Antibiotics 2024 PDF

Antimicrobia l Drugs Antimicrobial drugs: All agents that interfere with the growth of microbes. a). Synthetic (Man-made in the laboratory (non antibiotic) -sulfonamides and quinolones compounds and had limitations in terms of safety and efficacy) b). Natural antimicrobials....

Antimicrobia l Drugs Antimicrobial drugs: All agents that interfere with the growth of microbes. a). Synthetic (Man-made in the laboratory (non antibiotic) -sulfonamides and quinolones compounds and had limitations in terms of safety and efficacy) b). Natural antimicrobials. (i). From plants (alkaloids) or animals (lysosyme). (ii). Antibiotics: From bacteria and fungi. Antibiotic: Any of a large group of natural chemical substances produced by various microorganisms and fungi, having the capacity in dilute solutions to inhibit the growth of or to destroy bacteria. c). Semisynthetic antibiotics: Examples -methicillin and amoxicillin. Are molecules produced by a microbe that are subsequently modified by an organic chemist to enhance their antimicrobial properties Synthetic antimicrobials 1910 - Salvarsan or compound 606 a drug that was introduced at the beginning of the 1910s as the first effective treatment for syphilis. -Sulfonamides (Prontosil) -1932. A Prodrug: metabolised (i.e., converted within the body) into a pharmacologically active drug. No activity against bacteria in the test-tube. Natural source of antimicrobials Antibiotics 1928: Fleming discovered penicillin 1940: Howard Florey and Ernst Chain performed first clinical trials of penicillin.  Spectrum of antimicrobial activity Broad spectrum antibacterials are active against both Gram-positive and Gram-negative organisms. Examples: Tetracyclines, fluoroquinolones. Inhibit protein synthesis by preventing the attachment of aminoacyl-tRNA to the ribosomal acceptor (A) site. Narrow spectrum antibacterials have limited activity and are primarily only useful against particular species of microorganisms. Example: 1. Bacitracin is only effective against Gram-positive bacteria.  Selective toxicity: Drug kills pathogens without damaging the host. Why do antimicrobials have selective toxicity? Biochemical pathways in the bacteria are in some way different from those in the animal cells. This permits interference with bacteria biochemical pathways without effecting the host. Antimicrobials can be; General 1. Bactericidal kills the pathogen. Features of 2. Bacteriostatic effect (stops bacteria reproducing) and allows the Antimicrobial immune system to overcome the pathogen. Prevents toxic shock syndrome. Drugs In treating urinary tract infections and preventing staphylococcal wound infections, studies have shown that bacteriostatic drugs work as well as bactericidal drugs. In central nervous system infections, a rapidly bactericidal drug can release bacterial products that stimulate inflammation Properties of the ideal antimicrobial drug ? Have highly selective toxicity to the pathogenic microorganisms in host body. No toxicity or less toxicity to the host. Low tendency for the development of resistance to the agent by the microorganism. Should not induce hyper-sensitivities (immune response) in the host. Immune system should not mistakenly identifies a drug as a harmful substance(this can occur with penicillin). Have rapid and extensive tissue distribution. (the transfer of a drug from one location to another within the body). Must reach sufficient concentrations at the site of infection. This is dependent on vascular permeability, regional blood flow, cardiac output etc. Be free of interactions with other drugs. Be relatively inexpensive. ANTIBIOTICS … Important factor for consideration when prescribing antibiotics Stability, Tissue distribution, metabolism, and excretion. 1. Stability in acid. (Determines oral or intravenous administration). 2. Distribution of the antimicrobial. Describes the transfer of drug from one location to another within the body. 3. Metabolism. Sulfonamides -No effect observed in the test tube, ONLY exerting antibacterial action in live animals, WHY? the drug must be metabolised into two parts inside the body, releasing the active compound called sulfanilamide. (PRODRUG). 