Lecture 1 and Lecture 2 Dr Mustapha 2024 (PDF)

11/25/2024 PHARMACEUTICAL MICROBIOLOGY ANTIMICROBIALS PROF. DR. MUSTAPHA ELNAKEEB COURSE 1 A NTIMICROBIALS ARE BROADLY DIVIDED INTO : 1. A NTIBIOTICS 2. ANTIVIRALS 3. ANTIFUNGALS...

11/25/2024 PHARMACEUTICAL MICROBIOLOGY ANTIMICROBIALS PROF. DR. MUSTAPHA ELNAKEEB COURSE 1 A NTIMICROBIALS ARE BROADLY DIVIDED INTO : 1. A NTIBIOTICS 2. ANTIVIRALS 3. ANTIFUNGALS 4. ANTIPARASITIC 5. N ON - ANTIBIOTIC ANTIMICROBIALS 2 1  11/25/2024 Antibiotics and Antimicrobials ANTIBIOTICS: Greek words anti (against) and biotikos (concerning life) refer to substances produced by microorganisms, which selectively suppress the growth of or kill other microorganisms at deficient concentrations. CHEMOTHERAPEUTIC AGENTS: It uses drugs (chemical entities) with selective toxicity against infections/ viruses, bacteria, protozoa, fungi, and helminths. ANTIMICROBIALS: Derived from the Greek words anti (against), mikros (little), and bios (life) and refers to all agents of natural, synthetic, or semi-synthetic origin that at low concentrations kill or inhibit the growth of microorganisms but cause little or no host damage. antimicrobials include both chemotherapeutic agents + antibiotics. 3 ANTIBACTERIAL AGENTS An antibacterial is an agent that interferes with the growth and reproduction of bacteria An antibiotic is a product produced by a microorganism or a similar substance produced wholly or partially by chemical synthesis, which in low concentrations, inhibits the growth of other microorganisms. Bactericidal : The organism is lysed or killed by direct damage on susceptible cell targets. Bacteriostatic : These agents exert their influence by inhibiting growth and reproduction of the bacteria usually by inhibiting protein synthesis. 4 2  11/25/2024 Narrow Spectrum Antimicrobial - An antimicrobial that acts on a limited number of microbial species, e.g. Metroimidiazole derivatives etc Broad Spectrum Antimicrobial - An antimicrobial that acts on a wide range of species, e.g., erythromycin for Gram positive. Gram negative, Legionella, Mycoplasma, etc. An antibiotic is a selective poison. It has been chosen so that it will kill the desired bacteria, but not the cells in your body. Each different type of antibiotic affects different bacteria in different ways. 5 CLASSIFICATION OF ANTIMICROBIALS Antimicrobials are classified in several ways A. Chemical structure B. Mechanism of action C. Type of organisms (against which primarily active) D. Spectrum of activity E. Type of action (bacteriostatic and bactericidal) F. Source of antibiotics 6 3  11/25/2024 7 RESISTANCE Antimicrobial Resistance (AMR) occurs when bacteria, viruses, fungi, and parasites no longer respond to antimicrobial medicines. As a result of drug resistance, antibiotics and other antimicrobial medicines become ineffective and infections become difficult or impossible to treat, increasing the risk of disease spread, severe illness, disability, and death. AMR is a natural process that happens over time through genetic changes in pathogens. Its emergence and spread is accelerated by human activity, mainly the misuse and overuse of antimicrobials to treat, prevent or control infections in humans, animals and plants. 8 4  11/25/2024 Antimicrobial resistance can be divided into Unresponsiveness of a microorganism to an antimicrobial agent – Natural resistance – Acquired resistance Natural resistance: Some microbes have resistant to certain AMAs. E.g.: Gram negative bacilli not affected by penicillin G; M. tuberculosis insensitive to tetracyclines. Acquired resistance: Development of resistance by an organism (which was sensitive before) due to the use of AMA over a period of time. E.g.:  Staphylococci, tubercle bacilli develop resistance to penicillin (widespread use for >50 yr).  Gonococci quickly developed resistant to sulfonamides in 30 yr. Acquired Resistance is mainly developed by either MUTATION Or GENE TRANSFER. 9 Mechanisms of resistance  Permeability: -Some microbes → alteration in chemical nature of outer membrane → change cell wall permeability to drug - Eg: Tetracyclin resistance by Pseudomonas aeruginosa  Production of enzymes: - enzymes which can act on drug - Eg: β-lactamase produced by certain bacteria destroy penicillin  Altered structure target: -Aminoglycosides act by attaching to 30S subunit but resistant bacteria develop altered receptor  Altered metabolic pathway: - Drugs inhibit certain pathways - Resistant bacteria → bypass the reaction 10 5  11/25/2024 Antimicrobial Resistance COMMON MODES e.g. aminoglycosides & tetracyclines OF ANTIMICROBIAL RESISTANCE e.g. aminoglycosides chloramphenicol & penicillins e.g. Penicillins e.g.tetracyclines 11 Site and Mechanism of action of Antibiotics 12 6  11/25/2024 MECHANISM OF ACTIONS Disruption of Inhibition of Inhibition of Inhibition of Inhibition of cell cytoplasmic bacterialprotein Nucleic Acid Folic Acid wall synthesis membrane synthesis Synthesis Synthesis  Penicillins  Polymyxin B  Aminoglycosides  Fluoroquinolones  Sulfonamides  Cephalosphorins  Colistin  Chloramphenicol  Rifampin  Trimethoprim  Imipenem  Daptomycin  Macrolides  Pyrimethamine  Meropenem  Tetracycline  Aztreonam  Streptogrmins  vancomycin  linezolid 24 June 2016 NNRG SCHOOL OF PHARMACY 13 13 1. Inhibition of protein synthesis 14 7  11/25/2024 Overview of Protein Synthesis 15 protein synthesis inhibitors According to this target of action, antibiotics follow several pathways 1. amino acid activation 2. Initiation complex formation 3. Peptide bond formation and elongation 4. Translocation and termination 5. Release and recycling ribosome. so, most of the antibiotics in this group are bacteriostatic Antibiotics on Amino acid activation step we have 20 amino acids usually in L-form. Each amino acid has a specific t-RNA (transfer amino acid) from the cytoplasm to the ribosome. AA1+ ATP…… AA1+AMP (energized amino acid)+PPi t-RNA synthetase AA1+AMP+ tRNA ……………………….. AA1 +tRNA (acyl tRNA)+enzyme or amino acid synthetase 16 8  11/25/2024 – Structure of prokaryotic ribosome acts as target for many antimicrobials of this class Differences in prokaryotic