Microbiology Lecture Notes PDF Outline - Exam 1 2025

From: Tortora, et al., Microbiology: An Introduction, 11th Edition, Pearson Education, Inc. Science of Microbiology dates back only a few hundred years, but… DNA of Mycobacterium tuberculosis, c.a. of TB, found in 3000 year-old Egyptian mummies Endospore-forming bacteria may have been revived...

From: Tortora, et al., Microbiology: An Introduction, 11th Edition, Pearson Education, Inc. Science of Microbiology dates back only a few hundred years, but… DNA of Mycobacterium tuberculosis, c.a. of TB, found in 3000 year-old Egyptian mummies Endospore-forming bacteria may have been revived from 200-million-year old salt crystals We can get an idea how current concepts in microbiology developed by looking at a few historic milestones in microbiology that have changed our lives… Robert Hooke 1665 Englishman; built the first crude microscopes Observed thin slices of cork and described “little boxes” – cells Proposed that cells were the smallest structural units of life – the beginning of Cell Theory: “All living things are composed of cells” Anton von Leeuwenhoek 1670s Dutch Merchant, amateur scientist Built the first effective microscopes; over 400 of them Excellent lens grinder; a single lens: 50 – 300 x magnification Observed, described and drew: Protozoa Bacteria Yeasts Molds Algae RBCs First to describe the shapes of bacteria; often called the “Father of Bacteriology” Most bacteria exist as one of the three shapes described by Leeuwenhoek Spherical-shaped bacteria are called cocci (single, coccus) Rod-shaped bacteria are bacilli (bacillus) Spiral-shaped bacteria are termed spirilla (spirillum) or vibrios, depending on the number of twists and turns Spontaneous Generation Debate During the time Hooke and Leeuwenhoek lived, debate regarding the origins of life: Spontaneous Generation (aka abiogenesis) was active. A concept started by Aristotle (346 B.C.) who said: …life can appear spontaneously from non-living matter Francesco Redi 1668 Italian Physician; set out to disprove the theory of S.G. Demonstrated that maggots did not arise spontaneously from decaying meat – a common belief Set up an experiment with three jars containing decaying meat: From: Tortora, et al., Microbiology: An Introduction, 11th Edition, Pearson Education, Inc. John Needham mid 1700’s Redi’s experiments served to shift the S.G. proponents’ claims to be valid for microbes only – simple John Needham, Englishman, Catholic priest Conducted experiments he believed proved bacteria could arise spontaneously from organic matter Spontaneous Generation Debate – Needham Designed an experiment using boiled chicken broth: Claimed that boiling killed the microorganisms; but that new microbial life arose from the organic matter where there was none before: spontaneous generation Lorenzo Spallanzani 1760’s Italian scientist Answered Needham’s claim; Spallanzani said that open flasks allowed airborne microbes to contaminate the broth: Designed an experiment using boiled chicken broth: Needham’s answer: sealing the flask cut off oxygen and prevented the “vital force” from getting in the broth and generating new life… From: Tortora, et al., Microbiology: An Introduction, 11th Edition, Pearson Education, Inc. Louis Pasteur 1861 French microbiologist and chemist Finally put the argument to rest by demonstrating that microbes are present in the air and can contaminate sterile solutions: Designed an experiment using boiled beef broth: the “swan-neck flask” experiment The design allowed air into the boiled broth, but microbes settled in the curved neck Only when the stem was broken and microbes could fall into the flask did growth occur Pasteur Microorganisms are present in the air – As free cells Adhered to dust particles Attached to skin cells shed by humans and other animals Hitching a ride on insect parts Attached to many other particles Pasteur showed that microbes can be present in/on non-living matter Solids Liquids Gases (air) They are ubiquitous – occurring virtually everywhere Pasteur demonstrated that: Microbes are ubiquitous Microbes can be destroyed by heat Microbes can be blocked from the environment His discoveries formed the basis of Aseptic Technique – a means of containment and exclusion of microbes; now the standard practice in laboratories and medical environments Pasteur also responsible for: Fermentation of fruits and grains to produce ethanol is the result of microbial growth From: Tortora, et al., Microbiology: An