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Resistance to anti-microbial agents

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Anti-microbial resistance has become a world health issue today. Therefore it is imperative to know about the methods of acquiring resistance and ways to deal with the situation and prevent resistance.

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Resistance to anti-microbial agents

  1. 1. Antimicrobial Resistance By: Dr. Manjeeta Gupta 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 1
  2. 2. •History •Introduction •What is antimicrobial resistance •Why antibacterial resistance is a concern •Mechanisms of resistance •NDM-1 •Factors causing antimicrobial resistance •Present status of development of antimicrobials •Strategies to contain resistance 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 2 Objective s
  3. 3. In his 1945 Nobel Prize lecture, Fleming himself warned of danger of resistance – “It is not difficult to make microbes resistant to penicillin in laboratory by exposing them to concentrations not sufficient to kill them, and same thing has occasionally happened in body… …and by exposing microbes to non-lethal quantities of drug make them resistant.” History (Nobel 1945) Sir Alexander Fleming 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 3
  4. 4. • Magic Bullets (Miracle cures) •Present scenario Resistance to every major class of antimicrobial agents •Golden age  End of Antimicrobial Era Introduction 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 4
  5. 5. 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 5 “Concentration of drug at site of infection must inhibit organism & also remain below level that is toxic to human cells.“ Antimicrobial resistance
  6. 6. 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 6 Resistance in malaria The emergence of P. falciparum multidrug resistance, including resistance to ACTs, in the Greater Mekong subregion is an urgent public health concern that is threatening the ongoing global effort to reduce the burden of malaria. Routine monitoring of therapeutic efficacy is essential to guide and adjust treatment policies. It can also help to detect early changes in P. falciparum sensitivity to antimalarial drugs. Resistance in HIV HIV drug resistance emerges when HIV replicates in the body of a person infected with the virus who is taking antiretroviral drugs. Even when antiretroviral therapy (ART) programmes are very well-managed, some degree of HIV drug resistance will emerge. Available data suggest that continued expansion of access to ART is associated with a rise in HIV drug resistance. In 2013, 12.9 million people living with HIV were receiving antiretroviral therapy globally, of which 11.7 million were in low- and middle-income countries. HIV drug resistance may rise to such a level that the first-line and second-line ART regimens currently used to treat HIV become ineffective, jeopardizing people’s lives and threatening national and global investments in ART programmes. As of 2010, levels of HIV drug resistance among adults who had not begun treatment in countries scaling up ART were found to be about 5% globally. However, since 2010, there are reports suggesting that pre-treatment resistance is increasing, peaking at 22% in some areas. Continuous surveillance of HIV drug resistance is of paramount importance to inform global and national decisions on the selection of first and second-line ART and to maximize overall population level treatment effectiveness. Resistance in influenza Over the past 10 years, antiviral drugs have become important tools for treatment of epidemic and pandemic influenza. Several countries have developed national guidance on their use and have stockpiled the drugs for pandemic preparedness. The constantly evolving nature of influenza means that resistance to antiviral drugs is continuously emerging. By 2012, virtually all influenza A viruses circulating in humans were resistant to drugs frequently used for the prevention of influenza (amantadine and rimantadine). However, the frequency of resistance to the neuraminidase inhibitor oseltamivir remains low (1-2%). Antiviral susceptibility is constantly monitored through the WHO Global Surveillance and Response System. Present situation (Updated April 2015) Resistance in bacteria WHO’s 2014 report on global surveillance of antimicrobial resistance revealed that antibiotic resistance is no longer a prediction for the future; it is happening right now, across the world, and is putting at risk the ability to treat common infections in the community and hospitals. Without urgent, coordinated action, the world is heading towards a post-antibiotic era, in which common infections and minor injuries, which have been treatable for decades, can once again kill. •Treatment failure to the drug of last resort for gonorrhoea – third-generation cephalosporins – has been confirmed in several countries. Untreatable gonococcal infections result in increased rates of illness and complications, such as infertility, adverse pregnancy outcomes and neonatal blindness, and has the potential to reverse the gains made in the control of this sexually transmitted infection. •Resistance to one of the most widely used antibacterial drugs for the oral treatment of urinary tract infections caused by E. coli – fluoroquinolones – is very widespread. •Resistance to first-line drugs to treat infections caused by Staphlylococcus aureus – a common cause of severe infections acquired both in health-care facilities and in the community – is also widespread. •Resistance to the treatment of last resort for life-threatening infections caused by common intestinal bacteria – carbapenem antibiotics – has spread to all regions of the world. Key tools to tackle antibiotic resistance – such as basic systems to track and monitor the problem – reveal considerable gaps. In many countries, they do not even seem to exist. Resistance in tuberculosis In 2013, there were an estimated 480 000 new cases of MDR-TB in the world. Globally, 3.5% of new TB cases and 20.5% of previously treated TB cases are estimated to have MDR-TB, with substantial differences in the frequency of MDR-TB among countries. Extensively drug-resistant TB (XDR-TB, defined as MDR-TB plus resistance to any fluoroquinolone and any second-line injectable drug) has been identified in 100 countries, in all regions of the world.