and eukaryotic ribosomes responsiblefor selective toxicity Antibiotics acting on amino acid activation usually act on tRNA synthetase 24 June 2016 NNRG SCHOOL OF PHARMACY 17 17 1. Mupirocin Mupirocin is a naturally synthesized antibiotic agent that is isolated from a strain of Pseudomonas fluorescens It is used as topical treatment of primary or secondary bacterial infections (to avoid plasma inactivation) Mupirocin is bactericidal agent at high concentrations while it is bacteriostatic at low concentrations. It is widely used for the treatment of complications related to the skin caused by bacteria i.e. Impetigo (blisters or sores on the face, neck, hands, and diaper area), Furuncle (a deep folliculitis, infection of the hair follicle) and open wounds. Mupirocin is also used to treat infection caused by gram-positive bacteria, Staphylococcus aureus, and beta-hemolytic streptococci including Streptococcus pyogenes. Mupirocin is highly effective in the treatment of methicillin- resistant Staphylococcus aureus (MRSA). Mupirocin is also used for the treatment of nasal infections. 18 9  11/25/2024 Mechanism of action Mupirocin is an antibiotic that acts by reversible binding to the bacterial Isoleucyl-tRNA synthetase Bacterial Isoleucyl-tRNA synthetase enzyme is involved in the formation of Isoleucyl-tRNA from tRNA and isoleucine. Inhibition of bacterial Isoleucyl-tRNA synthetase enzyme results in the inhibition of RNA and protein synthesis. 19 Mechanism of resistance 1. Target-site modification: Resistance arises from alterations in bacterial isoleucyl-transfer RNA, resulting in the loss of mupirocin side chain recognition and the subsequent failure of mupirocin insertion at the binding site. a. High-level resistance resulting from plasmid-mediated gene mutations (mupA, mupB, or ileS2) b. Low-level resistance associated with chromosomal point mutations in isoleucyl-tRNA synthetase (IRS), causing alterations in ileS gene. 2. Efflux pumps: In some cases, bacteria can pump the drug out of the cell. Resistance is more common with long-term or widespread use. 20 10  11/25/2024 Antibiotics on Initiation complex formation step Ribosomal Subunits and "S" Value 1. Svedberg Unit (S): The "S" in 70S or 80S stands for the Svedberg unit, a measure of sedimentation rate during ultracentrifugation. It reflects the size, shape, and density of the ribosome, not just its molecular weight. 2. Bacterial Ribosome (70S): The bacterial ribosome consists of two subunits: a. 50S (large subunit): Contains 23S rRNA, 5S rRNA, and ~34 proteins. b. 30S (small subunit): Contains 16S rRNA and ~21 proteins. When assembled, these subunits sediment at 70S due to their combined shape and structure. The 30S initiation complex is a crucial intermediate in the initiation of bacterial translation. It forms on the small ribosomal subunit (30S) and ensures proper positioning of the start codon and initiator tRNA before the large subunit (50S) joins to form the complete 70S ribosome. 21 Ribosomal subunits Structure of the 30S Initiation Complex The assembled 30S initiation complex consists of: 1. 30S ribosomal subunit: The scaffold for assembly. 2. mRNA: Positioned correctly via Shine-Dalgarno interaction. 3. fMet-tRNAᶠᴹᵉᵗ: Aligned in the P site with the start codon. 4. Initiation factors: o IF-1: Blocks the A site (acceptance site) o IF-2: Guides fMet-tRNAᶠᴹᵉᵗ into the P site (Protein release site)and hydrolyzes GTP for energy. o IF-3: Prevents premature association of the 50S subunit and aids mRNA alignment. 5. GTP: Bound to IF-2, providing energy for subsequent steps. 22 11  11/25/2024 Ribosomal subunits Transition to the 70S Initiation Complex 1. 50S Subunit Joining: o The 50S ribosomal subunit associates with the 30S initiation complex. o IF-3 is released to allow this association. 2. GTP Hydrolysis: o IF-2 hydrolyzes GTP, providing the energy needed for proper assembly. o IF-1 and IF-2 are released. 3. Final Assembly: o The resulting structure is the 70S initiation complex with the start codon and fMet- tRNAᶠᴹᵉᵗ in the P site, ready for elongation. 23 24 12  11/25/2024 25 2. Aminoglycosides Older group aminoglycosides are naturally occurring substances produced by actinomycetes. Features of aminoglycosides include amino sugars bound by glycosidic linkages to a relatively conserved six-membered ring that itself contains amino group substituents (Aminosugars). Complex heterocyclic compound Aminocyclitol group + one/more Amino Sugars group. 26 13  11/25/2024 Examples of Aminoglycosides 1. Streptomycin: (natural) An older agent, used as part of combination therapy for tuberculosis. 2. Kanamycin: (natural) 3. Neomycin: Topical use or oral administration for bowel decontamination. 4. Gentamicin (Garamycin): (natural) Widely used for serious Gram-negative infections. 5. Amikacin, Sisomicin, and Tobramycin: (derivatives of gentamicin, semisynthetic) Effective against resistant strains, including ESBL-producing organisms. Tobramycin is preferred for Pseudomonas aeruginosa infections. 6. Plazomicin: (semisynthetic )A newer aminoglycoside, effective against multidrug- resistant bacteria. 7. Arbekacin: (semisynthetic )A newer aminoglycoside. 27 Mechanism of Action Aminoglycosides bind to the 30S subunit of the bacterial ribosome, thereby inhibiting bacterial protein synthesis (translation): 1. Interfere with formation of initiation complex of peptide formation 2. Binds to Ribosome and alter its shape in sucha way that there is misreading of mRNA causing incorporation of incorrect amino acid. 3.Causes breakup of polysomes into nonfunctional monosomes 4. Interfere with attachment of tRNA to mRNA-Ribosome complex protein Requires oxygen uptake, therefore ineffective against anaerobes. Bactericidal 28 14  11/25/2024 Important notes 16SrRNA is the RNA in 30S while 23SrRNA is the RNA in 50S The result of the interaction between aminoglycoside antibiotic and initiation complex is the formation of a misleading condone which causes inactive or fold protein (Toxic effect). Aminoglycosides’ effect on the bacterial membrane is by attraction; aminoglycosides are polycationic and bacteria are –vely charged. Aminoglycosides have poor oral absorption so used only if needed for their local action on GIT in cases of diarrhea (e.g neomycin) It is usually administered topically, intravenously or intramuscularly. 