Introduction, 11th Edition, Pearson Education, Inc. The process of gentle heating that kills many microorganisms while preserving the flavor of foods: named for him – Pasteurization Improving the consistency of wine by selecting specific microbes for the fermentation process while excluding others – Pure Culture Technique The realization that microbial activity changes the physical characteristics of organic material got scientist to wondering if microbes can cause changes in plant/animals…specifically, can they cause disease? Experiments in this area led to the Germ Theory of Disease – “germs” cause certain diseases Evidence: 1865 – Pasteur showed that Silkworm Disease was caused by and infection of silkworm moths by a protozoan Evidence for germ theory of disease, cont: 1860s – Joseph Lister English Surgeon Began treating surgical wounds with phenol…a cmpd. he knew killed microorganisms Greatly reduced the number of surgery-related infections and deaths. His methods were soon adopted by other surgeons Proof of the germ theory of disease came from Robert Koch, 1876: German Physician Discovered a rod-shaped bacterium in the blood of cattle dying of anthrax Proposed that the bacterium was the causative agent of anthrax Since humans are also susceptible to anthrax…this was the first report of a microorganism causing a human disease Koch thus established a set of principles known as Koch’s Postulates, which state that: 1. A specific organism can always be found with a specific disease 2. The organism can be isolated (recovered) from the diseased animal/plant and grown in pure culture in the laboratory 3. When inoculated back into a healthy, susceptible host, the pure culture will produce the disease 4. The organism can be recovered from the newly-infected host and grown in the lab in pure culture Koch also: Invented solid culture media Developed dyes to stain microorganisms for microscopy Discovered the bacterium that causes tuberculosis Edward Jenner 1796 First experiments to impart immunity from a specific disease From: Tortora, et al., Microbiology: An Introduction, 11th Edition, Pearson Education, Inc. Used scrapings from cowpox (mild disease) blisters to inoculate healthy volunteers who became mildly sick, but were now immune to smallpox…a killer of 400,000/yr. in Europe He named the process: vaccination (vacca,: Latin for “cow”) Paul Ehrlich 1910 German Physician Developed the first drugs to effectively treat infectious diseases Salvarsan – arsenic drug used to treat syphilis Established the practice of chemotherapy – using chemical compounds to fight microbial infections without harming the host Alexander Fleming 1928-29 Scottish physician Observed that a pure culture of a bacterium he had grown on solid culture media was contaminated with a mold colony Growth of bacterial colonies near the mold was inhibited Alexander isolated the inhibitory substance and named it penicillin, after the mold (Penicillium sp.) Alexander Fleming 1928-29 One of the greatest discoveries in science It led to the search for “substances produced by one microorganism that kill or inhibit the growth of another microorganism” – Antibiotics 1950’s - magnification and resolution of electron microscopes (EM; first built in the late 1930s) improved greatly Many cell structures were described that were previously unseen with the light microscope (LM) LM limit = 1500x EM limit = 400,000x (early models) With increase in magnification and resolving power: two very different cell types seen, and called: Prokaryotic Cell Eukaryotic Cell PROKARYOTIC CELL More primitive cell type No nuclear membrane No mitochondria or other complex organelles Structural unit of bacteria and archaea More complex cell type Membrane-bound nucleus From: Tortora, et al., Microbiology: An Introduction, 11th Edition, Pearson Education, Inc. Many organelles - mitochondria, golgi apparatus, endoplasmic reticulum, etc. Structural unit of fungi, algae, protozoa, higher plants and animals Microbial Classification PROKARYOTES Bacteria Archaea ACELLULAR FORMS Viruses EUKARYOTES Fungi Protozoa Algae Helminths Prokaryotic & Eukaryotic Cells PROKARYOTIC CELL More primitive cell type No nuclear membrane No mitochondria or other complex organelles Structural unit of bacteria and archaea EUKARYOTIC CELL More complex cell type Membrane-bound nucleus Many organelles - mitochondria, golgi apparatus, endoplasmic reticulum, etc. Structural unit of fungi, algae, protozoa, higher plants and animals Prokaryotic and Eukaryotic