Introduction to Pharmaceutical Microbiology

1. بسم الله الرحمن الرحيم

2. Pharmaceutical Microbiology I Dr. Magdy Muharram Associate professor of Microbiology

3. Course Description:This course is an introductory study of microbiology. This course emphasizes on the structure of bacteria, viruses and fungi. Also it describes the mechanism of infection and pathogenicity • Teaching Objectives • To introduce and cover the following topics: • Classification of microorganisms • Structure of, bacteria, fungi and viruses. • Sterilization, physical methods, chemical methods and antiseptics and preservatives. • Bacterial growth and physiology. • Bacterial genetics. • Mechanism of infection and pathogenicity of microorganisms • Acquired resistances. • Introduction of virology and classification of viruses. • Basic structure and ultra structure of the viruses, viral growth and physiology and viral genetics • Viral pathogenesis.

5. Tests and Exams: Midterm 1 15% Midterm 2 15 % Practical 25% Exam 40 % • Practical will be introduced by Mr. Moatasem • Safety FIRST • Lab coat – only in the laboratory • No open shoes • Permanent Marker • No eating or drinking • Hand washing • Request • Work hard and Please BE ON TIME !!!!!!!!

6. Microbiology And microorganisms Microbiology is the study of all organisms that are invisible to the naked eye - Its subjects are bacteria, viruses, fungi, many algae and protozoa. - The importance of microbiology and microorganisms can not be overemphasized. - Microorganisms are necessary for the production of bread, cheese, antibiotics, vaccines, vitamins, enzymes, etc. - Modern biotechnology rests upon a microbiological foundation. • Microorganisms are everywhere; almost every natural surface is colonized by microbes, from body to ocean. Some microorganisms can live hot springs, and others in frozen sea ice. - Most microorganisms are harmless to humans; You swallow millions of microbes every day with no ill effects. In fact, we are dependent on microbes to help us digest our food. - Microbes also keep the biosphere running by carrying out essential functions such as decomposition of dead animals and plants. They make possible the cycles of carbon, oxygen, nitrogen and sulfur that take place in terrestrial and aquatic systems. - They sometimes cause diseases in man, animals and plants. They are involved in food spoilage.

7. - Antony van Leewenhoek (1632 – 1723) who invented the first microscope (50 – 300x), was the first to accurately observe and describe microorganisms.

8. Microbial World • Viruses • Bacteria • Fungi (Yeasts and Molds) • Protozoa • Microscopic Algae

9. Benefits • Basis of food chain • Nitrogen fixation and other cycles • Photosynthesis • Digestion, synthesis of vitamins • Manufacture of food and drinks • Genetic engineering and Biotechnology • Synthesis of chemical products • Recycling sewages • Bioremediation: use microbes to remove toxins (oil spills) • Use of microbes to control crop pests Harmful Effects • Cause disease (basis for bioterrorism) • Food spoilage

10. Discovery of Microorganism and the Pioneers of Microbiology • Antony van Leewenhoek (1632 – 1723) who invented the first microscope (50 – 300x), was the first to accurately observe and describe microorganisms. • Edward Jenner 1796 – First vaccine (smallpox)

11. Louis Pasteur (1822-1895) • Developed vaccines for Chickenpox, anthrax, rabies • Demonstrated that all fermentations were due to the activities of specific yeasts and bacteria. • - Developed Pasteurization to preserve wine during storage. Important: Foods • - Discovered that fermentative microorganisms were anaerobic and could live only in absence of oxygen. • Proposed aseptic techniques (prevent contamination by unwanted microbes) - Bassi – showed that silkworm disease was caused by a fungus. - Berkeley and Pasteur showed that Microorganisms caused disease. - Charles Chamberland (1851-1908) discovered viruses and their role in disease.