29 Antimicrobial Spectrum 1. Gram-negative bacteria: o Highly active against aerobic Gram-negative bacilli, including Pseudomonas aeruginosa, Klebsiella pneumoniae, and Escherichia coli. 2. Gram-positive bacteria: o Synergistic activity when combined with cell-wall-active agents (e.g., beta-lactams or glycopeptides) against Enterococcus spp. and Staphylococcus aureus. 3. Mycobacteria It is ineffective against Anaerobic bacteria due to the requirement for oxygen- dependent transport into bacterial cells. 30 15  11/25/2024 31 Adverse Effects All members are Nephrotoxic and Ototoxic but vary in their ability to cause this adverse effect. 1. Ototoxicity (esp. with loop diuretics) Irreversible damage to auditory or vestibular branches of the cranial nerve leads to hearing loss or balance issues. 2. Nephrotoxicity(esp. with cephalosporins) Reversible damage to renal tubules; risk increases with prolonged use or high doses. Neomycin, Tobramycin, Gentamicin ( most nephrotoxic) 3. Neuromuscular Blockade: It causes NM blockade by interfering with acetyl choline release from motor nerve by blocking Ca++ ion 4. Allergic Reactions: Rare, but hypersensitivity reactions may occur, especially with topical formulations. 32 16  11/25/2024 Mechanism of Resistance Bacterial resistance to aminoglycosides occurs via one of four mechanisms that prevent the normal binding of the antibiotic to its ribosomal target: 1. Modification or inactivation of the antibiotic "enzymatically“ such as; a. acetyltransferase (ATC, AAC) which adds an acetyl group to the antibiotic to form acetyl CoA b. Adenylyl transferase or nucleo adenyl transferase which adds adenine from ATP c. Phosphoryl transferase Which adds phosphate to OH of the antibiotic. 2. On target initiation Complex: modification or mutation al 16SrRNA "30S“ 3. Efflux pumps: protein in the cytoplasmic membrane can exclude or egress or remove antibiotics to the outside. They prevent the accumulation of the acetyl aminoglycoside in the cytosol of the bacterium. 4. production of Ribosomal protective proteins 'RPPs" 33 Plazomicin Plazomicin is a semi-synthetic Bactericidal aminoglycoside developed to overcome resistance mechanisms that limit the efficacy of older aminoglycosides It is particularly effective against multidrug-resistant (MDR) Gram-negative bacteria, including carbapenem-resistant Enterobacteriaceae (CRE). Plazomicin evades most aminoglycoside-modifying enzymes (e.g., acetyltransferases, phosphotransferases, and nucleotidyltransferases). Resistance Mechanisms  Although plazomicin resists most aminoglycoside-modifying enzymes, it may still be affected by ribosomal mutations (16srRNA) or efflux pumps in certain bacterial strains. 34 17  11/25/2024 Spectrum of Activity  It is effective against Gram-negative bacteria. It is mainly potent against CRE and other extended-spectrum beta-lactamases (ESBL)-producing bacteria causing urinary tract infections (UTI)  Klebsiella pneumoniae (CRE), Escherichia coli, Enterobacter spp, Proteus mirabilis, Serratia marcescens, Shigella, and Salmonella.  It has limited or no activity against Pseudomonas aeruginosa since it has a highly impermeable outer membrane, limiting drug entry. Moreover, It possesses multiple efflux pumps that actively expel aminoglycosides, including plazomicin.  Reduced activity against Acinetobacter baumannii compared to other aminoglycosides.  Limited activity against Gram-positive and anaerobic bacteria. 35 Arbekacin A semisynthetic aminoglycoside developed to combat antibiotic-resistant bacteria, particularly methicillin-resistant Staphylococcus aureus (MRSA) and multidrug-resistant Gram-negative bacteria. Mechanism of Action  Arbekacin, acts on both 30S and 50S ribosomal subunits, interfering with bacterial protein synthesis. Besides, It has a damaging effect on bacteria membranes (double activity on the bacteria)  Arbekacin resists inactivation by many aminoglycoside-modifying enzymes that render other aminoglycosides ineffective.  Against MRSA infection, an effective dosage of arbekacin was reported to induce dramatic changes in the biofilm membranous structure as well as in the inflammatory response, resulting in eradication of the biofilm structure and resolution of inflammation. 36 18  11/25/2024 Spectrum of Activity 1. Gram-positive Bacteria: o Highly effective against MRSA. o Useful against other resistant Staphylococcus and Enterococcus species. 2. Gram-negative Bacteria: o Active against some resistant strains, including Pseudomonas aeruginosa and Acinetobacter baumannii. (highly resistant gentamycin organisms) o Limited activity against certain aminoglycoside-resistant Gram-negative organisms. Notes:  Tobramycin/Gentamycin/Arbekacin: Effective against Pseudomonas.  Streptomycin/Kanamycin/Amikacin: Effective against Mycobacterium tuberculosis. 37 Combination Therapy Aminoglycosides are often combined with other antibiotics to enhance efficacy:  Beta-lactams or glycopeptides: Synergistic effect against Gram-positive bacteria by disrupting cell walls, facilitating aminoglycoside entry (improving the permeability). Ex: cephalosporins or Penicillin  Reduces the risk of resistance emergence when treating serious infections. 38 19  11/25/2024 3. OXAZOLADINONES: Linezolid Mechanism of Action Linezolid inhibits bacterial protein synthesis by binding to the 23S rRNA of the 50S ribosomal subunit. This prevents the formation of the 70S initiation complex, which is essential for bacterial translation and protein synthesis. Bacteriostatic for most bacteria (e.g., Staphylococcus and Enterococcus). Available as oral and intravenous (IV) formulations, with excellent bioavailability (~100%), making the oral form as effective as the IV. Resistance 1. Mutations in 23S rRNA: Decreases binding affinity of linezolid. 