  7. 7. 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 7
  8. 8. 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 8 ICMR programme on Antibiotic Stewardship, Prevention of Infection & Control (ASPIC) Indian J Med Res. 2014 Feb; 139(2): 226–230. Combating Antibiotic-Resistant Bacteria—issued by President Barack Obama on September 18, 2014 — National Action Plan FDA is partnering with Center for Disease Control and Prevention (CDC) on "Get Smart: Know When Antibiotics Work." In November 2012 an important paper from India was published that was ‘Chennai Declaration’. J Antimicrob Chemother doi:10.1093/jac/dkt062.
  9. 9. 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 10 Selection pressure
  10. 10. 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 11 Mechanism of Resistance ”Some microorganisms are ‘born’ resistant, Some ‘achieve’ resistance by mutation and Some have resistance ‘thrust upon them’ by plasmids.”
  11. 11. Intrinsic Resistance 1. Lack target : M. tuberculosis resistant to common antibiotics 2. Innate inability to cross outer membrane or bind to target: E. coli, P. aeruginosa 3. Drug inactivation: Cephalosporinase in Klebsiella 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 12
  12. 12. 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 13
  13. 13. Acquired resistance 1. Mutation (Vertical transfer) • Occurs in 1/10 million cells • Single or multiple step 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 14
  14. 14. 2. Plasmid mediated (Horizontal transfer) • Most common • Extra chromosomal genetic elements can replicate independently & freely in cytoplasm • R-plasmids Carry resistant genes ( r-genes) 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 15
  15. 15. Mechanisms of Resistance Gene Transfer  Conjugation  Transduction  Transformation  Transposons  Integrons 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 16
  16. 16.  Conjugation : (Main mechanism for spread of resistance) Conjugative plasmids makes a connecting tube between 2 bacteria through which plasmid itself can pass  Transduction : (Less common) Plasmid DNA is enclosed in bacteriophage  transferred to another bacterium of same species. Seen in Staphylococci , Streptococci  Transformation : (Least common) Free DNA is picked up from environment 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 17
  17. 17.  Insertion sequences of mobile DNA  Cannot self replicate  Resistance gene can ‘hitch hike’ 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 18 Transposons
  18. 18.  Large DNA packed with multiple gene cassettes  Located within transposon, plasmid or mobile  DO NOT SELF REPLICATE  Encode as Integrase provide specific site for gene cassettes integration  Multidrug resistance & virulence06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 19 Integrons
  19. 19. • When subset of total microbial population is resistant, despite total population being considered susceptible on testing • Vancomycin in S. aureus, E. faecium Anti-tubercular drugs in TB Fluconazole in Cryptococcus neoformans, Candida albicans 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 20 Hetero- resistance
  20. 20. • Viral replication: more error prone •Viral evolution under drug & immune pressure: relatively easy •Quasi species (drug resistant subpopulations) •Failure of antiretroviral therapy 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 21 Viral quasi species
  21. 21. • Ability to protect genetic information & allow changes by causing defects in repair mechanisms: •Insertion of correct base pair by DNA polymerase III •Proof reading by polymerase •Post replicative repair  Adaptation & Emergence of multidrug-resistant strains of M. tuberculosis 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 22 Mutator (Mut) phenotypes Hypermutable phenotypes
  22. 22. • Prevention of drug accumulation in bacteria • Modification of target site • Use of alternative pathways for metabolic / growth requirements • By producing an enzyme that inactivates antibiotic • Quorum sensing 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 23 Biochemical mechanisms
