Microbiology Notes free essay sample

His masterpiece sculptor â€Å"David†, was made between 1501 and 1504 C. He was a French sculptor and poet D. Created the famous fresco on the ceiling of the Sistine Chapel in Rome Branches of Microbiology Medical Microbiology Industrial Microbiology Environmental Microbiology Agricultural Microbiology Legumes +N2-fixing microorganisms Microorganisms have important role in cycling plant nutrients (N, P, C, S) Cattle production depends on rumen microorganisms Biotechnology Food production (Dairy products (ogurt, cheese) sauerkraut, pickles, sausage, bread, beer, wine etc. Bioremediation (detoxification by use of microorganisms Genetic Engineering (manipulation of genes in order to create new products) * Who started Microbiology? Antoine van Leeuwenhoek a Dutch merchant The first person to see microbes as being ALIVE! Father of bacteriology * * What Leeuwenhoek could see? Different shapes of bacteria Red blood cells, spermatozoa, plant and animal cells Magnification of his micro scope was 32X Robert Hooke’s Microscope Compared to Leeuwenhoek’s Robert Hook introduced the compound microscope (objective lens + ocular lens) Robert Hook (1665) reported that the smallest biological structural unites were â€Å"little boxes†-cells CELL THEORY- all living things are composed of cells Golden Age of Microbiology (1830-1900) Scientists searched for answers to the following questions: Is spontaneous generation of microbial life possible? What causes fermentation? What causes disease? How can we prevent infection and disease? Is spontaneous generation of microbial life possible? Aristotle proposed spontaneous generation-living things can arise from non-living matter Toads and snakes could be born of moist soles Maggots could arise from corpses Francesco Redi’s Experiment, 1670s Flies with meat 1765 Lazzaro Sallanzani conducted experients to disprove the doctrine of spontaneous generation biolied infusion, hermetically sealed the container. We will write a custom essay sample on Microbiology Notes or any similar topic specifically for you Do Not WasteYour Time HIRE WRITER Only 13.90 / page Nly a small crack in the neck was sufficient to allow penetration and development of â€Å"animalcules† in the infusion. Nicholas Appert patented his invention and established a business to preserve a variety of food in sealed bottles-canning Finally! Arguments about spontaneous generation resolved by Louis Pasteur Read about Louis Pasteur’s experiment How he really did it? Pasteur’s S-shaped flask kept microbes, but let in (Swan Neck Flask) Joseph Meister First person to be treated by Louis Pasteur against rabies Pasteur’s Discoveries Performed the most convincing experiment disproving the theory of spontaneous generation Role of yeast in alcohol fermentation Devised the process of pasteurization and basis of aseptic techniques Disease of vine-cold be prevented by heating the wine for a short time to a temp of 55-60 degrees Celsius The first preventive treatment for rabies What causes disease? Pasteur developed germ theory of disease, but had no proof†¦began working on Anthrax, but could not get a cure Robert Koch Studied causative agents of disease Gave the first proof that bacteria causes disease (by isolating the anthrax bacillus in pure culture)-germ theory of disease Perfected the technique of isolating bacteria in pure culture-solid media-boiled potato Discovered Mycobacterium tuberculosis-the organism causing the tuberculosis. Tuberculin-substance used for diagnosis of tuberculosis Excreted into liquid culture and if there is an allergic reaction than the person has tuberculosis Introduced the staining procedure for M. tuberculosis (acid fast staining) First to stain bacterial smears Discovered the causative agent of cholera 1905 received the Nobel Prize for Medicine Koch’s postulates Identify criteria for proving that a specific type of a microorganism causes a specific disease: 1. The microorganism should be constantly present in a diseased animal 2. The microorganism must be cultivated in pure culture 3. When inoculated into a healthy animal, such microorganism should cause characteristic disease symptoms 4. The microorganism should be re-isolated from the experimental animal, and it should have the same characteristics as the original microorganisms Ignaz Semmelweis e implemented that if doctors wash their hands they would reduce spreading germs Joseph Lister (1827-1912) Father of modern surgery Became aware of Louis Pasteur’s work (rotting and fermentation) Applied antiseptic (carbolic acid) treatment for prevention and care of would infection Introduced the procedure of disinfection of operating rooms Edward Jenner Performed the first vaccinat ion against small