2. Efflux Pumps 39 Mechanism Of Action 40 20  11/25/2024 Spectrum of Activity Broad Gram-positive, aerobic, and anaerobic bacteria Coverage  Effective primarily against Gram-positive bacteria, including: o MRSA (Methicillin-resistant Staphylococcus aureus). o VRE (Vancomycin-resistant Enterococcus). o Penicillin-resistant Streptococcus pneumoniae. o Clostridium perfringens and Clostridium tetani.  Ineffective against Gram-negative bacteria due to efflux pumps and inability to penetrate the outer membrane. 41 Tedizolid Tedizolid is a second-generation oxazolidinone antibiotic, It is approved for the treatment of multidrug-resistant Gram-positive bacteria, including MRSA (methicillin-resistant Staphylococcus aureus) and VRE (vancomycin-resistant Enterococcus). Mechanism of Action  Tedizolid, like linezolid, inhibits bacterial protein synthesis by binding to the 23S rRNA of the 50S ribosomal subunit, preventing the formation of the 70S initiation complex. Note: more curing effect than linezolid (better and faster therapeutic ratio). Note: active against linezolid-resistant organisms. Advantages of Tedizolid Over Linezolid 1. Improved Potency: Tedizolid is more potent than linezolid, requiring lower doses. It is long-acting (I dose/day) 2. Shorter Treatment Duration 3. More efficient (less resistance) 42 21  11/25/2024 4. Tetracyclines The name "tetracycline" refers to several bacteriostatic antibiotics of either natural or semi- synthetic origin They are so named for their four (“tetra-”) hydrocarbon rings  Examples are: Tetracycline, Oxytetracycline, Lymecycline, Methacycline, Doxycycline, and Minocycline. A newer generation is Tigecycline The structure of Novel Third-Generation Tetracyclines: tetracycline Eravacycline, Omadacycline, and Sarecycline: 43 Spectrum of activity Bacteriostatic (almost always) Broad spectrum: Wide range of aerobic and anaerobic Gram-positive and Gram-negative bacteria including Rickettsia, V. cholera, M. pneumonia, Chlamydia, Shigella, H. pylori, P. tularensis, P. pseudomallei, Brucella, Psittacosis, Borrelia Selectivity results from transfer into bacterial cells but not mammalian cells Active against multiplying bacteria Active against Propionibacterium acne Effect is reversible 44 22  11/25/2024 Mechanism of Action Penetrates bacterial cells passively (due to hydrophilicity) and actively (energy- dependent). They reversibly bind to the 30S ribosomal subunit of bacteria, blocking the binding of aminoacyl-tRNA to site A (Acceptance site)on the mRNA ribosome complex. This prevents the addition of amino acids to the growing peptide, resulting in the inhibition of protein synthesis Quickly bacteriostatic drugs, but at high dosages, they are also bactericidal. 45 Tetracyclines: Mode of Action Reversible binding to 30S ribosome subunit blocking aminoacyl-tRNA access to acceptor site (A site) 50S aa-tRNA tRNA X mRNA P A 30S 46 23  11/25/2024 Resistance Bacterial resistance to tetracyclines is mainly due to the following mechanisms: 1. Decreased intracellular accumulation owning to either impaired influx or increased efflux by an active transport protein pump; Production of novel cytoplasmic membrane protein that mediates active efflux of the drug so that inhibitory levels are not maintained within cells 2. Enzymatic inactivation: Involves specific enzymes (e.g., oxidoreductase, monooxygenase enzyme). 3. Target modification: Alteration of ribosomal binding sites. 4. Ribosome protective proteins (RPPs): Shield the ribosome from tetracycline binding. Newer derivatives, such as glycylcyclines like tigecycline, have been developed to overcome some resistance mechanisms. 47 Toxicity Gastrointestinal discomfort/distress: Anorexia, epigastric pain, abdominal distention Hepatotoxicity (Liver toxicity): Hepatic injury- Increased during pregnancy  Renal toxicity (Nephrotoxicity)  Discoloration of enamel (teeth) and hypoplasia of teeth Deposition in fetal and growing bones, inhibits bone growth in children (stunted growth)  Photosensitization  Severe sunburn in sun; doxy/demeclocycline  Potentially teratogenic  Vestibular toxicity Chelation Properties: o Tetracycline chelates divalent ions like Mg²⁺, Al³⁺, and Ca²⁺. o Should not be taken with milk, antacids, or iron-containing products as they reduce absorption. ( may lead to osteoporosis, separate intake at least 2 hours) 48 24  11/25/2024 Third generation tetracyclines Eravacycline, Omadacycline, and Sarecycline are all antibiotics belonging to the tetracycline class, specifically third-generation tetracyclines, with Sarecycline sometimes considered a fourth-generation due to its narrow spectrum of activity. Eravacycline Usage: Primarily used for treating complicated intra-abdominal infections (anaerobes). Administration: Intravenous (IV) only. Activity: Effective against a broad range of bacteria, including some resistant strains (Acinetobacter spp.) Resistance: due to upregulation of the multidrug efflux system AcrA-AcrB-TolC presence of adeB genes in VRE and CRE 49 Omadacycline Usage: Approved for community-acquired bacterial pneumonia and acute bacterial skin and skin structure infections. Administration: Available in both oral and IV formulations. Activity: Broad-spectrum activity, active against anaerobes (Clostridium difficile), designed to overcome tetracycline resistance. Resistance: Ribosome protective proteins (RPPs) and mutations in 16SRNA Sarecycline Usage: Specifically used for treating moderate to severe acne vulgaris. Administration: Oral. Activity: Narrow spectrum, which helps reduce the impact on gut microbiota, leading to fewer gastrointestinal side effects. 50 25  11/25/2024 5. Macrolides Characterized by Macrocyclic lactone rings + deoxy sugars Prototype: Erythromycin (Natural), given to patients allergic to penicillins Semisynthetic derivatives: clarithromycin, ketolides and azithromycin Active against Pneumococci, Streptococci, Staphylococci, H. Pyroli, and atypical bacteria (Rickettsia spp and Chlamydia spp.) while Hemophilus influenza and Campylobacter are less susceptible 51 Mechanism of action Macrolides bind tightly to the P site of 50S subunit of the bacterial ribosome, thus blocking the Aminoacyl translocation (exit of the newly synthesized peptide and formation of initiation complex Thus, they are interfering with bacterial translation. Mechanism of resistance Resistance is usually plasmid-encoded. Several mechanisms have been identified:  Reduced permeability of the cell membrane  Active efflux  production (by Enterobacteriaceae) of esterases that hydrolyze macrolides  Modification of the ribosomal binding site (so-called ribosomal protection) by chromosomal mutation or by a macrolide-inducible or constitutive methylase. 