MicroPara Midterms

Introduction to Microbiology

• Microbiology involves the study of microorganisms
• The word "microbiology" comes from the Greek words micros (small), bios (life), and logos (study)
• Parasitology originates from the Greek words para (with), site (food), and logos (study), focusing on living organisms (hosts) and how other organisms get food from them

History and Scope of Microbiology

• Initially focused on infectious diseases, it has expanded to include practical applications of the science.

Discovery of Microorganisms

• Robert Hooke (1665): Discovered cells while observing a piece of cork, noted for seeing strands of fungi
• Anton Van Leeuwenhoek (1673-1723): Observed live microorganisms (animacules), giving accurate descriptions of protozoa, fungi, and bacteria.

Debate over Spontaneous Generation

• Spontaneous generation: The belief that life could arise from nonliving matter
• Aristotle (~350 BC) introduced the idea that life could arise from dew, putrid matter, or dirty hay.
• Francesco Redi (1668): Challenged spontaneous generation by showing that fly maggots come from flies, not decaying meat
• John Needham (1745): Observed microorganisms forming after boiling meat, supporting spontaneous generation
• Lazzaro Spallanzani: Air might have entered Needham's experiment; sterilized covered boiled broth that did not produce microbes
• Laurent Lavoisier: Oxygen is absorbed from the air when something is burned
• Rudolf Virchow (1858): Theory of Biogenesis

Other Discoveries

• Edward Jenner (1798): Introduced the smallpox vaccine using cowpox, called "vacca"
• Agostino Bassi: Discovered that microorganisms cause disease (germ theory of disease), specifically muscardine disease in silkworms caused by the fungus Beauveria bassiana

Golden Age of Microbiology

• Louis Pasteur: Discovered that yeast converts sugars to alcohol in the absence of air during fermentation
• Pasteur addressed the souring of wine and dairy products by bacteria via pasteurization
• 1861: Determined that microorganisms are in the air, but air doesn't create microbes, forming the basis for aseptic techniques
• Disproved spontaneous generation by using swan-necked flasks filled with broth, encouraging the germ theory of disease, stating that microorganisms cause infectious diseases

Robert Koch

• Isolated Bacillus anthracis, the spore-forming bacteria
• Isolated the bacteria from a cow, the cause of Anthrax.
• Developed Koch's postulates: Specific microorganisms cause specific diseases by proof through cultivating anthrax bacteria and injecting pure cultures into mice, which invariably caused anthrax

Joseph Lister

• Discovered that phenol or carbolic acid can kill bacteria

Birth of Modern Therapy

• Focus shifted to discovering substances that could destroy pathogenic organisms
• Chemotherapy: Synthetic drugs, with Paul Ehrlich's "salvarsan" being the first synthetic drug, an arsenic derivative effective against syphilis, and developing theories on immunity
• Quinine - malaria
• Sulfonamides
• Antibiotics: Alexander Fleming discovered penicillin from Penicillium notatum
• Rene Dubos (1939): Discovered gramicidin and tyrocidine from Bacillus brevis

Modern Developments in Microbiology

• Development of microbiology branches: bacteriology, mycology, parasitology, immunology, and virology
• Heide Schulz (1997): Discovered large bacterium Thiomargarita namibiensis
• Genomics
• Recombinant DNA Technology, or genetic engineering, for manufacturing human hormones (Paul Berg)
• Microbial genetics and molecular biology
• Hans A. Kreb (1953): Developed chemical steps of the Kreb cycle in carbohydrate metabolism
• James Watson and Francis Crick: Identified the physical structure of DNA as helical, allowing for replication

Developments of the 1940s-1960s

• Electron microscope perfected
• Cultivation methods introduced for viruses
• Rapid knowledge development of viruses
• Vaccines developed for polio, measles, mumps, and rubella

Microorganisms

• Microorganisms are small, living things that can only be visible with a microscope
• They are found everywhere on Earth
• They include bacteria, archaea, fungi, unicellular algae, protozoa, and viruses
• Play important roles in numerous industries like drugs, food, bioremediation, genetic engineering, and gene therapy

Significance of Microorganisms

• Important for maintaining the balance of chemical elements in the natural environment
• Break down remains of living organisms
• Recycle carbon, nitrogen, sulfur, phosphorus, and other elements
• Some species cause infectious diseases, overwhelm body systems, and produce powerful toxins
• Viruses cause damage by replicating within tissue cells, leading to tissue degeneration