4. Rate of elimination of drug from body -expressed in half-life of the antibiotic. Half-life: Time it takes for the body to eliminate one half the original dose in serum. Half-life dictates frequency of dosage. How many capsules/day? Patients with liver or kidney damage tend to excrete drugs more slowly. 5. Adverse effects of certain antimicrobials – Allergic reactions, Allergies to penicillin – Toxic effect, Aplastic anemia -Body cannot make RBC or WBC – Suppression of normal flora, Antibiotic associated colitis Clostridium difficile bacteria – Intolerance -diarrhoea, rash.  Antimicrobial stewardship The key elements of antimicrobial stewardship are to ensure you: – prescribe the right antibiotic, antiviral, antifungal for the patient – consider age, medical conditions, pregnancy, or long-term care resident. – choose the right dose, duration, and route for the condition you are treating The main factors affecting antibiotic prescription were a high work pressure, insufficient staff resources, uncertainties regarding clinical decisions. Treatment expectations from patients and next of kin, suboptimal microbiological testing, and limited time for infectious disease specialists to offer advisory services also affected the antibiotic choices.  Factors Influencing Antibiotic Prescription Patient related factors Patient expectation of receiving antibiotics Economic status of patient affects both the actual and perceived need for antibiotics Patient social characteristics (education, occupation) have impacted prescription trends Patients self medicating and self diagnosis request specific antibiotics Physician related factors Physicians who actively refresh their expertise, medical training, workshops and journals are less likely to prescribe antibiotics. Time pressure, limited consultation time results in increased prescription Physicians who only practice outpatient medicine more likely to prescribe than in patients. Health system related factors Availability of evidence based clinical guidelines and protocols within a setting Pharmaceutical industry pressure and availability of over the counter antibiotics. Intra and Inter physician variability in antibiotic prescription The potential adverse effects of antibiotics have a limited influence on physicians' decision- making Physicians-in-training are strongly influenced by the antibiotic prescribing behavior of their supervising staff physicians Although other physicians' prescribing decisions are questioned, there is reluctance to provide critique, feedback, or advice  Testing antimicrobial drugs in the laboratory (Broth Method) Minimal inhibitory concentration (MIC) and Minimum bactericidal concentration (MBC). M.I.C: Quantitative test to determine lowest concentration of antimicrobial drug needed to prevent growth of a specific organism. The MIC is the minimal drug concentration at which no visible growth of bacteria is observed There will be no visual turbidity in a liquid medium. MBC: (Minimum Bactericidal Conc.). Concentration at which bacteria will have lost the ability to reproduce. (16ug/ml below). A microorganism may show no turbidity in culture but yet may not be killed at the M.I.C. MBC is therefore calculated by sub-culturing some liquid from the concentrations showing no turbidity in the M.I.C experiment, onto solid media, incubating to detect growth. The concentration which is considered as MBC is usually higher than the concentration for MIC. (16ug/ml below).  MBC determination Test organism grown with increasing concentrations of an antibiotic. Organism stop growing But not killed. Loopful from broths added to plates with no antibiotic present. Organisms killed at this concentration Microbiological, pharmacological, and clinical situations where an interpretation of the antibiotic concentration in relation to the MIC determination would be useful. TDM: therapeutic drug monitoring; MIC: minimal inhibitory concentration; ESBL: extended spectrum β-lactamase; MDR: multidrug-resistant. Determining Susceptibility of Bacterial to Antimicrobial Drug  Plate method: Disc diffusion method Kirby-Bauer disc diffusion assay routinely is used to qualitatively determine microbial susceptibility to antibiotics. Gives an indication of whether or not the antibiotic will be effective against the organism ‘in vivo’. A Standard concentration (McFarland 0.5 conc) of a bacterial strain is uniformly spread with a swab on standard media (Muller Hinton agar, The test is performed by applying a bacterial inoculum of approximately 1–2×10 8CFU/mL to the surface of a large (150 mm diameter) Mueller-Hinton agar plate.) of standard depth, incubation at a standard temperature/time. Discs impregnated with specific concentration of antibiotic are placed on plate and incubated. Examples of antibiotic discs are the Mastring discs. see photo above of Mastring discs of antibiotics and clear zones of inhibition of microbial growth around the effective discs.  