Chapter 12Chapter 12

Drugs, Microbes, Host – The Drugs, Microbes, Host – The Elements of ChemotherapyElements of Chemotherapy

IntroductionIntroduction

TerminologyTerminologyBacteriostatic (inhibits growth) vs. Bactericidal (kills Bacteriostatic (inhibits growth) vs. Bactericidal (kills organisms)organisms)Broad spectrum vs. Narrow spectrumBroad spectrum vs. Narrow spectrumGeneric (chemical composition of drug) vs. Trade name Generic (chemical composition of drug) vs. Trade name (name given to a drug by a manufacture(name given to a drug by a manufactureSources of antimicrobial agentsSources of antimicrobial agentsAntibiotics - growth products of organismsAntibiotics - growth products of organismsChemosynthetic agents - synthesized in a laboratoryChemosynthetic agents - synthesized in a laboratorySemi-synthetic agents - growth products of organisms Semi-synthetic agents - growth products of organisms that have been chemically altered in laboratorythat have been chemically altered in laboratory

Colony of Colony of StreptomycesStreptomyces, one of nature’s , one of nature’s most prolific antibiotic producersmost prolific antibiotic producers

Major targets of drugs acting on Major targets of drugs acting on bacterial cellsbacterial cells

Mode of action of antimicrobial agents: Mode of action of antimicrobial agents: competitive inhibition competitive inhibition

Antibacterial drugsAntibacterial drugsInvolves inhibiting Involves inhibiting synthesis of a critical synthesis of a critical metabolitemetaboliteBacteriostatic in activityBacteriostatic in activityExamplesExamples

Sulfa - competition based Sulfa - competition based on chemical similarity on chemical similarity between sulfas and PABA between sulfas and PABA which is needed for folic which is needed for folic acid synthesis; folic acid acid synthesis; folic acid critical in protein and NA critical in protein and NA synthesis; side effect = synthesis; side effect = possible kidney damage possible kidney damage from crystalsfrom crystals

Mode of action of antimicrobial agents: Mode of action of antimicrobial agents: inhibition of cell wall synthesisinhibition of cell wall synthesis

Inhibition of cell wall Inhibition of cell wall synthesissynthesisInterfere with one of Interfere with one of multiple steps in CW multiple steps in CW synthesissynthesisGreater effect Greater effect demonstrated against demonstrated against G(+) than G(-) – most G(+) than G(-) – most act against synthesis of act against synthesis of peptidoglycan; many peptidoglycan; many drugs cannot penetrate drugs cannot penetrate LPSLPSGenerally considered to Generally considered to be bactericidal - cause be bactericidal - cause lysislysis

Mode of action of antimicrobial agents: Mode of action of antimicrobial agents: inhibition of cell wall synthesisinhibition of cell wall synthesis

Drugs with beta lactam ring - prevent Drugs with beta lactam ring - prevent cross-linking in last step of synthesiscross-linking in last step of synthesis

PenicillinsPenicillins Original penicillins effective only Original penicillins effective only

against G(+) - esp. staph & strepagainst G(+) - esp. staph & strep Semisynthetic penicillins Semisynthetic penicillins

developed to overcome developed to overcome penicillinase (beta lactamase) penicillinase (beta lactamase) producing strains (i.e.methicillin, producing strains (i.e.methicillin, oxacillin & nafcillin) and broaden oxacillin & nafcillin) and broaden spectrum to G(-) (i.e. ampicillin & spectrum to G(-) (i.e. ampicillin & carbenicillin)carbenicillin)

Toxicity involves hypersensitivityToxicity involves hypersensitivity

Mode of action of antimicrobial agents: Mode of action of antimicrobial agents: inhibition of cell wall synthesisinhibition of cell wall synthesis

CephalosporinsCephalosporins Broad spectrum antibioticsBroad spectrum antibiotics Used when individuals are Used when individuals are

allergic to penicillinallergic to penicillin Modifications to original drug Modifications to original drug

increased spectrum increased spectrum (generations 1, 2, and 3)(generations 1, 2, and 3)

MoxalactamsMoxalactamsOther drugs - interfere at other Other drugs - interfere at other stepssteps

CycloserineCycloserine Bacitracin - used primarily as Bacitracin - used primarily as

topical (on the skin - not taken topical (on the skin - not taken internally because of toxicity)internally because of toxicity)

VancomycinVancomycin

Mode of action of antimicrobial agents: Mode of action of antimicrobial agents: Inhibition of protein synthesis Inhibition of protein synthesis

