Pharmaceutical Microbiology Lecture 1-2 PDF

Pharmaceutical Microbiology 2024/2025 Pharmaceutical Microbiology Prepared by: Noha Mahmoud Gamaleldin, PhD. Associate Professor of Microbiology and Biotechnology, Department of Microbio...

Pharmaceutical Microbiology 2024/2025 Pharmaceutical Microbiology Prepared by: Noha Mahmoud Gamaleldin, PhD. Associate Professor of Microbiology and Biotechnology, Department of Microbiology, Faculty of Pharmacy, The British University in Egypt, Cairo, Egypt. Faculty of Pharmacy- The British University in Egypt (FP-BUE). Page 1  Pharmaceutical Microbiology 2024/2025 Lecture (1) What Is Microbial Death? ▪ death in microscopic organisms that are composed of just one or a few cells is hard to detect, because such organisms often reveal no remarkable vital signs to detect. ▪ Lethal agents (such as radiation and chemicals) do not necessarily alter the appearance of microbial cells. Even the loss of movement in a motile microbe cannot be used to indicate death. ▪ The destructive effects of chemical or physical agents occur at the cellular and molecular level. As a cell is exposed to an agent such as intense heat or toxic chemicals, various cell structures break down, and the entire cell can show irreversible damage. ▪ At present, the most practical way to detect this damage is to determine if a microbial cell can still multiply when exposed to a suitable environment. ✔▪ In most situations of testing, the permanent loss of reproductive capacity, even under optimal growth conditions, has become the working definition of microbial death. Factors That Affect Death Rate ▪ Death of the whole population is not sudden but begins when a certain threshold of an antimicrobial agent (some combination of time and concentration) is met. ▪ Death continues in a logarithmic manner as the time of exposure is increased. ▪ When a point is reached at which survival of any cells is highly unlikely; this point is equivalent to sterilization. Faculty of Pharmacy- The British University in Egypt (FP-BUE). Page 2  Pharmaceutical Microbiology 2024/2025 ▪ The effectiveness of a particular agent is governed by several factors besides time. These additional factors influence the action of antimicrobial agents: 1. The number of microorganisms (bioburden). A higher load of contaminants requires more time to destroy. 2. The nature of microorganisms in the population. In most actual conditions of disinfection and sterilization, the target population is not a single species of microbe but a mixture of bacteria, fungi, spores, and viruses, presenting a broad spectrum of microbial resistance. 3. The temperature and pH of the environment. 4. The concentration (dosage, intensity) of the agent. 5. The mode of action of the agent. How does it kill or inhibit microorganisms? 6. The presence of solvents, interfering organic matter, and inhibitors. Saliva, blood, and feces can inhibit the actions of antimicrobial agents and even of heat. Faculty of Pharmacy- The British University in Egypt (FP-BUE). Page 3 Pharmaceutical Microbiology 2024/2025 Part 1. Antimicrobial Therapeutic Agents Important terminology of chemotherapy: Chemotherapeutic Drug Any chemical used in the treatment, relief, or prophylaxis of a disease. Prophylaxis Use of a drug to prevent developing a disease or infection in a person at risk. Antimicrobial Chemotherapy The use of chemotherapeutic drugs to control infections. Antibiotics Substances naturally produced by some microorganisms and can inhibit or destroy other microorganisms. Semi Synthetic Drugs Drugs that are chemically modified in the laboratory after being isolated from natural sources. Synthetic Drugs Drugs that are synthesized entirely in the laboratory by chemical reactions. Note: The use of the term “antibiotic” has changed over the years, to any substance (regardless its origin) which in low concentration can inhibit or kill microorganisms. This is because several antibiotics are now produced wholly or partially by chemical methods. ❖ Historical background The priests used natural antimicrobial compounds of plant origin since ancient Egyptians. In 1619, it was reported that quinine obtained from cinchona bark could be used for treatment of malaria. Paul Ehrlich in the beginnings of the 20th century used the