  23. 23. Interior of organism Cell wall Porin channel Antimicrobial Antimicrobials normally enter bacterial cells via porin channels in cell wall 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 24 Decreased permeability: Porin Loss
  24. 24. Interior of organism Cell wall New porin channel Antimicrobial New porin channels in the bacterial cell wall do not allow Antimicrobials to enter cells 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 25
  25. 25. • Cytoplasmic membrane transport proteins 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 26 Efflux pumps
  26. 26. 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 27 Interior of organism Cell wall Target siteBinding Antimicrobial Antimicrobials normally bind to specific binding proteins on bacterial cell surface Structurally modified antimicrobial target site
  27. 27. 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 28 Interior of organism Cell wall Modified target site Antimicrobial Changed site: blocked binding Antimicrobials are no longer able to bind to modified binding proteins on bacterial cell surface
  28. 28. 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 30 Interior of organism Cell wall Antimicrobial Target siteBinding Enzyme Inactivating enzymes target Antimicrobial Antimicrobial inactivation
  29. 29. 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 31 Interior of organism Cell wall Antimicrobial Target siteBindingEnzyme Enzyme binding Enzymes bind to Antimicrobial molecules
  30. 30. 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 32 Interior of organism Cell wall Antimicrobial Target siteEnzyme Antimicrobial destroyed Antimicrobial altered, binding prevented Enzymes destroy antimicrobials or prevent binding to target sites
  31. 31. •Microbes communicate with each other & exchange signals (Autoinducers) •Single autoinducer Incapable of induction •When its colony reaches a critical density (quorum), threshold of auto-induction is reached gene expression •Allows bacteria to coordinate gene expression for virulence, conjugation, apoptosis, mobility & resistance 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 33 Quorum sensing
  32. 32. • Some of the best-known examples of quorum sensing come from studies of bacteria. Bacteria use quorum sensing to coordinate certain behaviors such as biofilm formation,virulence, and antibiotic resistance, based on the local density of the bacterial population. Quorum sensing can occur within a single bacterial species as well as between diverse species, and can regulate a host of different processes, in essence, serving as a simple indicator of population density or the diffusion rate of the cell's immediate environment. A variety of different molecules can be used as signals. Common classes of signaling molecules are oligopeptides in Gram-positive bacteria, N-Acyl Homoserine Lactones (AHL) inGram- negative bacteria, and a family of autoinducers known as autoinducer-2 (AI-2) in both Gram-negative and Gram-positive bacteria.[1] • Bacteria that use quorum sensing constitutively produce and secrete certain signaling molecules (called autoinducers or pheromones). These bacteria also have a receptor that can specifically detect the signaling molecule (inducer). When the inducer binds the receptor, it activates transcription of certain genes, including those for inducer synthesis. There is a low likelihood of a bacterium detecting its own secreted inducer. Thus, in order for gene transcription to be activated, the cell must encounter signaling molecules secreted by other cells in its environment. When only a few other bacteria of the same kind are in the vicinity, diffusion reduces the concentration of the inducer in the surrounding medium to almost zero, so the bacteria produce little inducer. However, as the population grows, the concentration of the inducer passes a threshold, causing more inducer to be synthesized. This forms a positive feedback loop, and the receptor becomes fully activated. Activation of the receptor induces the up-regulation of other specific genes, causing all of the cells to begin transcription at approximately the same time. This coordinated behavior of bacterial cells can be useful in a variety of situations. For instance, the bioluminescent luciferase produced by Vibrio fischeri would not be visible if it were produced by a single cell. By using quorum sensing to limit the production of luciferase to situations when cell populations are large, V. fischeri cells are able to avoid wasting energy on the production of useless product. 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 34
