactam Previously formed cross-link Growth Beta-lactam interferes with the linking enzymes, and NAM subunits remain unattached to their neighbors. However, the cell continues to grow as it adds more NAG and NAM subunits. The cell bursts from osmotic pressure because the integrity of peptidoglycan is not maintained. Osmotic pressure causes water to move in and burst the cell
13 Beta-lactams (Penicillin) action Prevent bacteria from increasing amount of peptidoglycan Have no effect on existing peptidoglycan layer Effective only for growing cells Dormant cells are not affected Semisynthetic derivatives of beta-lactams More stable in acidic environments (methicillin) More readily absorbed in intestine Less susceptible to deactivation by bacteria More active against more types of bacteria
14 Other Inhibitors of synthesis of bacterial walls Vancomycin (MRSA) and cycloserine Interfere with particular Ala-Ala bridges that link NAM subunits in many Gram-positive bacteria Bacitracin Blocks transport of NAG and NAM from cytoplasm Isoniazid and ethambutol Disrupt mycolic acid formation in mycobacterial species Generation time is hrs so therapy extends for months or years
15 Lysozyme
16 Inhibitors of synthesis of fungal walls Fungal cells composed of various polysaccharides not found in mammalian cells (1,3 D-glucan) Echinocandins (caspofungin) inhibit the enzyme that synthesizes glucan
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18 Disruption of Fungal Cytoplasmic Membranes Polyenes - (Nystatins/Amphotericin B) attaches to ergosterol in fungal membranes form channel through cytoplasmic membrane and damage its integrity (Humans somewhat susceptible because cholesterol similar to ergosterol) Azoles (fluconazole) and allylamines (terbinafine) inhibit ergosterol synthesis
19 Disruption of Cytoplasmic Membranes in bacteria or protozoans Polymyxin disrupts cytoplasmic membranes of Gramnegative bacteria Pseudomonas Some parasitic worm drugs act against cytoplasmic membranes (praziquantel and ivermectin)
20 Mechanisms of Antimicrobial Action Inhibition of Protein Synthesis Prokaryotic ribosomes are 70S (30S and 50S) Eukaryotic ribosomes are 80S (40S and 60S) Mitochondria of animals and humans contain 70S ribosomes Side-effect (liver and bone marrow affected) WHY?
21 Figure 10.4 The mechanisms by which antimicrobials target prokaryotic ribosomes to inhibit protein synthesis. Incorrect amino acids 30S Ribosome Streptomycin/gentamicin mrna Some aminoglycosides cause change in 30S shape; mrna is misread. Lincosamides or macrolides bind to 50S subunit, blocking proper mrna movement through ribosome. Synthesis stops. erthromycin 50S mrna 50S Tetracycline and some aminoglycosides block docking site of trna. mrna 50S Antisense nucleic acid fomiversen mrna Amino acids Chloramphenicol binds enzymatic site 30S 50S mrna fmet 50S Mupirocin binds Isoleucine trna 30S Oxazolidinone mrna
22 Mechanisms of Antimicrobial Action Inhibition of Metabolic Pathways Antimetabolic agents can be effective when pathogen and host metabolic processes differ e.g. Inhibition of Nucleic Acid Synthesis Actinomycin D binds DNA block DNA replication or RNA transcription Not normally used to treat infections WHY? Atovaquone interferes with electron transport in protozoa and fungi
23 Antiviral agents Antiviral agents can target unique aspects of viral metabolism Amantadine (Influenza type A), rimantadine, and weak organic bases prevent viral uncoating Protease inhibitors interfere with an enzyme that HIV needs in its replication cycle
24 Inhibition of Viral Nucleic Acid Synthesis Nucleotide or nucleoside analogs Interfere with function of nucleic acids Most often used against viruses 1) viral DNA pol proof reading not as effective as human 2) rapid synthesis
25 Inhibition of Viral Nucleic Acid Synthesis Reverse transcriptase inhibitors Act against an enzyme HIV uses in its replication cycle NO SIDE-EFFECT Why?
26 Mechanisms of Antimicrobial Action Prevention of Virus Attachment Attachment antagonists (peptide/sugar analogs) block viral attachment or receptor proteins E.g. Arildone and pleconaril poliovirus and cold virus
27 Clinical Considerations in Prescribing Antimicrobial Drugs Ideal Antimicrobial Agent Readily available Inexpensive Chemically stable Easily administered Nontoxic and nonallergenic Selective toxicity
28 Clinical Considerations in Prescribing Antimicrobial Drugs Spectrum of Action/Effectiveness Routes of Administration/Safety and Side Effects Resistance to Antimicrobial Drugs The Development of Resistance in Populations Mechanisms of Resistance Multiple Resistance and Cross Resistance Retarding Resistance
29 Clinical Considerations in Prescribing Antimicrobial Drugs Spectrum of Action Number of different pathogens a drug acts against Narrow-spectrum effective against few organisms Broad-spectrum effective against many organisms Killing of normal flora reduces microbial antagonism Superinfection is a possibility
