Quest 1

Agar

A gelatinous substance derived from red algae, commonly used as a culture medium in microbiology to grow bacteria and fungi.

Archae

A group of organisms often linked to prokaryotes. Have no defined nucleus for DNA.

Candida albicans

fungal yeast

Dictostelium (Slime Mold)

Eukaryotic. Cells can clump together and for 1-2mm “fruiting bodies”

Escherichia coli

A rod shaped gram-negative bacteria, known to stain pink in gram stains. Contains flagella (motile), and is a facultative anaerobe.

Lymphocyte

White blood cell that produces antibodies or t-cells

Macrophage

White blood cell that kills microorganisms.

mm, μm, nm

1mm = 1000 μm,

1 μm = 1000nm

Prokaryote

Single-celled organism lacking a membrane-bound nucleus and membrane-bound organelles. Do contain ribosomes.

Saccharomyces cerevisiae

Baker’s yeast. This is a eukaryotic organism that is a facultative anaerobe.

Staphylococcus epidermidis

A circular (coccus), gram-positive bacteria that colors purple in a gram stain. Immotile.

Acid-Fast Stain

Used to visualize mycobacteria.

1. Stain w/ Fuschin (pink, lipid-soluble) over boiling water. Heat drives through lipids into the cell wall. Fuschin binds to mycolic acid(extremely hydrophobic).

2. Cool, then rinse with acid-alcohol (A-F negative lose color)

3. Counterstain A-F negative cells w/ methylene blue (no heat, don’t want to push color into A-F + cells)

Cell Wall

Composed of hydrophilic peptidoglycans in bacteria. Creates rigid structure to resist osmotic pressure.

Resolution

Ability to distinguish between two objects. Function of lens and wavelength. Limit of resolution with 100x objective = 0.2 μm

Contrast

Ability to differentiate between cells and the medium. Use dyes

Crystal Violet

Cationic dye used for staining. Binds to DNA. Forms CV-I clumped complexes when introduced to iodine. CV-I remains in G+ cells (can’t escape pores in peptidoglycan), washes out in ethanol in G- cells.

Plasma/Cytoplasmic Membrane

Phospholipid bilayer. Hydrophilic heads, hydrophobic tails.

DAPI

Fluorescent dye that binds to DNA double helix. Visualized via fluorescence microscopy

Differential Staining

Procedure for staining different bacteria to be different colors because of their chemical properties.

Electron Microscopy

Illuminate sample with electrons (shorter wavelength), allows better resolution. See smaller. High energy, so vaporizes cell. must be coated in heavy metals.

Fluorescence Microscopy

Illuminate with known wavelength of light, measure wavelength emitted by sample (different)

Fuschin

Pink, lipid-soluble dye

Immersion Oil

Used bc it has about the same refractive index as glass. Will prevent light from reflecting and refracting away from lens, allowing full resolution.

Glycosyltransferase

Enzyme that catalyzes glycosidic bond formation btwn 1,4 Carbons on sugar ring. Forms sugar backbone of peptidoglycan layer.

Gram Stain

Classic differential stain allowing one to observe the cell wall / membrane characteristics of the cells, and differentiate two bacteria based on their gram characteristics.

1. Stain w/ Crystal Violet

2. Add Iodine (clumps form CV-I Complexes)

3. Ethanol wash (CV-I stays G+, leaves G-)

4. Safranin counterstain (both stain, G+ covered by CV so still purple, G- now pink)

Gram-negative Bacteria

Thinner peptidoglycan layer, encompassed by a second plasma membrane known as the outer membrane. The area in between is known as the periplasm. Outer membrane contains LPS (lipopolysaccharide). Stain Pink. ex. E.coli

Gram-positive Bacteria

Thick peptidoglycan layer (cell wall). Only one membrane. Outside contains teichoic acids and lipoteichoic acids. Stain Purple. ex. S. epidermidis

Isoniazid

Antibiotic that inhibits mycolic acid synthesis. Necessary for cell survival. Works for mycobacterium (ex. tuberculosis)

Lipopolysaccharide

Attach to and branch outward from Gram-negative outer membranes. Protects from antibiotics.

Lysozyme

Enzymes that break trans-peptide bonds (peptide crosslinks) in peptidoglycan. Weakens cell walls creating susceptibility to osmotic pressure and cell lysis.

Methylene Blue

Common cationic dye.

Mycobacterium genus

Genus of bacteria with mycolic acid in their cell walls (ex. tuberculosis, leprosy). Capsule cell wall covered in polysaccharide, so most dyes don’t penetrate. Need acid-fast stain.

Mycolic Acid

Present in cell walls of Mycobacterium. Extremely hydrophobic. Critical to cell function. Vulnerable to isoniazid drugs which inhibit their synthesis.

