Microbiology Exam #1 (Chapter 1,3,4,5,6) Part 2

Distinguishes or separates two adjacent objects or points from one another

Degree of contrast from the surroundings. Iris diaphragm controls the amount of light entering the condenser

The objective lens forms the initial image of the specimen which is the real image. The virtual image occurs when the initial image is projected up through the microscope body to the plane of the eyepiece to the ocular lens

Longer wavelengths of light offer less resolution than short wavelengths. Oil prevents the scattering of light rays and increases the numerical aperture and resolution

Bright-field, Dark-field, Phase contrast, and Differential interference microscopes use visible light

Fluorescence and Confocal microscopes

An electron microscope uses beams of electrons instead of rays of visible lights like light microscopes do

Scanning tunneling microscope ( 100,000,000x)

Phase contrast: most useful for observing intracellular structures such as bacterial spores, granules, and organelles, as well as locomotor structures of Eukaryotic cells such as cilia
Differential interference: detailed view of unstained, live specimens by manipulating the light. Also adds contrasting colors rot the image and two beams of light rather than a single one
Confocal: delivers sharper image focusing on just a single plane. Captures highly focused view at any level, ranging from the surface to the middle of the cell
We use a Bright field microscope which produces an image brightly illuminated. Used for both live, unstained material, and preserved stained material

Fluid maintains viability and priories a medium for movement. Also provides a true assessment of size, shape, arrangements, color, and motility of cells

Positive charged dye sticks to the cell, gives it color, and is attracted to negatively charged cell walls. Negative charged dye is repelled by negatively charged bacterial cell walls.

- Simple: require a single dye, uncomplicated procedure
- Differential: uses 2 differently colored dyes to contrast cell types, or cell parts. Complex staining technique
- Special: Emphasize certain cell parts that are not revealed with conventional staining procedures

Differential stain

Cell membrane
Ribosomes
Cytoplasm
Bacterial chromosomes in the nucleoid region

Cocci
Spirilla
Bacilli

Amphitrichous

Gram positive

Gram negative

70S

Endospores

Archaea

Determinative Bacteriology

Composed of proteins.
Three parts:
- Basal Body: a stack of rings firmly anchored through the cell wall to the body
- Hook: anchored to the cell by the basal body
- Filament: helical structure, composed of proteins, approximately 20 nanometers in diameter. Inserted into a curved, tubular hook

- Monotrichous polar: single flagellum
- Lophotrichous polar: several tufts emerging from same site
- Amphitrichous polar: flagella at both ends
- Petritichous: flagella randomly dispersed over the entire surface of the cell

Movement in response to chemical signals.

Positive: Run - counterclockwise movement of the flagella, cells swim in a smooth, linear direction toward a stimulus
Negative: Tumble - flagella reverses direction, causing the cell to stop

- Two or more long coiled threads found in spirochetes
- Internal flagellum enclosed between the cell wall and cell membrane
- Impart twisting or flexing motion to the cell

- Small bristle-like fibers sprouting off the surface of certain species of bacteria
- Composition varies, but most contain protein
- Have the inherent tendency to stick to each other and to surfaces
- E. Coli and the gonococcus use fimbriae to adhere to epithelial cells

- Also known as sex pious
- Long, rigid tubular structure made of pilin protein
- Only found in gram negative bacteria
- Used in conjugation, the partial transfer of DNA from one cell to another
- Production of Pili is controlled genetically

Slime layer - Forms loosely around the cell, protects the cell from loss of water and nutrients
Capsule - More tightly bound than the slime later, denser and thicker, formed by pathogenic bacteria, protects bacteria against phagocytic white blood cells
Biofilms - Can infect long-term in dwelling artificial devices such as plastic catheters, intrauterine devices, and metal pacemakers

Peptidoglycan is found in the cell walls of bacteria. It is a unique macromolecule composed of gylcan chains cross linked with short peptide fragments. Also provides a strong but flexible support framework

Gram positive cells have a thick layer of peptidoglycan 20-80nm thick and gram negative cells have a thin layer of peptidoglycan 1-3nm

Cell wall.
The most important species is

- Contains specialized polysaccharides and proteins
- Lipopolysaccharide
- Polysaccharide chains function as antigens and receptors

- The cell membrane is a phospholipid belayer embedded with proteins completely around the cytoplasm
- The cell membrane transports the nutrients into the cell and discharges wastes.
- It is selectively permeable, water and small uncharged molecules diffuse freely, and special carrier mechanisms may exist.
- Secretes discharge of metabolic products into the extra cellular environment