52 26  11/25/2024 53 54 27  11/25/2024 Azithromycin It is effective against respiratory infections, skin infections, ear infections, eye infections, and sexually transmitted diseases. Roxithromycin (second generation) It treats various bacterial infections, including respiratory tract, urinary, and soft tissue infections. Roxithromycin is effective against a range of bacteria, including Streptococcus, Staphylococcus, and Mycoplasma. Common side effects include gastrointestinal issues such as diarrhea, nausea, and abdominal pain Clarithromycin Acts on H. pylori Telithromycin and solithromycin (fluoro compounds) Telithromycin and Solithromycin are both Ketolide antibiotics used to treat bacterial infections, particularly community-acquired bacterial pneumonia (CABP). Side Effects: gastrointestinal issues (diarrhea, nausea, abdominal pain), headache, and disturbances in taste. Rare but severe side effects include liver damage. Solithromycin is a Fluoroketolide antibiotic Usage: Also treats CABP, including macrolide-resistant strains. Side Effects: Infusion-site reactions and transient liver enzyme elevations. Leucomycin and kitasamycin Animal use but induces resistance to humans 55 6. Lincosamides Lincomycin and Clindamycin Lincomycin/ Clindamycin are members of the lincosamide series of antibiotics Clindamycin is a chlorine-substituted derivative of lincomycin Clindamycin binds to the 50S subunit of the ribosome to inhibit protein synthesis by interfering with the formation of initiation complexes and translocation reaction. Clindamycin is more clinically used than Lincomycin 56 28  11/25/2024 Spectrum: Narrow Gram (+) spectrum Excellent activity against anaerobic bacteria; strep, pneumococci, staphylococci Resistance: Mutation of the ribosomal receptor site Modification of the receptor (23S) by a constitutively expressed methylase enzymatic inactivation Toxicity of Clindamycin Clindamycin kills many components of the gastrointestinal flora, leaving only Clostridium difficile. This can result in overgrowth by C. difficile, which is resistant - Antibiotic associated diarrhoea and caused by toxigenic C. difficile Diarrhea, nausea, skin rashes, impaired liver function and neutropenia 57 Uses of Clindamycin Main utility is in treatment of Gram-positive bacteria and anaerobic bacteria Active against Staphylococcus, including some strains of MRSA Not useful against Gram-negative bacteria Clindamycin is indicated for treatment of anaerobic infection caused by bacteroides and other anaerobes that often participate in mixed infections. Clindamycin, sometimes in combination with an aminoglycoside or cephalosporin, is used to treat penetrating wounds of the abdomen and the gut; infections originating in the female genital tract, eg, septic abortion and pelvic abscesses; and aspiration pneumonia. Clindamycin is now recommended rather than erythromycin for prophylaxis of endocarditis in patients with valvular heart disease who are undergoing certain dental procedures. 58 29  11/25/2024 7. Pleuromutilin Pleuromutilin (Semi-Synthetic /Natural), is primarily produced by a fungus. They include the licensed drugs Lefamulin (for systemic use in humans), retapamulin (approved for topical use in humans), valnemulin, and tiamulin (approved for use in animals) Mechanism They inhibit protein synthesis in bacteria by binding to the peptidyl transferase component of the 50S subunit of ribosomes. 59 Clinical uses: acute bacterial skin and skin structure infections, sexually transmitted infections, hospital- acquired bacterial pneumonia, osteomyelitis, and prosthetic joint infections. Spectrum Lefamulin exhibits excellent potency against a broad spectrum of Gram-positive bacteria, especially multidrug-resistant isolates that cause skin and ocular infections. They have also demonstrated potent activity against Chlamydia trachomatis, the leading cause of blindness in the world, and Propionibacterium acnes, the causative agent of acne. Resistance They are associated with low rates of resistance development. Additionally, they display minimal cross- resistance with other antibiotics that target the bacterial ribosome. Three mechanisms of resistance to pleuromutilin have been identified; 1. Mutations in 23S rRNA and rplC genes encoding the ribosomal protein L3, 2. Methylation of the nucleotide A2503 by Cfr methyltransferase which is activated by Erm gene (responsible for several macrolides resistance) 3. Drug efflux by ATP-binding cassette. 60 30  11/25/2024 8. Chloramphenicol Chloramphenicol was the first broad-spectrum antibacterial developed. Chloramphenicol binds to the bacterial 50S ribosomal subunit and inhibits protein synthesis at the peptidyl transferase reaction. Attaches at P sites of 50 S subunit of microbial ribosomes and inhibits the functional attachment of aminoacyl end of AA-t-RNA to 50S subunit (A site). Failure to properly align prevents the Peptidyl transferase enzyme from transferring the growing chain from the "P" site to the bound charged tRNA in the "A" site. This stops protein synthesis. 61 Antimicrobial Activity Broad spectrum antibiotic. Chloramphenicol is primarily bacteriostatic, but it may be bactericidal to some strains of microorganisms even at lower concentrations: H. influenzae, N. meningitidis N. Gonorrhea, and Bacteroides fragilis Bacteriostatic for – S. epidermidis, S. aureus, M. pneumonia, L. monocytogenes, Corynebacterium diphtheriae, P. multocida, Salmonella sp., Shigella sp., E. coli, Rickettsia and Anaerobes Ineffective for Chlamydial infections More effective than Tetracyclines against Typhoid Fever and other Salmonella infections. 