Transcript ANTIBIOTIC RESISTANCE

MECHANISMS OF ANTIBIOTIC
RESISTANCE
 Dr T. Aswani Ndonga
Msc TID I
April 2010
Definition
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 Definition:-Relative or complete lack of effect of
antibiotic against a previously susceptible microbe
MECHANISMS OF RESISTANCE
 Enzymatic inhibition
 Alteration of bacterial membranes
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Outer membrane permeability
Inner membrane permeability
 Rapid ejection of the drug [efflux] or reduced drug influx.
 By pass of antibiotic inhibition.
 Alteration of target sites
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Altered ribosomal target sites
Altered cell wall precursor targets
Altered target enzymes
Molecular genetics of antibiotic
resistance
 Genetic variability is essential for microbial evolution.
It may occur in a variety of ways :o Micro evolutionary changes by point mutations-in
nucleotide base pair
o Micro evolutionary changes [whole scale changes] like
-Inversions
 Duplications
 Deletions
 Transposition
o Acquisition of foreign Dna –plasmid
mediated/bacteriophages/transposons
MECHANISMS OF RESISTANCE
1.Enzymatic inhibition
Enzymes inactivating antibiotics
 Beta-lactamases-split amide bond of the beta lactam ring.
 There are many types-characterised by amino acid and
nucleotide sequencing
Class A MWT 29000-Preferentially hydrolyze penicillins e.g. TEM-1
prevalent in many gram neg
 Class B-metalloenzymes have a zinc –binding thiol group required
for beta lactamase activity
 Class C mwt 39000 –mainly cephalosporinases
 Class D-oxacillin –hydrolyzing enzymes
Many beta lactamases are plasmid mediated all are produced
constitutively there are 6 main groups
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Those that hydrolyze benzylpenicillin
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Beta lactamases
Those that hydrolyze oxacillin and related penicillins
Carbenicillinases
Those that break extended spectrum beta lactams like
aztreonam
4. Those that break down oxyimino B lactams
5. Carbapenemases-found in pseudomonas
Most cephalosporinases are inhibited by clavulanate,sulbactam
or tazobactam. Carbapenamases are metalloenzymes
inhibited by EDTA but not clavulanate or sulbactam
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Production of enzymes modifying antibiotics
 . Aminoglycosides, chloramphenicol-coded by plasmids or
chromosomal genes
Beta lactamases site of action
Modifying enzymes
 Reactions are N-acetylation
 O nuleotidylation
 O phosphorylation
 Chloramphenicol acetyltransferase-inactivates the drug by
3-o-acetylation-plasmid mediated/chromosomal
 Erythromycin esterase-seen in E coli-hydrolyze lactone
ring thus deactivating it-limits utility of oral erythromycin
in reducing the aerobic gram neg flora of the GIT prior to
Gi surgery.
ENZYMES
Degrading enzymes will bind to
the antibiotic and essentially degrade it
or make the antibiotic inactive
Blocking enzymes attach side chains to
the antibiotic that inhibit its function.
E.g. -lactamases
Alteration of bacterial membranes
 Outer membrane permeability—outer membrane of
gram neg acts as a barrier to antibiotics esp
hydrophobic ones.
 Inner membrane permeability- rate of entry of
aminoglycosides into bacterial cells is a function of
them binding to a non saturable anionic
transporter,where they retain their positive charge and
are pulled across the cytoplasmic membrane by the
internal charge of the cell.This is an energy dependent
process. The energy generation or proton motive force
may be altered through mutation
Alteration of bacterial membranes
continued
 Promotion of antibiotic efflux-major mechanism for
tetracycline resistance in gram negplasmid/chromosomal/transposone mediated
.Efflux /influx mechanism
 Bacterial cells have an intrinsic capacity to restrict the
entry of small molecules especially gram neg-outer
membrane is protective,gram pos no outer membrane
hence more antibiotic sensitive
 Restriction of influx is a physiological way to reduce
toxixity to bacterial cell.
 The most wellstudied efflux system in E. coli is the
AcrAB/TolC system this system comprises of an inner
membrane proteinAcr B, and an outer membrane
protein, Tol C, linked by a periplasmic protein, Acr A
Influx/efflux
 When activated, the linker protein is believed to fold
on itself, bringing the AcrB and Tol C proteins in close
contact, thus providing an exit path from the inside to
the outside of the cell. Antibiotics are pumped out
through this channel.
Efflux pump
The efflux pump is a
membrane bound protein
that "pumps" the
antibiotic out of the bacterial
cell.
3. Modification of target sites
 Alteration of ribosomal target sites-hence failure to
inhibit protein synthesis and cell growth.
 Affected antibiotics are aminoglycosides
,tetracylines,macrolides,lincosamides.
Altered cell wall precursor targets
 Glycopeptides like vancomycin-bind D-alanine-D-
alanine which is present at the termini of
peptidoglycan precursors.
 The large glycopeptide molecules prevent the
incorporation of the pre cursors into the cell wall
Alteration of target enzymes
 Alteration of PBPs in B lactams
 SMX/TMP-production of a dihdropteroate synthetase
that is resistant to binding by sulphonamides-plasmid
mediated
 Quinolones-DNA gyrase is made up of gyr Aand gyr B
genes-mutations in gyr A result in resistance
By pass inhibition
 Development of auxotrophs-have growth factor
requirements different from those of wild strain these
mutants require subtrates that normally are
synthesized by the target enzymes and if these are
present in the environment the organisms grow
despite inhibition by synthetic enzymes
Modification of AB
target sites:
disruption in
protein synthesis
Some terminologies
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. VRE . vancomycin-resistant enterococci
. 70% of E. faecium strains in USA
. GISA . glycopeptide intermediately susceptible
S.aureus
. VISA . vancomycin intermediately susceptible
S.aureus
. VRSA & VRSE . vancomycin-resistant S.aureus and
S.epidermidis
. (MIC> 32 mcg/ml; 1st clinical case described in 2002 in USA)
. ESBL producing K.pneumoniae . extended
spectrum -lactamase producing K. pneumoniae
. PRSP penicillin-resistant S. pneumoniae
Antibiotic resistance
Factors promoting drug resistance
 Exposure to sub-optimal levels of antimicrobial
 Exposure to microbes carrying resistance genes
 Inappropriate drug 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 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
Antibiotics and mechanisms of
resistance
ANTIBIOTIC
TARGET
MOA
MECHANISM
OF
RESISTANCE
b-Lactams
Transpeptidas
es/transglycos
ylases (PBPs
Blockade of
cross linking
enzymes in
peptidoglycan
layer
b-Lactamases,
PBP mutants
Vancomycin
D-Ala-D-Ala
termini of
peptidoglycan
and of lipid II
Sequestration
of substrate
required for
cross linking
Reprogrammi
ng of D-AlaD-Ala to DAla-D-Lac od
D-Ala –D-ser
CELL WALL
PROTEIN
SYNTHESIS
END