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.

12
Propolis

13
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. ..

14
Ehrlichs Magic Bullets
Gerhard Domagk - Prontosil
Fleming and Penicillin
Selman Waksman

18
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

16
Number of agents approved

14

12

Resistance
10

2
0
0
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

28
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

31
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 ( lactam ring)
Natural penicillins
Narrow range of action
Susceptible to penicillinase ( 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 + -lactamase inhibitors
Penicillinase ( 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

Inhibitsproteins 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.
70
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
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 over-
the-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 Prevent
Transmission Infection

Antimicrobial Infection
Resistance
Effective
Optimize Diagnosis
Use and Treatment

Antimicrobial Use
 12 Steps to Prevent
Antimicrobial Resistance:
Hospitalized Adults
12 Contain your contagion
11 Isolate the pathogen
10 Stop treatment when cured Prevent
9 Know when to say no to vanco
8 Treat infection, not colonization Transmission
7 Treat infection, not contamination Use Antimicrobials
6 Use local data
5 Practice antimicrobial control Wisely
4 Access the experts Diagnose and Treat
3 Target the pathogen
2 Get the catheters out Effectively
1 Vaccinate Prevent Infection
 Antimicrobial Resistance Among
Pathogens Causing Hospital-Acquired
Infections
Methicillin (oxacillin)-resistant Vancomycin-resistant
Staphylococcus aureus 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 -
157