Transcript Reduction in cellular permeability to the antibiotic

 In the clinical context an organism is said to be resistant:
 If it is not killed or inhibited by drug concentrations
readily attainable in the patient; this usually means
blood and tissue concentrations.
 However, an organism resistant to these may of course
be sensitive to the higher concentrations attainable in
urine or by topical application.
 Even the broadest of broad-spectrum antibacterial drugs
is ineffective against some bacterial genera, against
some species of other genera, and usually against some
strains of species that are in general sensitive to it.
Resistance
Inherent
(non specific)
Acquired
Inherent (non specific) Resistance
 Certain bacteria are, and as far as we know always have
been, more or less resistant to some antibiotics.
 For
example,
gram-negative
bacteria,
especially
Ps. aeruginosa, are inherently resistant to a number of
antibiotics that are very effective against gram-positive
bacteria such as penicillin G, erythromycin, lincomycin…….
Acquired Resistance
 When a bacterial population adapts to the presence of
an antibiotic, sensitive cells are gradually replaced by
resistant cells as in the presence of antibiotic.
 Resistant cells continue to grow at the expense of
sensitive cells.
 When a new antibiotic is introduced into clinical practice
for the treatment of infections caused by bacteria that
are not inherently resistant to the drug, the majority of
infections respond to the new drug.
 But following months or years of continuous use,
resistant strains are reported.
 The degree of resistance and the speed with which it
develops varies with:
 The organism &  The drug.
 Generally, the development of acquired bacterial
resistance is common and must be usually expected
with some exceptions.
 Streptococcus pyogenes has remained sensitive to
Penicillin G after 40 year's exposure to the drug.
 Staph aureus develops slow or multisteps resistance to
penicillin, chloramphenicol and tetracycline.
 While
Mycobacterium
tuberculosis
and
various
organisms develops sudden or one step resistance to
Streptomycin.
Biochemical Mechanisms of Resistance

Production of druginactivating enzymes

Change in the
antibiotic target site

. Reduction in cellular
permeability to the
antibiotic:

Switch to alternative
metabolic pathways
unaffected by the
drug:

Increased production
of essential
metabolite
Production of drug-inactivating

enzymes
Inactivation of
Inactivation of
Aminoglycosides
ß-lactams
Inactivation of
Chloramphenicol
Inactivation of Aminoglycosides
 Inactivation of aminoglycosides by plasmid controlled
adenylating , phosphorylating or acetylating intracellular
enzymes of drug resistant gram negative bacteria.
 Although the inactivating enzymes vary considerably in
their substrate and specificities, known modifications
are restricted to acetylation of amino groups and
adenylylation or phosphorylation of hydroxyl groups.
Inactivation of ß-lactams
 Inactivation
of β-lactams by β -lactamases
( penicillinase, cephalosporinase ) into penicilloic or
cephalosporoic acid.
 β -lactamases are:
Gram-positive
Gram-negative
Chromosomally
Plasmid
Constitutive
Inducible
Penicilloic acid
 The synthesis of gram positive β-lactamase is induced by
the antibiotic themselves and is released extracellularly
and destroy antibiotic in the external environment.
 Most strains of gram-negative cells, by contrast,
synthesize β-lactamases constitutively. i.e. continuously
and are not released into the external environment
(cell-bound or intracellular).