Inhibit one of many steps in protein synthesisInhibit one of many steps in protein synthesisExamplesExamples

Aminoglycosides - includes streptomycin, Aminoglycosides - includes streptomycin, kanamycin, tobramycin, gentamicin, kanamycin, tobramycin, gentamicin, neomycin, and amikacin; causes neomycin, and amikacin; causes misreading of mRNA; bactericidal; misreading of mRNA; bactericidal; synergistic with penicillinssynergistic with penicillins

Tetracyclines - block binding to tRNA; Tetracyclines - block binding to tRNA; bacteriostatic; broad spectrumbacteriostatic; broad spectrum

Chloramphenicol - prevent peptide bond Chloramphenicol - prevent peptide bond formation; bacteriostatic; excellent formation; bacteriostatic; excellent penetration of CNS (useful for treating penetration of CNS (useful for treating meningitis); side effect = aplastic anemiameningitis); side effect = aplastic anemia

Erythromycin - antimicrobial spectrum Erythromycin - antimicrobial spectrum similar to penicillin; used especially in similar to penicillin; used especially in penicillin-allergicpenicillin-allergic

Lincomycin and clindamycin - Lincomycin and clindamycin - bacteriostatic; clindamycin useful for bacteriostatic; clindamycin useful for anaerobes; may cause anaerobes; may cause pseudomembranous colitispseudomembranous colitis

Structure of aminoglycoside, tetracycline, Structure of aminoglycoside, tetracycline, chloramphenicol, and erythromycinchloramphenicol, and erythromycin

Effects of drugs on bacterial cell Effects of drugs on bacterial cell membranemembrane

Act directly on cell Act directly on cell membranes (do not need membranes (do not need to enter cell to cause to enter cell to cause damage)damage)

Bactericidal - produce Bactericidal - produce irreversible damage to irreversible damage to membrane permeabilitymembrane permeability

Example = polymyxins Example = polymyxins and colistinand colistin

Highly toxic to kidneys Highly toxic to kidneys and nervesand nerves

Antifungal drugs Antifungal drugs Damage cell membranesDamage cell membranes

Bind or interfere with ergosterol Bind or interfere with ergosterol (unique fungal sterol in (unique fungal sterol in membrane)membrane)

ExamplesExamplesPolyenes - nystatin and Polyenes - nystatin and amphotericin Bamphotericin BImidazoles - clotrimazole, Imidazoles - clotrimazole, miconazole (topical) and miconazole (topical) and ketoconazole (systemic)ketoconazole (systemic)

Inhibition of nucleic acid Inhibition of nucleic acid synthesissynthesis

Griseofulvin - interferes with Griseofulvin - interferes with mitosis; selectively binds to mitosis; selectively binds to keratin in skin, hair & nails; keratin in skin, hair & nails; used primarily with fungi used primarily with fungi classified as dermatophyticclassified as dermatophytic

5-fluorocytosine5-fluorocytosine

Antiviral drugs Antiviral drugs

Interference with uptake or uncoating of virus - Interference with uptake or uncoating of virus - amantadine (used to prevent influenza A)amantadine (used to prevent influenza A)

Inhibition of nucleic acid synthesisInhibition of nucleic acid synthesis

Ribavirin - effective in vitro against a wide range Ribavirin - effective in vitro against a wide range of viruses; highly toxicof viruses; highly toxic

Acyclovir - Herpes-specific (genital herpes, cold Acyclovir - Herpes-specific (genital herpes, cold sores, chickenpox)sores, chickenpox)

Azidothymidine (AZT) - treatment of HIV Azidothymidine (AZT) - treatment of HIV infectionsinfections

Antibiotic resistance in bacteria Antibiotic resistance in bacteria

Sites of resistanceSites of resistance

Membrane transport - Membrane transport - LPS layer of G(-) LPS layer of G(-) prevent entry of many prevent entry of many drugsdrugs

Targets of Targets of antimicrobial agents antimicrobial agents (e.g. ribosomes)(e.g. ribosomes)

Presence of antibiotic-Presence of antibiotic-destroying enzymes destroying enzymes (e.g. beta lactamase)(e.g. beta lactamase)

