Antibiotics: Mode of

Action and resistance

Antibiotic
 Chemical substance
 Natural, semi synthetic and wholly synthetic
 Effective at low concentration (μ/ml)
 Bacteria static or bactericidal

Dr Ellabib MS
 Criteria for
antibiotic
 Selectively toxic for bacteria
Bactericidal (killing)
Bacteriostatic (growth inhibition)

 No harm to patient
 Destroy structures
Present in bacteria
Not present in host
 Antibiotics work together with immune
system
 Minimal inhibitory concentration

 Lowest level stopping growth

e.g. zone of inhibition around a disk
impregnated with antibiotic
Antibiotics inhibit cell wall are bactericidal
Without cell wall osmotic pressure cause
bacteria to burst
Mode of Action
 Inhibitors of cell wall synthesis (Peptidoglycan)
 Inhibit Peptidoglycan biosynthesis at various stages
 Stage one: inhibit UDP –N-Acetylmuramic acid through
inhibition of tranferase enzyme
 Fosfomycin
 Stage two: inhibit building pentapeptides side chain through
inhibition of synthetase enzyme
 Cycloserine
 Stage three: inhibit reaction leading to formation of a linear
Peptidoglycan polymer (pyrophosphtase enzyme)
 Vancomycin & Bacitracin
 Stage four: preventing cross-linking and formation of
Peptidoglycan (transpeptidase enzyme)
 Bound covalently to various proteins called penicillin binding
proteins (PBPS)
 B-lactam antibiotics
Chemical modification changes of B-
lactam and their biological activity
 Early B-lactam antibiotics
 Penicillin G &V
 Inactive against gram negative
 No penetration of outer membrane
 Unstable to B-lactamases enzyme
 Active against streptococcus pyogens
 Pencilliinase-resistant penicillin
 More stable
 Staphylococcus species
 E.g. Methicillin and Oxacillin
 Aminopencillin
 activity against gram positive
 increased activity toward gram negative (E. coli)
 Ampicillin, Amoxil
Antipseudomonase penicillin
Carboxypenicillin
Increased activity against -ve and decreased activity
against +ve
Carbenicillin and ticarcillin
Uredopenicillin
Increased activity against –ve and preserved activity
against +ve
Piperacillin and mezalocillin
B-lactamases inhibitory
Binds strongly to beta-lactamase
Inhibit activity
Called suicidal agents
Clavulanic acid & Tazobactam
Cephalosporin's
 First generation
 Old and narrow spectrum cephalosporin's
 Cephaloridine, Cephalexin
 2nd generation
 -ve and some anaerobic bacteria
 Cefuroxime and cefoxitin
 3rd generation
 -ve such as pseudomonas
 Ceftriaxone and Ceftazidime
 4th generation
 Broad spectrum
 cefpime
Other B-lactam antibiotics
 Monobactams
 -ve
 Aztreonam
 Carbapenems
 -ve
 Imipenem

Antifungal agents
Antifungal
 Chitin synthetase
 Polyxin and nikkomycin
 Glucan synthesis
 cilofungin
Compounds inhibit cell membrane
 Concentration dependent
 Effect integrity of CM
 Leakage of K, proteins and nucleic acid
 Disinfectants, antiseptics and polypeptides

 Phenols
 Release compounds absorbed at 200nm
 Inhibits electron transport chains (metabolic activity)
 Alcohols
 Interact with ester fatty acid and thiol group of
proteins
 Used as 70% concentration
Chlorohexidine
 Inhibit adenosine triphosphotase (ATPase)
 Uptake of K+
 Polymyxin B
 Inhibit phospholipids of –ve but not +ve

 Antifungal agents (inhibit Ergosterol)

 Interact with phospholipids of CM directly
 Pores and leakage
 Amphotericin B and Nystatin
 Indirectly inhibit ergosterol biosynthesis
 Cytochrome P-450
 Azoles antifungal (Miconazole, itraconazole,
Clotrimazole )
 Other non azoles such as terbifine morpholin and
tolnaftate
Inhibitor of protein synthesis
 Mostly Bacteriostatic
 Selectivity due to difference in ribosome's
 Some toxicity –eukaryotic 70S ribosome's

 Classes of ribosome's subunit

 80s ribosome's (eukaryotic)
 Dissociated to 60s and 40s as Mg+ concentration
 Protein to RNA (50:50)
 70s (prokaryotic and eukaryotic)
 dissociate to 30s and 50s
 Protein to RNA (35:65)
 50-55s ribosome's (mammalian mitochondria)
Antimicrobial that bind to the 30s
Amino glycosides
 Such as Streptomycin, gentamicin, Amikacin
 Gram positive and –Ve
 Slight initial entry of the drug inside cell
 Interact with chain elongation (PC) at 12s
 Misreading of mRNA abnormal protein
 formation of abnormal channels
 Increase and irreversible entry through channels
 Blockage initiation of ribosome's
 Inhibit binding of aminoacyl-tRNA and peptide synthesis
inhibit protein synthesis
 Tetracycline's ( Rickettsias and
mycoplasma)