Chromosomally-mediaied
β-lactamases
of
gram
negative hydrolyze cephalosporins more rapidly than
penicillins and are inhibited by cloxacillin but not by
clavulanic acid.
 However, those of Aeromonas spp. and Klebsiella
spp. are more active against penicillins and not
inhibited by cloxacillin.
1
2
R1= cl , R2 = H (Cloxacillin)
 Inducible types are found in microorganisms such as
Pseudomonas spp., Proteus and other gram negative
bacteria but infrequently in E. coli in which, as many
enterobacter species, constitutive types could be
isolated.
 Different types of plasmid-mediated β-lactamases were
isolated.
 TEM type enzymes are present in almost all gram
negative bacteria.
 These enzymes were first isolated from E. coli strains
isolated, in Athens, from a young girl called Temoniera
and was referred to as TEM enzyme.
 Electrophoretically different type was then isolated from
Pseudomonas aeruginosa (TEM-2).
OXA
E. coli
SHV
Klebsiella spp.
ROB
H. influenza
AER
Aeromonas spp.
LCR
Ps. aeruginosa
Inactivation of chloramphenicol
 Inactivation of chloramphenicol by chloramphenicol
acetyl transferase (CAT).
 Usually they are a plasmid-mediated enzymes which are
inducible type in gram-positive bacteria but constitutive
in gram-negative bacteria.
 These enzymes acetylates the OH groups in the side chain
of the drug.
 Replacement of the terminal - OH group of this side
chain, which is normally the first to be acetylated by an
inert fluorine atom, yields a chloramphenicol derivative
that is not susceptible to attack by CAT.

Change in the antibiotic target
site
Chromosomal
Resistance to
Resistance to
Erythromycin
Aminoglycosides
Resistance to
Resistance- to some
Penicillins
Sulfonamides &
Trimethoprim
Chromosomal Resistance to
Aminoglycosides
 It is associated with the loss or alteration of a specific
protein in the 30S subunit of the bacterial ribosome that
serve as a binding site in the susceptible organisms.
Resistance to Erythromycin
 It is associated with alteration, of its receptor on the
5OS subunit of the ribosome.
Resistance- to some Penicillins
 Resistance- to some penicillins due to loss or alteration
of Penicillin Binding Proteins (PBPs).
Resistance to Sulfonamides & Trimethoprim
 Occurs by alteration of the tetrahydropteroat synthetase
and tetrahydrofolate reductase, respectively, that have
a much higher affinity for PABA than these drugs.
Reduction in cellular permeability

to the antibiotic
 Bacterial cells altering the permeability of their cell
membrane making it difficult for antimicrobials to enter.
 This type of resistance is found in bacteria resistant to:
a. Polymyxins.
b. Tetracyclines.
c. Amikacin & some aminoglycosides.
 Streptococci have a natural permeability barrier to
aminoglycosides.
 This can be partly overcome by combination with cell wall
active drug, e.g. (penicillin).
Switch to alternative metabolic

pathways unaffected by the drug:
 The organism develop an altered metabolic pathway
that bypasses the reaction inhibited by the drug
e.g. some sulfonamide-resistant bacteria do not require
extra-cellular PABA but, like mammalian cells, can utilize
preformed folic acid.
Increased production of essential