organic arsenical compound arsphenamine for treatment of syphilis. In 1932, a milestone development in antimicrobial chemotherapy started by Domagk who demonstrated that the red azo dye (Sulfonamidochrysoidine: the first commercially available antibiotic marketed under the brand name Prontosil) could cure streptococcal infections in human beings. It was later proved that Prontosil is metabolized in body fluids to give the highly active sulphonamide. Faculty of Pharmacy- The British University in Egypt (FP-BUE). Page 4 Pharmaceutical Microbiology 2024/2025 The second milestone in the antimicrobial therapy began with the observation of Fleming (1928) that a mold contaminant (Penicillium rubens) inhibited the growth of staphylococci in a culture plate. Through the work of Florey and Chain in 1940- 1945 benzyl penicillin was produced on industrial scale. It was later called the magic bullet that could be used for the treatment of infections. Later, several other antibiotics were discovered through screening programs, such as streptomycin (1944), chloramphenicol (1947), tetracyclines (1948), and many other antibiotics. ❖ The goal of antimicrobial drugs: The goal of antimicrobial drugs is either to disrupt the cell processes or structures of bacteria, fungi, and protozoa or to inhibit the virus multiplication cycle. Simply, administer a drug to an infected person that destroys the infective agent without harming the host’s cells. This goal is rather difficult to achieve. ❖ The characteristics of the ideal antimicrobial drug: 1. Selectively toxic to the microbe but nontoxic to the host cells: Selective toxicity of antimicrobial agents means that, they have the ability to kill or inhibit an invading microorganism without harming the cells of the host. This can be achieved by acting specifically on microbial structures or functions not found in host cells. In most instances, the selective toxicity is relative rather than absolute, requiring that the concentration of the drug be carefully controlled to attack the microorganism, while still being tolerated by the host. 2. Microbicidal rather than microbiostatic. Microbicidal: Any substance that kill the microbes. Microbiostatic: Any substance that inhibits the growth of microbes. Bacteriostatic drugs stop the growth and replication of bacteria, thus limiting the spread of infection until the immune system attacks and eliminates the pathogen. If the drug is removed before the immune system has cleaned the organisms, enough viable organisms may remain to begin a second cycle of infection. Bactericidal drugs kill bacteria at drug serum levels achievable in the patient. Bactericidal agents are often the Faculty of Pharmacy- The British University in Egypt (FP-BUE). Page 5 Pharmaceutical Microbiology 2024/2025 drugs of choice in seriously ill (endocarditis, meningitis…..) and immunocompromised patients. Effects of bactericidal and bacteriostatic drugs on the growth of bacteria in vitro 3. Remains potent long enough to act and is not broken down or excreted rapidly. 4. Doesn’t lead to the development of antimicrobial resistance. Antimicrobial resistance: an adaptive response in which microorganisms begin to tolerate an amount of drug that would usually be inhibitory. 5. Remains active in tissues and body fluids. 6. Easily delivered to the site of infection. 7. Reasonably priced. 8. Does not disrupt the host’s health by causing allergies or predisposing the host to other infections. In short, the perfect drug does not exist, but by balancing drug characteristics against one another, a satisfactory compromise can be achieved. ❖ Selection of antimicrobial agents Selection of the most appropriate antimicrobial agent requires knowing the following: 1) the organism’s identity. 2) the organism’s susceptibility to a particular agent. 3) the site of the infection. 4) patient factors. Faculty of Pharmacy- The British University in Egypt (FP-BUE). Page 6 Pharmaceutical Microbiology 2024/2025 5) the safety of the agent. 