Short acting Chlortetracycline

Intermediate Demeclocycline
Long acting Doxycycline
Blocking binding of aminoacyl-tRNA acceptor site on
the mRNA ribosome's complex
Prevent the addition of new amino acids to the
growing peptide chain
Antimicrobials bound to 50s subunit
Chloramphenicol
 Natural antibiotic
 Meningitis, typhoid fever
 Associated with bone marrow toxicity
 Reversible effect
 Bind to a region on 50s close to site bound aminoacyl-tRNA
in peptidyltransferase center
 Blocking addition of new amino acids
 Prevent growing of protein chain
 Inhibit peptide bound formation
Macrolides (erythromycin,
claritromycin and spiramycin)
 +ve bacteria, mycoplasma, legionella
 Resistant common
 Stimulate dissociation of peptidy-tRNA from
ribosome's during translocation step
 Interrupting completion of peptide chain formation
Lincosamides (Lincomycin & Clindomycin)
 Similar to chloramphenicol and erythromycin
 +ve cocci
 Resistant common
 Anaerobic bacteria
Inhibitors of protein synthesis
Fusidic acid

Bind to 70s ribosome's

Active against gram positive cocci
Inhibits polypeptide chain elongation
Inhibitor of nucleic acid synthesis
 To toxic
 Some are used to treat tumor, viral and serious
bacterial such as TB
 DNA inhibition
 result in cell division inhibition
 Effect Extra chromosomal elements of DNA
and plasmids
 Effect bacterial response to environmental
changes
 RNA inhibition
 Inhibit protein synthesis
 Nucleic acid inhibitors (Two
groups)

 Compound interfere with precursor of

nucleic acid (purine, pyrimidine)
Sulfonamides and trimethoprim

Compounds interfere with nucleic acid

synthesis at the polymerization stage
 RNA polymerase inhibitors

 rifampin
Mycobacterium tuberculosis
Inhibit DNA-dependent RNA polymerase
RNA polymerase

Minimal or core enzyme Sigma factor (RNA)

α, β, β1 and ω subunits
 Rifampin continuous
 Form a tight one to one complex with β subunit
 Prevent protein synthesis (chain initiation)

 DNA gryases or topoisomerase inhibitors

 quinolones
 Nalidixic acid, ciprofloxacin
 Called nick closing enzyme
 Enzyme play important role in supercoil strand DNA
 Supercoiling is completed the single strand DNA is
abolished by an enzyme that seals the nicked DNA
 The enzyme is known as DNA gryase or
Topoisomerase 2
DNA gryase or Topoisomerase 2
Four subunits and two were identified as A and β
 A subunit introduce the nick and seal the nick
they produced initially
Nalidixic acid
 β subunit
Responsible for supercoil
Norofloxacin & ciprofloxacin
May interfere with A and β subunit
 Inhibitors of Folic Acid
Synthesis
 Sulfonamides
 Trimethoprim
Anti- Mycobacterial Antibiotics

Para-aminosalicylic acid (PSA)

Para-aminosalicylic
 Bacteriostatic

 Dapsone
 Bacteriostatic
 treatment of leprosy (Mycobacterium leprae)

 Isoniazid (INH)
 bacteriostatic
 inhibits synthesis of mycolic acids.
 Furantoin
 Gram positive and negative
 Urinary tract infection
 Damaging DNA
Nitro-imidazole (metronidazole)
 Anaerobic bacteria and protozoa
 Reduced to Nitro radical compounds
 Acts as nuclease and damaging DNA
Griseofulvin
 Antifungal agent
 Dermatophytes infection only
 Effect nuclear function
 Interfere with microtubules during separation of
chromosomes in cell division at the metaphase
Antibiotic Mechanisms
of Action
 Microbial resistance to
antimicrobial agents
 Clinical resistance
 By mutation or acquisition of a plasmid
 Provides a selective advantage
 Single or multiple steps
 Cross resistance Vs multiple resistance
 Cross resistance
 single mutation
 closely related antibiotics
 Multiple resistance
 multiples mechanisms
 Unrelated antibiotics
Genetic basis of resistance
Mutation
 For the origin of some resistant variants
Acquisition
 Transfer of genetics material from R to S microorganism
 Confined by genes on Chromosomal or plasmid
 Chromosomal mediated resistant
 Remain with the particular bacterial cell
 And Offspring
 Plasmid mediated resistant
 Self replicating extra chromosomal DNA
 Widely distributed in nature
 Often transmissible
 Often Carry resistant determinants to many drugs
 Other function of plasmid

