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ANTIBIOTICS

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

Key Words
Sterilization/disinfection/antisepsis
Antibiotic
Selective toxicity
Bactericidal
Bacteriostatic

Minimal inhibitory concentration (MIC)


Susceptibility testing
Penicillin binding proteins

Penicillinase/beta lactamas
Resistance
3

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

11

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. Leavesheal ulcers in urinary passage,
emmenagogue, skin diseases. Fruitskin 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 biostimulator 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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Ehrlichs 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 for


Numbers of Approved Antibacterial Agents
18

14
12

Resistance

Number of agents approved

16

10
8
6
4
2

1983-87

1988-92

1993-97

1998-02

2003-05

2008

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

Azamulin

21

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

Tissue stability

Selective toxicity

Resistance
Acquisition

Stable toxicity
level

Shelf Life

Allergenicity

Cost

ANTIBIOTICS
Selectively toxic for bacteria
bactericidal (killing)
bacteriostatic (growth inhibition)
no harm to patient

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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

Antibacterial Antibiotics
Inhibitors of Cell Wall Synthesis

Penicillin (over 50 compounds)


Share 4-sided ring (b lactam ring)

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

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

44

Chemical structure of penicillins


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

Penicillins

Figure 20.6

Semisynthetic Penicillins

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

Extended-spectrum penicillins
Penicillins + b-lactamase inhibitors

Penicillinase (b Lactamase)

Other Inhibitors of Cell Wall


Synthesis

Cephalosporins
2nd, 3rd, and 4th
generations more
effective against
gram-negatives

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

Other Inhibitors of Cell Wall


Synthesis

Polypeptide antibiotics
Bacitracin
Topical application
Against gram-positives
Vancomycin
Glycopeptide
Important "last line" against antibiotic resistant S.
aureus

Inhibitors of Protein Synthesis

Broad spectrum, toxicity problems


Examples

Aminoglycosides: Streptomycin,
neomycin, gentamycin
Tetracyclines
Macrolides: Erythromycin
Chloramphenicol

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 penicillinresistant 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

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

Polymyxin B (Gram negatives)


Topical
Combined with bacitracin and
neomycin (broad spectrum) in overthe-counter preparation

Inhibitors of Nucleic Acid


Synthesis
Rifamycin
Inhibits RNA synthesis
Antituberculosis
Quinolones and fluoroquinolones
Ciprofloxacin
Inhibits DNA gyrase
Urinary tract infections

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

Competitive Inhibitors
Sulfonamides (Sulfa drugs)
Inhibit folic acid synthesis
Broad spectrum

Figure 5.7

Sulfonamides compete with PABA for the


active site on the enzyme.

The sulfonamide Sulfamethoxazole is


commonly used in combination with
trimethoprim

98

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

Antibiotic Resistance

Figure 20.20

Antimicrobial Resistance
Relative

or complete lack of
effect of antimicrobial against
a previously susceptible
microbe
Increase in MIC

Mechanisms of Antibiotic
Resistance
Enzymatic destruction of drug
Prevention of penetration of drug
Alteration of drug's target site
Rapid ejection of the drug

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 correctly


Antibiotics for viral infections
Antibiotics sold without medical
supervision
Spread 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

The Future of Chemotherapeutic


Agents

Antimicrobial peptides
Broad spectrum antibiotics from
plants and animals
Squalamine (sharks)
Protegrin (pigs)
Magainin (frogs)

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

Susceptible Pathogen
Antimicrobial-Resistant Pathogen

Prevent
Transmission

Prevent
Infection

Infection

Antimicrobial
Resistance

Effective
Diagnosis
and Treatment

Optimize
Use
Antimicrobial Use

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 experts
3 Target the pathogen
2 Get the catheters out
1 Vaccinate

Prevent
Transmission
Use Antimicrobials
Wisely
Diagnose and Treat
Effectively
Prevent Infection

Antimicrobial Resistance Among


Pathogens Causing Hospital-Acquired
Infections
Methicillin (oxacillin)-resistant
Staphylococcus aureus

Vancomycin-resistant
enterococci

Non-Intensive Care Unit Patients


Intensive Care Unit Patients
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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