6) the cost of therapy. However, some patients require empiric therapy (immediate administration of drug (s) prior to bacterial identification and susceptibility testing). A clinical specimen for microbial identification must be taken before the beginning of any empiric therapy. Note: Antibiotics can be used as: ▪ Prophylaxis: to prevent infection ▪ Empiric: Antibiotics intended to treat a suspected bacterial infection; causative microorganism is not identified yet. ▪ Definitive: causative microorganism is identified, culture based antibiotic selection. Antimicrobial therapeutic agents can be grouped according to the microorganisms they act primarily against into: ▪ Antibacterial agents. ▪ Antifungal agents. ▪ Antiviral agents. ▪ Antiparasitic agents. 1- Antibacterial Therapeutic Agents [I]Classification of antibacterial agents Antibacterial agents can be classified depending on various characteristics among which: 1. The function: being either bactericidal or bacteriostatic. 2. The chemical structure: as β lactams, quinolones, aminoglycosides, tetracyclines, …etc. 3. The spectrum of action: Spectrum: Range of bacteria against which the agent is typically active. The clinically important bacteria can be organized into groups based on Gram stain, morphology, and biochemical or other characteristic such as (Gram-positive, Gram-negative, Anaerobes, Spirochetes, Chlamydia, Mycoplasma, Mycobacteria and others. i. Narrow spectrum (limited spectrum) agents. Agents acting only on a single or a limited group of microorganisms. For example, isoniazid is active only Faculty of Pharmacy- The British University in Egypt (FP-BUE). Page 7 Pharmaceutical Microbiology 2024/2025 against Mycobacterium tuberculosis (limited), natural penicillins act mainly against gram-positive bacteria (narrow). ii. Extended (intermediate) spectrum agents. The term applied to antibiotics that are modified to be effective against wider range of bacteria. For example, ampicillin is considered to have an extended spectrum because it acts against gram-positive and some gram-negative bacteria. iii. Broad spectrum agents. Drugs that affect wide variety of microbial species (gram-positive, gram-negative, etc.) such as tetracycline, fluoroquinolones, carbapenems…... 4. The mechanism of drug action i. Inhibition of cell wall synthesis. ii. Breakdown of the cell membrane structure or function. iii. Interference with functions of DNA and RNA. iv. Inhibition of protein synthesis. v. Blocks on key metabolic pathways (antimetabolites). In this module, a survey on the most important types of antibacterial agents has been conducted depending on their mechanisms of action. The following figure outlines the mechanisms of action of antibacterial drugs. Faculty of Pharmacy- The British University in Egypt (FP-BUE). Page 8 Pharmaceutical Microbiology 2024/2025 Major targets of drugs acting on bacterial cells 1. Antibacterial drugs that affect the bacterial cell wall The cell wall of most bacteria contains a rigid layer of peptidoglycan, which protects the bacterial cell against rupture from hypotonic environment. Faculty of Pharmacy- The British University in Egypt (FP-BUE). Page 9  Pharmaceutical Microbiology 2024/2025 ▪ There are three stages of peptidoglycan biosynthesis. The first occurs in the cytoplasm where cell wall precursors (disaccharide pentapeptide) are synthesized and is then transported across the membrane on a lipid carrier. The second stage is the insertion of the disaccharide pentapeptide into the cell wall by a transglycosylase (transglycosylation). The third step is the fusion of these precursors through their peptides by a transpeptidase (transpeptidation). Basic structure of peptidoglycan layer ▪ Mammalian cells do not possess a cell wall. Consequently, antibiotics that interfere with peptidoglycan synthesis generally have excellent selective toxicity since the target is vital to the bacteria but absent from mammalian cells. ▪ To be maximally effective, inhibitors of cell wall synthesis require actively growing microorganisms. Antibacterial agents interfere with cell wall synthesis will cause the cell to develop weak points at growth sites and become osmotically fragile. Antibiotics that produce this effect are considered bactericidal, because the weakened cell is subject to cell lysis. Faculty of Pharmacy- The British University in Egypt (FP-BUE). Page 10 Pharmaceutical Microbiology 2024/2025 Effect of antibiotics on bacterial cell wall The most important members of this group of drugs are: β-Lactam antibacterials, Glycopeptides and miscellaneous cell wall inhibitors. 