CYTOPLASM: - Gelatinous solution contained by the cell membrane., - Prominent site for the cells biochemical and enzymatic activities, - 70-80% water, complex mixture of sugars, amino acids, and salts, - Also contains chromatin, ribosomes, granules, and fibers that act as the cytoskeleton

BACTERIAL CHROMOSOME: - Typically only one and found in a circular appearance, - Aggregated in a dense area called the nucleiod, - DNA is tightly coiled around protein molecules

PLASMIDS: - Non-essential pieces of DNA, - Separate, double stranded circles of DNA, - Confer positive traits, - Duplicated and passed onto offspring during replication, - Important in genetic engineering

RIBOSOMES: - Made of rRNA and protein, - Dispered throughout the cytoplasm, - Svedberg (S) units: Measurement of the relative size of cell parts through sedimentation during centrifugation, - Bacterial ribosomes: 70S, - Eukaryotic ribosomes: 80S

INCLUSION BODIES: - Storage sites for nutrients during period of abundance, - Single-layered membranes, - Vary in size, number, and content

MICRO-COMPARTMENTS: - Made of protein, - Packed full of enzymes and work together in pathways

CYTOSKELETON: - Long polymers of proteins similar to eukaryotic actin, - Arranged in helical ribbons around the cell just under the cell membrane, - Contributes to cell shape

- Bacterial endospores withstand hostile conditions and facilitate survival, - Stimulus for endospore formation: depletion of nutrients, especially carbon and nitrogen sources], - Can resist hear, drying, freezing, radiation, chemicals, - Constant intruders where sterility and cleanliness are important, - Resist ordinary cleaning methods, - Frequently contaminate cultures and media, - Hospitals must protect against endospores in wounds, - Destruction of endospores important in the food-canning industry, , LIFE CYCLE:, - Germination begins when favorable conditions arise, - Exposure to water and a germination agent, stimulates the formation of hydrolytic enzymes that break down the cortex, - Core rehydrates and takes up nutrients and bacterium grows out of the endospore coats, , PRIMARY DYE:, - Malachite green, , MORDANT:, - Steam, , DECOLORIZER:, - Water, , COUNTERSTAIN:, - Safranin, , COLORS:, - Vegetative cells stain red because they lack affinity for malachite and a cortex, - Endospores stain green because they have an affinity for malachite and a cortex

- Closely related to the Domain Eukarya
- Share ribosomal RNA (rRNA) sequences not found in bacteria
- Protein synthesis and ribosomal subunit structures are similar
- Certain genetic sequences are only found in their rRNA
- Unique method of DNA compaction
- Unique and chemically distinct cell walls
- Most primitive of all life forms
- Closely related to cells that originated 4 billion years ago

BERGEY'S MANUAL:
- Definitive published source for bacterial and archaea classification
- Basis for early classification was phenotypic traits: shape, cultural behavior, and biochemical reactions
BERGEY'S MANUAL OF SYSTEMATIC BACTERIOLOGY:
- Comprehensive view of bacterial and archaeal relatedness
- Five volume set, combines phenotypic information with rRNA sequencing for classification
BERGEY'S MANUAL OF DETERMINATIVE BACTERIOLOGY:
- Based entirely on phenotypic characteristics
- Categorizes bacteria by traits assayed in clinical, teaching, research labs
- Identify bacteria not based on evolutionary backgrounds
- Useful for students of medical microbiology

- Gracilicutes: Gram-negative cell walls, thin-skinned
- Firmicutes: Gram-positive cell walls, thick and strong
- Tenericutes: Lack a cell wall, soft
- Mendosicutes: Archaea

First eukaryotic cells appeared on Earth 2 - 3 billion years ago
- Bacteria and eukaryotes evolved from a precursor called the Last Common Ancestor
• Endosymbiosis
- Large precursor cell engulfed small bacterial cells that lived and reproduced inside the large cell

1. Protozoa
2. Fungi and Algae
3. Helminths
4. Plants and animals are considered unicellular

Ribosomes

The flagella and cilia

- Eukaryotic flagella
• 10x thicker
• Structurally more complex
• Covered by an extension of the cellmembrane
• Long, sheathed cylinder containing regularly spaced hollow microtubules: 9+2 arrangement
• Certain protozoa and algae
- Eukaryotic cilia
• Similar in overall structure to eukaryotic
flagella
• Shorter and more numerous - up to
several thousand in some cells
• Found only in a single group of protozoa and certain animal cells
• Function as feeding and filtering
structures on some cells

