Evolution of Beta Lactamases Specificity: Carbapenemases and Beta-lactamase inhibitors

Question 1

Name the Class A Beta=lactamases

Answer 1

BlaZ (S. Aureus, penicillinase)
TEM, SHV, CTX-M (ESBLs, enterobacteriaeceae
KPC (Carbapenamse: eneterobacteriaceae)

Question 2

Name the Class C Beta-lactamases

Answer 2

AmpC, CMY

Question 3

Name the Class D Beta-lactamases

Answer 3

OXA (carbpaenase resistance in A. Baumaani and Enterbacteriaceae)

Question 4

ame the Class B beta-lactamases

Answer 4

IMP, VIM (found in non-fermenting bacteria e.g. P, aueginosa)
NDM (in enterbacteriaceae)

These are generally broach spectrum including carbapenemases and whislt most are chromosomal there are the above worrying examples on plasmids

Question 5

Describe teh structure of pencillins

Answer 5

All have the beta lactam ring but fused to a 5 member S containing rign

Question 6

Describe the structure of cephalosporins

Answer 6

Beta lactam ring fused to a 6 member S containing ring and double bond

Question 7

Describe the structure of carbapenems

Answer 7

Beta lactam ring fused to a 6 member S containing ring with a double bond

Question 8

Descrbie the structure of monobactams

Answer 8

Beta-lactam ring but not fused to anything

Question 9

Describe the structure of Clavulanate (inhibitr)

Answer 9

Bi-cyclic beta lactam containing molecule reminescent to pencillins

Poor antimicrobila activity but good inhibitor action

Question 10

What has happened to resistance spectrum in Class A beta-lactamses

Answer 10

Resistance has been progressively increasing particularly in class A
Carbapenamase activity has even been seen now

Question 11

Summarise evolution of beta-lactams in response to b-lactamases

Answer 11

1) Penicillins produced –> only had gram+ve activity and BlaZ conveyed resistance
2) Aminopenicillins made –> meant that had activity against gram -ves. But then TEM and SHV conveyed resistance
3) Cephalosporins introduced –> BUT TEM and SHV mutations conveyed resistance them
4) Oxy-amino 3rd Generation cephalosporins introduced to combat TEM and SHV resistance –> mutations in TEM and SHV to convey resistance . Also emergenec of CTX-M

Question 12

What is the solution to Class A Beta Lactamases

Answer 12

Combination therapy with inhibitors!

Clavulanic acid, tazobactam, sulbactam

Question 13

How do beta-lactamase inhibitors work

Answer 13

They have a bicylcic- beta lactam structure hence from an acyl-enzyme species by reacting with the active site serine in the Beta-lactamases

Enzyme cant break down the same was as a beta lactam

Get a range of fragmentation reactions – > throguh complex intermediates the result is a covalently modified species where the active site serine has reacted with a fragmentation product

The beta lactamase is permantily nhibited

Question 14

Resistance to beta-lactamse inhibitors in TEM and SHV has arisen how

Answer 14

244 - mutation inactivates the water moleule that catalyses the fragmentation reactoin

130 - mutation modifies the active site structure to improve inhibitor binding

HOWEVER the mutations conveying resistance to inhibitors are mutually exclusive to those conveying ESBLs

Question 15

What are the mecahnisms of carbapenem resistance

Answer 15

1) Permeability modification & weak enzymes
2) Acquired serine carbapenemases e.g. KPC and OXA48
3) Metallo-B-Lacatmases: IMP, VIM, NDM

Question 16

How can permeability modification and weak enzymes convey carbapenem resistance

Answer 16

Reduction in permeability with a weak enzyme can be enough to convey resistance without the enzymes strictly being a carbapenemase. E.g. ESBL/ AmpC

P. Aureginosa is an example of this: Loses OprD porin and combo with a weak enzyme can provide resistanct

Question 17

What is the KPC carried in

Answer 17

Acquired serine carbapenemase on Tn 4401b composite transposon
Only carried on a few restricted strains of Klebsiella
Global enzymes –> particular problem in Northern England

Freeuqnelty get multirestance

Question 18

What is the normal action of carbapenems

Answer 18

Carbapenems have a hydroxyethyl substituent unique to them
They can acylate and inhibit both PBP and Serine Beta-Lactamases to cause their effects –> the OH bond on the carbapenem forms a H bond to water, hence deactivating it and preventing it from deacylating and inactivating carbapenems

Question 19

How does KPC convey carbapenem resistance

Answer 19

Active site modification in the B-lactamase –> binds carbapenem in a slightly different orientation –> the hydroxyl carbapenem group is too far from the water to form a H bond –> hence water can still deacylate the beta lactam ring and hence degrades the antibiotics

Question 20

name inhibitors that can target KPC

Answer 20

Avibactam

Vaborbactam

Question 21

How does avibactam work

Answer 21

Has a beta-lactam like structure
Reacts with the serine of beta-lactamases reversibly to form acyl intermediate enzymes that cant be hydrolysed
Active against class A and class C

Question 22

How does vaborbactam work

Answer 22

It is a cyclic boronate inhibitor of class A and C
Has a Beta-lactam like structure
Contains a boron and carboxylic acid –> can form an acyl enzyme to the active site serine attached to the Boron
The Boron is conducive to a tetrahedral sturcuter –> mimics the tetrahedral transition state
Approved in combo with eropenem

Question 23

What is OXA48

Answer 23

It is an acquired Class D serine carbapenemase
Causing increased numebrs of carbapenem resistance in the UK
May overtake KPC as the primary method of carbapenem resistant plasmid carried

Question 24

What are the most common MBLs

Answer 24

NDM and VIM

Question 25

Where is NDM found

Answer 25

Particularly found in clinical strains of enterbacteriaceae e.g. Klebsiella –> however NOT restcited to a few strains unliike KPC and is present on many plasmid types

Question 26

Where is VIM 2 found

Answer 26

Important in non-fermenting bacteria e.g. P. auerginosa

Question 27

What p. eurginosa clone is associated with VIM B-lactamses

Answer 27

ST235 strain !!

Question 28

How can we tackle this carbapenem resistance

Answer 28

There was the option of avibactam and vaborbactam inhibitors
HOWEVER CLass D are insusceptible to these inhibitors
Colisitn is a last resort but is effective against many MBL producers

Question 29

What can MBLs hydrolyse

Answer 29

Virtually anything with a beta-lactam ring