1.1. β-Lactam Antibacterials: The β-Lactam antibacterials comprise the penicillins, cephalosporins, carbapenems, and monobactams. Their name derived from the presence of a β-lactam ring in their structure; this ring is essential for antibacterial activity. Faculty of Pharmacy- The British University in Egypt (FP-BUE). Page 11 Pharmaceutical Microbiology 2024/2025 Differences in the side chain (s) attached to the single or double ring can have a significant effect on the pharmacologic properties and spectrum of any β-lactam. β-lactams interfere with the transpeptidation reactions that seal the peptide crosslinks between glycan chains. They do so by interference (through irreversible binding) with the action of the transpeptidase enzymes, which carry out this cross- linking. These targets of all the β-lactams are commonly called penicillin-binding proteins (PBPs). β-lactam antibiotics act by inhibiting penicillin-binding proteins, which normally catalyze cross-linking of bacterial cell walls 1.1.1. Penicillins: The penicillin antibiotics is a large group of compounds most of which end in the suffix- cillin. Their name is for the parent compound benzyl penicillin (Penicillin G), whose action is restricted to gram-positive bacteria. Natural penicillins (penicillin G and penicillin V) are obtained from fermentations of the fungus Penicillium rubens. Semisynthetic penicillins, such as amoxicillin and ampicillin are created by chemically attaching different R groups to the main chemical nucleus. The following table summarizes the characteristics of selected penicillin drugs: Faculty of Pharmacy- The British University in Egypt (FP-BUE). Page 12 Pharmaceutical Microbiology 2024/2025 Agent Spectrum Uses, Advantages Disadvantages Penicillin G Narrow Best drug of choice when Can be hydrolyzed by bacteria are sensitive; low β-lactamases and acids; toxicity; low cost rapidly excreted in the urine; causing allergy. Penicillin V Narrow Good absorption from Hydrolysis by β- intestine; otherwise, as lactamases; allergies penicillin G Oxacillin, Narrow Not susceptible to β- Allergies; expensive dicloxacillin lactamases; good (antistaphylococcal) absorption Methicillin, Narrow Not usually susceptible to Poor absorption; nafcillin β-lactamases allergies; growing (antistaphylococcal) resistance Ampicillin Extended Works on G-negative Can be hydrolyzed by bacilli but not β-lactamases; allergies; Pseudomonas species only fair absorption Amoxicillin Extended As ampicillin; good Hydrolysis by β- absorption (taken with lactamases; allergies food) Carbenicillin Very Same as ampicillin but Poor absorption; used (antipseudomonal) extended active against only parenterally Pseudomonas species Piperacillin, Very Effective against P. Allergies; susceptible ticarcillin extended aeruginosa and other to β-lactamases (antipseudomonal) enterococci. Notes: 1- β-lactamases: Enzymes produced by bacteria that capable of destroying the β-lactam ring also named penicillinases. 2- Staphylococcus aureus: it is a gram-positive bacteria, it is a common cause of skin infections such as abscesses, respiratory infections such as sinusitis, and food poisoning. The emergence of antibiotic-resistant forms of S. aureus such as MRSA (Methicillin Resistant S. aureus) is a worldwide problem in clinical medicine. Faculty of Pharmacy- The British University in Egypt (FP-BUE). Page 13 Pharmaceutical Microbiology 2024/2025 3- Pseudomonas aeruginosa: Medically important gram-negative bacteria highly resistant to antibiotics, cause skin, respiratory, and urinary infections. Important considerations: ❑ Variable semisynthetic penicillins are made to overcome problems associated with the naturally occurring penicillins such as narrow spectrum of activity, hydrolysis by gastric acids (i.e: cannot be taken orally) and inactivation by β- lactamases. ❑ Generally, as the activity against gram-negative bacteria increases, the activity against gram-positive bacteria decreases. ❑ The major disadvantage of penicillins is hypersensitivity (allergy), which is estimated to occur in 1-10% of patients. The hypersensitivity reactions include anaphylaxis, skin rashes, hemolytic anemia, nephritis, and drug fever. Anaphylaxis, the most serious complications, occurs in 0.5% of patients. Anaphylaxis is a serious allergic reaction that is rapid in onset and may cause death. It typically causes a number of symptoms including an itchy rash, throat swelling, and low blood pressure. 