ANTIBIOTICS

Antibiotics

Topics- Antimicrobial Therapy- Selective Toxicity - Survey of Antimicrobial Drugs- Microbial Drug Resistance- Drug and Host Interaction

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Key WordsKey WordsSterilization/disinfection/antisepsis

Antibiotic

Selective toxicity

Bactericidal

Bacteriostatic

Minimal inhibitory concentration (MIC)

Susceptibility testing

Penicillin binding proteins

Penicillinase/beta lactamas

Resistance

Selective Toxicity

Drugs that specifically target microbial processes, and not the human host cellular processes.

Improved Patient Outcomes Associated With Proper Hand Hygiene

Semmelweis

Chlorinated lime hand antisepsis

Antibiotics

Naturally occurring antimicrobials– Metabolic products of bacteria and

fungi– Reduce competition for nutrients and

space Bacteria that produce them:

– Streptomyces, Bacillus, Molds

– Penicillium, Cephalosporium

History

Ancient remedies

Ehrlich

Domagk

Fleming

Neem Plant

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Neem Plant

Uses: Arthritis, blood purifier and detoxifier, convalescence after fever, cough, diabetes, eczema, fever (used with black pepper and gentian), inflammation of muscles and joints, jaundice, leukorrhea, malaria, mucus membrane ulcerations, nausea, obesity, parasites, rheumatism, skin diseases/inflammations, cleanses liver, syphilis, thirst, tissue excess, tumors, vomiting, worms, drowsiness, loss of appetite. Leaves—heal ulcers in urinary passage, emmenagogue, skin diseases. Fruit—skin diseases, bronchitis. Kernel powder —washing hair. Effective as a pesticide.

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Propolis

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Propolis

Propolis is plant resin compound, different fabric compositions, wax, essential oils, iron, microelements – copper, zinc, manganese, cobalt, plus pollen, flavonoids, salivary gland secretions of bees. Propolis is used as a bio-stimulator which enhances endurance and eliminate fatigue. Because its antiviral properties, antitoxic and anti-inflammatory propolis finds more and more uses. Recovery is a good stimulator of affected tissue injuries, cuts... 

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Ehrlich’s Magic Bullets

Gerhard Domagk - Prontosil

Fleming and Penicillin

Selman Waksman

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Between 1962 and 2000, no major classes of antibiotics were introduced

Fischbach MA and Walsh CT Science 2009

A Changing Landscape forNumbers of Approved Antibacterial Agents

Bars represent number of new antimicrobial agents approved by the FDA during the period listed.

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6

8

10

12

14

16

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1983-87 1988-92 1993-97 1998-02 2003-05 2008

Infectious Diseases Society of America. Bad Bugs, No Drugs. July 2004; Spellberg B et al. Clin Infect Dis. 2004;38:1279-1286;New antimicrobial agents. Antimicrob Agents Chemother. 2006;50:1912

Resistan

ce

Azamulin

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Daptomycin chemical structure.

Steenbergen J N et al. J. Antimicrob. Chemother. 2005;55:283-288

JAC vol.55 no.3 © The British Society for Antimicrobial Chemotherapy 2005; all rights reserved

Daptomycin mechanism of action.

Steenbergen J N et al. J. Antimicrob. Chemother. 2005;55:283-288

JAC vol.55 no.3 © The British Society for Antimicrobial Chemotherapy 2005; all rights reserved

Linezolid

Ideal Antimicrobial Attributes

Solubility

Selective toxicity

Stable toxicity level

Allergenicity

Tissue stability

Resistance Acquisition

Shelf Life

Cost

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ANTIBIOTICSANTIBIOTICS

• Selectively toxic for bacteriaSelectively toxic for bacteria– bactericidal (killing) bactericidal (killing) – bacteriostatic (growth inhibition)bacteriostatic (growth inhibition)

no harm to patientno harm to patient

Antibiotic/Antimicrobial

Antibiotic: Chemical produced by a microorganism that kills or inhibits the growth of another microorganism

Antimicrobial agent: Chemical that kills or inhibits the growth of microorganisms

Microbial Sources of Antibiotics

Administration of Antibiotics

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Spectrum of Activity

Determining Microbial Sensitivities

Disk Diffusion Method

Dilution Method

Serum Killing Power

Automated Methods

Drug Mechanisms of Action

Penicillin (over 50 compounds)– Share 4-sided ring ( lactam ring)

Natural penicillins• Narrow range of action• Susceptible to penicillinase ( lactamase)

Antibacterial Antibiotics Inhibitors of Cell Wall Synthesis

Prokaryotic Cell Walls

Cell wall synthesis

Bactericidal– Penicillin and cephalosporins – binds

and blocks peptidases involved in cross-linking the glycan molecules

– Vancomycin – hinders peptidoglycan elongation

– Cycloserine – inhibits the formation of the basic peptidoglycan subunits

Antibiotics weaken the cell wall, and cause the cell to lyse.

