Antibiotic Resistance

Mechanisms of Molecular Antibiotic Resistance

Objectives History of antibioticsDefinition of antibiotic resistance Why antibiotic resistance is a concern ?Why is it important?How antibiotics work ?Mechanisms of resistance to antibiotics ? Control

Introduction Throughout history there has been a continual battle between human beings and multitude of micro-organisms that cause infection and disease

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-1928: Penicillin first discovered by Alexander Fleming

-Chain and Florey, helped develop penicillin into a widely available medical product

History Nobel Lecture, December 11, 1945Sir Alexander FlemingThe Nobel Prize in Physiology or Medicine 1945

Antimicrobial agents were viewed as miracle cure when introduced into clinical practice. However it became evident rather soon after the discovery of penicillin that resistance develops quickly terminating the miracle. This serious development is ever present with each new antimicrobial agents and threatens end of antimicrobial area. Today even major class of antibiotics are resistant 4

Fleming himself warned of the danger of resistance It is not difficult to make microbes resistant to penicillin in the laboratory by exposing them to concentrations not sufficient to kill them, and the same thing has occasionally happened in the body and by exposing his microbes to non-lethal quantities of the drug make them resistant.

History Nobel Lecture, December 11, 1945Sir Alexander FlemingThe Nobel Prize in Physiology or Medicine 1945

Antimicrobial agents were viewed as miracle cure when introduced into clinical practice. However it became evident rather soon after the discovery of penicillin that resistance develops quickly terminating the miracle. This serious development is ever present with each new antimicrobial agents and threatens end of antimicrobial area. Today even major class of antibiotics are resistant 5

Antibiotic ResistanceDefined as micro-organisms that are not inhibited by usually achievable systemic concentration of an antimicrobial agent with normal dosage schedule and / or fall in the minimum inhibitory concentration (MIC) range.

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Mechanisms of action of antibiotics

Drug resistance occurs in :

BACTERIAANTIBIOTIC RESISTANCE Endoparasites VirusesResistance to antiviral drugs Fungi Cancer cellsDrug Resistance

Why resistance is a concern ? Resistant organisms lead to treatment failure .Increased mortality .Resistant bacteria may spread in Community.Low level resistance can go undetected. Added burden on healthcare costs .Threatens to return to pre-antibiotic era .

Antibiotic ResistanceSome microorganisms may born resistant, some achieve resistance by mutation or some have resistance thrust upon them by plasmids

Intrinsic ResistanceLack target : No cell wall; innately resistant to penicillin Innate efflux pumps:Drug blocked from entering cell or export of drug (does not achieve adequate internal concentration). Eg. E. coli, P. aeruginosa Drug inactivation: Cephalosporinase in Klebsiella

It occurs naturally.

Acquired resistanceMutations It refers to the change in DNA structure of the gene.Occurs at a frequency of one per ten million cells.Eg.Mycobacterium.tuberculosis,Mycobacterium lepra , MRSA. Often mutants have reduced susceptibility

Presence of few mutants not sufficient to produce resistance Single step : E.coli & staph to Rifampicin Multistep : erythromycin, tetracyclines, chloramphenicol

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Plasmids Extra chromosomal genetic elements can replicate independently and freely in cytoplasm.Plasmids which carry genes resistant ( r-genes) are called R-plasmids.These r-genes can be readily transferred from one R-plasmid to another plasmid or to chromosome.Much of the drug resistance encountered in clinical practice is plasmid mediated

Mechanisms of Resistance Gene Transfer Transfer of r-genes from one bacterium to anotherConjugationTransductionTransformationTransfer of r-genes between plasmids within the bacteriumBy transposonsBy Integrons

Transfer of r-genes from one bacterium to another

Conjugation : Main mechanism for spread of resistance The conjugative plasmids make a connecting tube between the 2 bacteria through which plasmid itself can pass.Transduction : Less common method The plasmid DNA enclosed in a bacteriophage is transferred to another bacterium of same species. Seen in Staphylococci , StreptococciTransformation : least clinical problem. Free DNA is picked up from the environment (i.e.. From a cell belonging to closely related or same strain.

The new DNA is then incorporated into the genome of the bacteria which becomes resistant.16

Mechanisms of Resistance Gene Transfer Transposons

Transposons are sequences of DNA that can move around different positions within the genome of single cell.

The donor plasmid containing the Transposons, co-integrate with acceptor plasmid. They can replicate during cointegration

Both plasmids then separate and each contains the r-gene carrying the transposon.

Eg ; Staphylococci,Enterococci

They have insertion sequence at end of gene.17

Mechanisms of Resistance Gene Transfer IntegronsIntegron is a large mobile DNA that can spread Multidrug resistance

Each Integron is packed with multiple gene casettes, each consisting of a resistance gene attached to a small recognition site.

These genes encode several bacterial functions including resistance and virulence.

They cannot promote self transfer

They are commonly associates with Transposons.

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Biochemical mechanisms of antibiotic resistance

Prevention of drug accumulation in the bacterium

Modification/protection of the target site

Use of alternative pathways for metabolic / growth requirements

By producing an enzyme that inactivates the antibiotic

Decreased permeability: Porin Loss

Interior of organismCell wallPorin channel into organismAntibiotic

Antibiotics normally enter bacterial cells via porin channels in the cell wall

Decreased permeability: Porin Loss

Interior of organismCell wallNew porin channel into organismAntibiotic

New porin channels in the bacterial cell wall do not allow antibiotics to enter the cells

Structurally modified antibiotic target site

Interior of organismCell wallTarget siteBindingAntibiotic

Antibiotics normally bind to specific binding proteins on the bacterial cell surface

Structurally modified antibiotic target site

Interior of organismCell wallModified target siteAntibiotic

Changed site: blocked binding

Antibiotics are no longer able to bind to modified binding proteins on the bacterial cell surface

Antibiotic inactivation

Interior of organismCell wallAntibiotic

Target siteBinding

Enzyme

Inactivating enzymes target antibiotics

Antibiotic inactivation

Interior of organismCell wallAntibiotic

Target siteBinding

Enzyme

Enzymebinding

Enzymes bind to antibiotic molecules

Antibiotic inactivation

Interior of organismCell wallAntibiotic

Target site

Enzyme

AntibioticdestroyedAntibiotic altered,binding preventedEnzymes destroy antibiotics or prevent binding to target sites

Timeline of Antibiotic Resistance

Strategy to Prevent antibiotic Resistance Develop new antibiotics. Appropriate use of the existing antibiotics Phage therapy. Use of the lytic enzymes found in mucus and saliva.

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References

-Medicinska Mikrobiologija Sa Imunologijom i Parazitologijom-Mirsada Hukic -http://en.wikipedia.org/wiki/Scientific_method-http://www.slideshare.net/nasertadvi/antibiotic-resistance-14709382?related=2-http://www.who.int/drugresistance/amr_q&a.pdf-http://www.powershow.com/view4/53f288-NDBiN/ANTIMICROBIAL_RESISTANCE_powerpoint_ppt_presentation

They are commonly associates with Transposons.

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Worked by :

Adnan Fojnica