Cells Groups of Microorganisms 1. Bacteria Saprophytes or Parasites (some photosynthesis) Saprophytes obtain nutrients from dead organic material Parasites derive nutrients from living host Very small - usually < 5m in diameter Micrometer = 1 millionth of a meter Complex cell wall containing peptidoglycan Peptidoglycan = unique structural molecule made of repeating units of sugars and amino acids Unicellular Primary form of reproduction = binary fission Budding, fragmentation, low order of sexual reproduction Groups of Microorganisms 2. Archaea From: Tortora, et al., Microbiology: An Introduction, 11th Edition, Pearson Education, Inc. Saprophytes Usually only cell membrane If cell walls - no peptidoglycan Three main groups: Methanogens - produce methane as waste product Extreme Halophiles - live in extremely salty environments Extreme Thermophiles - sulfur hot springs Many are anaerobes - do not require O2 to grow Not sensitive to antibiotics No known human disease Groups of Microorganisms 3. Fungi Yeasts and Molds Saprophytes or Parasites Eukaryotic cell structure No photosynthesis Unicellular or multicellular Larger than bacteria Cell wall of chitin Chitin - nitrogen-containing polysaccharide Sexual and asexual reproduction Groups of Microorganisms 4. Protozoa Unicellular Eukaryotes Larger than bacteria Move by Pseudopods, Flagella or Cilia Pseudopod = false foot Saprophytes or Parasites Tremendous morphological diversity Found in a wide variety of environments with H2O Some photosynthetic species Reproduce sexually or asexually Groups of Microorganisms 5. Algae Photosynthetic Eukaryotes Use light as an energy source, CO2 as carbon source Cell walls composed of cellulose Cellulose = linear chain of glucose, ß(1-4) linkage Single cells or multicellular Abundant in fresh water, salt water, soil From: Tortora, et al., Microbiology: An Introduction, 11th Edition, Pearson Education, Inc. Asexual and sexual reproduction Groups of Microorganisms 6. Viruses Acellular Ultra small “filterable units” Most can be seen with EM only Obligate intracellular parasites; many pathogens Bacteria, Plants, Animals Simple structure: DNA or RNA plus protein coat DNA = deoxyribonucleic acid RNA = ribonucleic acid Reproduce only using host cell “machinery” Viruses are inert outside the host cell Groups of Microorganisms 7. Helminths Flatworms and Roundworms Eukaryotic cell structure Multicellular, animal parasites Some stages of their life cycle are microscopic Parasitic forms: Often lack a digestive system (absorb) Have a reduced nervous system (food search) Can lack a means of locomotion (host-to-host transfer) Possess complex reproductive cycle Nomenclature 1735: Carolus Linnaeus, Swedish physician and biologist, established system still in use today Scientific names of organisms are “latinized” Latin was the language of scholars Each organism assigned two names: Genus - first name, always capitalized (plural: “genera”) Species (specific epithet) - second name, lower case Both names are underlined or italicized Can abbreviate the genus name after the full name is first mentioned Pseudomonas aeruginosa becomes P. aeruginosa Nomenclature Scientific names can: Describe an organism’s appearance, physiology, etc. Honor a researcher, scientist (usually in the same field) From: Tortora, et al., Microbiology: An Introduction, 11th Edition, Pearson Education, Inc. Identify the habitat of the organism Examples Staphylococcus aureus - staphylo clustered arrangement coccus spherical shape of cells aureus L. “golden”, colony color on agar plates Escherichia coli - Named in honor of Theodor Escherich, German pediatrician who established the relationship between intestinal bacteria and digestion in infants (1886) The Scientific Method Why Use the Scientific Method? The scientific method attempts to minimize the influence of bias or prejudice in the experimenter (even the best-intentioned researchers can't escape bias from personal beliefs, cultural beliefs, etc. In the scientific community, where results have to be reviewed and duplicated, bias must be avoided. That's the job of the scientific method. It provides an objective, standardized approach to conducting experiments and, in doing so, limits the influence of personal, preconceived notions of the researcher. The Scientific Method The process begins with the formulation of a question Then, facts regarding the question are gathered through observation or experimentation Next, a hypothesis (well-educated guess) is