pox Chemotherapy (treatment of disease by using chemical substances) the first antibiotic was discovered by accident was penicillin Alexander Fleming-1928 discovered penicillin Produced by mold: Penicillin Crysogenum Commercial production started in 1945 in the US Problems with Antibiotics Can be toxic to humans Spread of new varieties of resistant microorganisms Development of techniques that facilitated further progress of microbiology Pure cultures (clone) is a culture consisting of only one type of microorganisms Use of Petri dishes Aseptic techniques Bacteria as distinct species Development of microbiology in the 20th century Bacteriology study of bacteria. Mycology study of fungi. Fungal infections rising Parasitology study of protozoa and parasitic worms Immunology study of immunity. Virology study of viruses. Cannot be seen by a microscope, you need an electro microscope (Dimitri Iwanowski, reported on the presence of infectious agents much smaller than bacteria. He discovered the tobacco mosaic virus) Recombinant DNA technology fragments of foreign DNA can be incorporated into bacterial genome The Establishment of the Scientific Method Scientific method-based on an experimental system It includes: 1. Observation and description a phenomenon 2. Formulate a hypothesis to explain the phenomena 3. Performance of experimental tests (to confirm hypothesis) If supported hypothesis becomes theory Joseph Lister He was born in 1827 in Upton, England Attended London and Edinburgh Universites Joseph Lister was educated in Edinburgh. Edinburgh is located in Scotland Scotland is an independent state Scotland is part of Great Britain Scotland is a province in France Scotland is part of Canada Why Chemistry? Bacteria subcellular structures complex molecules simple molecules atoms Atoms Building blocks of molecules Subatomic particles Shell Electrons rotate about the nucleus in their orbitals Nucleus Protons Neutrons Atoms do not carry charge Electron Orbitals and Shells Electrons rotate around the nucleus in orbitals (pathways) Low level energy electrons Nearest to the nucleus 1st shell-1 orbits (2e) 2nd shell- 4 orbitals (up to 8 e) 3rd shell-9 orbitals (up to 18e) Elements Each element contains characteristic number of protons and electrons-position in the table There are 118 known elements Elements in groups of similar properties Biologically important atoms 11 elements are of biological importance The Fathers of the Periodic Talbe Dmitri Mendeleev Lothar Meyer Isotopes Have a different number of neutrons; same physical properties Some isotopes have unstable nuclei-emit energy-radioactivity Significance in Biology Tracing atoms and molecules in metabolic reactions Diagnostic purposes Sterilization (gamma irradiation) Video Notes: Charges of protons and electrons are equal in magnitude Isotopes are identical to each other but sometimes vary in properties Carbon 12 and 13 are stable Carbon 11, 14, 15 are not Radiactivity Henri Becquerel-mysterious rays * Molecules Two or more atoms combined form a molecule-a new compound has new characteristics Compounds are formed when atoms are, donate or lose electrons Number of electron in outer shell-valence Determines the readiness of an element to react with other element How the atoms bind together? Through the chemical bonds Covalent bonds Ionic bonds Hydrogen bonds Covalent Bonds Bonds between atoms that share electrons Electrons are not shared equally- change of polarity Hydrogen molecule H2 Water molecule is polar Ionic Bonds Electrons are transferred from one atoms to another When valences complement each other Ex- NaCl (table salt) 7 is not stable Ionization Dissociation of molecules (atoms) into charge particles: Cations (+) Anions (-) When added to water, crystals of NaCl get ionized-Na+ Cl- Electrolytes-conduct electricity: acids, bases, salts When two electrodes are put together Each ion becomes hydrated-surrounded by water molecules Hydrophilic molecules attracts water Hydrophobic molecules repel water (benzene- a non-polar molecule) Hydrogen Bonds Attractive forces between nearby molecules Ex. Water molecules; proteins and DNA Represented with dotted line Oxidation-Reduction Reactions Oxidation-losing electron Reduction-receiving electrons Chemical formulas Atomic symbols and number of atoms in a molecule: H2O Do not provide information on position of bonds between atoms C6H12O6 glucose and fructose Reactions Synthesis reaction Equation must be balanced Decomposition reactions Larger molecule is broken up in two smaller units 2H2+Ox 2H2O Solutions Mixture of substances solute- (solid, gaseous, liquid) in the solvent (liquid) Water is the most common solvent * * Acidity, Alkalinity, pH * Solutions can be acidic or basic H2H+ + OH- ionization of water H+H+ + OH- access of H+ acidic pH pH=_log [H+] pH1 =0. 