metabolite
:
 That is competitively antagonized by the drug in
sensitive cells e.g. resistance to sulfonamides may be
associated with high level of bacterial synthesis of PABA.
The origin of drug resistance
Non Genetic Origin
Genetic Origin
Non Genetic Origin
 This involves metabolically inactive cells or loss of target
sites.
a. Most antimicrobial agents act effectively only on
replicating cells.
• Mycobacteria survive for many years in tissue yet are
restrained by the host's defenses and do not multiply.
• Such persisting organisms are resistant to treatment
and cannot be eradicated by drugs.
• When they start to multiply they are fully susceptible to
the drugs.
b. Loss of a particular target structure, often induced by
the drug, may result in antimicrobial resistance.
• Exposure of some gram-positive bacteria to penicillin
results in the formation of cell lacking cell wall
(i.e. L-forms).
• These cells then are penicillin resistant, having lost the
structural target site of the drug.
• When these organisms revert to their bacterial parent
forms resuming cell wall production, they are again fully
susceptible to penicillin.
Genetic Origin
 Most drug-resistant microbes emerge as a result of
genetic change and subsequent selection processes by
antimicrobial drugs.
The mechanisms by which genetic
change occur are:
Chromosomal
Resistance
Extra
Chromosomal
Resistance
Chromosomal Resistance
 This develops as a result of mutation in a gene locus that
controls susceptibility to a given antimicrobial drug.
 The presence of the drug serves as a selecting
mechanism to suppress susceptible organisms and favor
the growth of drug resistant mutant.
 Spontaneous mutation occurs at a frequency of 10
10 12.
7
to
 Chromosomal mutants are most commonly resistant by
virtue of a change in a structural receptor for a drug as
in bacterial resistance to erythromycin, lincomycin,
aminoglycosides and others by alteration of their target
site in susceptible cells.
 Prevention of the emergence of resistant mutants is one
of the main indications for the clinical use of
combinations of drugs.
 But provided that the mechanisms of action of the two
drugs are unrelated.
 Therefore if both drugs are given in adequate dosage,
the risk of the emergence of a resistant strain is very
much less than if either is used alone.
Extra Chromosomal Resistance
Plasmids
Transposons
Plasmids
 Bacteria often contain extra chromosomal DNA units
known as plasmids.
 Some of which alternate between being free and being
integrated into the chromosome.
 R factors are a class of plasmids that carry genes for
resistance to one and often several antimicrobial drugs
and heavy metals.
 Plasmid genes for antimicrobial resistance often control
the
formation
of
enzymes
that
inactivate
the
antimicrobial drugs such as β-lactamases, CAT and
enzymes that inactivates aminoglycosides; or enzymes
that determine the active transport of tetracyclines
across the cell membrane, and for others.
Transposons
 The drug resistance (R) genes are often part of highly
mobile short DNA sequences known as transposons
(Transposable elements or jumping genes) that is able to
move, from one position to another, between one
plasmid and another or between a plasmid and
a portion of the bacterial chromosome within a bacterial
cell.
 Thus, transposons are able to insert themselves into
many different genomic sites with no homology with
them.
 Simple
transposons
(IS)
only
carry
information
concerned with the insertion function.
 Simple transposons (IS) (i.e. insertion sequences) have
no known effects beyond transposition and inactivation
of the gene (or operon) into which they may insert.
 Complex or composite transposons (Tn) contain
additional genetic material unrelated to transposition,
such as drug-resistance genes.
 Such
as
sulfonamides,
trimethoprim.
penicillin,
kanamycin,
tetracyclines,
streptomycin,
chloramphenicol
,and
Mechanisms of Transmission of Genetic
Material and Plasmids
Transduction
Transformation
Conjugation
Transposition
Transduction
 This is the main mechanism for transmission of antibiotic
resistance between gram-positive cocci, and occurs in
other bacterial groups.
 Plasmid DNA is enclosed in a bacteriophage and
transferred by the virus to another bacterium of the same
species e.g. the plasmid carrying the gene for β-lactamase
production can be transferred from a penicillin-resistant
to a susceptible staphylococcus if carried by a suitable
bacteriophage.
Transformation
 Naked DNA passes from one cell of a species to another
cell, thus altering its genotype.
 This can occur through laboratory manipulation and
perhaps spontaneously.
Conjugation
 This is the commonest method by which, multi-drug
resistance
spreads
among
different
genera
gram-negative bacteria.
 But also occurs among some gram-positive cocci.
of
 The usual plasmid found in resistant gram-negative
bacteria consists of two distinct but frequently linked
elements:
a. One or more linked genes each conferring resistance
to
a
specific
antibacterial
drug
