Antibiotics & Antibacterial Resistance PDF

Antibiotics. Antibacterial resistance. Dr Kārlis Rācenis Department of Biology and Microbiology Resistance of microorganisms in environment 1. Biological factors: 2. Chemical factors Antimicrobials 3. Physical factors Bacterial antagonism...

Antibiotics. Antibacterial resistance. Dr Kārlis Rācenis Department of Biology and Microbiology Resistance of microorganisms in environment 1. Biological factors: 2. Chemical factors Antimicrobials 3. Physical factors Bacterial antagonism Bacteriophages  Terminology (I) Aseptic technique – is a combination of practices that precludes the presence of microorganisms and other infectious agents in the environment or particular objects. (sterilization and disinfection methods are used) Antiseptic substances are chemicals thar are used locally when treating the skin, mucosa or a wound. They are chemical agents that affect living tissue and inhibit microorganisms.  Terminology II Decontamination - is the extermination of microorganisms in an object. It is a relative term (can be achieved by disinfection and sterilization) Disinfection – is the extermination of most bacteria or bacterial count reduction. Sterilization – complete and permanent extermination of all microorganisms in an environment and on inanimate objects. In comparison to disinfection, sterilization ensures the extermination of endospores, resistant bacteria and viruses. Terminology III Biocidal activity (bactericide, virucide, fungicide, sporocide) is inherent to substances that kill microbes, viruses, fungi and spores Biostatic activity (bacteriostatic, fungistatic) stops the growth of microbes and fungi accordingly, however, does not kill them. After biostatic activity has ceased, the growth of bacteria and fungi can resume. http://basicbacteriology.blogspot.com/p/mode-of-actions-of-antibiotics.html Terminology IV Antimicrobial substance: is any substance of natural, semisynthetic or synthetic origin that kills or inhibits the growth of microorganisms but causes little or no damage to the host. These include antibacterial, antiviral, antifungal, antiprotozoal substances. Antibiotics: are produced naturally by microorganisms and kill or inhibit the growth of other microorganisms, mainly bacteria. All antibiotics are antimicrobials, but not all antimicrobials are antibiotics. Terminology V Antibacterial spectrum: is defined as the range of different microorganisms that an antibiotic or antimicrobial agent inhibits or kills. Broad spectrum: inhibit several bacterial groups Narrow spectrum: inhibit or kill limited species of bacteria Minimum inhibitory concentration: is the lowest concentration of an antibiotic that inhibits the growth of a given strain of bacteria Terminology VI Multidrug-resistant bacteria – resistant to one or more classes of antimicrobial agents and usually are resistant to all but one or two commercially available antimicrobial agents (excluding M. tuberculosis) History  Paul Erlich Several chemical compounds act against microorganisms, including antibacterial substances. Paul Erlich was the first who observed and detected such phenomena (in the end of XIX century) Introduced the term of selective toxicity Alexander Fleming 1928.g. Alekxander Fleming (Scottish microbiologist) discovered first antibiotic. Observed that the growth of S.aureus was inhibited in the area surrounding the colony of a mold (Penicillium notatum) that had contaminated a Petri plate. https://www.svt.steber.fr/wp-content/uploads/2014/04/?C=S;O=D Antibiotic era in medicine http://www.newworldencyclopedia.org/entry/Penicillin  Classification of antibacterial substances Classification principles of antibiotics I 1. According to their origin: a) Microorgansims: Fungi – against bacteria (penicillins, cephalosporins) Main group of Atypical bacteria Actinomyces – against bacteria, fungi, tumor cells a/b producers Bacteria – against other bacteria b) Human and animal cells c) Plant cells Antibiotics from bacteria Bacterial produced antibiotics (a/b) are proteins that are toxic to human organism therefore these substances are of limited use. Mainly used locally (topical application), e.g. in chronic wound infection. 