Antibiotic resistance in bacteriaAntibiotic resistance in bacteriaMechanisms of changing Mechanisms of changing resistanceresistanceMutations - permanent Mutations - permanent changes in chromosomes; changes in chromosomes; not caused by antibioticsnot caused by antibioticsAcquisition of new genetic Acquisition of new genetic information - methodsinformation - methodsTransformation = naked Transformation = naked DNADNATransduction = via virusesTransduction = via virusesConjugation = via sex pili Conjugation = via sex pili (sexual recombination) – (sexual recombination) – most rapid methodmost rapid methodSelective pressures of Selective pressures of antimicrobial therapy - use antimicrobial therapy - use of antibiotics select for of antibiotics select for bacteria that are resistant - bacteria that are resistant - sensitive bacteria are sensitive bacteria are destroyeddestroyed

Antibiotic susceptibility testing Antibiotic susceptibility testing

Susceptibility no longer predictableSusceptibility no longer predictableVariables affecting outcome of therapyVariables affecting outcome of therapyCondition of host (immune status - Condition of host (immune status - underlying diseases)underlying diseases)Site of infection (can drugs get to site?)Site of infection (can drugs get to site?)Properties of antimicrobial agentProperties of antimicrobial agentOther drugs taken concurrentlyOther drugs taken concurrentlySusceptibility of organism to drugSusceptibility of organism to drug

The role of antimicrobials in disrupting The role of antimicrobials in disrupting microbial flora and causing superinfectionsmicrobial flora and causing superinfections

Broth dilution methods (Minimal Inhibitory Broth dilution methods (Minimal Inhibitory Concentration) – Quantitative MethodConcentration) – Quantitative Method

ProcedureProcedure Uses decreasing concentrations of antimicrobial agents prepared Uses decreasing concentrations of antimicrobial agents prepared

in 2-fold dilutions of broth that will support growth of test organismin 2-fold dilutions of broth that will support growth of test organism Standard inoculum is added to broth containing dilutions of Standard inoculum is added to broth containing dilutions of

antimicrobial agent, incubated overnight and then examined for antimicrobial agent, incubated overnight and then examined for growth (turbidity)growth (turbidity)

Lowest concentration of agent that inhibits growth as detected by Lowest concentration of agent that inhibits growth as detected by lack of visible turbidity = Minimal Inhibitory Concentration (MIC)lack of visible turbidity = Minimal Inhibitory Concentration (MIC)

Susceptibility and resistance determined by break point of Susceptibility and resistance determined by break point of drug (highest conc. of drug in the blood that can be drug (highest conc. of drug in the blood that can be achieved with maximal therapy); if MIC is lower than achieved with maximal therapy); if MIC is lower than breakpoint, organism = susceptible; if MIC is higher than breakpoint, organism = susceptible; if MIC is higher than breakpoint, organism = resistant breakpoint, organism = resistant

Minimum Inhibitory ConcentrationMinimum Inhibitory Concentration

Disk diffusion (Kirby/Bauer) - QualitativeDisk diffusion (Kirby/Bauer) - Qualitative

Advantage - rapid testing of several antibiotics Advantage - rapid testing of several antibiotics simultaneouslysimultaneouslyUses antimicrobial agents incorporated into filter paper Uses antimicrobial agents incorporated into filter paper disks placed on agar media causing drug to diffuse disks placed on agar media causing drug to diffuse creating a concentration gradient (conc. highest closest creating a concentration gradient (conc. highest closest to disk)to disk)Susceptibility/resistance determined by measuring Susceptibility/resistance determined by measuring diameter of zone of inhibition around disk and comparing diameter of zone of inhibition around disk and comparing to established zones for each antibioticto established zones for each antibioticStandardization - Bauer, Kirby, Sherris & Turck Standardization - Bauer, Kirby, Sherris & Turck (correlated with MIC’s using large numbers strains & (correlated with MIC’s using large numbers strains & regression analysis)regression analysis)

Techniques for preparation and Techniques for preparation and interpretation of disc diffusion testsinterpretation of disc diffusion tests

Antimicrobial gradient strip method (E Antimicrobial gradient strip method (E test)test)

Disk diffusion method that Disk diffusion method that allows determination of MIC in allows determination of MIC in agaragarConsists of plastic strip Consists of plastic strip containing gradient of containing gradient of antimicrobial agent along with antimicrobial agent along with an interpretive scalean interpretive scalePerformed similar to disk Performed similar to disk diffusiondiffusionOrganisms grow in elliptical Organisms grow in elliptical zone of inhibition around strip zone of inhibition around strip relative to concentration of relative to concentration of antibiotic along its lengthantibiotic along its lengthMIC determined by reading MIC determined by reading scale at point where zone of scale at point where zone of inhibition intersects stripinhibition intersects strip