Molecular mechanism behind development of antibiotic resistant strains of bacteria

1. Molecular Mechanism of Multi Drug
Resistance (MDR)
Geetanjali Baruah

2.  In 2015, an estimated 4,80,000 people
worldwide developed MDR-TB
 An additional 1,00,000 people with rifampicin-
resistant TB were also newly eligible for MDR-
TB treatment
 India, China, and the Russian Federation
accounted for 45% of the 5,80,000 cases
 It is estimated that about 9.5% of these cases
were XDR-TB (WHO, March 2017)
 In Europe 25,000 people die each year due to
MDR bacterial infections, costs €1.5 billion
annually
The concern is real

3. The concern is real

4. The history and benefits

5. Reasons behind the crisis
 Overuse
 Inappropriate Prescribing
 Extensive agricultural use
 Availability of few new antibiotics
 Regulatory barriers
Multiple Drug resistance (MDR): MDR was defined as
acquired non-susceptibility to at least one agent in three or
more antimicrobial categories
Extensive Drug resistance (XDR): XDR was defined as
non-susceptibility to at least one agent in all but two or
fewer antimicrobial categories (i.e. bacterial isolates remain
susceptible to only one or two categories)
Pan-drug resistant (PAN) : PDR was defined as non-
susceptibility to all agents in all antimicrobial categories

6. How an antibiotic works
Types:
Bactericidal: Bactericidal antibiotics kill bacteria
e.g. Beta-lactam antibiotics (penicillin
derivatives), cephalosporins, monobactams,
carbapenems and vancomycin
Bacteriostatic: Bacteriostatic antibiotics slow
their growth or reproduction
e.g. Chloramphenicol, clindamycin, and linezolid

7. Intrinsic resistance in bacteria
Examples:
oTriclosan
- Unable to inhibit gram-negative
Pseudomonas
Reason: carries an insensitive allele
of fabI (encodes an additional enoyl-
ACP reductase enzyme)
oDaptomycin
- Active against G+ not G-
Reason: G- bacteria have low anionic
phospholipids in cytoplasmic membrane
(Blair et al., Nature reviews, 2015)
E. coli genes responsible:
thioredoxin (TrxA), thioredoxin
reductase (TrxB), FabI, RecQ, SapC,
DacA
if inhibited, can greatly promote the activity
of existing drugs
Understanding about genetic basis
of intrinsic bacterial resistance
and the spectrum of activity of an
antibiotic are key to new
combinations with improved or
expanded activity against target
species

8. 3 Basic ways of acquired resistance:
minimize the intracellular concentration of
antibiotic by poor penetration or efflux
Modification of target by mutation or post-
translational modification
Inactivation of antibiotic by hydrolysis or
modification
Mode of action & acquired resistance

9. 1. Inhibition of Cell Wall Synthesis
Beta-Lactams: Inhibition of peptidoglycan synthesis
Resistance: fails to cross membrane, fails to bind to altered PBP’s &
hydrolysis by beta-lactamases
2. Inhibition of Protein Synthesis
30S Ribosome site:
Tetracyclines: Block tRNA binding to 30S ribosome-mRNA complex (b-
static)
Resistance : decreased penetration, active efflux , protection of 30S
ribosome
50S Ribosome site:
Chloramphenicol: Binds peptidyl transferase of 50S ribosome, blocks
peptide elongation
Resistance: plasmid-encoded chloramphenicol transferase, altered
outer membrane
Mode of action & acquired resistance

10. 3. Alteration of Cell Membranes
Bacitracin: Disrupt cytoplasmic membranes
Resistance: inability to penetrate outer membrane
4. Inhibition of Nucleic Acid Synthesis
DNA :
Quinolones: Inhibit DNA gyrases or topoisomerases; bind to alpha subunit
Resistance: alteration of alpha subunit of DNA gyrase, decreased uptake by
alteration of porins
RNA:
Rifampin: Binds to DNA-dependent RNA polymerase inhibiting initiation &
Rifabutin of RNA synthesis
Resistance: altered beta subunit of RNA polymerase , intrinsic resistance
(decreased uptake)
5. Antimetabolite Activity
Sulfonamides: Compete with p-aminobenzoic acid (PABA) preventing
synthesis of folic acid
Resistance: permeability barriers
Mode of action & acquired resistance

11. Some recent facts
Prevention of access to target
Reduced permeability
- Hydrophilic antibiotics diffusing through outer-membrane porin proteins
- Major porins OmpF and OmpC of E. coli, previous evidence said drug-
binding sites present within which was proven wrong
-reducing the permeability of the OM by the downregulation or by replaceing
with more-selective channels
Enterobacteriaceae, Pseudomonas spp. and Acinetobacter spp., reductions
in porin expression significantly contribute to resistance to newer drugs
Increased efflux
-When overexpressed, efflux pumps confers high levels of resistance
-Some efflux pumps have narrow substrate specificity (for example, the Tet
pumps), but many transport a wide range of structurally dissimilar substrates
and are known as multidrug resistance (MDR) efflux pumps

12. Recent understandings on acquired resistance
(Blair et al., Nature reviews, 2015)

13. Changes by mutation
modification through the
formation of ‘mosaic’ genes. E.g.
penicillin resistance in S.
pneumoniae, by mosaic
penicillin-binding protein
(encodes penicillin-insensitive
enzymes), arisen by
recombination with
Streptococcus mitis
Modification of targets
qnr families of quinolone
resistance genes encode
pentapeptide repeat proteins
(PRPs), which bind to and
protect topoisomerase IV and
DNA gyrase from the lethal
action of quinolones

14. Direct modification of antibiotics
Inactivation of antibiotics by
hydrolysis
The early β lactamases, which were‑
active against first-generation
β lactams, were followed by extended-‑
spectrum β lactamases (ESBLs)‑
Inactivation of antibiotic by
transfer of a chemical group
-addition of chemical groups to
vulnerable sites on the antibiotic
molecule by bacterial enzymes causes
antibiotic resistance by preventing the
antibiotic from binding to its target
protein as a result of steric hindrance
-Aminoglycoside antibiotics are
particularly susceptible

15. Cell Reports| 2017
On the basis of RNA seq and Tn seq data Jensen et al. suggested
nutrient and antibiotic stresses respectively lead to coordinated
and uncoordinated responses. Network models are thus key to
understanding cellular responses, thereby aiding in predicting bacterial
behavior

16. THANK YOU