 Carry genes allow bacteria to attach to mucosal

surface
 Produce toxins
 Invade and colonize host cells
Transformation of plasmid resistant
 Conjugation (direct cell to cell contact)
 Transduction (bacteriophage vector)
 Transformation (uptake DNA from environment)
 Transposition (transformation via transposons
Transduction ( bacteriophages
vector)
 Transposons
 So called jumping genes
 Movable DNA elements
 Carrying resistant genes
 Jump or hop from plasmid to plasmid
 From plasmid to chromosomal
 Found in many bacteria
 Carrying resistant to many antibiotic
 Main cause of hospital and community outbreaks
resistant
 Plasmid from Epidemiological
viewpoint
 Most important type of resistant
 Transmissible
 Usually highly stable
 Convert resistant to different class of antibiotics
 Often associated with other characteristics
 Requirements for antibiotic activity and mechanisms resistant
 Properties of antibiotic required for efficacy
 Penetration to target site in sufficient amount
 Evade inactivation enzymes by microorganism
 Interaction with target molecules to initiate an effect
Steps of resistant mechanisms
 Cellular barrier to drug penetration
 Reduce antibiotic activity
 Altered target molecules
 Interact less effectively with drug with retain function
 May use additional pathway insensitive to the drug
 Modifying or inactivating enzymes
 Rendering antibiotic ineffective
Two or more of these mechanisms give rise to high level
of resistant than any one mechanisms alone
 Resistant due to permeability
 Beta lactam
 Poor permeability
 change in porin channels
 Mutation
 Such as Neisseria gonorrhoeae
 Outer membrane composition
 Lipoplysaccharides, lipids and proteins
 Pseudomonas aeruginosa
 Usually display resistant to other antibiotics
E. coli deficient mutant in porin channels
 Was sensitive to imipenem because
 Due to its compact zwitterionic structure
 Permit rapid penetration
 PBP2 per cell 20
 Require only few drug molecules for antimicrobial
Aminoglycosides
 loss or reduction in outer membrane proteins
 Enterobacteriacae
 Appearance of new outer membrane proteins
 Or alteration in surface Lipoplysaccharides
 Affect intracellular uptake of Aminoglycosides
 Ps. aeruginosa
 MLS Antimicrobials
 Intrinsic low level resistant to erythromycin
 Limited drug permeability
 Enterobacteriacae
 Reduced drug uptake
 Staph. epidermidis
 Chloramphenicol
 Loss of outer membrane proteins
 Diminish uptake of drug in Ps. aeruginosa (acquired)
 Tetracycline's
 Poor penetration
 Change in outer membrane (proteins, porin)
 Alteration in Lipoplysaccharides
 Ps. Aeruginosa (intrinsic)
 Poor penetration
 plasmid-mediated changes in transport system
 Acquired
 Anaerobic bacteria
 Quinolones
 Decrease level of major outer membrane proteins
 Enterobacteriacae
 Cross resistant with unrelated antibiotics
Folate pathway inhibitors
 Permeability barriers
 Intrinsically
 Ps. aeruginosa
 Trimethoprim
 Acquired
 Diminished level of putative proteins
 Enterobacteriacae
 Trimethoprim
 Cross resistant to other unrelated drugs classes
 Decrease intracellular penetration
 Intrinsic or acquired
 Gram negative bacteria
 sulphonamides
 Resistant due to altered target
molecules
 Beta-lactam antibiotics
 Altered penicillin-binding proteins (PBPS)
 Pneumococci
 Staph. aureus
 Methicillin Resistant Staph. aureus (MRSA)
 PBP2 alteration
 Lower affinity to drug
 Resistant gene located on transposable genetic element
 Should be consider resistant to all beta-lactam drugs
 Aminoglycosides
 Mutation affecting binding to ribosomal target
 Streptomycin and kanamycin
 Alteration of 12s protein
 Control binding of drugs to 30s ribosomal subunit
 Staph. aureus, Ps. aeruginosa and mycobacterium
Macrolides-Lincosamides
 Barrier to penetration
 Intrinsic In Enterobacteriacae
 Alteration in 50s ribosomal subunit
 Plasmid-mediated
 Found on transposons
 Staph. aureus., streptococci, Pneumococci, Bacteroides
 Demethylation of adenine group of 23s rRNA
 Decrease binding to 50s ribosomal subunit
Continuous
 Inducible or constitutive
 Inducible bacteria
 Show resistant to erythromycin and sensitive to
Clindomycin
 Sub inhibitory concentration of erythromycin
 Increase methylase enzyme
 Induced cells became resistant to all MLS
 Constitutive cell
 Resistant to MLS without prior antibiotic exposure
Quinolones
 Mutation in DNA gryase A and B subunit
 E. coli
 Rifampin
 Single step mutation in B-subunit RNA
polymerase
 E. coli
 Folate pathway inhibitors
 Low affinity to dihydropteroate synthetase
 Hyper production of enzyme
 Gram negative bacteria
 Transposons
 sulphonamide
Folate pathway
inhibitors
 Low affinity to dihydrofolate reductase
 Hyper production of enzyme
 Trimethoprim
Resistant due to inactivation of
antimicrobial