made, based on the observations, that might explain the facts The hypothesis is tested by experimentation If supported by the experimental data (accept/reject), the hypothesis becomes a theory…tested and retested by other scientists The Scientific Method - Example Edward Jenner, a British Physician and pioneer of the smallpox vaccine. Jenner’s scientific method: Question: Can people be immunized against smallpox? Observation: People who have had cowpox do not become ill with smallpox Hypothesis: If a person has been intentionally infected with cowpox, then that person will be protected from becoming ill after a purposeful exposure to smallpox Test: Infect a person with cowpox. Then try to infect the person with smallpox. From: Tortora, et al., Microbiology: An Introduction, 11th Edition, Pearson Education, Inc. Conclusion: Infecting a person with cowpox protects from infection with smallpox Microbes and Human Welfare Most microorganisms are beneficial to humans, other animals, and plants Beijerinck and Winogradsky: Described the role of microbes in recycling vital elements between soil and atmosphere Microbes convert carbon, nitrogen, phosphorous, sulfur and other elements into chemical forms plants and animals can use Examples of this process: Rhizobium sp. for N2 fixation…only bacteria Bacteria and fungi decompose organic wastes and return CO2 to the atmosphere Microbes and Human Welfare Microbes recycle water Microorganisms, esp. bacteria, are source for the conversion of organic wastes to minerals in waste water Sewage treatment Microbes clean up pollutants Diversity of microbial enzymes and physiology makes them useful for cleaning up toxins and other pollutants - bioremediation Underground wells, industrial pollution sites Microbes help control pests Bacterium known as Bacillus thuringiensis produces protein crystals that are toxic to insects that damage crops Microbes are used in biotechnology and gene therapy research Recombinant DNA technology is used to turn microbes into biochemical factories Vaccines, proteins, enzymes for practical use - biotechnology Gene therapy uses viruses to transfer new or missing genes to host cells Ex: Adenosine deaminase (ADA) deficiency – SCID Microbes and Human Welfare Advantages of Using Microorganisms in Research Keep a large breeding stock in a small space (millions/mL of broth) They carry out some chemical processes 10 to 1 million times faster than larger forms of life Cheap to grow and maintain - cost efficient compared to plants and animals Genetics experiments can take place in a few days vs. years for plants or animals - rapid reproduction To illustrate: From: Tortora, et al., Microbiology: An Introduction, 11th Edition, Pearson Education, Inc. 70% earth is covered in water - 330 million cubic miles of H2O By continued progression of multiplication, the bacteria which spring from a single organism would take less than five days to fill the earth’s seas Bacteria and other microbes possess an amazing array of enzymes and biochemical processes they can carry out Microscopy and Specimen Preparation - Chapter 3 Microscopy Light Microscopes: Bright Field - type of light microscope showing darker objects on a light background Dark Field - light microscope with dark background; objects appear light Phase Contrast - used to study living organisms; special condenser that helps distinguish objects of similar density such as internal structures Fluorescence - allows us to see fluorescence (natural and artificial) in cells; Mycobacterium tuberculosis stained with Auramine O will fluoresce bright yellow under the scope Microscopy Electron Microscopy (EM) To view objects < 0.2 m (bacteria 1- Viruses Internal structures in cells EM uses a beam of electrons instead of light to create an image Wavelengths of electrons are 100,000 times smaller than those of visible light Allows for much greater magnification and object resolution Microscopy Transmission Electron Microscopy (TEM) Beam of electrons passes through an ultrathin section of the specimen The beam continues through a series of electromagnetic lenses onto a fluorescent screen or a photographic plate The final image, the transmission electron micrograph, is composed of light and dark areas, much like a black-and-white photograph TEM excellent for viewing different layers of a specimen; internal structure Electron Microscopy Scanning Electron Microscopy (SEM) The specimen must be sectioned for TEM, which: Kills the organism Can cause distortion and other