Kirby Bauer Disc Diffusion Method Clear zone of inhibition around disc reflects susceptibility depending on zone diameter. Size of the zone (mm diameter) of clearing around the antibiotic impregnated disk is critical. It must be measured and interpreted using a table that permits interpretation of results as susceptible (S), intermediate resistant (I) or resistant (R). The method and performance of Mastring discs should be routinely checked using quality control strains. Results valid only if all test parameters are standardised and evaluated through QC tests. Using reference strains that always give known diameter size under standardised experimental conditions of agar type, agar depth, incubation temperature etc, for method validation.  Zone diameter above for example >/=18mm. ( S ). What does it mean? ‘S’ means a High probability of clinical success in vivo. 'The "susceptible" category implies isolates are inhibited by the usually achievable concentrations of antimicrobial agent when the recommended dosage is used for the site of infection.’ The intermediate category ‘I’ is defined as uncertain therapeutic success for the individual species/drug combination tested by EUCAST and is intended for compounds for which dosing can be increased. CLSI defines the intermediate category as a lower response rate than for susceptible isolates, but clinical efficacy if the drug accumulates at the site of infection. The intermediate category represents the ‘grey zone’ regarding therapeutic success. https://www.youtube.com/watch?v=ATBoj5jWJhg&t=46s&ab_channel=RejoJacobJoseph Etest Etest® strips – are considered a gold standard for determining the M.I.C of antibiotics, antifungal agents and anti-mycobacterial agents. Etest® is a predefined, stable gradient of 15 antimicrobial concentrations on a plastic strip. Simple, cost-effective tool that offers results when you need more precision than what automated or Kirby-Bauer antimicrobial susceptibility tests (AST) tests provide. Etest® has an extensive range of over 100 antimicrobial references available in the following categories: Antibiotics, Antifungals, Antimycobacterials, and Antimicrobial Resistance Detection (ARD). The MIC is determined by the intersection of the lower part of the ellipse shaped growth inhibition area with the test strip. Etest results have correlated well with MICs generated by broth or agar dilution methods  Five Basic Mechanisms of Antibiotic Action against Bacterial Cells  A. Inhibition of cell wall synthesis  B. Inhibition of functions of cellular membrane  C. Inhibition of protein synthesis  D. Inhibition of nucleic acid synthesis  E. Inhibition of folic acid synthesis.  95% of drug resistant strains are gram negative bacteria Inhibition of bacteria by antibiotics Summary. Figure 20.2  Bacterial cell wall Bacterial cells are surrounded by a cell wall made of peptidoglycan, which consists of long sugar polymers. NAM (N- acetylglucosamine)and NAG (N-acetylglucosamine) molecules The NAM molecule consists of a pentopeptide with a D- Alanine as terminal amino acid Penicillan binding proteins  Penicillin binding proteins Group of transmembrane proteins which bind NAG and NAM molecules. 