30 Figure 10.8 Spectrum of action for selected antimicrobial agents. Which are narrow/broad spectrum?
31 Clinical Considerations in Prescribing Antimicrobial Drugs Effectiveness Ascertained by 3 tests Diffusion susceptibility test (KIRBY BAUER) Minimum inhibitory concentration test (MIC) Minimum bactericidal concentration test (MBC) (for bacteriocidal/static check)
32 Figure 10.9 Zones of inhibition in a diffusion susceptibility (Kirby-Bauer) test. Bacterial lawn Zone of inhibition Kirby-Bauer test Diffusion susceptibility test
33 Figure Minimum inhibitory concentration (MIC) test in test tubes. Turbid tubes Clear tubes MIC smallest amount that inhibits growth Increasing concentration of drug
34 Figure A minimum bactericidal concentration (MBC) test. MBC Concentration of antibacterial drug ( g/ml) Clear MIC tube IC refinement test 8 g/ml 16 g/ml 25 g/ml Bacterial colonies No colonies No colonies Drug-free media
35 Figure An Etest, which combines aspects of Kirby-Bauer and MIC tests. Etest = MIC+Kirby-Bauer type
36 Clinical Considerations in Prescribing Antimicrobial Drugs Routes of Administration Topical/local application of drug for external infections Oral route requires no needles and is self-administered Intramuscular administration delivers drug via needle into muscle Intravenous administration delivers drug directly to bloodstream
37 Figure The effect of route of administration on blood levels of a chemotherapeutic agent.
38 Clinical Considerations in Prescribing Antimicrobial Drugs Safety and Side Effects - 3 Toxicity (1/3) Cause of many adverse reactions poorly understood Drugs may be toxic to kidneys, liver, or nerves Consideration needed when prescribing drugs to pregnant women Therapeutic index is the ratio of the dose of a drug that can be tolerated to the drug's effective dose (higher TI = safer) Therapeutic Range/Window (safe range of drug conc.)
39 Figure Some side effects resulting from toxicity of antimicrobial agents. Metronidazole antiprotozoan Black hairy tongue hemoglobin breakdown products Accumulate in tongue Tetracyline-Calcium complexes get incorporated in bones and teeth
40 Clinical Considerations in Prescribing Antimicrobial Drugs Safety and Side Effects Allergies (2/3) Allergic reactions are rare but may be life threatening Anaphylactic shock (0.1 % American penicillin 300 deaths/year) Disruption of normal microbiota (3/3) May result in secondary infections by opportunistic pathogens Overgrowth of normal flora causing superinfections Of greatest concern for hospitalized patients
41 Methods for Evaluating Disinfectants and Antiseptics Development of Resistant Microbes Little evidence that products containing antiseptic and disinfecting chemicals add to human or animal health Use of such products promotes development of resistant microbes
42 FDA bans antibacterial soaps - Sep2016 Rule removes triclosan and triclocarban from overthe-counter antibacterial hand and body washes The agency after some data suggested that longterm exposure to certain active ingredients used in antibacterial products for example, triclosan (liquid soaps) and triclocarban (bar soaps) could pose health risks, such as bacterial resistance or hormonal effects.
43 Figure The development of a resistant strain of bacteria. Drug-sensitive cells Drug-resistant mutant Exposure to drug Population of microbial cells Sensitive cells inhibited by exposure to drug Remaining Population grows over time Most cells now resistant
44 Resistance to Antimicrobial Drugs The Development of Resistance in Populations Some pathogens are naturally resistant Resistance by bacteria acquired in two ways New mutations of chromosomal genes Acquisition of R plasmids via transformation, transduction, and conjugation.
45 Resistance to Antimicrobial Drugs Mechanisms of Resistance At least seven mechanisms of microbial resistance Production of enzyme that destroys or deactivates drug Slow or prevent entry of drug into the cell Alter target of drug so it binds less effectively Alter their own metabolic chemistry Pump antimicrobial drug out of the cell before it can act Bacteria in biofilms can resist antimicrobials
46 Figure 10.3c-e Bacterial cell wall synthesis and the inhibitory effects of beta-lactams on it. Beta-lactams Functional groups are beta-lactam rings Penicillin G (natural) Penicillins Methicillin (semisynthetic) Cephalothin (natural) Cephalosporins New cross-links inhibited by beta-lactam Previously formed cross-link Growth Beta-lactam interferes with the linking enzymes, and NAM subunits remain unattached to their neighbors. However, the cell continues to grow as it adds more NAG and NAM subunits. The cell bursts from osmotic pressure because the integrity of peptidoglycan is not maintained. Osmotic pressure causes water to move in and burst the cell
47 Figure How -lactamase (penicillinase) renders penicillin inactive. 1) Production of enzyme that destroys or deactivates drug Lactam ring Penicillin -lactamase (penicillinase) breaks this bond Inactive penicillin 200 lactamases known many coded by R plasmids
48 Resistance to Antimicrobial Drugs Multiple Resistance and Cross Resistance Pathogen can acquire resistance to more than one drug Multi-drug-resistant pathogens are resistant to at least three antimicrobial agents Cross resistance resistance to one confers resistance to similar drugs e.g. streptomycin and other aminoglycoside-related drugs
49 Resistance to Antimicrobial Drugs 4 ways for Retarding Resistance 1) Maintain high concentration of drug in patient for sufficient time (Complete the dose) Inhibit the pathogen so immune system can eliminate 2) Use antimicrobial agents in combination Synergism penicillin and streptomycin
50 Figure An example of synergism between two antimicrobial agents. Disk with semisynthetic amoxicillin-clavulanic acid Disk with semisynthetic aztreonam
51 Resistance to Antimicrobial Drugs Retarding Resistance 3) Use antimicrobials only when necessary 4) Develop new variations of existing drugs
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