artifacts With SEM, No sectioning Primary and secondary electron beams From: Tortora, et al., Microbiology: An Introduction, 11th Edition, Pearson Education, Inc. Image of the sample surface 3-D effect SEM provides a view of the surface of the object; gives depth to the object. Specimen Preparation - LM Preparing Bacterial Smears for Staining Light microscopy requires staining Thin film of microbial growth applied to a glass slide This smear is allowed to air dry Must “fix” the organisms onto the slide – heat Basic dyes commonly used because bacteria are negatively charged at pH 7 Examples of typical basic dyes: Crystal violet Methylene blue Malachite green Safranin Simple and Differential Stains Simple Stains Single, alkaline dye used Colors the entire cell Highlights basic shape and arrangement of cells Dye applied to fixed smear for a specified time and then washed off Slide allowed to air dry, then examined by LM Simple and Differential Stains Differential Stains Multiple dyes or reagents are used Can color all, or just part, of the cell Can react differently with different species Common Differential Stains: Gram Stain 1st step in bacterial ID Acid Fast Stain Mycobacterium tuberculosis The Gram Stain Light Microscopy Stain Developed by Hans Christian Gram – 1884 Danish bacteriologist The Gram Stain Light Microscopy Stain Developed by Hans Christian Gram – 1884 Danish bacteriologist From: Tortora, et al., Microbiology: An Introduction, 11th Edition, Pearson Education, Inc. Classifies bacteria into two large groups Gram-positive (purple or blue) Gram-negative (red or pink) First stain used in ID of unknown bacterium Shape Arrangement Relative size Gram reaction The Gram Stain Steps of the Gram Stain: Heat-fixed slide is covered with an alkaline purple dye, the primary stain. All cells stain purple. Purple dye is washed off and iodine (mordant) is added to the slide. Iodine intensifies the stain...binds to the crystal violet. Slide is washed with alcohol, the decolorizing agent, which removes the purple dye from the cell walls of some species. The alcohol is washed off, and a red counter stain is added…stains only those species decolorized by the alcohol. The Gram Stain Typical dyes and contact times Primary Stain: Crystal Violet – 1 min Mordant: Gram’s Iodine – 1 min Decolorizing Agent: 95% ethanol – 15 sec Counter Stain: Safranin – 1 min Choice of dyes and contact times vary somewhat between laboratories/researchers The Gram Stain Classifying bacteria by Gram Stain: Basic to bacterial identification Critical technique for medical microbiology Bacterial cultures 18-24 hours old are used Most accurate results Older cultures can lose ability to resist decolorizing Some species of bacteria are Gram-variable: Even under ideal conditions, Gram stain will result in both Gram-positive and Gram-negative cells Relatively few bacterial species The Gram Stain Gram reaction is a function bacterial cell wall structure and chemical composition From: Tortora, et al., Microbiology: An Introduction, 11th Edition, Pearson Education, Inc. Gram-positive bacteria: cell walls composed of a thick layer of peptidoglycan (structural molecule of saccharides and amino acids) Gram-negative bacteria: cell walls have thin layer of peptidoglycan and an outer layer of lipopolysaccharide Crystal Violet-Iodine complex trapped in Gram-positive cells…purple dye remains Alcohol (decolorizing agent) disrupts polysaccharide layer in Gram-negative cells…purple dye leaks out Counter stain (pink) colors Gram-negative cells The Acid-Fast Stain Differential for bacteria in two genera: Mycobacterium Mycobacterium tuberculosis - Tuberculosis Mycobacterium leprae – Leprosy Nocardia Nocardia asteroides – opportunistic pathogen immunocompromised patients The Acid-Fast Stain Steps in the Acid-Fast Stain: Carbolfuchsin - primary stain Heated to help dye penetrate cell wall Acid alcohol - the decolorizing agent Methylene blue - counter stain The Acid-Fast Stain Mycobacterium and Nocardia species Cell walls contain large amounts of waxes and glycolipids Causes cell to retain primary dye and resist decolorizing with acid-alcohol Cells of Mycobacterium sp. and Nocardia sp. are said to be acid-fast Special Stains Used to color and isolate specific parts of the bacterial cell Capsule stain Endospore stain Flagellum stain Structure and Function of Prokaryotic and Eukaryotic Cells All living cells: two groups Prokaryotes Eukaryotes Macroscopic plants and animals, fungi, algae, protozoa are