62 31  11/25/2024 Clinical uses It may be considered for treatment of serious rickettsial infections such as typhus and Rocky Mountain spotted fever. It is an alternative to beta-lactams for the treatment of meningococcal meningitis occurring in patients who have major hypersensitivity reactions to penicillin or bacterial meningitis caused by penicillin-resistant strains of pneumococci. Used as a backup drug for severe infections caused by Salmonella Typhi and Paratyphi Infections caused by anaerobes like B. fragilis Commonly used as a topical agent Occasionally used topically in the treatment of eye infections for its good penetration to ocular tissues and the aqueous humor 63 Mechanism of Resistance Chloramphenicol resistance has been attributed to decreased membrane permeability due to the loss of an OMP that results in impaired penetration of the drug to the target site. - Production of chloramphenicol acetyltransferase (CAT); / chloramphenicol transacetylase, a plasmid-encoded enzyme that inactivates the drug. Adverse effects Chloramphenicol causes a dose-related reversible suppression of red cell production at high dosages causing Aplastic anemia (a rare consequence) or in Premature infants: Gray baby syndrome gray baby syndrome Newborn infants lack an effective glucuronic acid conjugation mechanism for the degradation and detoxification of chloramphenicol (Mainly due to deficiency of hepatic glucosyl transferase) Symptoms: vomiting, flaccidity, hypothermia, gray color, shock. 64 32  11/25/2024 9. STREPTOGRAMINS Group A streptogramins contain 23-membered unsaturated rings with lactone and peptide bonds. Group B streptogramins are cyclic hexa- or hepta-depsipeptides produced by various members of the Streptomyces genus. They have been used to treat a range of bacterial infections, including those due to vancomycin-resistant Staphylococcus aureus (VRSA) and vancomycin-resistant enterococci (VRE). They include pristinamycin, (quinupristin + dalfopristin), and virginiamycin Quinupristin (Streptogramin B)/Dalfopristin (Streptogramin A) (30:70) (Synercid) 65 Mechanism of action The streptogramins block protein synthesis by attaching to the 50S portion of the ribosome. Dalfopristin binds to the 23S portion of the 50S ribosomal subunit, changing the conformation of it and enhancing the binding of quinupristin by a factor of about 100. In addition, it inhibits peptidyl transfer. Quinupristin binds to a nearby site on the 50S ribosomal subunit and prevents elongation of the polypeptide, as well as causing incomplete chains to be released. Activity: -Action is similar to macrolides except bactericidal for Staphylococcus and most organisms except Enterococcus faecium -Prolonged post-antibiotic effect up to 10 h for Staphylococcus aureus 66 33  11/25/2024 Nascent polypeptide 50S DALFOPRISTIN A chain QUINUPRISTIN (MACROLIDE) Transferase site aa mRNA template P 30S 67 Resistance Resistance to streptogramins B is due either to 1. Hydrolysis of the antibiotic mediated by the vgb gene initially reported in Staphylococcus aureus 2. Modification of the ribosomal target by a 23S rRNA methylase encoded by the ermB gene resulting in MLSB phenotype Notice Three main mechanisms of resistance to MLS (Macrolides-Lincosamides- Streptogramins B) antibiotics have been described: 1. Methylation of rRNA (target modification) Target modification is achieved through the action of the protein product of one of more than 42 different erm (erythromycin rRNA methylase) genes. They confer cross-resistance between macrolides, lincosamides, and streptogramin B (so-called MLSB resistance) 2. active efflux. (M and L only) 3. inactivation of the antibiotic. (M and L only) Based on the mechanisms of resistance, various resistant phenotypes are expressed. The most prevalent phenotypes are ΜLSB (constitutive or inducible), which is associated with the presence mainly of ermA and ermC genes. 68 34  11/25/2024 10. Fusidic acid Fusidic acid is also sometimes known as sodium fusidate. It is from a natural origin. It works by stopping bacteria from growing. It is a narrow-spectrum antibiotic mainly against gram-positive. Fusidic acid is active in vitro against Staphylococcus aureus, most coagulase- positive staphylococci, and Beta-hemolytic streptococci. Fusidic acid has no known useful activity against enterococci or most Gram- negative bacteria. It's used to treat bacterial infections, such as skin infections including cellulitis and impetigo, and eye infections including conjunctivitis (red, itchy eyes). Fusidic acid is only available on prescription. It comes as a cream, ointment, or eye drops. It's also available with a steroid as a combined cream. 69 Mechanism of action Fusidic acid binds to EF-G after translocation and GTP (guanosine-5'- triphosphate) hydrolysis. This interaction prevents the necessary conformational changes for EF-G release from the ribosome, effectively blocking the protein synthesis process. Fusidic acid can only bind to EF-G in the ribosome after GTP hydrolysis. Since translocation is a part of elongation and ribosome recycling, fusidic acid can block either or both steps of protein synthesis. It also can inhibit chloramphenicol acetyltransferase enzymes. 70 35  11/25/2024 Mechanism of resistance 1. Alteration in EF-G: point mutations in fusA, the chromosomal gene that codes for EF-G. The mutation alters EF-G so that fusidic acid is no longer able to bind to it. 2. FusB type resistance genes mediated by fusB, fusC, and fusD genes found primarily on plasmids, and have also been found in chromosomal DNA. Active efflux of the drug Increased permeability of the bacterial membranes 71 2.Inhibition of DNA synthesis 72 36  11/25/2024 1.Fidaxomicin It is the first member of a class of narrow-spectrum macrocyclic (18 carbon) antibiotic Fidaxomicin is minimally absorbed into the bloodstream when taken orally for GIT infections Dose:1-2 doses /day. Effective for 5-10 hours after stopping the dose (long-acting) Spectrum: (Aerobic and Anaerobic Gram +ve) 1. it is bactericidal and selectively used in CDI to eradicate pathogenic Clostridium difficile with relatively little disruption to the multiple species of bacteria that comprise the normal, healthy intestinal microbiota. C. difficile is an anaerobic Gram-positive that causes severe ulcerative colitis. When used orally for 3 days it prevents sporulation of bacteria. 