  33. 33. 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 35
  34. 34. Antimicrobials Mechanisms Effect Beta lactams (Penicillins, Cephalosporins, Carbapenems) Altered high mol. wt. PBP (MecA gene) Altered no. & size of porins Active efflux pumps β lactamase enzyme (Gram +veInducible, Gram –veConstitutive) ↓ affinity ↓ permeability Efflux of drug Cleavage of β lactam ring Inactivation Quinolones Altered DNA gyrase (gyrA gene, gyrB gene) Altered Topoisomerase IV (parC gene, parE gene) Efflux Pumps ↓ Affinity Efflux of drug Aminoglycosides (Gentamicin, Streptomycin) Drug modifying enzymes (acetylase, phosphorylase & adenylase) Altered ribosomal structure Drug inactivation ↓ permeability (Gram +ve & Gram –ve) Antibacterial agents 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 36
  35. 35. Antimicrobials Mechanisms Effect Chloramphenicol Acetyltransferase enzyme Altered ribosomal target Inactivation (Gram -ve constitutive, Gram +ve inducible) ↓ permeability & binding Macrolides (Azithromycin, Erythromycin) Esterases (Enterobactericeae) Altered ribosomal target (ermA, B, C gene) Efflux pumps (mefA, msrA, mefE gene) Hydrolysis ↓ binding ↓ drug concentration 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 37 Tetracyclines Efflux pumps (tetK gene) Altered ribosomal target (tetM gene) Drug modifying enzymes (tetX) ↓ drug concentration ↓ binding Inactivation
  36. 36. Sulphonamides Point mutations in DHPS gene Altered metabolic pathway ↑ efflux Overproduction of PABA ↓ affinity & ↓ bacterial permeability ↓ inhibition of DHPS ↓ drug concentration ↓ drug effect 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 38 Antimicrobials Mechanisms Effect Lincosamide (Clindamycin) Streptogramins (Quinupristin, Dalfopristin) Altered ribosomal target (erm gene) Altered ribosomal target (ermA, B, C gene) Lactonases (vgbB gene) Acetyltransferases (vatB, C, D, satA gene) Efflux pumps (msrA gene) ↓ binding ↓ binding Inactivation ↓ Drug concentration Glycopeptides (Vancomycin) Altered target (D-alanyl-D-alanine to D-alanyl-D- lactate/serine) ↓ binding
  37. 37. Antimicrobials Mechanisms Effect Pyrazinamide Point mutation (pncA gene) ↓ affinity of Pyrazinamidase Isoniazid (1 in 106 bacilli) Mutation (katG/kasA gene) Overexpression of inhA gene & ahpC promoter gene Induction of efflux pumps Inactivation ↓ binding of INH-NAD inhibitor ↓ killing ↓ drug concentration Ethambutol Mutation (embB gene) ↑ activity of efflux pumps Inactivation ↓ drug concentration Streptomycin Mutation (rpsL & rrs, gidB gene) ↑ activity of efflux pumps Inactivation ↓ drug concentration Anti tubercular agents Rifampicin (1 in 107 to 108 bacilli) DNA polymerase target altered (rpoB gene) Induction of dnaE2 gene ↓ binding Error prone DNA repair 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 39
  38. 38. Antimicrobials Mechanisms Effect Atovaquone Single point mutation (cyt b gene) (mitochondrial chromosome) Inhibits binding of drug to cyt bc1 complex Pyrimethamine Proguanil Multiple mutation in plasmodium DHFR gene ↓ binding affinity Chloroquine (common) Mutation (K76T) in pfCRT polymorphic gene Efflux of drug, ↓ MOA Mefloquine Quinine Gene amplification & point mutation of pfmdr1 Mutation of PfNHE Antimalarial agents 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 40
