CC10 Antimicrobial Modes of action & mechanisms of resistance

What are the requirements for antimicrobial activity

Adsorbs to cell surface

Passage into cell

Interacts with specific target e.g. ribosome/nucleoid

Disinfectant mode of action

Has a specific mode of action

multiple cellular targets

e.g.

oxidation

crosslinking

coagulation

disruption of structures

Describe oxidation

Disruption and breakage of chemical bonds in macromolecule

How does oxidation break bonds through DNA/RNA

Strand breakage, binding to DNA or RNA

Disrupts transcription, translation and replication

e.g. peroxide penetrates spores and reacts with DNA

How does oxidation occur through fatty acids

Reaction and degradation of unsaturated fatty acids in cell membranes

Disrupts membrane

Leakage of cytoplasmic contents

How does oxidation occur through disulfide bonds

Modification of S-S bonds

loss of protein structure

Disrupts enzyme function and results in cell death

Describe X-linking and coagulation

Interactions between macromolecules

such as clumping

leads to loss of function

X linking

Aldehydes cause the cross linking of lysine residues to other amino acids

This change in protein structure can cause protein aggregation (exposure of hydrophobic residues)

Cross linking of DNA/RNA/PROTEINS/PEPTIDOGLYCAN

Describe coagulation

Occurs due to macromolecule denaturation

Can lead to coagulation and precipitation

Chlorohexidine

Phenols

Ethanol

QAC’s

What is precipitation

disruption of cytoplasm, lipid membranes

Describe the mode of action for disruption of cellular structures

Disruption of functional structures

for example cell wall and cytoplasmic membrane

How does this disruption occur with reference to ions

+VE ions have affinity for -VE microbial membranes

Disrupts proton motive force

Disrupts cell-membrane associated activities e.g. energy generation

What is the proton motive force

A gradient of electrochemical charge across a membrane

Movement of protons across a membrane

Cells cannot function without

How can heavy metals disrupt cellular structures

Directly bind to and damage lipid membrane

Indirectly bind by charge to LPS (lipopolysacchride)

Describe the effect of concentration

At low concentrations:

• reversible enzyme inhibition

• permeability changes

structural damage

leakage

At high concentrations:

• Autolysis

• Lysis

• Cytoplasm coagulation

Describe the effect of increasing exposure time

Short time:

Bacteriostatic

Inability to repair

Long time:

Bactericidal- LETHAL

Biocides and cellular targets

Different biocides can target different cellular target

Can have multiple targets e.g. bacteria, fungi, viruses, protozoa, helmets

Outer membrane targets

• Agents: QACs, Mercury (Hg), Silver ions (Ag+), EDTA.

• Effect: Disrupts the outer membrane.

Cell wall targets

• Agents: Low concentration of phenol, formaldehyde, hypochlorite, EDTA.

• Effect: Damages the cell wall.

Cytosol targets

• Agents: Aldehyde, phenols, biguanides, QAC.

• Effect: Causes coagulation of cellular contents.

Ribosome targets

• Agents: Oxidizing agents, peroxygen.

• Effect: Damages ribosomes, impacting protein synthesis.

Protein targets

• Agents: Alcohols.

• Effect: Denatures proteins.

Coagulation in Cytoplasm targets

• Agents: High concentration chlorhexidine, phenol, mercury (Hg) salts.

• Effect: Leads to protein coagulation.

Membrane Proton Motive Force targets

• Agents: Parabens, phenols.

• Effect: Disrupts proton motive force, impacting energy production

Membrane Permeability targets

• Agents: Cetrimide, chlorhexidine, phenol, ethanol.

• Effect: Increases permeability, damaging the cell membrane.

Membrane associated activity targets

• Membrane ATPase: Chlorhexidine disrupts ATP synthesis.

• Electron Transport System: Hexachlorophane interferes with electron transport.

• Enzymes with -SH Groups: Mercury and silver disrupt these enzymes.

Nucleic acid targets

• Agents: Aldehyde, peroxygen, biguanides, phenols, QACs.

• Effect: Damages DNA/RNA, preventing replication and transcription.

What are the 5 mechanisms of resistance

Intrinsic/innate resistance : A natural chromosomal encoded property

Extrinsic/acquired resistance: organism becomes resistant

Phenotypic resistance: Response to mode of growth- reversible

Genetic resistance: Mutation or genetic transfer- irreversible

Co-resistance: multiple resistance genes

What factors affect innate resistance

Composition of outer layer

Efflux pumps

What are efflux pumps

Antimicrobial pumped out of cell via pumps in the cell wall

So the agent doesn’t reach optimum concentration

What are the types of efflux pumps

Chromosomal encoded pumps

which contributes a natural low level of resistance at all times

What is a biocide

a chemical, mixture, or microorganism that controls harmful organisms in a way that's not purely physical or mechanical

List the 3 mechanisms of innate resistance to biocides

• physical barrier to protection e.g. spores, waxy fatty acids, outer membrane, peptidoglycan

• Decreased accumulation of biocide through efflux pumps, degradation

• Absence of target/ metabolic pathway

What are the two types of acquired phenotypic resistance

• intracellular bacteria: A parasite inside protozoa, isolated = more resistant

• Biofilms: Physical barrier, cell-cell communication, 3D community, increased genetic exchange

What are the 2 acquired genetic resistance mechnaisms

• Chromosomal mutation/gene transfer

• Efflux

How do mutations affect resistance

• Changes proteins, fatty acids, phospholipids

• Changes O-chain lengths of LPS

• Changes number and size of porins

• Modifies target

• alters metabolic pathway

• Increases efflux

ALL CAN DECREASE EFFICACY OF ANTIMICROBIAL AGENT

How can Efflux affect

Creates widespread resistance of multiple solvents, detergents, antibiotics

Controlled by Multiple antibiotic resistance operon

mar A - activator

mar - repressor

How can we increase efficiency of pump

inactivate mar R

over express mar R

This up regulation can be produced by mutation

Describe multi-drug resistant plasmids

Different plasmids code for different resistance

Co-resistance

Selecting resistance for one antimicrobial selects resistance for every compound encoded by plasmid e.g. both antibiotics and biocides

How can we reduce development of resistance

What is meant by the ‘Chain of infection’

• Infectious agent

• Reservoir

• Portal of exit: the way the infectious agent leaves the reservoir

• Mode of transmission

• Portal of entry

• Susceptible host (any person in healthcare)

Disinfectant vs antimicrobial