1 mole H=/liter ph 2+0. 01 mole H+/liter Inorganic and Organic Compounds Compounds can be: Inorganic ( do not have C and H combined) NaCl, CaCO3†¦ Organic (CH4)-complex compounds with C boned to other atoms The Chemistry of Carbon Carbon is the fundamental element of life- Why? Molecular skeleton Have 4 electrons in outer orbital-can form 4 bonds Most often forms stable bonds with C H O N S and P Functional Groups Carbon binds to other atoms via molecular groups-functional groups Functional groups determin characteristics of a molecules Carbodroxyl-fatty acids, proteins, organic acids Amino- proteins, nucleic acids Macromolecules Smaller molecules (monomers) are assembled into larger compounds-macromolecules (polymers) **Table 2. 3 and Table 2. 4 ** Classes of Carbohydrates Monosaccharide Have 3-7 atoms Pentose (5 Carbons) Hexose (6 carbons) Glucose Disaccharides Sucrose Polysaccharides Glycosidic Bonds Bonds between two sugars Carbons from two molecules are bonded via oxygen with release of H2O molecule Function of Polysaccharides Provide structural support (cell wall) Nutrient and energy storage Some examples Cellulose Agar Chitin-present in insects Peptidoglycan-polysaccaharide makes up cell wall of bacteria Glycocalyx- surrounds bacterial cell and provides protection Lipids Not soluble in water Classes of Lipids Triglycerides Phospholipids Steroids Waxes Synthesis of Triglycerides Glycerol bonds with three molecules of fatty acds through ester bonds Dehydration (need more) Biological Significance of Lipids Storage material (triglycerides) Membrane lipids Phospholipids Hydrophilic and hydrophobic portion Cholesterols Provides support to the cell wall of some bacteria Proteins Composed of amino acids Assembled together through peptide bonds Peptide: short chain of amino acids Polypeptide (proteins): long chains of amino acids Protein structure determines its functionality: Primary-long chain of amino acids Secondary Tertiary Quaternary- complex molecule * * Peptide Bonds Bond between amino group of one AA and carboxyl groups of another AA Proteins are made up of 20 AA Look at list the Waterstreet gave us The Nucleic Acids DNA and RNA; informational molecules Contain genetic information Composed of nucleotides Nucleotides composed of: Bases Sugars Phosphate The Double Helix of DNA Held together by nucleotides ATP: the energy molecule of cells Adenosine triphosphate contains Adenine Ribose Three phosphates Gives off energy when the bond is broken and one phosphate group removed Marie Curie Pioneered research on radioactivity Theory of radioactivity Discovered two elements: polonium and radium The first female professor at the University of Paris The first Nobel st Nobel prize- in physics 2nd Noble prize in chemistry Marie Curie was born in France England Poland Austria General Laboratory Techniques-5 I’s Inoculation Incubation Isolation Inspection Identification Inoculation Introducing a sample (the inoculum) into a container with a nutrient medium The medium contains appropriate nutrients that sustains the growth of microorganisms Some micro bes have to be inoculated into a living organism Isolation: Separating one species from another Obtaining pure culture Cultures composed of cells arising from a sing cell-PURE CULTURES Obtaining Pure Culture Streak plate method Petri plate Material picked up from somewhere and put at the tip of inoculation needle Dilution method Transfers to build bacteria in the petri plates Types of Media Physical State Liquid Solid (agar) Chemical composition Synthetic Nonsynthetic (complex) Functional type General purpose Enrichment Selective Differential Measures to be taken when working with microbiological media Needs to be sterilized Prevent contamination Synthetic media Known chemical composition (NaNO3/I; glucose 2g/l) Non-synthetic (complex) media Contains chemically undefined components (Pepton, beef extract†¦) Enrichment medium- supports the growth of a specific group of microorganisms (Ex. N2-fixing) Selective media-favor specific microorganisms and inhibits the others (methylene blue inhibits the growth of Gram+ bacteria) Differential media-contain substances that permit detection of microorganisms with specific metabolic activity E. coli (red) Incubation Microbiological cultures are placed temperature-controlled chamers-incubators Temperature: 20-40 C Pathogenic: 37 C MICROSCOPE-The Instrument Microscopes are the instruments that magnify the cell (object) to extent at which the cell details become visible Leeuwenhoek’s microscope had one lens Robert Hooke invented the compound microscope Microscope- The Basic Principle The specimen is magnified with the objective