Penicillin Penicillin chrysogenum A diverse group (1st, 2nd , 3rd generations)

– Natural (penicillin G and V)– Semisynthetic (Ampicillin, Carbenicillin)

Structure– Thiazolidine ring– Beta-lactam ring– Variable side chain (R group)

-4C – 28C - 38C water activity 0.98

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The R group is responsible for the activity of the drug, and cleavage of the beta-lactam ring will render the drug inactive.

Chemical structure of penicillins

Penicillins

Figure 20.6

Penicilinase-resistant penicillins• Carbapenems: very broad spectrum• Monobactam: Gram negative

Extended-spectrum penicillins Penicillins + -lactamase inhibitors

Semisynthetic Penicillins

Penicillinase ( Lactamase)

Cephalosporins– 2nd, 3rd, and 4th

generations more effective against gram-negatives

Other Inhibitors of Cell Wall Synthesis

Figure 20.9

Cephalosporin Cephalosporium acremonium (mold) Widely administered today

– Diverse group (natural and semisynthetic) Structure

– similar to penicillin except

• Main ring is different

• Two sites for R groups

The different R groups allow for versatility and improved effectiveness.

Other Inhibitors of Cell Wall Synthesis

Mycobacteria: interfere with mycolic acid synthesis or incorporation

– Isoniazid (INH)

– Ethambutol

Polypeptide antibiotics– Bacitracin

• Topical application• Against gram-positives

– Vancomycin• Glycopeptide• Important "last line" against antibiotic resistant S.

aureus

Other Inhibitors of Cell Wall Synthesis

Broad spectrum, toxicity problems Examples

– Aminoglycosides: Streptomycin, neomycin, gentamycin

– Tetracyclines

– Macrolides: Erythromycin

– Chloramphenicol

Inhibitors of Protein Synthesis

Aminoglycosides From Streptomyces Inhibit protein synthesis

Streptomyces synthesizes many different antibiotics such as aminoglycosides, tetracycline, chloramphenicol, and erythromycin.

Sites of inhibition on the procaryotic ribosome

Tetracycline

Inhibits proteins synthesis Broad spectrum and low cost Commonly used to treat sexually

transmitted diseases Minor side effect – gastrointestinal

disruption

Tetracyclines (bacteriostatic)tetracycline, minocycline and doxycycline

Mode of action - The tetracyclines reversibly bind to the 30S ribosome and inhibit binding of aminoacyl-t-RNA to the acceptor site on the 70S ribosome.

Spectrum of activity - Broad spectrum; Useful against intracellular bacteria

Resistance - Common

Adverse effects - Destruction of normal intestinal flora resulting in increased secondary infections; staining and impairment of the structure of bone and teeth. Not used in children.

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Spectinomycin (bacteriostatic)

Mode of action - Spectinomycin reversibly interferes with m-RNA interaction with the 30S ribosome. It is structurally similar to the aminoglycosides but does not cause misreading of mRNA. Does not destabilize membranes, and is therefore bacteriostatic

Spectrum of activity - Used in the treatment of penicillin-resistant Neisseria gonorrhoeae

Resistance - Rare in Neisseria gonorrhoeae

Erythromycin

Inhibits protein synthesis Broad-spectrum Commonly used as prophylactic drug

prior to surgery Side effects - low toxicity

Streptomycin - treat Plague

Chloramphenicol

Broad-spectrum Treat typhoid fever, brain abscesses Rarely used now due to side effects –

aplastic anemia

Chloramphenicol

UDP-glucuronyl transferase

Aminoglycoside

Polymyxin B (Gram negatives)– Topical– Combined with bacitracin and neomycin (broad

spectrum) in over-the-counter preparation

Injury to the Plasma Membrane

Polymyxin B (Gram negatives)

– Topical

– Combined with bacitracin and neomycin (broad spectrum) in over-the-counter preparation

Injury to the Plasma Membrane

Rifamycin

– Inhibits RNA synthesis

– Antituberculosis Quinolones and fluoroquinolones

– Ciprofloxacin

– Inhibits DNA gyrase

– Urinary tract infections

Inhibitors of Nucleic Acid Synthesis

Inhibition of Nucleic Acid Synthesis

Rifampin binds to DNA-dependent RNA polymerase and inhibits intiation of RNA synthesis

Antibacterials — Antimetabolites

Sulfonamides

Isoniazid

Ethambutol

Nitrofurans

Folic acid synthesis

Sulfonamides (sulfa drug) and trimethoprim

– Analogs

– Competitive inhibition of enzymes

– Prevents the metabolism of DNA, RNA, and amino acid

– Sulfonamides (Sulfa drugs)• Inhibit folic acid synthesis• Broad spectrum

Competitive Inhibitors

Figure 5.7

Sulfonamides compete with PABA for the active site on the enzyme.