Types of Microorganisms: Bacteria

• Unicellular organisms whose genetic material is not enclosed in a nuclear membrane (prokaryotic)
• Cells have rodlike (Bacillus), spherical or ovoid (Coccus), or spiral (corkscrew or curved) shapes
• Form pairs, chains, clusters, or other groupings
• Enclosed in cell walls made of peptidoglycan
• Reproduce by binary fission
• Most use organic material from dead or living organisms; some make their own food or derive it from inorganic material
• Have flagella
• Exist in most environments on Earth, with temperature ranges from 0°C to 100°C in oxygen-rich or oxygen-free conditions

Types of Microorganisms: Archaea

• Often found in extreme environments
• Include methanogens, halophiles, and thermophiles
• Prokaryotic cells do not have peptidoglycan in their cell walls

Types of Microorganisms: Fungi

• Eukaryotic organisms, either unicellular or multicellular
• True fungi have cell walls made of chitin
• Large multicellular fungi include mushrooms
• Unicellular fungi: Yeasts, slightly larger than bacteria, used in alcoholic fermentations and bread making, some can be pathogenic (e.g., Candida albicans)
• Molds: Filamentous, branched fungi that use spores for reproduction
• Prefer acidic environments and live at room temperature under oxygen-rich conditions
• Are most typical fungi: molds
• Reproduce sexually or asexually
• Obtain nourishment by absorbing organic material from their environment

Types of Microorganisms: Protozoa

• Unicellular, eukaryotic microbes
• Move by pseudopods, flagella, or cilia
• Have a variety of shapes
• Live as free entities or parasites
• Reproduce sexually or asexually
• Exist in an infinite variety of shapes because they have no cell walls
• Cause human diseases like malaria, sleeping sickness, dysentery, and toxoplasmosis, slime molds, amoebas, flagellates, and ciliates

Types of Microorganisms: Algae

• Photosynthetic eukaryotes (plant-like organisms)
• A variety of shapes
• Reproduce sexually or asexually
• Abundant in fresh water, salt water, soil, and in association with plants
• Many algae have cellulose/cell walls
• The diatoms and dinoflagellates inhabit the oceans and are foundations of marine food chains

Types of Microorganisms: Viruses

• Ultramicroscopic genetic material (DNA or RNA) enclosed in a protein shell (sometimes with a membrane envelope)
• Have no metabolism, making it difficult to use drugs to interfere with their structures or activities
• Multiply in living cells by using the cells' chemical machinery, destroying the cells through replication
• Acellular

Types of Microorganisms: Multi-cellular Animal Parasites

• Not strictly microorganisms, but are of medical importance
• Include helminthes
• Some stages of their life are microscopic

Significance of Microorganisms

• Modern microbiology reaches into many fields of human endeavor
• Microbes and Human Welfare
• Indigenous microflora
• Opportunistic pathogens
• Decomposers
• Development of pharmaceutical products, the use of quality-control methods in food and dairy product production as well as other fermented foods.
• Control of disease-causing microorganisms in consumable waters and the industrial applications of microorganisms
• Use to produce vitamins, amino acids, enzymes, and growth supplements
• Biotechnology is a major area of applied microbiology, in which microorganisms produce pharmaceuticals
• Bacteria can be reengineered to increase plant resistance to insects for pest control, and also used in sewage treatment
• Bioremediation utilize some bateria to clean the environment, and some perform recycling of vital elements, such as fixing atmospheric nitrogen, and carbon and oxygen cycles

Microbes and Human Disease

• Cause spoilage of food
• Emerging infectious diseases: mad cow disease (1988), bloody diarrhea (E.coli O157:H7, 1982),Ebola Hemorrhagic fever; Streptococcus-IGAS; SARS; AIDS; DENGUE fever

The Spectrum of Microbiology

• Taxonomy: The science of classifying living organisms to show relationships and differences
  • Provides a common reference for identifying organisms
  • Provides a universal language of communication among scientist
  • Displays the unity and diversity among living things, including microorganisms.
• Early taxonomist: Carolus Linnaeus, who classified all known plants and animals and set rules for nomenclature
• Organisms placed in taxonomic categories (taxon) to show degrees of similarity
• Systematics or phylogeny: The study of the evolutionary history of a group of organisms