1. Polymixin B – produced by Bac.polymixa, bactericidal effect, inhibit CM synthesis – mainly against Gr- bacteria 2. Bacitracin – produced by Bac.subtilis, inhibit CM synthesis – mainly against Gr+ (MRSA decontamination) 3. Gramicidin - produced by Bac.brevis, broad spectrum – Gr+, Gr-, also anaerobes, very toxic, topical use 4. Mupirocin – produced by Pseudomonas fluorescens, bacteriostatic effect, inhibit protein synthesis. Against Gr+ cocci, Gr- rods. 5. Colistin – produced by E.coli (antagonism) Antibiotics form human and animal cells 1. Lysozyme or muramidase – break down peptidoglycan: Discovered by A.Fleming in 1929. Found in perspiration, tears, saliva, nasal secretions, tissue fluid Against Gr+ and Gr- cocci Regulation of microflora 2. Spermine – from animal semen 3. Defensins – from leukocytes Antibiotics form plant cells Volatile substances with bactericidal effect. Most active substances are found in onions, garlic, horseradish, in practical medicine are not used Examples: Alicin – from garlic Allilcep – from onion Raphanin – from radish Chlorophyllipt – from eucalyptus Classification principles of antibiotics II 2. According to their spectrum: a) Broad spectrum (e.g. tetracycline) b) Narrow spectrum (e.g. oxacillin) 3. According to the mode of action: a) Bactericidal: b) Bacteriostatic: 4. According to the route of administration: a) Enteral b) Parenteral Classification principles of antibiotics III 4. According to their mechanism of action: Cell wall inhibitors Protein synthesis inhibitors Nucleic acid synthesis inhibitors Antimetabolites Plasma membrane inhibitors Action mechanism of antibiotics Cell wall inhibitors BETA LACTAMS PENICILLINS CEPHALOSPORINS CARBAPENEMS MONOBACTAMS  β-lactam antibiotics: Main structure β-lactam ring https://www.medsafe.govt.nz/profs/PUarticles/September2016/antibiotics.jpg  β-lactam antibiotics: Mechanism of action β-lactam antibiotics bind to transpeptidase and inhibit peptidoglycan synthesis! Transpeptidase s. PBP (penicillin binding protein) BACTERICIDAL effect https://basicmedicalkey.com/penicillins-cephalosporins-and-other-%CE%B2-lactam-antibiotics-3/  β-lactam antibitoics are not working against: Metabolically inactive bacterial cells (chronic infections) Bacteria which have no CW Bacteria which do not have certain transpeptidase Main inhibitor of β-lactam antibiotics: β-lactamase β-Lactamases – bacterial produced enzymes that cleave (inactivate) β-lactam ring (penicilinase, cephalosporinase, carbapenemase) β-lactam a/b - PENICILLINS 1st generation penicillins – natural penicillins synthesized by Penicillium chrysogenum (Penicillium notatum) (acid unstable) Benzylpenicillin (Penicillin G) – short acting Parenteral Benzathine penicillin G – long acting β-lactamase sensitive (acid stable) Enteral Phenoxymethylpenicillin (penicillin V) SPECTRUM: narrow, Gr+ and Gr- cocci, Spirochetes, Actinomyces. (Neisseria meningitidis, Enterococcus faecalis, E. faecium, Corynebacterium diphtheriae) β-lactam a/b – SEMISYNTHETIC PENICILLINS Semisynthetic penicillin: Common penicillin nucleus is retained, and other side chains are chemically added Stable against low pH (can be used p/o) Wider antimicrobial spectrum β-lactam a/b - PENICILLINS 2nd generation penicillins – semisynthetic / antistaphylococcal / β-lactamase resistant penicillins Methicillin Oxacillin β-lactamase resistant Nafcillin SPECTRUM: narrow similar to first generation, Gr+ and Gr- cocci, Spirochetes, Actinomyces - !very effect against Staphylococci! β-lactam a/b - PENICILLINS 3rd generation penicillins (semisynthetic) - aminopenicillins Ampicillin β-lactamase Amoxicillin sensitive Ampicillin + Sulbactam β-lactamase Amoxicillin + Clavulanate resistant SPECTRUM: broad spectrum (especially in combination with β-lactamase inhibitors): Gr+ and Gr- cocci, Gr+ and !Gr- rods! + anaerobes (except Pseudomonas spp.) β-lactam a/b - PENICILLINS 4th generation penicillins (semisynthetic) - carboxypenicillins (antipseudomonal penicillins) Ticarcillin β-lactamase sensitive Carbenicillin Ticarcillin + Clavulanate β-lactamase resistant SPECTRUM: broad spectrum - Gr+ and Gr- cocci, Gr+ and Gr- rods (especially !Pseudomonas spp.!)