2 enzymatic functions – Glycoslytransferase (GT) activity Linking of NAG and NAM dissaccarhides to form long polysaccarhides – Transpeptidase (TP), crosslinking of the long polysacchardies to strengthen cell wall PBPs, bind to the pentopeptide of the NAM Remove D-Alanine from pentopeptide Link two peptides  Beta lactams and Glycopeptides Both classes of antibiotics prevent the joining of the polysaccharide strands. Different targets. Beta Lactams: bind to PBPs, this – prevents PBPs from removing NAMS terminal D-Alanine – prevents cross linking of polypeptide strand – weakens cell wall Glycopeptida ses Bind to PBPs transpeptidase active site. Alanine cannot bind to PBP No cross linking of peptidoglycan strands Weakens cell wall  Antibacterial Antibiotics A. Inhibitors of Cell Wall Synthesis: Inhibition of cell wall synthesis (by Beta Lactam antibiotics) –Penicillins demonstrate excellent selective toxicity –no peptidoglycan in eukaryotes. Microbial resistance to penicillins: Resistance Mechanism 1. Some bacteria produce enzymes called Penicillinases (β-lactamases enzymes). These bacterial enzymes can destroy or disable antibiotics. Example, b -lactamase hydrolyses b -lactam ring of penicillins. Without a b -lactam ring, penicillins are ineffective. https://www.youtube.com/watch?v=qBdYnRhdWcQ&t=5s&ab_channel=MechanismsinMedicin e On Canvas Resistance mechanism 2. Some bacteria have acquired altered transpeptidases (enzymes involved in wall synthesis), the altered/mutated transpeptidases will not bind to beta-lactam antibiotics and therefore can cross-link the peptidoglycan layers of the cell wall rendering the bacterium resistant to penicillins. Enzymes in wall synthesis and repair ( Penicillin Binding Protein) 1. Transpeptidase. 2. Glycoslytransferase  Inhibition of functions of Cellular membrane (poor B. selective toxicity) Polymyxins (antibiotics) combine with phospholipid in cell membranes and cause an increase in membrane permeability. They have a positive charge that interfere with the negatively charged of the outer membrane, of Gram negative cells destabilising the membrane and causing leakage. EFFECT Bind to lipopolysaccharide (LPS) in the outer membrane of Gram-negative bacteria, polymyxins disrupt both the outer and inner membranes Because cell membranes are found in both eukaryotic and prokaryotic cells, the action of this class of antibiotic are often poorly selective and can often be toxic for systemic use in the mammalian host. ( can cause kidney damage in host) Most clinical usage is therefore limited to topical applications. Examples: Polymyxin B and Polymyxin E (colistin). (PEK Ps, E. coli, K. pneumonia) They have become the last line of treatment for infections that are resistant to other antibiotics. They are useful in treating infections of the urinary tract, meninges, and bloodstream by susceptible strains of Pseudomonas aeruginosa, Enterobacteriaceae, and Acinetobacter baumannii. Polymyxins are used off label in adults via an inhaled form for patients with cystic fibrosis who have chronic pulmonary infections with Pseudomonas aeruginosa. Also, aerosolized polymyxins have been used as adjunctive antibiotic therapy for the treatment of hospital-acquired and ventilator-associated pneumonia caused by multidrug resistance organisms such as Pseudomonas aeruginosa  C. Inhibition of protein synthesis 3 methods – Selective toxicity due to differences in ribosome structure. Eukaryote 80S v 70S bacteria. – Eucaryotic cells have 80S (60S + 40S subunits) ribosomes. – Procaryotic cells have 70S (50S + 30S subunits) ribosomes. Examples below of the different mechanisms by which different antibiotics inhibit protein synthesis in bacteria Treat conjunctivitis, meningitis, plague, cholera, and typhoid fever G. negative. T.B. brucellosis, UTI, RTI, treatment of chlamydia  Figure 20.4 Most antibiotics that bind to the ribosome have been shown to interact directly with ribosomal  One of the side-effects of tetracyclines is incorporation into tissues that are calcifying at the time of their administration. They have the ability to chelate calcium ions and to be incorporated into teeth, cartilage and bone, resulting in discoloration of both the primary and permanent dentitions. D. Inhibition of nucleic acid synthesis (Quinolones & Fluoroquinolones –active primarily against Gram negative bacteria) This mechanism interfere WITH proteins/enzymes in bacterial cells that are involved in DNA synthesis (DNA gyrase (topoisomerase 11), DNA topoisomerase, DNA polymerase). Example: 1. Interference with bacterial DNA replication. Interferes with DNA gyrase. An enzyme in bacteria that relieves strain while double- stranded bacterial DNA is being unwound by helicase during DNA replication. Basis for selectivity is -DNA GYRASE ABSENT in higher Eukaryotes Examples: Nalidixic acid 1. inhibit DNA gyrase and topoisomerases). 2. Interference with DNA polymerase. 