Classification Schemes

• Species: For organisms such as animals and plants, a species is defined as a population of individuals that breed among themselves. For microorganisms, a species is defined as a group of organisms that are 70 percent similar from a biochemical standpoint.
• Genus: Grouping of various species in the classification, for e.g. species Shigella boydii and Shigella flexneriare are in the genus Shigella
• Family: Grouping of various genera due to similarities
• Order: Placement of various families
• Class: Grouping of orders
• Phylum or division: Categorizing multiple classes
• Kingdom: The broadest classification entry

Historical Classifications

• Aristotle: Classified as plant and animal
• 1857 Carl von Nageli: proposed classifying bacteria and fungi in the plant kingdom
• 1866 Ernst Haeckel: Kingdom Protista proposed, included bacteria, protozoa, algae, and fungi
• 1937 Edward Chatton coined the term prokaryote for cells that do not have a nucleus
• 1961 Roger Stanier defines prokaryote as cells with no nuclear membrane
• 1959 Fungi were placed in their own kingdom
• 1968 Robert Murray proposed the kingdom prokaryote
• 1969 Robert H. Whittaker founded: 5-kingdom system where prokaryotes placed into Kingdom Prokaryote or Monera

Rules For Binomial Nomenclature

• The first letter of the genus name is capitalized, the rest of the genus and the species are lowercase
• The full binomial name is either italicized or underlined
• The genus name can be shortened to its first letter, for example, Escherichia coli can be abbreviated as E. coli
• Species criteria and broader classification use morphology, structure, cell, biochemical, genetic, and immunological properties.

Whittaker's Five Kingdom Classification

• Devised by Robert Whittaker
• Includes:
• Kingdom Plantae - multicellular, autotrophic, eukaryotic
• Kingdom Animalia - multicellular, heterotrophic eukaryotes that do not have cell walls or plastids
• Kingdom Fungi - eukaryotic Thallophytes, soft bodied, non-photosynthetic, lacking chloroplasts, saprophytic or parasitic.
• Kingdom Protista - mostly unicellular eukaryotes, can be colonial, and have animal-like and plant-like characteristics.
• Kingdom Monera - unicellular bacteria with prokaryotic cells.

The Three Domains

• 1978, Carl R. Woese proposed above kingdom, called domain: Bacteria, Archaea, and Eukarya
• Basis: The ribosomal RNA and its nucleotides among the three domains are distinct from each other, in addition to membrane lipid structure, transfer RNA molecules, and antibiotic sensitivity

Evolution

• Oldest known fossils: prokaryotes that lived more than 3.5 billion years ago
• Eukaryotic cells evolved more recently (~1.4 billion years ago)
• Endosymbiotic theory: Eukaryotic cells derived from prokaryotes living inside one another

Evolution of Eukaryotes - Endosymbiotic Theory

The Three Domains

Bacterial Morphology

• Probably the smallest living organisms, ranging in size from 0.15 µm (mycoplasmas) to 2.0 µm (many of the bacteria)
• Cocci, spherical forms
• Bacilli, rodlike forms: Bacilli
• Some bacteria have comma, flexible, wavy, or corkscrew shapes
• Some prokaryotes have a variety of shapes and sizes: Pleomorphic
• Form diplococcus of pair cocci: Streptococcus
• Form streptococcus of chain cocci: Tetracoccus
• Form tetracoccus of four cocci arranged in a cube is: Staphylococcus
• Form staphylococcus of grapelike cluster cocci: Streptobacilli
• Form streptobacilli of long chains shape

Preparing Smear For Staining

• A process of spreading the microorganism over the surface of the glass slide
• Heat fixing kills the organisms and ensures they adhere to the slide, so they can accept the stain

Microscopic Preparations

• Wet mount: Placing the microorganisms directly on the slide to be studied
• Hanging drop technique: Placing the microorganisms between a hollowed slide, coverslip, and droplet of cell to be studied

Cell Wall Composition

• With exception of mycoplasmas, all bacteria have semi rigid cell wall:
• Provides cell shape and protects against osmotic pressure
• Gram-positive bacteria have thick peptidoglycan able to be stained
• Gram-negative bacteria have thin peptidoglycan, outer-membrane containing lipopolysaccharide (endotoxins), lipoproteins
• Archaea have pseudomurein

