Determination of Antibiotic Resistance

Question 1

What is antimicrobial resistance?

Answer 1

A natural phenomenon accelerated by use of antimicrobial medicines
Resistant strains survive and aggregate

Question 2

How does AMR develop?

Answer 2

Microbes acquire mutations
Microbes acquire genetic information from other microbes to develop resistance

Question 3

What are the types of AMR?

Answer 3

Antibacterial resistance
Antiviral resistance
Antiparasitic resistance
Antifungal resistance

Question 4

Why is AMR a global concern?

Answer 4

Increases mortality
Challenges control of infectious diseases
Threatens a return into the pre-antibiotic era
Increases costs of healthcare
Jeopardises healthcare gains to society

Question 5

What are the 3 main mechanisms that lead to drug resistance?

Answer 5

Modify drug target
Modify drug
Modify efflux and influx of drug

Question 6

How does modification of drug target lead to drug resistance?

Answer 6

gyrase A mutation cause fluoroquinolone resistance

Question 7

How are drugs modified?

Answer 7

Aminoglycoside modifying enzymes
Beta lactamases degrade B-lactam antibiotics

Question 8

How does modification of efflux and influx of the drug lead to drug resistance?

Answer 8

Efflux pumps pump drug out the cells
Influx via porins

Question 9

What are genetic mechanisms of resistance?

Answer 9

Chromosome-mediated - due to spontaneous mutation in the target molecule or the drug uptake system - mutants are selected, not induced
Mobile gene-mediated - common in gram -ve rods, multiple mechanism of transfer e.g. plasmids, carry genes conferring multidrug resistance

Question 10

What is meant by selection of mutants?

Answer 10

Random mutation occurs causing resistance in that mutant
When antibiotic added to the population, the mutants are selected for and survive, creating a resistant species

Question 11

Describe genetic transfer in bacteria

Answer 11

Mechanism for genetic heterogeneity and evolution
Rapid, cross-species
Virulence (toxins), drug resistance, antigens (immune evasion)

Question 12

Give examples of fluoroquinolones

Answer 12

Ciprofloxacin
Moxifloxacin
Ofloxacin

Question 13

What do fluoroquinolones do?

Answer 13

Bind to DNA gyrase (gyrA) and topoisomerase IV (parC)
Inhibits DNA replication

Question 14

How does resistance to fluoroquinolones occur?

Answer 14

Mutations in the target gyrA and parC genes
Mutations usually in quinolone resistance-determining region (QRDR)

Question 15

Give examples of aminoglycosides

Answer 15

Tobramycin
Gentamicin
Amikacin

Question 16

What di aminoglycosides do?

Answer 16

Irreversibly bind to 16S rRNA in 30S subunit of ribosome
Inhibits protein synthesis

Question 17

How does resistance to aminoglycosides occur?

Answer 17

Mutations in target rrs gene (16S rRNA)
Aminoglycoside modifying enzymes

Question 18

What are the aminoglycoside modifying enzymes?

Answer 18

N-Acetyltransferase (AAC)
O-Adenyltransferase (ANT)
O-Phosphotransferase (APH)

Question 19

What does AAC do?

Answer 19

Catalyses acetyl CoA-depedent acetylation of an amino group

Question 20

What does ANT do?

Answer 20

Catalyses ATP-dependent adenylation of hydroxyl group

Question 21

What does APH do?

Answer 21

Catalyses ATP-dependent phosphorylation of a hydroxyl group

Question 22

Give examples of beta-lactams

Answer 22

Penicillins
Carbapenems
Cephlasporins

Question 23

Give examples of penicillins

Answer 23

Ampicillin
Amoxicillin

Question 24

Give examples of cephalosporins

Answer 24

Cefazolin
Ceftriaxone

Question 25

Give examples of carbapenems

Answer 25

Meropenem Ertapenem

Question 26

Give examples of monobactams

Answer 26

Aztreonam Tigemonam

Question 27

What is the active structure in penicillin?

Answer 27

Beta-lactam ring

Question 28

What do beta-lactams do?

Answer 28

Bind to penicillin binding proteins (PBPs) The bound PBPs are then unable to crosslink peptidoglycan chains Bacteria unable to synthesis cell wall and bacteria are lysed

Question 29

How does resistance to beta-lactams develop?

Answer 29

Mainly due to beta-lactamase enzymes

Question 30

What do beta-lactamases do?

Answer 30

Hydrolyse and break the beta-lactam ring, inactivating beta-lactam Can be overcome by adding beta lactamase inhibitors

Question 31

Give examples of beta-lactamase inhibitors

Answer 31

Tazobactam Clavulanic acid

Question 32

What are extended spectrum beta-lactamases (ESBLs)?

Answer 32

Carried on plasmids that have other resistance genes - results in multidrug resistance

Question 33

Give an example of an ESBL

Answer 33

Carbapenemases - they inactivate carbapenems, penicillins and extended spectrum cephalosporins

Question 34

What do efflux pumps do?

Answer 34

Transport proteins that can expel toxic substances from the cell

Question 35

Where are efflux pumps found?

Answer 35

In gram +ve and -ve species

Question 36

What are the 5 families of efflux transporters in bacteria?

Answer 36

Major facilitator (MF) Multidrug and toxic efflux (MATE) Resistance nodulation division (RND) Small multidrug resistance (SMR) ATP binding cassette (ABC)

Question 37

What is the genetic basis of antibiotic resistance in Neisseria gonorrhoeae?

Answer 37

Acquired genes Mutations Gene mosaics with or without superimposed mutations

Question 38

What mutations of Neisseria gonorrhoeae provide resistance to penicillin?

Answer 38

penA - PBP2 ponA - PBP1 penB - outer membrane porin mrR regulator - controls mtrCDE efflux pump operon penC - secretin PilQ of the type IV pilin

Question 39

What acquired genes in Neisseria gonorrhoeae provide resistance to penicillin?

Answer 39

plasmid-mediated TEM-1 penicillinase - blaTEM-1

Question 40

How are N. gonorrhoeae resistant to tetracycline?

Answer 40

Acquisition of plasmid-mediated tet(M) ribosomal protection protein A combination of mtrR, penB and rpsJ gene mutations

Question 41

What makes N. gonorrhoeae resistant to fluroquinolones?

Answer 41

Mutations in gyrA and parC in QRDR

Question 42

What makes N. gonorrhoeae resistant to cephalosporin?

Answer 42

Multiple alterations in penA (PBP2), often + mosaic gene exchange with other Neisseria species High level ceftriaxone resistance - point mutations superimposed upon mosaics

Question 43

What makes N. gonorrhoeae resistant to azithromycin?

Answer 43

Mutations to multiple 23S rRNA gene copies - single mutation + internal recombination (high level resistance)

Question 44

What gives N. gonorrhoeae low level resistance to azithromycin?

Answer 44

mtr mutations acquired mphA mefA-E (efflux pumps) ermABCF (ribosomal methylases)

Question 45

How can drug resistance phenotype be determined?

Answer 45

Culture based methods Molecular diagnostics Genome sequencing