Outermost boundary that comes into direct
contact with the environment
• Also called an extracellular matrix
• Composed of polysaccharides
• Appearance
-Network of fibers
-Slime layer
- Capsule

Fungi/Yeast, & Algae.
- Rigid and provide structural support andshape
- Different chemical composition than bacterial
cell walls

Yes, all eukaryotic cells posses a cell membrane.
Typical bilayer of phospholipids embedded with proteins
- Contain sterols
- Selectively permeable barrier
- Sophisticated mechanisms for transporting materials in and waste and other productsout

No, they do not have a nucleus.

Eukaryotic cells do.
• Nucleus: the control center
- Compact sphere, most prominent organelle
- Separated from the cytoplasm by thenuclear envelope
- Nucleolus: Stains more intensely due to its RNAcontent
• Site for ribosomal RNA (rRNA)synthesis

They are both Microscopic series of tunnels used in transport and storage
-Rough endoplasmic reticulum (RER)
• Ribosomes attached to its surface
• Proteins held for packaging and transport

- Smooth endoplasmic reticulum (SER)
• Does not contain ribosomes
• Synthesis and storage of nonproteinmolecules (lipids)

- Site of protein modification and transport
• Consists of a stack of flattened, disc-shape sacs
• Closely connected to the endoplasmic reticulum
• Receives transitional vesicles and forms condensing vesicles

Nucleus, ER, and Golgi apparatus: Nature's Assembly Line (the secretory pathway or transport process)
- Genetic information originates from the nucleus
- Proteins are synthesized on ribosomes and deposited into the RER
- Proteins are then transported to the Golgi apparatus to be modified and packaged into vesicles

Lysosomes
• Originate in the Golgi apparatus and contain a variety of hydrolytic enzymes
• Involved in intracellular digestion of food and protection against invading microorganisms

- Aerobic respiration, also known as cellular respiration
- Supply the bulk of the energy of a cell
- Outer membrane, inner membrane/cristae, matrix, circular DNA, 70S ribosomes (like bacterial ribosomes)
- Divide independently of the cell

- Found in algae and plants
- Capable of converting the energy of sunlight into chemical energy
- Primary producers of all organic nutrients
- Primary producers of oxygen gas
- Outer membrane, inner membrane/thylakoid, stroma, circular DNA, 70S ribosomes

Eukaryotic ribosomes are 80S in size
• Similar in structure to prokaryotic ribosomes but different in size
- Distributed throughout the cell
• Scattered freely in the cytoplasm (80S)
• Attached to RER (80S)
• Inside mitochondria and chloroplasts (70S)

- Functions
• Anchoring organelles
• Moving RNA and vesicles
• Permitting shape changes and movement

- Three main types
• Actin filaments
• Intermediate filaments
• Microtubules

Algae and Protozoa

• Roundworms
- Nematodes
• Flatworms
- Trematodes (flukes) and Cestodes (tapeworms)
• Sometimes large enough to be seen with the naked eye: 1 mm - 25 m in length
• Multicellular animals equipped with organs and organ systems
- Reproductive tract is the most developed
- Primitive digestive, excretory, nervous, and muscular systems
• Majority of helminths derive nutrients and reproduce sexually in the host's body

Latin for poison. A tiny infectious particle, or obligate intracellular parasite. A virus is different from a cell because it can infect every type of cell

Early discoveries
- Pasteur developed a vaccine for rabies
• Coined the term "virus" (Latin for poison)
- Ivanovski and Beijernick: tobacco disease caused by a virus
- Loeffler and Frosch: foot-and-mouth disease caused by a virus
• Filterable virus
- Infectious fluids were passed through filters designed to trap bacteria
• Cell-free filtered fluid remained infectious
- Proved that an agent smaller than bacteria was the cause of disease

• Viral size range
- Ultramicroscopic size
• Smaller than the average bacterium
• Electron microscopes are required to detect them
• Parvoviruses: 20 nm in diameter
• Mimiviruses: 450 nm in length - larger than some small bacteria
• Cylindrical viruses: 800 nm long, but 15 nm in diameter

• Viruses bear no resemblance to cells
• Need only those parts required to invade andcontrol a host cell

Viruses can infect every type of cell
- Bacteria, algae, fungi, protozoa, plants, animals