lens (real image) This image is magnified by ocular lens (virtual image) An enlarged and inverted image is received by retina Basic features of microscopy Magnification Resolution Contrast Magnification Magnification is the result of light refraction Mag=Objective powerx Ocular lens power Ex: Objective lens: low power lens=10x Ocular lens power=10x Mag=10x 10=100x * * Your objective lenses are color coded 4x- red 10c-yrlloe 40x-blue 100-black Use of immersion oil with high power objectives Immersion oil has the same refractive index as the glass Between glass cover slip and slide Used only with 100x lens magnification Refractive index is a measure of relative velocity at which light passes through material Resolution Resolution (resolving power) is the ability of a lens to distinguish two adjacent points as two separate objects. In light microscopes resolution is 0. 2 micrometers (limit 2,000x) High resolution-further apart Low resolution-closer together Which images can be resolved? Lower than 0. 2 micrometers Bigger than 0. 2 micrometers can be seen with microscope How does the resolution depend on the wavelength? Resolving distance=wavelength of light/w xNA (numerical aperture) The shorter the wavelength- the greater resolution Contrast Specimen must contrast with their background This can be achieved by: Changing the refractive index of specimen Stain the specimen Refractive index- mesures of light bending ability * * Types of Microscopes * What is the illuminating sources? * Light Bright field Phase contrast Fluorescent Dark filed Differential interference Confocal * Electron beam transmission scanning Light Microscopy-Compound Microscope Optical microscope parts: Illuminator Condenser Objective lens Ocular lens (eyepiece) Dark-Field Microscopes Best for observing pale objects Only those light rays scattered by specimen enter objective lens Specimen appears light against dark background Increases contrast and enables observation of more details Bright filed vs. Dark field Microscopy When light reaches the ocular lens dark objects are seen easier Fluorescent Microscopy Fluorescence is the ability of certain substancces to absorb short wavelengths of light and emit light at a longer wavelength Fluorescence Microscopy Sputum mucus Same mucus showing Mycobacterium tuberculosis stained with a fluorescent dye Immunofluorescence Diagnostic procedure: Antibody produced against a specific bacterium Conjugate antibody and fluorochrome Treat the unknown bacterium If suspected bacteria are indeed present they will bind the tagged antibodies Ultraviolet (or near) light is used as a light source Phase Microscopes Provides better contras and more details in the cell The light rays that hit he specimen travel a different path than the rays, which do not hit the specimen Bright field looks very light Phase looks better Differential Interference Microscopy (Nomarsky) Uses two beams of light Higher resolution -D images Confocal Microscopy Uses fluorescent dyes and UV lasers to illuminate the sample An image is taken in a single plane that is not thicker than 1. 0micrometer Resolution is increased by up to 40% because emitted light passes through pinhole aperture Computer constructed 3-D images Bacillus cereus Live cells-red Dead cells-green Composite image of 10 images Each 0. 2 micrometers thickness HIV (in green) travels down the microtubules (in red) Toward the nucleus (Blue) Bacteria are seen in light microscopes Electron Microscopy Two types of electron microscopes: Transmission (TEM) Scanning (SEM) Source of illumination electron beam Advantage of using EM Resolving distance=wavelength of lig/2 Wavelength of visible light=400A Resolution (light microscopy): 2000 A (0. 2 micrometers) EM uses an electron beam as a source of illumination (1000,000 times shorter wavelength than visible light) Resolution (EM): 2 A Magnification up to 100,000 Transmission electron microscope (TEM) Image formed by the elctrons transmitted through a specimen A specimen is a thin section of material (fixed, embedded, and sliced-never alive) TEM is used for objects smaller than 0. 2 mm * The first electron microscope Constructed by Ernst Ruska in 1931 He demonstrated that a magnetic coil could act as an electron lens In 1986, he was awarded the Nobel Prize in Physics for his many achievements in electron optics Ruska was born in Heidelberg Heidelberg is city in France Heidelberg University is one of the oldest Universities in Europe Heidelberg is a city in Germany B and C are the correct answers Scanning electron microscope Used to study the surface of the cell/tissue Image formed by the electrons reflected from the surface 3D view image is obtained on TV screen Preparation of specimen for optical microscopes