2. MRSA(Methicillin-resistant Staphylococcus aureus ) 3. VRSA (Vancomycin-resistant Staphylococcus aureus ) 4. Mycobacterium tuberculosis. It was approved by FDA in 2011 and used in 2012. Not 1st line treatment because it is costly. Can be taken or without food In CDI, Fidaxomicin + Vancomycin → more effective → increase curing rate, decrease rate of recurrence. 73 Mechanism of action of Fidaxomicin It acts on DNA-dependent RNA polymerase (RNA polymerase consists of five subunits: two alpha (α) subunits, a beta (β), a beta prime subunit (β′), and a small omega (ω) subunit. Resistance:-poly-mutation of DNA-dependent RNA polymerase (specifically in the beta (β) subunit {RPB} 74 37  11/25/2024 2. Rifamycins  Natural and Semisynthetic derivative of Rifamycin B  Rifampicin, rifabutin, rifapentine, rifaximin  Bactericidal to M. Tuberculosis and many other Gram(+) and Gram (-) bacteria like S. aureus, N. meningitidis, H. influenza, E. coli, Klebsiella, Pseudomonas, Brucella, Proteus and Legionella (broad spectrum) Mechanism of action  Interrupts RNA synthesis by binding to β subunit of mycobacterial DNA-dependent RNA polymerase (encoded by rpoB gene )  Therefore, it prevents the elongation of RNA  These drugs should be used in combinations to prevent resistance 75 Mechanism Of Resistance Rifampin resistance is nearly always due to mutation in the rpoB gene reducing its affinity for the drug No cross-resistance with any other antitubercular drug Rifaximin not absorbed orally  Imp. in diarrhea (GIT) (Travel’s diarrhea) - especially because of E. coli Used in combinations with Fluoroquinolones for IBS (Irritable bowel Syndrome) Oral - injections – suspension  brown color in stools or urine– brown teeth of children and also contact lenses (permanent)  Multiple rug interactions especially (cardiac and neuro drugs) Prophylaxis in Meningitis (endogenous bacteria) → due to lower immunity Combination of doxycycline and rifampin is first-line therapy for brucellosis 76 38  11/25/2024 3. 5-Nitroimidazoles Bactericidal against anaerobes especially Bacteroides fragilis, clostridia species and other colon diseases.Has no aerobic activity Metronidazole (flagyl) - Tinidazole -Minordiazole _Megazole They are prodrugs activated by nitro-reductase enzyme by anaerobic bacteria. Taken either alone or in combination. Highly Resistant because of OTC use Resistance by decreasing uptake of drug and decreasing reduction efficiency Effective against protozoa: E. histolytica, Giardia lamblia, trichomonas. Used in surgeries to prevent sepsis Not good for UTI (not absorbed) For Gardinella infections, it should be by injection Dimetridazole: was used for veterinary purposes banned in Europe (carcinogenic) 77 MECHANISM OF ACTION 78 Antimicrobial action is due to the accumulation of short- lived intermediate compounds (produced by reduction inside anaerobic bacteria after uptake) or free radicals that damage DNA and other cellular macromolecules Nitro gp converted to nitrite radical by low redox potential. Metabolites covalently bind to DNA, disturb its helical structure, fragmentation of microbial chromosomes. Finally inhibits nucleic acid biosynthesis bacterial death Resistance: -differential expression of reductase enzymes (Fe and O2 will prevent activation) - Alteration of target 78 39  11/25/2024 4. 5-Nitrofurans Nitrofurans enter bacterial cells and interact with several enzymes; thereby inhibiting bacterial growth. They are metabolized by bacterial nitro reductases, which convert nitrofurans to a highly reactive electrophilic intermediate that attacks bacterial ribosomal proteins, causing complete inhibition of protein synthesis. 79 NITROFURANTOIN It is a synthetic, orally active, bactericidal, antibacterial agent used for the treatment of urinary tract infections (UTIs). Nitrofurantoin is active against many Gram-positive and Gram-negative bacteria. It is particularly useful in treating urinary pathogens, including Escherichia coli, Enterococci, Staphylococcus aureus, and susceptible strains of Klebsiella and Enterobacter. However, Pseudomonas aeruginosa is almost always resistant. Resistance has been recorded. Step-wise mutations of different nitro-reducing activities are identified as the reason behind the progressive increase in resistance patterns. These genes are mapped as nfsA and nfs B genes. However, like many antibiotics, it can disrupt the natural balance of bacteria and yeast in the body, potentially leading to yeast infections, such as candidiasis. Moreover, Long-term (3 months or longer) nitrofurantoin increases the risk of pulmonary toxicity and hepatic toxicity. If receiving long-term, liver function and lung function should be monitored. 80 40  11/25/2024 NITROFURAZONE NITROFURAZONE is an effective drug that acts on several Gram-positive and Gram-negative microorganisms (staphylococci, streptococci, dysentery bacillus, colon bacillus, paratyphoid bacillus, and others). It is generally used externally for treating and preventing the pro-inflammatory processes, and internally for treating bacterial dysentery. Wounds are irrigated and wet bandages are synthesized using nitrofurazone. It is used in the form of eye drops for practically all suppurative processes that require the use of antibacterial drugs 81 Furazolidone Furazolidone is a nitrofuran antibacterial agent with both antiprotozoal and antibacterial properties. It is primarily used to treat bacterial and protozoal diarrhea and enteritis caused by susceptible organisms. It has been used in both human and veterinary medicine. In humans, it has been used to treat conditions such as traveler's diarrhea, cholera, and infections caused by Giardia lamblia and Helicobacter pylori. 