12. Branches of Microbiology • Bacteriology: study of bacteria • Mycology: study of fungi • Immunology: study of immunity • Edward Jenner, UK: developed vaccination (1798) • Metchnikoff, RU: discovered phagocytes (1884) • Paul Ehrlich, DE: theory of immunity (1890) • Virology: study of viruses • Beijerinck, NE: discovered intracellular reproduction of TMV; coined the term “virus” (1899) • Parasitology: study of protozoa and parasitic worms • Chemotherapy • Treatment of disease by using chemical means • Antibiotics produced naturally • Synthetic drugs Chemotherapy • Alexander Fleming, Scotland (1928) discovered penicillin • Selman Waksman, Ukraine (1944) discovered streptomycin

13. Branches of Microbiology • Problems • Toxicity of drugs => Selective toxicity • Resistance of bacteria to drugs • Recombinant DNA Technology • Recombinant DNA • Genetic engineering/biotechnology • Microbial genetics – mechanism by which microbes inherit genes • Molecular biology – structure and function (expression) of genes • Molecular epidemiology/diagnostics • Biotechnology • GMOs for industrial, pharmaceutical and agricultural applications • Improvements of agriculture (plants and animals) • Gene therapy: inserting a missing gene or replacing a defective one in human cells

14. Classification of Microorganisms In 1930s electron microscopy made it clear that bacterial cells lacked a nucleus. The term procaryote was introduced in 1937. • In 1959 Kingdom Fungi was established. • In 1961 the current definition of the term procaryote was established. • In 1968 the KingdomProcaryotae was accepted by biologists. • In 1969 Robert Whitaker proposed a five-kingdom system of biological classification for all living organisms. .Kingdom Procaryotae (Monera): Lack a nucleus and membrane bound organelles. The other four kingdoms are eucaryotes. Have a true nucleus and membrane bound organelles. 2. Kingdom Protista: Mostly unicellular, lack tissue organization. Most have flagella during life. 3. Kingdom Fungi: May be unicellular (yeasts) or multicellular (molds). Many are saprotrophs. 4. Kingdom Plantae:Multicellular, photosynthetic • 5. Kingdom Animalia:Multicellular, heterotrophs that ingest food through a mouth or oral cavity.

15. Differences Between Eucaryotic and Procaryotic Cells Procaryotes Eucaryotes Cell size 0.2-2 um in diameter 10-100 um in diameter Nucleus Absent Present Membranous Organelles Absent Present Cell Wall Chemically complex When present, simple Ribosomes Smaller (70S) Larger (80S) in cell 70S in organelles DNA Single circular Multiple linear chromosome chromosomes (histones) Cell Division Binary fission Mitosis

16. Procaryotes: Lack Nucleus and Membrane-Bound Organelles

19. Scientific Nomenclature • Scientific nomenclature: Universal system for naming and classifying living organisms. Initially developed in the 18th century by Carl Linnaeus. • Binomial nomenclature: Each organism (species) has a two part name. Names are either italicized or underlined. • Genus name: Always capitalized, always a noun. May use initial. • species name: Always lower case, usually an adjective. • Names are usually derived from Latin (or Greek) or may have latinized endings. Examples: • Penicillium notatum (P. notatum): Mold that produces penicillin • Staphylococcus aureus and Escherichiacoli

20. Classification of Organisms Hierarchy of Taxonomic Categories DOMAIN Kingdom • Phylum or Division (Bacteria) • Class • Order • Family • Genus species

21. Shapes and arrangements: • A- COCCI—arranges as • 1-bunches 2-diplococci 3- chains, 4 tetrads • B- BACILLI arranged as • 1- single, diplobacilli, 3- chains • C- vibrios • D- spirals or coils ( spirochetes)

24. Gram negative bacteria

26. Simple stain by Methylene Blue

27. Mixed cultures of Gram negative and Gram positive bacteria

35. Structure of Bacteria • All cells have 3 main components: • DNA (‘nucleoid”) • genetic instructions • cytoplasmic membrane • limits access to the cell’s interior • cytoplasm, between the DNA and the membrane • where all metabolic reactions occur • especially protein synthesis, which occurs on the ribosomes • Bacteria also often have these features: • cell wall • resists osmotic pressure • flagella • movement • pili • attachment • capsule • protection and biofilms