(resistance determinants).
b. A resistance transfer factor (RTF) that enable the cell
to conjugate with a sensitive bacterium and to transfer
to It a copy of the entire plasmid.
R-factor
 The entire linked complex of RTF and resistance
determinants is known as R-factor and takes the form of
a double stranded circular molecule of DNA.
 A proportion of R-factor-bearing cells (R* cells) possess
hair-like structures that extend out from the bacterial
surface known as pili.
 The pili, whose synthesis is under the control of the
RTF component of the R-factor are essential to the
conjugation phenomenon with R' bacteria and the
transfer of an R- factor.
Transposition
 A transfer of short DNA sequences (transposon) occur
between a plasmid and another or between a plasmid
and a portion of the bacterial chromosome within
a bacterial cell.
Transferable or infective drug resistance is important for the
following reasons
i. The
transferable plasmids commonly
resistance to several unrelated drugs.
determine
ii. Such plasmids are transferable not merely to related
strains of the same species but to strains of other species
and genera; for example, antibiotic-resistant but
harmless organism in human or animal intestine
(E. coli) can confer antibiotic resistance, by plasmid
transfer,
on
pathogenic
but
previously
antibiotic-sensitive bacteria of other genera which the
host happens to ingest (such as typhoid or dysentery
bacilli).
iii. It is possible for multiple-resistant enterobacteria to
develop in farm animals and be transmitted to man.
Development of this resistance is due to the
widespread use of antibiotics especially cheap
types, as food supplements for young animal to
accelerate their growth by partial suppression of
their intestinal flora.
Specific and Cross Resistance
a. Specific
resistance:
When
the
organism
acquire
resistance to a certain drug but Its susceptibility to other
drugs is unaffected.
b. Cross resistance: Microorganisms resistant to a certain
drug may also be resistant to other drugs that share
a mechanism of action.
 Such relationship exist mainly between agents that are
closely related
 Polymyxin B and Polymyxin E.
 Erythromycin and Oleandomycin.
 Neomycin and kanamycin.
 However, it may also exist between unrelated chemicals
 Erythromycin-Lincomycin.
 When the active nucleus of the chemicals is so similar,
extensive cross-resistance is to be expected e.g.
resistance to one of the tetracyclines imparts resistance
to the other members of the group e.g. resistance to
one sulfonamide cause resistance to hundreds of other
sulfonamides.
Antibiotic Policies
 Abuse of antibiotics, is avoided for many reasons
including the following:
a. To prevent the emergence of antibiotic resistance.
b. To reduce the cost of antibiotic use.
c. To prevent antibiotic toxicity.
General Principles of Optimal
Antibacterial Therapy
1
Unless there is a valid reason for giving an antibiotic,
the patient would probably be better off without it.
Treatment of
Known or
Suspected Infection
Prevention of
Bacterial Infection
Peri-operative
Prophylaxis
Patients at Special
Risk
2
In cases when immediate drug treatment is
necessary.
3
It is bad treatment to use broad-spectrum antibiotics
when an infective condition can be treated with
a more specific agent.
4
It is essential to use
bacteriostatic therapy.
5
It is essential to use combination of antimicrobial
drugs in certain situations.
bactericidal
and
not
6
For treatment of superficial infections it is important
to use either antiseptic or antibiotics which are rarely
or never used systemically.
7
Give enough, for long enough, and then stop
treatment with the antibiotic.
8
To reduce the spread of microbial resistance, avoid
the use of antibiotics as food supplement for animals
or for preservation of human food stuffs, avoid
liberation of antibiotic powders and solutions into
the environment.
9
To reduce the emergence of antibiotic-resistant
strains, an antibiotic policy has to be introduced for
a hospital or area e.g., using antibiotics in rotation,
keeping a particular antibiotics and permitting their
use only on rare and special occasions, or insisting on
combined therapy.
Drug combination
1. To provide broad coverage.
2. For initial (blind) therapy when the patient is seriously
ill and results of cultures are pending.
3. To provide synergism when organisms are not
effectively eradicated with a single agent alone e.g., in
enterococcal
endocarditis
both
penicillin
and
an aminoglycoside are given because their combined
effect is greater than the sum of their independent
activities.
4. To prevent emergence of resistance, as in the
treatment of tuberculosis.
 Inappropriate use of combinations could result in:
 Antagonism it occurs when a bactericidal agent is used
with a bacteriostatic one as in penicillins plus
tetracycline or Chloramphenicol.
Sulphonamides do not antagonize penicillins, possibly
because their bacteriostatic action is too low.
 An increase in the number or severity of adverse
reactions.
 Increased coast.