The sulfonamide Sulfamethoxazole is commonly used in combination with trimethoprim

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Necrotizing Fasciitis

Group A hemolytic streptococci and Staphylococcus aureus, alone or in synergism, are frequently the initiating infecting bacteria. However, other aerobic and anaerobic pathogens may be present, including Bacteroides, Clostridium, Peptostreptococcus, Enterobacteriaceae, coliforms, Proteus, Pseudomonas, and Klebsiella.

Summary of Targets

Figure 20.20

Antibiotic Resistance

Antimicrobial Resistance

Relative or complete lack of effect of antimicrobial against a previously susceptible microbe

Increase in MIC

• Enzymatic destruction of drug

• Prevention of penetration of drug

• Alteration of drug's target site

• Rapid ejection of the drug

Mechanisms of Antibiotic Resistance

Antibiotic Selection for Resistant Bacteria

What Factors Promote Antimicrobial Resistance?

Exposure to sub-optimal levels of antimicrobial

Exposure to microbes carrying resistance genes

Inappropriate Antimicrobial Use

Prescription not taken correctlyAntibiotics for viral infectionsAntibiotics sold without medical

supervisionSpread of resistant microbes in

hospitals due to lack of hygiene

Inappropriate Antimicrobial Use

Lack of quality control in manufacture or outdated antimicrobial

Inadequate surveillance or defective susceptibility assays

Poverty or war Use of antibiotics in foods

Antibiotics in Foods

Antibiotics are used in animal feeds and sprayed on plants to prevent infection and promote growth

Multi drug-resistant Salmonella typhi has been found in 4 states in 18 people who ate beef fed antibiotics

Consequences of Antimicrobial Resistance

Infections resistant to available antibiotics

Increased cost of treatment

Multi-Drug Resistant TB

MRSA “mer-sah”

Methicillin-Resistant Staphylococcus aureus

Most frequent nosocomial (hospital-acquired) pathogen

Usually resistant to several other antibiotics

Proposals to Combat Antimicrobial Resistance

Speed development of new antibiotics

Track resistance data nationwide Restrict antimicrobial use Direct observed dosing (TB)

Proposals to Combat Antimicrobial Resistance

Use more narrow spectrum antibiotics

Use antimicrobial cocktails

Antimicrobial peptides

– Broad spectrum antibiotics from plants and animals

• Squalamine (sharks)

• Protegrin (pigs)

• Magainin (frogs)

The Future of Chemotherapeutic Agents

Side Effects

Resistance to Drugs

Chromosomal

Plasmid borne

Mechanisms of Drug Resistance

Mutations in Target molecules

Alterations in membrane permeability

Enzyme development

Mechanisms of Drug Resistance

Enzyme Activity Changes

Alterations in Anabolic Pathways

Generations of Drugs

First/Second/Third Line Drugs

Cross Resistance

Limiting Drug Resistance

Effective Drug Concentrations

Simultaneous Drug Administration

• Synergism

• Antagonism Restricting Drug Prescriptions

Antibiotic Resistance

Inactivation of the antibiotic by a microbial enzyme

Prevention of the antibiotic from reaching its target cell structure

Alteration of the target cell structure so that it is no longer affected by the antibiotic

Mechanisms of Resistance

Failure of the antibiotic to penetrate the outer membrane

Failure to bind to the target site (penicillin binding protein)

Hydrolysis of the antibiotic by beta lactamases

Drug Resistance

Intrinsic as well as acquired Intrinsic drug resistance exists naturally

and is not acquired through specific genetic changes

How does drug resistance develop?

The genetic events most often responsible for drug resistance are either chromosomal mutations or transfer of extrachromosomal DNA from a resistant species to a sensitive one.