36. Cell Envelope • The cell envelope is all the layers from the cell membrane outward, including the cell wall, the periplasmic space, the outer membrane, and the capsule. • All free-living bacteria have a cell wall • periplasmic space and outer membrane are found in Gram-negatives • the capsule is only found in some strains

41. Cell Membrane • The cell membrane (often called the plasma membrane) is composed of 2 layers of phospholipids. • Phospholipids have polar heads and non-polar tails. • “Polar” implies that the heads are hydrophilic: they like to stay in an aqueous environment: facing the outside world and the inside of the cell. • “non-polar” means that the tails are hydrophobic: they want to be away from water, in an oily environment. The tails are in the center of the membrane • A pure phospholipid membrane only allows water, gasses, and a few small molecules to move freely through it.

42. Membrane Proteins • Proteins float in the membrane like ships on the surface of the sea: the fluid-mosaic model. • Peripheral membrane proteins are bound to one surface of the membrane. • Some attached to the cell membrane by a fatty acid covalently attached to one of the protein’s amino acids • Others are attached by stretches of hydrophobic amino acids of the protein’s surface • Integral membrane proteins are embedded in the membrane by one or more stretches of hydrophobic amino acids. Many of these proteins transport molecules in and out of the cell. The transport proteins are very selective: each type of molecule needs its own transporter.

43. Functions of the cell membrane • Selective permeability to different molecules. • Active transport of nutrient substances through special enzymes • It provides enzymes needed for the cell wall synthesis • It supplies cell with energy because it is the site of respiration • Mesosomes: • Are invaginations of the cytoplasmic membrane that play a role in cell division and respiration as they are the site of electron transport.

44. Cell Wall • Gram-positive vs Gram-negative are defined by the structure of the cell wall • the Gram stain binds to peptidoglycan • Gram-positive: many layers of peptidoglycan, which is anchored to the cell membrane by teichoic acid. • Gram-negative: 1-2 layers of peptidoglycan = thin • The periplasmic space is between the cell membrane and the cell wall. It contains enzymes and other proteins, such as chemoreceptors for sensing the environment. • Outside the peptidglycan layer is the “outer membrane”. It is pierced by porins: protein channels, and its out surface is covered with lipopolysaccharides (sugars linked to membrane lipids), which are often antigenic and or toxic.

45. Capsule Some bacteria (often pathogens) are surrounded by a thick polysaccharide capsule. This is a loose jelly-like or mucus-like layer. It helps prevent immune system cells from reaching the bacteria, and it forms part of biofilms.

46. Membrane Structures • Pili (singular = pilus) are hairs projecting from the surface. They are composed of pilin protein. There are several types: • DNA can be transferred between bacteria by conjugation, which is initiated when sex pili on the donor cell attach to and draw in the recipient cell. • Fimbriae (singular = fimbria) are pili used to attach the bacteria to target cells ( in infection) or to surfaces, where they form a biofilm. • Flagella are long hairs used to propel the cells. They are composed of flagellin protein. • lophotrichous = several flagella all clustered at one end. • Monotrichous= a single flagellum at one end • Amphitrichous= one or more flgellae at each end • Peritrichous= flgellae are distributed around the cell

49. Spores • Some bacteria can form very tough spores, which are metabolically inactive and can survive a long time under very harsh conditions. • some bacterial spores that were embedded in amber or salt deposits for 25 million years have been revived. These experiments are viewed skeptically by many scientists. • Spores can also survive very high or low temperatures and high UV radiation for extended periods. This makes them difficult to kill during sterilization. • Anthrax • Spores are produced only by a few genera • Bacillus species including anthracis (anthrax) and cereus (endotoxin causes ~5% of food poisoning) • Clostridium species including tetani (tetanus), perfringens (gangrene), and botulinum (botulism: food poisoning from improperly canned food)