SUPERBUGS: MECHANISMS, CONTROL & UTILIZATION

Presented by:Noor-ul-Ain

WHAT ARE SUPERBUGS?

• Bacteria which have acquired increased resistance towards the antibiotic class used for their treatment

• Multi-drug resistance acquired by bacteria through various mutations which enhance its morbidity and mortality levels

Two type of resistance offered by bacteria:• Innate Resistance• Acquired ResistanceAcquired resistance may be through:• Chromosomal mutations• Gene Transfer among strains

Resistance Mechanisms Adopted by Superbugs

• Modification/ Inactivation of Antibiotics

• Target Site Modification

• Membrane Permeability & Efflux Pumps

Resistance Mechanisms Adopted by Superbugs

1. Modification/ Inactivation of Antibiotics• Three enzymes involved areβ-lactamases, Chloramphenicol acetyltransferases and Aminoglycoside-modifying enzymes• Transferases work by binding phosphoryl, adenylyl or acetyl

groups to the antibiotic molecule • Aminoglycoside modifying enzymes reduce affinity of a modified

molecule and hinder binding to the 30S ribosomal subunit • β-lactamases break open β-lactam ring through hydrolysis

containing ester and amide linkages

Resistance Mechanisms Adopted by Superbugs

2. Target Site Modification• Fluoroquinolone resistance linked to the mutation among genes responsible for

encoding the target sites• Genes undergo mutation at Quinolone Resistance-Determining Region• Mutations in genes amino acid substitutions modify the target

protein structure fluoroquinolone-binding affinity of the enzyme decreases• For instance, Amino acid substitution may include: substitution at serine 83 with Leucine in GyrA for P. stuarti substitution at serine 80 with Arginine in ParC for K. pneumoniae

Resistance Mechanisms Adopted by Superbugs

2. Membrane Permeability & Efflux Pumps

• Double membrane structure of gram negative bacteria resists the uptake and transfer of drug

• Certain strains acquire such genes which produces altered bacterial cell walls

• Efflux pumps pump the antibacterial agent out of the cell • Bacteria which are resistant towards tetracycline secrete membranous

proteins which act as efflux system of antibiotics

Resistance Mechanisms Adopted by Superbugs

Resistance Mechanism in Clostridium difficile

C. difficile produces actin-ADP–ribosylating toxin (C. difficile transferase)

CDT adds ADP ribose to the actin protein, causes actin depolymerization

CDT also produces Microtubule-based protrusions

Actin depolymerization increases secretion of fibronectin (ECM Protein)

Changes in intracellular calcium level occurs

Increased concentration of ECM protein & microtubule based protrusions forms a meshwork at host cell surface

C. difficile is adhered tightly to the host cell surface

Salmonella enterica confers resistance by the emergence of qnrS genotype with increased mobility

Resistance Mechanism in Salmonella enterica

Qnr proteins provide resistance to the strain by protecting DNA-gyrase from quinolones

Strains carrying qnr alleles can withstand elevated concentrations of fluoroquinolones

Plasmid-mediated Quinolone Resistance, increased possibility of transfer to other strains

MRSA CM05 strain confers resistance against linezolid

Resistance Mechanism in MRSA

Resistance caused by qnr gene which releases Cfr methyltransferase

Enzyme leads to the modification in adenosine located at position 2503 in 23S rRNA

Cfr adds extra methyl group to A2503

Leads to changes in molecular conformation

Ways to Combat Superbugs

Ways to Combat Superbugs

1. Use of Nanoshuttles

• Nanoparticles used for efficient delivery of drug into the cell

• Increase potential of therapeutic treatment

• Reduce the concentration of antibiotic into the surrounding serum

• For example, ciprofloxacin loaded zinc doped hydroxyapatite increased

antimicrobial activity against Staphylococcus aureus

2. Use of Filamentous Phage

Ways to Combat Superbugs

• Combination of certain drug resistant strains with filamentous phage resulted in resensitization of previously

resistant strains• Filamentous phage is secreted through aqueous channels

in outer membrane • Phage protein pIV creates channels with high conductivity • Opening of pIV channels results in increased susceptibility

of host bacteria

2. Use of Filamentous Phage

Ways to Combat Superbugs

3. Antimicrobial peptides as Anti-MRSA agents

Ways to Combat Superbugs

• Antimicrobial assays of natural antimicrobial peptides isolated demonstrated the inhibition of Staphylococcal growth

• Novel peptides can also be designed which have significant antibacterial activity against bacterial strains

• A unique peptide sequence was designed i.e. DFTamP1 (1K, 2G, 2S, and 8 L) through database filtering technology

• DFTamP1 inhibited MRSA USA300 because of its high hydrophobicity and low cationicity

Ways to Combat Superbugs

Database Filter Technology

Ways to Combat Superbugs

Anti-MRSA strategy of designed peptide sequence

POTENTIAL USE OF SUPERBUGS

Superbugs for Bioremediation

• Research today is focused on Design of superbugs with modified degradative genes through genetic

engineering OR Discovery of naturally tolerant bacteria found in contaminated sites and

their utilization for bioremediation • Hybrid strain of Pseudomonas putida was engineered by replacing gene

encoding for bphA1 with the gene encoding for toluene dioxygenase (todC1)

• Hybrid strains were found to have enhanced ability to grow on wide variety of hydrocarbons, also had the ability to degrade xenobiotic compounds

• Genetically modified superbugs can play a significant role in improving the power output efficiency of fuel cells

• Superbugs are designed to over-express genes can increase the prospect of electron flow to an electrode

• Certain bacteria secrete redox-active mediators which can transfer electrons to conductive surfaces

• High flow rates require large biofilm surface area and increased biofilm metabolic rate

Superbugs for Bioremediation

Superbugs for Bioremediation

Convective-flow membrane-less MFC with dual anodes and dual cathodes for wastewater treatment