Osmotic Pressure

Water flow towards area with most solutes. Hypertonic solution: more solute less water

Hypotonic solution: less solute more water

Outer Membrane

Present in Gram-negative bacteria, creating periplasm between inner and outer membranes. Help provide area for proton gradient to build at end of ETC for ATP Synthase. Also provide extra layer of selective permeability.

Penicillin

Inhibits transpeptidase activity, preventing the peptidoglycan layer from growing as the cell grows.

Peptidoglycan

Polymer of sugars and amino acids that form a cell wall of rigidity around bacterial cells due to glycosidic bonds on the sugar backbone, and peptide crosslinks (N bonds) between layers. Monomers an the polymer as a whole are quite hydrophilic. Bonds formed from glycosyltransferase and transpeptidase.

Safranin

Pink dye used in final step of gram-stain.

Teichoic Acid / Lipoteichoic Acid

Found in cell walls of gram-positive bacteria.

Transpeptidase

Enzyme catalyzing peptide crosslinks between layers of peptidoglycan. Accounts for rigidity of peptidoglycan in addition to glycosidic linkages from glycosyltransferase.

ABC Transporter

ATP Binding Cassette Transporter: ATP hydrolysis (-Pi) needed to facilitate transport across a membrane through this transporter protein. Active / facilitated transport.

Bacillus genus

Gram-positive, rod-shaped bacteria. SPORULATE.

Capsule

Polysaccharide coating around live bacteria, serving as protection from host’s immune system. Harder for immune systems to select against polysaccharides.

Chemotaxis

Movement towards or away from some molecule.

Clostridium genus

Sporulates.

Cytoplasmic/Plasma Membrane Function

Permeability barrier, selective for size and hydrophobicity. Protein anchor (transporter etc). Energy conservation (no mitochondria, so energy production occurs in cytoplasmic membrane.

Flagella / Flagellum

Structures external to cell wall that allow movement. Move toward resources, away from toxins / competition. Spins, moves like corkscrew. Flagellin is main structure providing motion.

Flagellin

Main flagellum structure providing motion.

Germinant Receptor

In cytoplasmic membrane. Bind specific target proteins and change shape. Others bind to amino acids.

Germination

Spore becomes living, growing, vegetative cell when conditions become favorable again. Nutrient (amino acid ie L-Proline availability).

Lac Permease

Protein responsible for transport of lactose into cells. Symporter that takes advantage of proton gradient.

Periplasm

Space between the outer and inner membranes of gram-negative bacteria. Helps gram negative bacteria in energy production by providing a space for a proton gradient to build.

Run/Tumble

Motion of random (tumble) followed by unidirectional (run) movement exhibited by cells with multiple flagella, and used for the processes of chemotaxis and phototaxis.

CCW: Bundle together, “run”

CW: Move all over, “tumble”

Spore Coat

Protective layer of proteins that surrounds endospores, protects them from harsh environmental conditions, and prevents dye from penetrating cells.

Endospore

Differentiated cells that can survive in harsher conditions. Have many more protective layers that provide resistance. “storage form” of bacteria. Mostly form from Gram-positive bacteria.

Sporulation (+ signals)

Process of forming spores (differentiated cells) from mostly Gram-positive cells (Bacillus, Clostridium).

signals: low moisture, low food/nutrient, high radiation, high heat, high oxygen concentration (more O2 signals dead host, less nutrients will become available)

Transporter Protein (Anti vs Symporter)

Sit in cytoplasmic membrane and allow transport of larger molecules into/out of the cell.

Uniporter: 1 target molecule

Antiporter: 1 in, 1 out

Symporter: 1 in alongside proton gradient

bacillus

rod-shaped

Etiology

cause of a disease or condition

Koch’s Postulates

set of 4 criteria that a microorganism is the causal agent of a disease.

1. constant association: always present in diseased, absent in healthy.

2. isolation and culture: must be isolated from diseased host in pure culture

3. re-infection: causes same disease when inoculated from pure culture

4. re-isolation: microorganism isolated from second infection must match original one

Leprosy Bacterium

Mycobacterium, very similar to tuberculosis bacterium.

Tuberculosis

Disease caused my mycobacterium tuberculosis. Infectious disease that causes growths (tubercles) in the lungs

Complex Media

Cannot write out a comprehensive lost of parts, but has enough for growth. “To grow non-fastidious organisms). ex. Nutrient agar or broth (beef/yeast, peptone, nacl, agar), Tryptic soy agar or broth (casein and soy peptone, nacl, agar), YPD agar or broth (yeast extract, peptone, dextrose (glucose), agar)

Defined Media

Can write exact concentrations and chemical formula of all components

Endergonic

Non-spontaneous reactions that require energy input to run. Must be coupled with exergonic reactions.