+ anaerobes β-lactam a/b - PENICILLINS 5th generation penicillins (semisynthetic) – acylampicillin (antipseudomonal penicillins) Mezlocillin β-lactamase Azlocillin sensitive Piperacillin Piperacillin + tazobactam β-lactamase resistant SPECTRUM: broad spectrum - Gr+ and Gr- cocci, Gr+ and Gr- rods (especially !Pseudomonas spp.!) + anaerobes β-lactam a/b - PENICILLINS 6th generation penicillins (semisynthetic)– amidinopenicillins (bind to different PBP) Parenteral Mecillinam β-lactamase resistant Enteral Pivmecillinam SPECTRUM: against multiresistant Gr- rods (E. coli, K. pneumoniae) (except Pseudomonas spp.) β-lactam a/b - CEPHALOSPORINS First released by fungus Cephalosporium acremonium Semisynthetic or synthetic antibacterials Contain β-lactam ring Resitant to penicilinase (β-lactamase) Senstive to cephalosporinase (β-lactamase) (no cross-reaction) Cephalosporins are classified according to generations and their antimicrobial spectrum, more effective against Gr- infections β-lactam a/b - CEPHALOSPORINS Susceptibility SPECTRUM against β-lactamases Active against Gr+ Gr+ +++ (streptococci and staphylococci) less Gr- Against Gr+, also Gr- (incl.Enterobacteriaceae) ++ Less Gr+, more against Gr- Enterobacteriaceae (Ceftazidime + is effective against Pseudomonas) Gr- Broad spectrum: Gr+, good against Gr- Enterobacteriaceae, Pseudomonas, anaerobes Effective against multiresistant m/o (MRSA, S. pneumoniae) β-lactam a/b – CARBAPENEMS and MONOBACTAMS CARBAPENEMS: Broad spectrum a/b, especially against β-lactamase resistant bacteria Meropenem Ertapenem Imipenem + cilastatin (to avoid rapid renal inactivation) SPECTRUM: Gr+ and Gr- bacteria (cocci + rods), aerobes and anaerobes MONOBACTAMS: Narrow spectrum, particularly active against Gr- aerobes (Pseudomonas) Aztreonam Cell wall inhibitors GLYCOPEPTIDES Glycopeptides – VANCOMYCIN Produced by Streptomyces orientalis Inhibit synthesis of peptidoglycan (bactericidal) Do not work against Gr-, can not pass through the outer membrane because of the size of vancomycin molecule Vancomycin SPECTRUM: Gr+ cocci (Staphylococcus, Streptococcus, Enterococcus) and bacteria (Clostridium spp.), multiresistant m/o (MRSA) FOSFOMYCIN Inhibit first steps of peptidoglycan (bactericidal) Great penetration in tissue and activity i/c Fosfomycin SPECTRUM: Broad, Gr+ /Gr- cocci and rods, multiresistant m/o (ESBL) Protein synthesis inhibitors Inhibit protein synthesis by binding to ribosomes in prokaryotic cells. Antibiotics can bind to ribosomes: o reversibly – bacteriostatic o irreversibly – bactericidal Protein synthesis inhibitors 50s subunit Macrolides First substance erythromycin (Streptomyces erythreus) Binds reversibly to ribosome 50s subunit: o Low dose –bacteriostatic o High dose – bactericidal Development of resistance is relatively rapid: Commonly used as an alternative: e.g. other a/b side effects (allergic reactions) Erythromycin Azithromycin Clarithromycin SPECTRUM: Relatively broad, Gr+ /Gr- bacteria, atypical bacteria (i/c): Mycoplasma, Chlamydia, Legionella Chloramphenicol Binds reversibly to ribosome 50s subunit: o Bacteriostatic effect SIDE EFFECT: bone marrow toxicity – may cause aplastic anemia Belongs to alternative antibiotics Chloramphenicol (natural) Levomycetin (synthetic) SPECTRUM: Broad, Gr+ /Gr- bacteria, atypical bacteria (i/c): Mycoplasma, Chlamydia, anaerobes Clindamycin Binds reversibly to ribosome 50s subunit: o Bacteriostatic effect o In very high concentrations – bactericidal Inhibit bacterial exotoxin synthesis (Staphylococcus, Streptococcus) Commonly cause Cl. difficile enterocolitis Clindamycin Linezolid Binds reversibly to ribosome 50s subunit: o Bacteriostatic effect No cross-reaction with other protein synthesis inhibitors: o used to treat multiresistant bacterial infections Belongs to alternative antibiotics Linezolid SPECTRUM: Broad, Gr+ /Gr- bacteria, especially Staphylococcus and Streptococcus – multiresistant strains (VRE, MRSA) Protein synthesis inhibitors 30s subunit Aminoglycosides Binds irreversibly to ribosome 30s subunit: o Bactericidal effect Induce nephrotoxicity and ototoxicity Due to weak cell wall penetration are used in combination with cell wall synthesis inhibitors Amycacin Tobramycin Neomycin – topical use Gentamycin Streptomycin (M. tuberculosis treatment) Tetracyclines Binds reversibly to ribosome 30s subunit: o Bacteriostatic effect Binds to metal ions of bacteria → inhibits bacterial enzymes Commonly cause dysbacteriosis and superinfections (Candida spp.) due to broad antimicrobial spectrum Tetracycline Doxycycline Minocycline Nucleic acid synthesis inhibitors Quinolones Rifamycin QUINOLONES or FLUOROQUINOLONES Synthetic antibacterial substances DNA gyrase inhibitors → inhibit DNA replication → bactericidal effect Widley used although in last years are used less: o Side effects (tendinitis, aortic dissection, wide QT (ECG)) + broad resistance Ciprofloxacin Norfloxacin Moxifloxacin RIFAMYCIN Inhibits DNA-dependent RNA polymerase activity → no RNA synthesis → bactericidal effect Induce hepatotoxicity Rifampicin  Antimetabolites Sulfanilamide (Sulfamethoxazole) Dihydrofolate reductase inhibitor (Trimethoprim) ANTIMETABOLITES Synthetic antibacterial substances Inhibit synthesis of folic acid in bacteria (necessary for DNA synthesis) (human cells do not synthesis folic acid) Can be used in combinations (have synergistic effect) Effect: o bacteriostatic (single use) o bactericidal (in combination) Trimethoprim/sulfamethoxazole SPECTRUM: Broad, Gr+, Gr- bacteria, P. jirovecii, some protozoa Most common side effects of a/b therapy 1. Allergic reactions: commonly from β-lactams penicillins and cephalosporines rash fever interstitial nephritis 2. Dysbacteriosis: mainly from broad spectrum antibiotics development of superinfections (e.g. Candida) 3. Toxic reactions: chloramphenicol – disturbances in red cell maturation fluoroquinolones – tendinitis, aortic dissection, wide QT aminoglycosides – nephrotoxicity and ototoxicity rifampicin – hepatotoxicity tetracycline – liver and kidney toxicity, tooth discoloration 4. Development of antibacterial resistance Principles of rational selection of antimicrobial drugs 1. A specific etiologic diagnosis must be formulated. 2. In vitro antibacterial susceptibility should be detected. 3. The most active and least toxic a/b should be used. 4. The optimal dose and administration of antibiotic should be chosen (per kg., i/v, p/o etc.). 5. In case of long-term treatment a/b change should be considered. 6. Side effects should be taken in consideration. Antibacterial resistance Uncontrolled use of antibiotics– creates antibacterial resistance! http://today.uconn.edu/2015/02/getting-ahead-of-antibiotic-resistant-bacteria/ Antibacterial resistance Antibacterial resistance https://www.cdc.gov/drugresistance/about.html  The list of resistant bacteria 1st priority group - critically high level Carbapenem – resistant strains: Acinetobacter baumanii Pseudomonas aeruginosa Enterobacteriaceae (e.g. E.coli, K.pneumoniae): Carbapenem - resistant ESBL (Extended spectrum beta-lactamases) producers Carbapenem – resistant A. baumanii strains in Europe in 2019 https://atlas.ecdc.europa.eu/public/index.aspx?Dataset=27&HealthTopic=4  