1.1.2. Cephalosporins: The cephalosporines antibiotics represent about one-third of all antibiotics administered. They are β-lactam drugs that act in the same manner as pencillins; i.e., they are bactericidal agents that inhibit the cross-linking of peptidoglycan. The generic names of these compounds are often recognized by the presence of the root cef, ceph, or kef in their names. Cephalosporins were first isolated in the late 1940s and came into general use in the 1960s. The cephalosporins are classified by generation to first, second, third or fourth. The “generation” term related to causing breakthroughs in expanding the spectrum through modification of the side chains. Faculty of Pharmacy- The British University in Egypt (FP-BUE). Page 14 Pharmaceutical Microbiology 2024/2025 Agents Characteristics Generation Cephalexin High activity against gram-positive cocci, Streptococci as Cefazolin (Streptococcus pneumonia) First Cephradine Moderate activity against gram-negative bacteria as E.coli, Klebsiella, and Proteus. Not active against Haemophilus and Pseudomonas. Cefuroxime As for 1st generation against gram-positive bacteria. Generation Cefaclor Expanded activity against gram-negative bacilli and Second Cefoxitin Neisseria meningitidis, Haemophilus influenzae. Cefamandol Active against anaerobes as Bacteroides fragilis. Not active against Pseudomonas. Cefixime Less active against gram-positive bacteria, specially S. Ceftriaxone aureus. Cefotaxime have wider and very higher activity against most gram- Generation Third Ceftazidime negative organisms compared with the 1st and 2nd generations. Only ceftazidime is constantly active against Pseudomonas aeruginosa. Active against most anaerobes. Cefepime Similar activity against gram-positive bacteria as the 1st Generation Generation Fourth generation. Retain the high activity of the 3rd generation including activity against Pseudomonas, Neisseria, H. influenzae. Active against (MRSA) and other gram-positive bacteria. Ceftobiprole Retain the activity of later-generation cephalosporins Ceftaroline Fifth Ceftolozane having broad-spectrum activity against gram negative bacteria. They are used for Community Acquired Pneumonia (CAP) and complicated skin infections. Important considerations: ❑ Cephalosporines generally have superiority over penicillins in having broader spectrum of activity, being more resistant to the action of β- lactamases and producing fewer hypersensitivity reactions than do penicillins. ❑ Both penicillins and cephalosporines are used as prophylactic agents, in the prevention of infections, as prior to surgery in particularly susceptible patients. Faculty of Pharmacy- The British University in Egypt (FP-BUE). Page 15 Pharmaceutical Microbiology 2024/2025 1.1.3. Carbapenems The carbapenems are β-lactam drugs, they have the broadest spectrum of all β-lactam antibiotics. This fact appears to be due to having the ability of easy penetration of gram-negative and gram-positive bacterial cells and high level of resistance to β-lactamases. ▪ This class include: imipenem, meropenem, and ertapenem. They are active against aerobic and anaerobic, gram-positive and gram-negative bacteria. ▪ Imipenem is particularly important for its activity against P. aeruginosa and the Enterococcus species. ▪ Ertapenem differs from other carbapenems in having a somewhat less broad spectrum of activity (not against Pseudomonas aeruginosa and Acinetobacter), and in that its extended serum half-life allows it to be administered once every 24 hours. 1.1.4. Monobactams: Monobactams are β-lactam compounds where the β-lactam ring is alone and not fused to another ring. The only commercially available monobactam antibiotic is aztreonam. It works only against aerobic gram-negative bacteria including Neisseria and Pseudomonas but is inactive against gram-positive and anaerobic bacteria. It has minimal side effects, and it is very useful in patients who are hypersensitive to penicillin, because there is no cross-reactivity. Faculty of Pharmacy- The British University in Egypt (FP-BUE). Page 16