Beta-lactam antibiotics
 Most common and understand mechanisms
 Dozens of enzymes
 Differing in their substrate profile
 Potential for inhibition by B-lactam inhibitors
 Physical characteristics
 Divided into Four groups based on
 Preferred antibiotic substrate
 Inhibition by Clavulanic acid
 Based on Bush Scheme classification
Classification schemes of
Bush
Enzyme class OR Groups characteristic
Example
One cephalosporinase not inhibited chromosomal
enzyme of Ps. by Clavulanic acid aeruginosa
and Ent. Cloacae
2a-2c Pencilliinases and cephalosporinase plasmid mediated Tem-
type
inhibited by Clavulanic acid Staph, Kleb, chromosm
Group 3 Metalloenzymes Imipenem hydrolyzing
enzyme of Ps. Maltophilia
Group 4 Pencilliinases not inhibited chromosom enzyme of
Clavulanic acid Ps. Cepacia
Gram positive
bacteria
 Presented by exoenzyme of Staph, Enterococcus
 Less active against cephalosporin and stable
penicillin's
 Inhibited by Clavulanic acid
NB: Border line Oxacillin resistant (hyper producer of
enzyme)
 Gram-negative bacteria
 Many has been described
 Plasmid or chromosomal mediated
 Common among nosocomial pathogens
 Enterobacteriacae
 Ps. aeruginosa and Acinetobacter spp
Plasmid-mediated B-
lactamase
Such as TEM (Hydrolyzing ampicillin)
TEM3 (CTX) hydrolyzing cefotaxime
TEM5 (CAZ) hydrolyzing Ceftazidime
Inhibited by B-lactamase inhibitors
Inducible chromosomally-mediated
 Produced at low concentration before antibiotic
exposure
 Hyper production after exposure
 Resistant to many B-lactam antibiotics
 Ps. aeruginosa, Enterobacter
Aminoglycosides
 Acetylation of amino groups
 Adenylation
 Phosphorylation of hydroxyl group
 Plasmid or chromosomal modifying enzymes
 Enzyme may be located on transposons
 Staph. aureus or Ps. Aeruginosa

 MLS
 Esterase enzyme
 Plasmid born
 Klebseilla pneumoniae
 E. coli
 Erythromycin
Chloramphenicol
 Drug inactivation by enzyme
 Major mechanisms of resistant
 Gram positive bacteria
 By acetyltransferase enzyme
 Plasmid and inducible in Staph. aureus and Enterococci
 Constitutive in gram negative and plasmid located on
transposons
 Anaerobic bacteria
 Enzyme identified as nitroreductase
 Bacteroides fragilis
Tetracycline's

 Transposons element mediated resistant

 Promoting active drug efflux
 Drug detoxification
 Bacteroides fragilis
Notes: active efflux I a mechanism involving certain protein by
which the cell pumping the drug out
Antimicrobial cross
resistant
 Between drugs from same class
 Beta-lactam (MRSA)
 Aminoglycosides
 Inactivating enzymes
 Ps aeruginosa
 MLS
 Staph. Aureus
 Different classes
 Permeability mutant
 Altered outer membrane protein
Different classes
 Gram negative bacteria
 Quinolones, trimethoprim, chloramphenicol
 Single or many plasmids
 Resistant to multiple classes of antibiotics

 Reduction of antimicrobial resistant and synergism

 Synergism
 Two drug combined together
 0.25 of MIC of each drug
 Exert higher activity than that achieved by each drug
alone
 Used to prevent resistant
 Rifampin and streptomycin for TB
Continuous
 Rifampin plus Vancomycin for endocarditis
 B-lactam plus aminoglycosides for –ve
 Amphotericin B plus 5-flourocytosine for
Cryptococcus neoformans
 Empirical for unknown organisms or mixed
infection
 Metronidazole plus gentamicin