Enriched Media

Media having components that promote the growth of particular microbes.

Eosin Methylene Blue (EMB)

Inhibits Gram-positive growth. Lactose fermenting bacteria give a green oily sheen. EMB is both selective and differential.

Exergonic

Spontaneous, reactions that release energy, can occur without external input. Couple to allow endergonic reactions to run.

Heterotroph

Organisms that obtain energy from organic molecules in their environments

Mueller-Hinton Agar

Used for official testing of antibiotic sensitivity (arbitrarily chosen). Beef extract, casein hydrolysate, starch, NaCl, Agar. Protein-free medium.

Nutrients

Carbon source to build molecules, energy source (ie sugar), nitrogen source (peptides, NH/ammonia), vitamins (small, aid enzyme function), salt

Phototrophic

Obtain energy from sunlight

Redox

Involve transfer of electrons, or electrons and protons. Loss of electrons = oxidation. Gain of electrons = reduction. Principles behind electron transport chain, obtaining energy from movement of electrons from sugar to terminal electron acceptor.

Reduction Potential

Potential for an oxidized form to become reduced.

Selective Media

Inhibits growth of some organisms

Tryptic Soy Agar

Medium that contains casein (milk protein) and soy peptone, NaCl, and agar

YPD

Medium that contains yeast extract, peptone, dextrose (glucose), and agar.

Acetyl-Coenzyme A

Forms in pyruvate oxidation when CoA accepts the acetyl group from pyruvate. Acetyl-CoA then delivers the acetyl group to begin the citric acid cycle.

Aerobic Respiration

Capturing energy from glucose with oxygen (from air) as the terminal electron acceptor. Releases the most energy, as shown via the redox tower.

Aerobic Respiratory Electron Transport Chain

Glucose → NAD+/NADH electron carriers → flavoprotein → Fe/S proteins → Quinone → Cytochrome C Reductase →Cytochrome C → Cytochrome C Oxidase → oxygen

Anaerobic Respiration

Capturing energy from glucose with nitrate as the terminal electron acceptor. Release less energy than aerobic respiration. Can proceed to Nitrite, or N2 as final product.

ATP Synthase

Protein utilizing a proton motive force to turn ADP and inorganic phosphate to ATP. Protons flow in towards/through membrane via protein, to generate ATP within the cell.

ATP / ADP

Adenosine Tri(Di)phosphate. Hydrolysis of ATP → ADP + Pi releases energy, used to power cell work like growth, transport, reproduction.

Citric Acid Cycle

Begins with Acetyl CoA, and generates CO2, energy source GTP, and electron carriers FADH2 and NADH. Occurs in mitochondria of euks / baker’s yeast, cytoplasm of bacterial cells.

Cytochrome

Electron carrying protein containing iron in the electron transport chain.

Cytochrome C

Electron carrier protein as are other cytochromes. Use oxidase test to determine if present.

FAD / FADH2

Electron carrier molecules. Electrons carried in N-H bonds that form / break as electrons are taken in or delivered.

Fermentation

Respiration pathway with pyruvate or acetaldehyde (yeast) as the terminal electron acceptor. Leads to buildup of lactate. Releases least amount of energy.

Glucose

Sugar, used as energy source by many different forms of bacteria. Captured electrons used for respiration

Glyceraldehyde-3 Phosphate

Midway product of glycolysis. 1 glucose → 2 G3P → 2 pyruvate

Glycolysis

“Sugar breaking”. Occurs in cytoplasm of cells. Prepares pyruvate from glucose to serve in citric acid cycle.

GTP / GDP

Like ATP but with guanine, used to power protein synthesis and other cellular processes.

Lactate

Product of pyruvate reduction in fermentation. Easily forms lactic acid.

NAD+ / NADH

Nicotinamide adenine dinucleotide. Electron carrier molecule. Coenzyme critical to respiration.

Oxidase Test

Tests for presence of cytochrome C in bacteria. Oxidase reagent donates electrons straight to cytochrome c, turns deep purple. If wait long enough, turns purple anyway as it donates electrons to oxygen instead.

Terminal Electron Acceptor

Molecule that accepts electrons at the end of the electron transport chain. Either pyruvate, nitrate, or oxygen.

Oxygen

Terminal electron acceptor in aerobic respiration. Releases the most energy compared to the other terminal electron acceptors.

Pyruvate

Product of glycolysis. Can be oxidized in formation of Acetyl CoA for citric acid cycle (aerobic respiration). Or can be reduced to form lactate (fermentation).

Aerotolerant Anaerobe

Does not use oxygen, but can survive in its presence.

Bromthymol Blue

pH indicator used in O-F medium to indicate respiration. Begins green. Turns blue when peptone is broken down (basic), and yellow in the presence of acid, produced via fermentation / aerobic respiration.