  39. 39. Antimicrobials Mechanisms Effect Benznidazole Mutation in β tubulin gene ↓ expression of NADH dependent mitochondrial nitroreductase ↓ affinity for β tubulin No activation of drug Ivermectin Mutation in ATP dependent P-glycoprotein, Glutamate/GABA- gated Cl - channels Loss of MOA Antihelminthic agents 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 41 Metronidazole ↓ transcription of the Ferredoxin gene ↑ expression of SOD ↑ expression of nim (resistance) genes Loss of function mutations in NADPH nitroreductase (rdxA gene) Low levels of PFOR Impaired O2 scavenging capability No formation of reactive nitroso group
  40. 40. 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 42 Miltefosine Point mutation in P-type ATPase (aminophospholipid translocase subfamily) ↓ drug uptake Pentamidine Melarsoprol Point mutation of P2 transporter Mutation of HAPT1 transporter Loss of transporter, no uptake of drug Antimicrobials Mechanisms Effect
  41. 41. Antifungal agents Flucytosine Loss of Permease ↓ activity of UPRTase/cytosine deaminase ↓ transport ↓ conversion to 5- FUMP Azoles Mutation (ERG11 gene) (codes for 14 α demethylase) & ERG3 (C5,6 sterol reductase) Efflux pumps (ABC) ↑ production of 14 α sterol demethylase ↓ binding, cross resistance ↓ drug concentration ↓ effect Antibiotic Mechanisms Effect 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 43 Amphotericin B Replaces Ergosterol with other precursor sterols ↓ binding Echinocandins Mutation of FKS1 gene (essential component of 1,3-β-D-glucan synthase complex) ↓ MOA
  42. 42. Antimicrobials Mechanisms Effect Lamivudine Mutation of HBV DNA polymerase No MOA Zidovudine NNRTIs Stavudine NRTIs Altered target site Multiple mutation of RT gene (TAMs) Extrusion of nucleoside analogue Promote excision of incorporated nucleotide by, cross resistance Protease inhibitors Multiple mutation of HIV protease gene No MOA & cross resistance Maraviroc Shift in tropism from CCR5  CXCR4 Specific mutation in V3 loop of gp120 ↑ IC50 & ↓ maximum % inhibition of virus replication Enfuvirtide Integrase inhibitors Specific mutation at binding domain of gp41 Mutation of integrase gene ↑ IC50 & Cross resistance Anti retroviral agents 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 44 Entry inhibitors
  43. 43. Acyclovir Ganciclovir Penciclovir Famciclovir Impaired production of thymidine kinase Altered thymidine kinase substrate specificity Altered viral DNA polymerase No MOA 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 45 Antimicrobials Mechanisms Effect Antiviral agents Cidofovir Foscarnet Point mutation in viral DNA polymerase No MOA, cross resistance Amantadine Rimantidine Mutation in RNA sequence encoding M2 protein No MOA Oseltamivir Zanamivir Neuraminidase mutation No MOA Adefovir Point mutation in HBV polymerase No MOA
  44. 44. 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 48 NDM-1 New Delhi metallo-beta-lactamase
  45. 45. 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 49 • Transmission from one strain of bacteria to another plasmid mediated • Resistant to all except: 1. Colistin 2. Tigecycline 3. Aztreonam
  46. 46. Factors of Antibiotic Resistance EnvironmentalDrug Related Patient Related Prescriber Related Antibiotic Resistance 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 50
  47. 47. 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 51
  48. 48. 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 52 New Antibiotic Development •Only 15 antibiotics of 167 under development had new MOA with potential to combat of multidrug resistance •Lack of incentive
  49. 49. 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 53
  50. 50. 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 54
  51. 51. 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 55
  52. 52. References 1. Goodman and Gilman’s The Pharmacological Basis of Therapeutics 12th Edition 2. Rang and Dale’s Pharmacology 7th Edition 3. K. D. Tripathi Essentials of Medical Pharmacology 7th Edition 4. K. K. Sharma Principles of Pharmacology 2nd Edition 5. Koneman’s Color Atlas and Textbook of Diagnostic Microbiology 6th Edition 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 56
  53. 53. 06-08-2015 MIMER Medical College Talegaon Department of Pharmacology 57 Thank You

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