The list of resistant bacteria 2nd priority group - high level Enterococcus spp., vancomycin - resistant (VRE) Staphylococcus aureus, methicillin - resistant (MRSA), vancomycin – resistant (VRSA) Helicobacter pylori, clarithromycin - resistant Campylobacter spp., fluoroquinolone - resistant Salmonellae, fluoroquinolone - resistant Neisseria gonorrhoeae, cephalosporin - resistant, fluoroquinolone - resistant MRSA in Europe in 2019 https://atlas.ecdc.europa.eu/public/index.aspx?Dataset=27&HealthTopic=4  The list of resistant bacteria 3rd priority group - medium level Streptococcus pneumoniae, penicillin - resistant Haemophilus influenzae, ampicillin - resistant Shigella spp., fluoroquinolone - resistant Penicillin - resistant S. pneumoniae strains in Europe in 2019 https://atlas.ecdc.europa.eu/public/index.aspx?Dataset=27&HealthTopic=4 Classification of antibacterial resistance Primary There is no target for antibacterial drug: o Mycoplasma does not have cell wall – penicillin does not work against it Secondary Nongenetic Genetic o Chromosomal (bacterial genes) o Extrachromosomal (e.g. plasmids) Nongenetic antibacterial resistance Main mechanisms: 1. May lose the specific target structure, e.g may change to L form: Therapy of penicillin and cephalosporins 2. Metabolically inactive during chronic infection M.tuberculosis can be metabolic inactive for several years 3. May be at site where antimicrobials are excluded or are not active Intracellular – the concentration is too low (e.g. penicillin, cephalosporins) biofilms https://microbewiki.kenyon.edu/index.php/File:Biofilm.png Genetic origin of drug resistance: chromosomal It develops as a result of spontaneous and random mutations of genes encoding the target site, or cell structures affecting access to the target site. For example: resistance to the aminoglycosides can result from alteration of the amino acid in the specific protein of the 30S ribosomal subunit lost or altered PBP (MRSA) altered permeability – a block in the transport of the drug into the cell Genetic origin of drug resistance: extrachromosomal Horizontal gene transfer – the process of swapping the antibiotic resistance genes between bacteria. Via 3 main mechanisms: Transduction (with temperate bacteriophage) Conjugation (with R-plasmids) Transformation (uptake of short fragments of naked DNA)  Genetic origin of drug resistance: Extrachromosomal – conjugation (R-plasmid) Bacteria can contain plasmids (extrachromosomal circular DNA molecules): Carry at least one gene. Many of genes could be beneficial for host cell, e.g. antibacterial resistance R-plasmids – carry genes for antibacterial resistance Genetic origin of drug resistance: Extrachromosomal – conjugation (R-plasmid) 1. Plasmid replication occurs independently from bacterial genetic material, therefore donor cell may have variety of analogue plasmids 2. Plasmid exchange may occur even between different species and genera 3. Plasmid encoded resistance develops rapidly 4. Plasmids may determine resistance against high a/b concentrations 5. One plasmid can carry genes that determine resistance against several antibiotics – development of multiresistant bacteria 6. Bacteria can have several different R-plasmids - development of multiresistant bacteria https://www.reactgroup.org/toolbox/understand/antibiotic-resistance/plasmids-and-co-selection/ Mechanisms of antibacterial resistance Drug excretion ↑ Uptake ↓ - change of permeability Efflux pumps (macrolides) Change the diameter of pore – the intracellular concentration is low (tetracyclines) Develop an altered metabolic pathway or an altered enzymes – still perform metabolic function but much less affected by the drug e.g. trimethoprim/sulfamethoxazole Enzymatic inactivation of drugs Modification on the drug target e.g. -lactamase MRSA - altered PBP (penicillin, cephalosporin, carbapenem) Thank you for your attention!