Resistance Factors – R Factors

Transferred through conjugation, transformation or transduction

Many bacteria also maintain transposable drug resistance sequences – tansposons that are duplicated and inserted from one plasmid to another or from a plasmid to a chromosome

Conjugation – plasmids and chromosomal elements, conjugative transposons plasmids

Conjugative plasmid – plasmids that transfer themselves by conjugation must carry a number of genes encoding proteins needed for the conjugation process itself (tra genes)

Self-transmissable plasmids (STP) are usually at least 25kb

Mobilize plasmids – much smaller than STP because they need only 1 or 2 genes (mob genes)

Resistance Genes

Acquire sequential transposon insertions Integrons are probably responsible for

evolution of many of the plasmids that carry multiple resistance genes

Integrons

Integrons like transposons are linear DNA segments that insert into DNA

Unlike transposons, integrons integrate at a single site and do not encode a transposase

Conjugative transposons – located in the bacterial chromosome, also integrate into plasmids

Mechanism of Transfer

Excise themselves from the donor genome to form a covalently closed circle that does not replicate

The circular intermediate transfers similarly to a plasmid

In the recipient, the circular intermediate integrates in the chromosome by a mechanism that does not duplicate the target site

Origin of Antibiotic Resistant Genes

First it was assumed that antibiotic resistance genes appeared only after antibiotics began to be widely used in medicine

The genetic diversity within some classes of resistance makes it clear that these genes have been evolving for a much longer time

Origin of resistance

Hypothesis: resistance genes first evolved in the antibiotic-producing bacteria such as Streptomyces spp. As a mechanism for protecting them from the antibiotics they produce.

Genes for antibiotic production are frequently found in the same gene clusters with genes encoding resistance proteins

Specific mechanisms of drug resistance

Bacteria lose its sensitivity to a drug by expressing genes that stop the action of the drug.

Gene expression

Synthesis of enzymes that inactivate the drug

Decrease in cell permeability and uptake of the drug

Change in the number or affinity of the drug receptor sites, or

Modification of an essential metabolic pathway

Resistance

Some bacteria can become resistant indirectly by lapsing into dormancy, or, in the case of penicillin, by converting to a cell-wall-deficient form (L form) that penicillin cannot affect.

Drug Inactivation Mechanisms

Produce enzymes that permanently alter drug structure (beta lactamases)

Decreased Drug Permeability or Increased Drug Transport

Prevent drug from entering the cell and acting on the target

Gram negative – natural blockade for some of the penicillin drugs

Resistance to the tetracyclines can arise fro plasmid-encoded proteins that pump the drug out of the cell

Resistance

Resistance to the aminoglycoside antibiotics is a specific case in which microbial cells have lost the capacity to transport the drug intracellulary

Multidrug Resistant (MDR) Pumps

Actively transport drugs and other chemicals out of the cell

These pumps are proteins encoded by plasmids and chromosomes

They are located in the cell membrane and expel molecules by a protonmotive force similar to ATP synthesis

PUMPS

Because they lack selectivity, one type of pump can expel a broad array of antimicrobic drugs, detergents and other toxic substances.

Change of Drug Receptors

Alter nature of target site On bacteria resistant to rifampin and

streptomycin, the structure of key proteins has been altered so that these antibiotics can no longer bind.

Changes in Metabolic Patterns

Sulfonamide and trimethoprim resistance develops when microbes deviate from the usual patterns of folic acid synthesis

Natural selection and drug resistance

When a population of bacteria is exposed to a drug, sensitive cells are inhibited or destroyed and resistant forms survive and proliferate

In ecological terms, the environmental factor has put selection pressure on the population, allowing the more fit microbe to survive, and the population has evolved to a condition of drug resistance.

Antimicrobial Resistance: Key Prevention Strategies

Optimize Use

PreventTransmission

PreventInfection

EffectiveDiagnosisand Treatment

Antimicrobial-Resistant Pathogen

Antimicrobial Resistance

Antimicrobial Use

Infection

Susceptible Pathogen

12 Steps to Prevent Antimicrobial Resistance: Hospitalized Adults

12 Contain your contagion 11 Isolate the pathogen

10 Stop treatment when cured 9 Know when to say “no” to vanco 8 Treat infection, not colonization 7 Treat infection, not contamination

6 Use local data 5 Practice antimicrobial control

4 Access the experts3 Target the pathogen

2 Get the catheters out 1 Vaccinate

Prevent TransmissionUse Antimicrobials WiselyDiagnose and Treat EffectivelyPrevent Infection

Methicillin (oxacillin)-resistantStaphylococcus aureus

Vancomycin-resistantenterococci

Non-Intensive Care Unit Patients

Intensive Care Unit Patients

Antimicrobial Resistance Among Pathogens Causing Hospital-Acquired Infections

Source: National Nosocomial Infections Surveillance (NNIS) System

Prevalence of Isolates of Multidrug-Resistant Gram Negative Rods Recovered Within The First 48 h After

Admission to the Hospital

Pop-Vicas and D'Agata CID 2005;40:1792-8.

Conjugative transposons

Responsible for at least as much resistance gene transfer as plasmids, especially among G+, and they have a broad host range

G+ G+ ; G - G - ; G+ G -

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