- Capsid
• Shell surrounds the nucleic acid
• Nucleocapsid: capsid and nucleic acid together

- Naked viruses
• Consists only of a nucleocapsid
- Envelope
• Usually a modified piece of the host cell membrane
• Enveloped virus: nucleocapsid and envelope

- Spikes
• Found on both naked and enveloped viruses
• Project from either the nucleocapsid or envelope
• Allow viruses to dock with their host cells

Viral capsid: the protective outer shell
- Capsomeres
• Identical protein subunits that spontaneously self assemble to form the capsid
- Helical capsid
• Rod-shaped capsomeres that form a continuous helix around the nucleic acid
- Icosahedral capsid
• Three-dimensional, 20-sided figure with 12 evenly spaced corners

- Found in bacteriophages, viruses that infect bacteria
• Capsid head, nucleic acid, collar, sheath, base plate, tail pins, tail fibers
- Have multiple types of proteins
- Take shapes that are not symmetrical

- Composed of the membrane system of the host
- Cell membrane or nuclear membrane
- Regular membrane proteins are replaced with viral proteins

- Informal classification system
• Animal, plant, or bacterial viruses
• DNA or RNA viruses
• Helical or icosahedral
- Formal classification system
• Structure
• Chemical composition
• Genetic makeup
• Virus families - end in viridae
• Virus genera - end in virus

• Multiplication cycle in animal viruses
- Adsorption is when spikes attach the virus to cell
- Penetration is when the virus enters the cell via endocytosis (engulfment) or fusion
- Uncoating the DNA or RNA into the cell
- Synthesis of viral components (nucleic acid, capsomers, spikes) in the nucleus (DNA virus) or cytoplasm (RNA virus)
- Assembly of viral components in nucleus (DNA virus) or cytoplasm (RNA virus)
- Release of virus via budding (enveloped virus) or lysis (naked virus)

- Cytopathic effects (CPEs), • Virus-induced damage to the cell that alters its microscopic appearance, • Cells can become disoriented, undergo major changes in shape or size, or develop intracellular damage, , - Inclusion Bodies, • Compacted masses of viruses or damaged cell organelles in the nucleus or cytoplasm, , - Synctia, • Fusion of multiple host cells into single large cells containing multiple nuclei, , • Persistent infection, - Can last from a few weeks to the remainder of the host's life, , - Provirus: Viral DNA is incorporated into the DNA of the host, , -Chronic latent state: Can reactivate under the influence of various stimuli and cause symptoms, , - Oncogenic (oncogenes: cancer causing genes), • Transformation results in increased growth rate, alterations in chromosomes, changes in cell's surface, molecules, capacity to divide for an infinite period of time, • Oncoviruses: mammalian viruses capable of initiating tumors

Lytic phase or lytic cycle: life cycle of bacteriophage that ends in destruction of the bacterial cell
• Lysogenic cycle: bacteriophage becomes incorporated into the host cell DNA (prophage)
(See book page 156)

In vivo methods: viral cultivation in lab animals
or embryonic bird tissues
In vitro methods: viral cultivation in cell or tissue
culture

• Prions, - Distinct abnormal protein fibrils deposited in brain tissue of affected animals, - Common feature of spongiform encephalopathies, • Implicated in chronic, persistent disease in humans and animals, • Brain tissue removed from affected animals resembles a sponge, • Creutzfeldt-Jakob Disease (CJD), • Affects the central nervous system of humans and causes gradual degeneration and death, • Several animals are victims of similar diseases, , • Satellite viruses, - Dependent on other viruses for replication and contains only genetic material, - Adeno-associated virus (AAV), • Originally thought that it could only infect cells infected with the adenovirus, • Now found to infect cells infected with other viruses or had their DNA disrupted through other means, , • Viroids, - Virus-like agent that parasitizes plants, - About one-tenth the size of an average virus, - Composed only of naked strands of RNA, - Significant pathogens in economically important plants: tomatoes, potatoes, cucumbers, citrus trees, and chrysanthemums

Most common cause of acute infections that do not result in hospitalization
- Colds, chickenpox, influenza, herpes, warts
• Viral infections that only occur in certain regions
- Dengue fever, Rift Valley fever, Yellow fever

No, because:
• Viruses mutate at a rapid rate
• Scientists focus on developing vaccines against viruses since few antiviral drugs are available and antibiotics are ineffective
• Interferon (IFN)
- Naturally occurring human cell product
- Used with some success in treating viral infections