Question

We are fighting what seems to be a losing battle against antibiotic-resistant Gram negative bacteria, such as “beta-lactamase-producing Enterobacteriaeceae” (BLPE). BLPE seemed to arise all of a sudden in response to the common use of penicillin to treat infections caused by Enterobacteriaeceae.

A. BLPE are resistant to all of the following antibiotics. For each, state the antibiotic’s mode of action, and explain the most likely resistance mechanism. Ceftriaxone Tetracycline Aminoglycosides Vancomycin

B. If we only overused Penicillin, how could that lead suddenly to the development of resistance to all of these different antibiotics, especially Tetracycline and Aminoglycosides?

C. BLPE can still be treated with Polymyxins. How do Polymyxins work, and why would physicians be reluctant to use them in a patient unless absolutely necessary?

Answer 1

Introduction:

What are BPLE?

Enterobacteriaceae are gram negative bacteria which include E.coli, Y.pestis, Klebsiella, Shigella, Proteus, Serratia etc. Mostly E.coli, Klebsiella produce Beta lactamase (also called as Penicillinase) which can hydrolyse the Beta-lactam ring in penicillins. Thus the antibiotics become ineffective. BPLE now became a major concern due to their resistance to many antibiotics.

Answer for A:

A. Mechanism of action of Ceftriaxone: Ceftriaxone acts similar to penicillins by inhibiting the cell wall synthesis in the susceptible organisms. The main difference lies with the targeted proteins in the cell wall, while Penicillins bind to the Penicillin binding proteins (PBP), ceftriaxone similar but different proteins. Eventually cell wall deficient forms of bacteria are developed, which swell, burst due to hyperosmosis (cytolysis occurs that leads to cell death)

Possible resistance mechanism: Possible causes of resistance would be due to i. alteration in target proteins in the cell wall which reduces affinity for ceftriaxone, ii. impermeability to the antibiotic or its efflux from the bacterium that reduces its concentration at site of action and iii. Elaboration of the Beta lactamase enzyme(Cephalosporinases- which cleaves the cephalosporin ring in Ceftriaxone) which destroy the ceftriaxone

Mechanism of action of Tetracycline: Tetracyclines are broad spectrum antibiotics which are primarily bacteriostatic and inhibit the protein synthesis in the microorganisms by binding to 30s subunit of ribosomes. As a result of this, aminoacyl t-RNA to the mRNA-ribosomal complex is interfered which results in failure of growth of peptide chain in the susceptible microbes

Possible resistance mechanism: E.coli, B-lactamase producing enterobacteriaceae develop resistance to tetracyclines by reducing its concentration through efflux. Another possible mechanism is plasmid mediated protection protein which offers protection against tetracycline by safegaurding the ribosomal binding site.

Mechanism of action of Aminoglycosides: Aminoglycosides also inhibit the protein synthesis in bacteria but by binding to 50s subunit of ribosomes (except streptomycin which binds to 30s subunit). They prevent the formation of polysomes by disaggregating them into monosomes. Thus only one ribosome is attached to each strand of mRNA.

Possible resistance mechanism: i. By acquisition of cell membrane bound inactivating enzymes which phosphorylate, adenylate or acetylate the enzyme. ii. High degree of resistance is confered by a mutation which can decrease the affinity of ribosomal proteins that normally bind with the aminoglycosides iii. Decreased efficiency of the aminoglycoside transporting mechanism. Pores in the outer coat of bacterium become less permeable or the active transport is inhibited.

Mechanism of action of Vancomycin: Vancomycin inhibits the bacterial cell wall synthesis by binding to the terminal dipeptide D-Ala-D-ala sequence of peptidoglycan units and its release from bactoprenol lipid is prevented which results in failure of cross linking

Possible resistance mechanism: Beta lactamase producing Enterobacteriaceae develop resistance by plasmid mediated alteration of the dipeptide target site, which reduces the affinity of vancomycin.

Answer for B.

As seen from the above discussion Ceftriaxone, Penicillin and Vancomycin all act by disrupting the cell wall synthesis while Tetracycline and Aminoglycosides act by inhibiting the protein synthesis.

BLPE should only be having resistance towards Ceftriaxone, Penicillin and Vancomycin in theory, but unfortunately they also developed resistance even to different mechanism of actions shown by antibiotics.

Conjugation mediated transfer, and Transformation mediated dissemination of resistance can be attributed as one of the main reasons for wide spread of BLPE.

Answer for C:

Polymyxins- Mechanism of action: Polymyxins are bactericidal agents with detergent like action on cell membrane. Polymyxins have high affinity for phospholipids, and orient in between phospholipids of cell wall and protein layers which causes membrane disruption and pseudopore formation. This results in leakage of ions, amino acids. The bacterial endotoxins are also said to be inactivated.

Polymyxins show synergistic effect with other antibiotics and improve their permeability.

Why physician are reluctant to use polymyxins?

Polymyxins are very less absorbed by oral route. When injected the systemic toxicity is high with symptoms of flushing and paresthesias (occurs due to release of histamine from mast cells), marked nephrotoxicity, neurological disturbances and neuromuscular blockade.

Hence, even though their action is high as expected in theory, they are equally damaging to the host. Hence, they are less used in practice unless absolutely necessary