3. Interfere with rRNA synthesis. Anti-tuberculosis antibiotic, Rifamycin: High affinity for the prokaryotic RNA polymerase. Binds to the beta subunit of RNA polymerase, prevents initiation complex form forming (changes its shape) , RNA polymerase cannot be initiated. Poor affinity for the analogous mammalian enzyme.  Rifampin - blocks transcription – can cause red man syndrome - a result of accumulation of metabolic products of the antimicrobic in secretions  E. Sulfonamides: Inhibition of folic acid synthesis. Bacteria require and must synthesise folic acid. Animals get folic acid from their diet. Folic acid cannot cross through bacterial cell wall. Bacteria therefore must synthesise it. Sulfonilamide ( a sulphonamide) is competitive inhibitor, it blocks folic acid synthesis by interfering with the action of a bacterial enzyme involved in the folic acid synthesis process. If A sulfa drug is present, the first bacterial enzyme is not too specific and uses the sulfonamide in the first step instead of PABA the normal substrate. STEP 1 leads to  Pteridine-Sulfa drug. The 2nd enzyme in the pathway is VERY SPECIFIC, will NOT use Pteridine-sulfa drug as a substrate to make folic acid, so the pathway is inhibited and no folic acid is produced. Targets Gram positive aerobes Gram negative aerobes  Folic acid synthesis Folic acid pathway PABA(para amino benzoic acid) + Pteridine + L-Glutamate Dihydrofolate synthase Dihydro folic acid +NADPH Dihydrofolate reductase Tetrahydrofolic acid Purine (A) Pyrimidine (T) Methionine Folic acid synthesis  Folic acid inhibition Sulfonamides (sulfamethoxazole) Very similar structure to PABA, competes to bind with Pteridine. Inhibits enzyme dihydrofolate synthase, inhibits folic acid pathway. Trimethoprim, similar to hydrofolic acid, competes with binding to NADPH. Tetrahydrofolic acid not produced. When used separately, bacteriostatic When used in combination, bactericidal  EMERGENCE OF ANTIBIOTIC RESISTANCE. 1. Natural selection of antibiotic resistant bacteria The starting point in this example is a large mixed bacterial population mainly consisting of bacteria that are susceptible to antibiotics and a few bacteria that are antibiotic-resistant by chance. (SPONTANEOUS MUTATION: Bacteria can mutate at a rate of 1 in 10 10 bases per generation 6 to 9 other bacteria higher mutation rates) (Might result in change od affinity of target for antibiotic or antibiotic accessibility ) A subset of bacterial cells derived from a susceptible population develop mutations in genes that affect the activity of the drug, resulting in preserved cell survival in the presence of the antimicrobial molecule. If a bactericidal antibiotic is added which kills most of the susceptible bacteria in the population the resistant mutant bacteria survives and proliferate. The end result is a population of mainly resistant bacteria that can transmit this new characteristic to the vext generation VERTICAL TRANSFER Using narrow-spectrum antibiotics (when possible) or combinations of antibiotics reduces the risk of selecting for antibiotic resistance in the commensal flora of the body.  This just resistance described-- can be Spontaneo passed on us by vertical mutation transmissi and on antibiotic resistance. These mutations can When bacterial cells make it difficult for the replicate, there is a antibiotics to enter the small chance the bacteria or stick to it, new bacterial cell will making the antibiotic less not be exactly the effective at killing the same as the original bacteria. bacterial cell. We call Bacteria can multiply these within hours, allowing errors in for more mutations to the copied occur over a shorter Mutations are key to cell a period of time. the idea of evolution, mutation. and all of the diversity you can see in nature came from a series of many mutations over hundreds of thousands of years.  