Mechanism of antibiotic resistance

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1 Mechanism of antibiotic resistance Dr.Siriwoot Sookkhee Ph.D (Biopharmaceutics) Department of Microbiology Faculty of Medicine, Chiang Mai University

2 Antibiotic resistance Cross-resistance : resistance in a group of drugs which closely related structure Co-resistance : concomitant resistance in a different group of drugs due to the different mechanisms or genes Multidrug resistance (MDR) : resistance in 1/3 of empiric drugs Extensive, extensively or extremely drug resistant (XDR) : resistance at least 1 drug in each group of empiric drug

3 Pan-drug resistance Pan-drug resistance (PDR) : resistant in all tested empiric drugs

4 Sources of resistance Intrinsic resistance : natural or spontaneous resistance insensitive Stenotrophomonas maltophilia Intrinsic resistance to carbapenem Sensitive to ceftazidime, fluoroquinolone, sulphonamide

5 Sources of resistance Acquired resistance : imported resistant genes - Mutational resistance : vertical gene transfer - Transmissible resistance : horizontal gene transfer - Plasmid borne resistance : rapid resistance - Chromosomal resistance : slow resistance

6 Drug target modification Alternation of protein target mutation of PBPs gene change of penicillin binding protein reduction of β-lactam binding

7 Resistance of MRSA : - alteration of PBP2s - staphylococcal cassette chromosome mec or SCCmec (meca gene) - correlated with hospital/community acquired infection

8 Altering the target receptor for the antibiotic to reduce or block its binding ANTIBIOTIC CELL WALL RECEPTOR CYTOPLASM Bacteria alters the shape of target receptor

9 Modification of peptidoglycan structural in VRE High level resistant genes : vana, vanb and vand Enterococcus faecalis, Enterococcus faecium D-alanyl-D-alanine D-alanyl-D-lactate

10 Gram Positive Cell Wall PEPTIDE CROSS-BRIDGE TEICHOIC ACID PEPTIDOGLYCAN LIPOTEICHOIC ACID CELL WALL CELL MEMBRANE

11 Gram Positive Cell Wall TP TP TP CELL WALL CYTOPLASM

12 v v Mechanism of Action VANCOMYCIN CELL WALL CYTOPLASM

13 VRE - Resistant mechanism Enterococcus spp. VRE v D-Alanine v D-Alanine D-Alanine D-Lactate

14 Vancomycin intermediate resistance (VISA) Thickening of peptidoglycan level Difficult to drug penetration

15 Heterogenous vancomycin intermediate resistance (hvisa) 1 intermediate resistant cell 999,999 susceptible cells antibiotic Survive and multiply die Predominate with intermediate resistant cells Change the susceptibility, zone diameter, MIC

16 Target alteration on ribosome Macrolides, lincosamides, streptogramine (MLS) : posttranslational modification at ribosome subunit 50s i.e. methylation

17 Alteration of DNA synthesis Mutation of gyra Alteration of DNA gyrase Reduction of drug binding Increase of drug MIC for inhibiting bacteria

18 Alteration of cell membrane structure Porin : protein structure at cell membrane responsible to import small drug molecule i.e. beta lactam and quinolone Porin encoding gene is not expressed in Pseudomonas aeruginosa Imipenem cannot import into cell Imipenem resistance

19 Preventing the entry of the antibiotic into the bacteria ANTIBIOTIC TRANSPORT PROTEIN PERIPLASM CYTOPLASM Altering transport (carrier) proteins in the cytoplasmic membrane

20 Preventing the entry of the antibiotic into the bacteria ANTIBIOTIC PORIN PERIPLASM Altering porins in the outer membrane of a bacterium CYTOPLASM

21 Expression of drug efflux pumps Drug efflux pumps : responsible to export drug decrease drug intracellular concentration Multidrug efflux pump : multidrug transporter involved the resistance of all drugs except polymyxin (colistin)

22 Actively transporting the antibiotic out of bacteria ANTIBIOTIC TRANSPORT PROTEIN NEW TRANSPORT PROTEIN PERIPLASM Producing a new transport molecule in the cytoplasmic membrane CYTOPLASM