Structures External to Cell Wall

• Glycocalyx is the gelatinous polymer external to the cell wall.
• Composed of polysaccharide/polypeptide
• Protects from changes in environment
• Tightly bound = Capsule
• Poorly bound = Slime Layer
• Flagella are long filamentous appendages that are involved with motility.
• Classified as monotrichous, amphitrichous, lophotrichous, or peritrichous
• Parts include filament, hook, and basal body
• Fimbriae are for attachment: Pili/pilus
• Pili/pilus is for conjugation, which is for DNA passage between cells
• Axial filament allows spirochetes to rotate in a corkscrew-like fashion

Cell Wall Structures

• Main components include plasma membrane with phospholipids and proteins which have selective permeability, cytoplasm 80% of water
• Ribosomes for cell synthesis composed of 70S: 50S, and 30S: protein and r-RNA: some bind and inhibit
• Nuclear information in nucleic area. nucleoid consist of double stranded DNA and chromosomes, typically arranged in a loop structure

Prokaryotic Inclusions

• Plasmid small loops of DNA
• Metachromatic granules/volutin, an inorganic phosphate
• Polysaccharide granules: glycogen and starch
• Lipid inclusion such as polymer poly-ẞ hydroxybutyric acid
• Sulfur granules: the inclusion of iron oxide

Endospores

• Found in Genera Clostridium and Bacillus
• A resting cell only appearing in harsh conditions that provide a highly durable, dehydrated cell with thick walls
• They have their process in sporulation that include:

1. DNA replication
2. Spore septum that forms to isolated components
3. Plasma that starts to enclose in those materials 4.Protein coat form inside protein and calcium to form forespore

Compartments of Eukaryotic Cell

• Eukaryotic Cell also contains flagella as well as:
• Microtubules of tubulin
• Cell Wall of cellulose, glucan, and mannan
• Nucleus and nucleoli with DNA
• Endoplasmic reticulum
• Ribosomes
• Golgi Complex
• Lysosomes
• Vacuoles and mitochondria
• Peroxisomes and Chloroplast, when present

Contrasts Between Prokaryotic and Eukaryotic Cells

• CHARACTERISTIC*|PROKARYOTIC|EUKARYOTIC
• --|---|--- Size of cell|Typically 0.2-2.0 μm in diameter|Typically 10-100 μm in diameter Nucleus|No nuclear membrane or nucleoli (nucleoid)|True nucleus, which consists of both nuclear membrane and nucleoli Membrane-enclosed organelles|Absent|Present
• CELL WALL*|PROKARYOTIC CELL|EUKARYOTIC CELL
• CELL WALL*|Presents as capsule or slime layer|Present only in cells that lack a cell wall Cell Wall|Usually chemically complex (peptidoglycan and polypeptide)|When present, it is made of cellulose /glucan and mannan and is chemically simple Plasma Membrane|Phospholipids, protein, without carbohydrates and sterols|Phospholipids, protein, carbohydrates which serves as protein Cytoplasm|Lacks a cytoskeleton and cytoplasmic streaming Chromosomes (DNA)|Single circular chromosome that lacks histones Chromosome (DNA)|Multiple linear chromosomes that has histones Cell Division|Binary fission|Mitosis Sexual Combination|Lacks meiosis transfer of DNA fragments only (conjugation)|Involves meiosis

Basic Microbiology Equipment

• Incubator for microorganisms, such as bacteria and fungi
• Petri dish under controlled conditions
• Oven for drying glassware as a means of dry sterilization
• Petri dish/ petri plates used to culture microorganisms in solid media and test tubes that contain aqueous media
• Erlenmeyer flask to heat test tubes, or hot plates with magnetic function
• Water steam and batch is used to heat liquids, use autoclave to sterilize equipment
• Laminar flow hood is a close chamber to reduce contaminations
• Digital Weighing Balance an electrical scale that use for weighing solids.
• Glass Slide and coverslip used to observe microscopic specimen
• Use micropipettes and pipette to measure liquids.
• The Wash bottle is contains liquids used to add in small qualities used to wash in slide and also staining
• Inoculating loops and needles used for microorganism transfer
• Graduated cylinder is used to measure quantity of liquids measure in chemical reactions
• An alcohol lamb is used to sterilize Inoculating loops and needles in preparation of slide preparation. Microscope used to observe objects by their minute objects.