Acquisition of antimicrobial resistance 1. Spontaneous Mutation Mutation is a change in the organisms DNA that can cause a change in the gene product (protein), which was the target of the antimicrobial. The target protein may no longer bind to the antibiotic and is therefore no longer susceptible to the antibiotic. Example: Spontaneous mutation in the Bacterial DNA gyrase protein DNA gyrase is an essential bacterial enzyme required for DNA replication, it can be target by fluroquinolones. When a susceptible bacterium comes into contact with a therapeutic concentration of an antibiotic like fluroquinolones it binds to the DNA gyrase blocking it’s activity leading to cell death. If a spontaneous mutations occur in the gene for DNA gyrase in a cell during replication , altering it slightly so that the antibiotic cannot bind to it, DNA gyrase may still function AND this organism is now resistant to fluroquinolone. This allows the bacterium to continue DNA replication. 2. Intrinsic resistance Certain bacteria may have natural Inherent/Intrinsic resistance due to Darwinian evolution. Intrinsic resistance is the natural ability of a bacterial species to resist an antimicrobial agent through its inherent structural or functional characteristics, which allow tolerance of a particular drug or antimicrobial class. EXAMPLES Natural lack of affinity of the drug for it’s target site in a particular organism. 2. Inaccessibility -inability of the drug to enter the bacterial cell. 3. Drug may enter the cell but the bacterium has an extrusion mechanism (maybe a protein pump encoded for by a gene on the chromosome) that pumps the drug out thus it cannot reach toxic concentrations in the bacterial cytoplasm. 4. Innate production of enzymes that inactivate the drug. 3. HORIZONTL GENE TRANSFER Acquisition of antimicrobial resistance https://www.youtube.com/watch?v=08Q-MVeNeTU&ab_channel=NeuralAcademy 2023 v.good BY a. Conjugation. (Occurs within species or across Gram positive and Gram negative species) b. Transformation. (Only occurs within closely related strains) c. Transfection. (Only occurs within closely related species) https://www.youtube.com/watch?v=7tLV20dk-FM&ab_channel=Henrik%27sLab Transformation, transduction and conjugation  Acquisition of antimicrobial resistance Conjugation Conjugation is mediated by a particular kind of circular DNA called a plasmid, which replicates independently of the chromosome. Many plasmids carry genes that confer resistance to antimicrobials. When two cells are in close proximity to each other, a hollow bridge-like structure, known as a pilus, forms between two cells. This allows a copy of the plasmid, as it is duplicated, to be transferred from one bacterium to another. This enables a susceptible bacteria to acquire resistance (via the genes on a plasmid) to a particular antimicrobial agent. (HFr strains –when the F plasmid integrates into the host genome. Host becomes HFr. The plasmid can mobilise the chromosome –the chromosome is mobilised at the origin of transfer of the F plasmid. Some of the F plasmid is at the beginning of transfer, some at the end, that which transfers becomes double stranded and may integrate. The recipient remains F-)..  Acquisition of antimicrobial resistance Transformation During this process, genes are transferred from one bacterium to another as “naked” DNA. When cells die and break apart, DNA can be released into the surrounding environment. Other bacteria in close proximity can scavenge this free-floating DNA, and incorporate it into their own DNA. This DNA may contain advantageous genes, such as antimicrobial resistant genes and benefit the recipient cell. Double stranded Naked DNA binds to receptor on recipient cells. 1 strand enters competent cells. The other strand is degraded. The single strand DNA base pairs with a homologous region on the recipients chromosome by a breakage and reunion mechanism called homologous recombination. Differences in nucleotide sequences are repaired by the host mismatch repair mechanisms. This system can either remove the donor DNA or the recipient DNA. We select for the Donor DNA in the laboratory by plating transformants on selective agars.  Transduction In this process, bacterial DNA is transferred from one bacterium to another inside a virus that infects bacteria. These viruses are called bacteriophages or phage. When a phage infects a bacterium, it essentially takes over the bacteria's genetic processes to produce more phage. During this process, bacterial DNA may inadvertently be incorporated into the new phage DNA. Upon bacterial death and lysis (or breaking apart), these new phage go on to infect other bacteria. This brings along genes from the previously infected bacterium.