23 Drug inactivation / modification Structural translocation : Aminoglycosides, macrolides by enzymes involved the group translocation - phosphoryltransferases - nucleotidyltransferases - Adenylyltransferases - acetyltransferases reduction of drug binding cannot inhibit bacterial protein synthesis

24 Drug inactivation / modification Structural translocation : Chloramphenicol by enzymes involved the transacetylase acetyl group hydroxyl group reduction of drug binding cannot inhibit bacterial protein synthesis

25 Drug inactivation by hydrolysis Beta lactamase can hydrolyze beta lactam ring Inactive drug (Penicilloic acid) Beta lactam cannot kill bacteria

26 Drug inactivation by hydrolysis ereb expression in Escherichia coli Erythromycin esterase II production Hydrolysis lactone ring of erythromycin

27 Producing an enzyme capable of destroying the antibiotic ANTIBIOTIC PERIPLASM ENZYME CYTOPLASM

28 Producing an enzyme capable of destroying the antibiotic INACTIVATED ANTIBIOTIC ANTIBIOTIC PERIPLASM ENZYME CYTOPLASM

29 Mechanism of Resistance to β-lactams Porin channels Efflux system PBPs B-lactamases The Gram Negative Cell Wall

30 Why Test for β-lactamases? Smart use/ Appropriate therapy Breakpoints do not reliably detect new β-lactamases Infection control Identify drugs causing resistance

31 -LACTAMASE 1. Class A β-lactamase 1.1 Extended spectrum β-lactamase or ESBL Hydrolyze - penicillins - cephalosporin 1 st, 2 nd, 3 th - monobactams Can be inactivated by beta lactamase inhibitors, clavulanic acid, tazobactam, sulbactam Active Inactive

32

33 CLSI Screening Test E. coli ceftazidime 22 mm cefotaxime 27 mm Detect key hole phenomenon

34 Combination Disc Method CLSI Approved Method Combination discs Test cefotaxime and ceftazidime +/- clavulanate > 5 mm zone increase CAZ/CLA CTX/CLA CAZ CTX

35 Commercial methods E-test : ceftazidime, cefotaxime (MIC 8 times) cefotaxime cefotaxime + clavulanate

36 Class A carbapenemase Hydrolyze - penicillins - cephalosporin - carbapenem Can be inactivated by beta lactamase inhibitors, clavulanic acid, tazobactam, sulbactam KPC enzymes : carbapenem-resistant Klebsiella pneumoniae

37

38 Class B β-lactamase : Metallo carbapenemase or class B carbapenemase or metalloenzymes Can be inhibited by EDTA Cause of high-level carbapenem resistance Pseudomonas aeruginosa, Acinetobacter baumannii

39 New Delhi Metallo or NDM enzyme Found in 2009 in Klebsiella pneumoniae isolated from Swedish patient at India : Superbug organism multidrug resistance to all drugs except colistin, phosphomycin, tigecycline Klebsiella pneumoniae and Escherichia coli : community-acquired pathogens

40 Class C β-lactamase : AmpC β-lactamases cannot be inactivated by clavulanic acid or EDTA but can be inactivated by cloxacillin, oxacillin

41 Class D β-lactamase : Oxacillinase or OXA-type carbapenemase can be inactivated by NaCl Found in multidrug-resistant Acinetobacter baumannii

42 Macrolide resistance Intrinsic resistance : hydrophobic macrolide have low permeability through outer membrane (Gram -ve) Acquired resistance - ribosomal modification - efflux pump - enzyme inactivation0

43 Aminoglycoside resistance Enzymatic modification of drug - high level resistance - genes encoding resistance located in plasmids - gene transfer occurs across species Reduced uptake or decreased permeability of bacterial cell wall

44 Tetracyclines resistance mechanism predominate: efflux and ribosomal protection bacteria survived producing an enzyme to destroy or inactivate the antibiotic blocking tetracyclines from binding to the ribosome (

45 Quinolones resistance mechanism Changes in protein targets - DNA gyrase - Topoisomerase IV Reduction in the accumulation - decrease in permeability - increase in active efflux system DNA gyrase and topoisomerase IV protection - qnr gene

46 Sulfonamides resistance mechanism altering the target site for the antibiotic to reduce or block its binding altered dihydropteroate synthase or dihydrofolate reductase

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