82 41  11/25/2024 5. Quinolones Bactericidal broad-spectrum antibiotics DNA gyrase inhibitors Increasingly used because of their relative safety, their availability both orally and parenterally and their favorable pharmacokinetics Excellent oral absorption * Absorption reduced by antacids * BASIC STRUCTURE 4-QUINOLONE o Nalidixic acid Pyridone ring * o Fluoroquinolones * *Required for antibacterial activity 83 Fluoroquinolones The fluoroquinolones are a family of synthetic, broad-spectrum antibacterial agents with bactericidal activity. The parent of the group is nalidixic acid. It was used orally for the treatment of infections caused by gram-negative organisms. The newer fluoroquinolones have a wider clinical use and a broader spectrum of antibacterial activity including gram-positive and gram-negative aerobic and anaerobic organisms Examples : ciprofloxacin, ofloxacin, levofloxacin, moxifloxacin 84 42  11/25/2024 Mechanism of Action The fluoroquinolones enter the bacterium by passive diffusion through water-filled protein channels (porins) in the outer membrane. Once inside the cell, they inhibit 2 essential bacterial enzymes 1. DNA gyrase {Topoisomerase II}(The main target in Gram negative bacteria) 2. Topoisomerases IV (The main target in Gram +ve bacteria) DNA gyrase is responsible for supercoiling of DNA This enzyme relaxes tightly wound chromosomal DNA, thereby allowing DNA replication to proceed Quinolones interfere with DNA gyrase and prevent conversion of relaxed DNA to super-coiled DNA. Topoisomerase IV decatenates or removes the interlinking of daughter DNA strands. Quinolones interfere with topoisomerse IV & prevent decatenation of DNA Basis for Selective Toxicity Quinolones have a relatively low affinity for mammalian DNA topoisomerase 85 86 43  11/25/2024 doi:10.1016/j.jmb.2005.06.029 87 MECHANISM OF RESISTANCE 1. Chromosomal mediated A. Alterations in the targets of quinolones Single step mutation in DNA gyrase & topoisomerase IV High level of resistance was resulted due to Mutation in quinolone resistance-determining regions (QRDR) including gyrA, gyrB, parC and parE genes B. Decreased accumulations of drug inside bacteria i. Impermeability of the membrane and/or ii. Decrease in level of expression of OmpF porin in E. coli iii. An over-expression of efflux systems. 88 44  11/25/2024 89 MECHANISM OF RESISTANCE 2. Plasmid mediated quinolone resistance (PMQR)  responsible for low level of resistance.  PMQR detected only in Enterobacteriacae  Three known PMQR mechanisms to date: A. Qnr determinants (qnrA, qnrB, qnrS, qnrD, qnrC)  Protect DNA gyrase & topoisomerase IV from quinolone inhibition. B. Aminoglycoside acetyltransferase (aac(6)-Ib-cr) gene.  aac(6)-Ib-cr is a variant of aac(6)-Ib and is responsible for reduced susceptibility to ciprofloxacin or norfloxacin by N-acetylation of a piperazinylamine C. QepA (quinolone efflux pump) 90 45  11/25/2024 Nalidixic acid, cinoxacin Levofloxacin, Gatifloxacin, Sparfloxacin, Grepafloxacin ciprofloxacin, lomefloxacin, and ofloxacin. Trovafloxacin, Moxifloxacin, Gemifloxacin 91 Clinical uses Empiric therapy of Community-Acquired Pneumonia Oral therapy of un-/complicated UTI or RTI Oral therapy of serious infections such as osteomyelitis, pneumonia or soft tissue infections Treatment of STD: gonorrhea, chancroid, chlamydial urethritis Empiric therapy of traveler's diarrhea Legionellosis Treatment of typhoid Therapy for multidrug-resistant tuberculosis 92 46  11/25/2024 Side effects -The fluoroquinolones as a class are generally well tolerated. Most adverse effects are mild in severity, self-limited, and rarely result in treatment discontinuation. However, they can have some serious adverse effects. -Fluoroquinolones are approved for use only in people older than 18. They can affect the growth of cartilage in a child or fetus. -The FDA has assigned fluoroquinolones to pregnancy risk category C, indicating that these drugs have the potential to cause teratogenic or embryocidal effects. -These agents are also excreted in breast milk and should be avoided during breast-feeding if at all possible 93 Gastrointestinal effects. The most common adverse events experienced with fluoroquinolone administration are gastrointestinal (nausea, vomiting, diarrhea, constipation, and abdominal pain), which occur in 1 to 5% of patients. CNS effects. -Headache, dizziness, and drowsiness have been reported with all fluoroquinolones. Phototoxicity. Tendon damage (tendon rupture). Hepatoxicity. Cardiovascular effects. Glucose homostasis abnormalities (Hypoglycemia or hyperglycemia). 94 47  11/25/2024 3. Inhibition of Folic Acid Synthesis 95 What is Folic Acid? Folic acid is necessary for synthesizing amino acids and hence for bacterial protein synthesis. i.e. Bacteria synthesize folic acid; however, humans should take it as a supplement. What is the mechanism of folic acid synthesis? Di hydro folic acid tetrahydro folic acid Further reduction 96 48  11/25/2024 2- Amino-4-hydroxy-6 - hydroxymethyl-7,8 - dihydropteridine Pyrophosphate P-amino benzoic Dihydropteroate synthetase acid 97 Anti-DHPS Specific to bacteria Sulfonamides  Bacteriostatic  Binds and blocks enzymes mainly folate synthase, and dihydrofolate reductase which is responsible for folic acid synthesis.  Taken orally or parenterally. - should drink water because it is nephrotoxic (makes crystals).  Active against many G+ and G- but inactive on P. aeruginosa and Enterococcus. MOA: Acts by competition on PABA on DHPS Resistance: 1. Modification of DHPS. 2. mutation and acquisition of Sul1, Sul2, and Sul3 genes (plasmid-mediated genes) Sulphonamides can be taken For animals (Veterinary) by ingesting these animals, humans can acquire sulphonamides resistance. 24 June 2016 98 49  11/25/2024 Anti-DHFR Trimethoprim (Synthetic, Bacteriostatic) Oral tablets, bind to DHFR in bacteria (quantitative selectivity) 1000 times more than humans resulting in very little toxicity in humans. Spectrum: on both G+ and G-.Used in UTI, and RTI but inactive against P. aeruginosa Resistance: Mutation of DHFR, or overproduction. Sulfonamides and trimethoprim, are synergistic. Sulphamethoxazole (tablets or suspension) Sulfadoxine + pyrimethamine → Antiprotozoal These combinations are with a broader spectrum, less toxic, less resistance pyrimethamine → same mechanism but more on protozoa, malaria, toxoplasmosis methotrexate (DHFR inhibitor, anti-cancer)→ very toxic. 24 June 2016 99 50