eukaryotes Bacteria and archaea are prokaryotes From: Tortora, et al., Microbiology: An Introduction, 11th Edition, Pearson Education, Inc. Prokaryotic and Eukaryotic Cells Comparing Prokaryotic and Eukaryotic Cells - An Overview Chemically Similar - Nucleic acids - Proteins - Lipids - Carbohydrates Same chemical reactions for catabolism anabolism Principal differences: - Structure of cell walls - Cell membrane presence and structure - Presence/absence of organelles Prokaryotes No nuclear membrane No histones with DNA No organelles Cell walls have peptidoglycan Cell division by binary fission Plasma membrane lack sterols, carbohydrates 70S ribosomes *Ribosome: Site of protein synthesis; amino acids are linked together Eukaryotes True nucleus with membrane Histones present Membrane-enclosed organelles No peptidoglycan Cell division by mitosis (replication of chromosomes into two separate nuclei) Sterols, carbohydrates in plasma membrane (serve as receptors) 80S ribosomes (70S in organelles) *Histones: alkaline proteins found in the nucleus, that package and order DNA Prokaryotic Cells - Functional Anatomy Prokaryotes: Bacteria and Archaea Majority are bacteria Species of bacteria differentiated by: Morphology Chemical composition (esp. cell wall) Nutritional requirements Biochemical activities Sources of energy Prokaryotic Cells - Functional Anatomy Size, Shape and Arrangement of Bacterial Cells From: Tortora, et al., Microbiology: An Introduction, 11th Edition, Pearson Education, Inc. Most are 0.2 to 2.0 m diameter; 2- 8 m length A few basic shapes Coccus (plural - cocci) - spherical to oval Bacillus (plural - bacilli) - rod-shaped cells; coccobacilli – very short rods Spirillum (plural - spirilla) - spirals; comma-shaped Prokaryotic Cells - Functional Anatomy Coccus arrangements: Diplococci - cocci remaining in pairs after dividing Streptococci - cocci occurring in chains after division Tetrads - cocci that divide in two planes; groups of four Sarcinae - cocci dividing in three planes; groups of eight Staphylococci - multiple planes of division, grape-like clusters Prokaryotic Cells - Functional Anatomy Bacillus arrangements: Most occur as single bacilli Diplobacilli - bacilli occurring in pairs after division Streptobacilli - occur in chains Coccobacilli - oval rods, almost spherical *Note: “Bacillus” Bacillus - describes the shape of a bacterium (rod-like) Bacillus - a genus name, as in Bacillus subtilis Prokaryotic Cells - Functional Anatomy Spirillum arrangements: Spiral bacteria have one or more twists Vibrio - curved rod, comma-shaped Spirillum - helical shape; corkscrew Spirochete - flexible rods, snake-like appearance Spirilla move by flagella: tail-like appendages, propel cells Spirochetes move by axial filaments: flagella-like, but contained within the cell’s flexible, external sheath Other Prokaryotic Cell Shapes: Genus Stella - star-shaped bacteria Genus Haloarcula (archaea) - rectangular cells Structures External to Cell Wall The Cell Wall Structures Internal to the Cell Wall Structures External to Cell Wall: From: Tortora, et al., Microbiology: An Introduction, 11th Edition, Pearson Education, Inc. Glycocalyx Flagella Axial Filaments Fimbriae Pili Structures External to Cell Wall: Glycocalyx Secreted by most bacteria to adhere to the cell surface; “sugar coat” Gelatinous polymer composed of polysaccharide, polypeptide, or both - organized and firm - Capsule - unorganized and loosely attached - Slime Layer Capsules: - Contribute to bacterial virulence Bacillus anthracis Streptococcus pneumonia Capsule makes the cells resistant to phagocytosis, viruses and certain toxins Structures External to Cell Wall: Glycocalyx – Responsible for formation of biofilms - Allows cells to adhere to each other and to external surfaces - Affords protection to cells (toxins, for example) - Allows cells to adhere to each other and to external surfaces - Helps colonize surfaces; medically important Medical implants, Pseudomonas aeruginosa Surface of teeth, Streptococcus mutans Cells of small intestine, Vibrio cholerae Structures External to Cell Wall: Flagella Some prokaryotes have flagella (singular: flagellum) Long, filamentous appendages that propel bacteria Number and location of flagella varies between species Structures External to Cell Wall: Flagella Some prokaryotes have flagella (singular: flagellum) Long, filamentous appendages that propel bacteria Described as thin, hair-like projections Number and location of flagella varies between species