MIP300

vegetative cell

A cell that has not formed spored or other resting stages

virulence factor

Ability to infect and cause disease

The exterior of the cells have what kind of charge; anionic or cationic?

Anionic

pili

Appendages that allow bacteria to attach to each other and to transfer DNA

What does a Gram stain tell me?

Reveals gram reaction of bacteria Gram positive (+) = PURPLE Gram negative (-) = PINK/RED Reveals morphology of bacteria Bacillus (rods), Cocci ( circular), or Coccobacillus Pairs, clusters, chains (GP pic)

lactose-fermenting bacteria

These bacteria grow pink colonies on MacConkey's agar. Examples include Citrobacter, Klebsiella, E. coli, Enterobacter, and Serratia. Lactose is KEE. Test with MacConKEE'S agar.

Why do you decolorize for 5 - 10 seconds only?

because too long would destroy the peptidoglycan layer in G+ cells and they may appear as G-

fimbrae

bristle like fibers on bacteria's surface. Help cells to cling together

Which layer of the endospore is resistant to antibacterials?

coat

smear

driec mixture of bactera and water on a slide

Bacillus subtilis

endospores were greeen

Bright field microsocpy

field is bright and specimen appears opaque

Dark field microscopy

field is dark and specimen appears bright

motility of bacteria

flagella axial filaments fimbrae/pili

Which gram-stained bacteria contain porin proteins in their cell walls?

gram-negative bacteria

Which gram-stained bacteria have a more permeable cell wall?

gram-positive bacteria

Which gram-stained bacteria have a thicker peptidoglycan layer?

gram-positive bacteria

Which gram-stained bacteria possess teichoic acids in their cell walls?

gram-positive bacteria

Gram negative cells peptidoglycan

have a thin layer

Storage granules that store carbon and energy.

inclusion bodies

Staphylococcus food poisoning

is a gastrointestinal illness caused by eating foods contaminated with toxins produced by Staphylococcus aureus

What is the purpose of iodine in gram staining

it is the mordant or fixative for the cv. it is insoluble

List 3 components that describe the chemical makeup of gram-negative cell walls.

lipopolysaccharide lipoprotein peptidoglycan

halotolerant

salt-tolerant Staphylococcus aureus

safranin

secondary stain which stains peptidoglycan pink for both gram negative and gram positive

MacConkey agar

selective medium which contains crystal violet and bile salts to inhibit the growth of gram-positive microorganisms while allowing most GRAM-NEGATIVE bacteria to grow *lactose-fermenting bacteria produce PINK to RED colonies; non-lactose fermenters remain colorless

Layer of glycocalyx that allows bacterial attachment to prosthetic plastic devices.

slime layer

Name the 2 main parts of the glycocalyx.

slime layer: poor organization, weak attachment to cell wall. capsule: organized, adheres to cell wall

steps of gram staining procedure in order

smear and heat fix 1. apply primary stain (crystal violet) 2. apply mordant (grams iodine) 3. decolorize with ethyl alcohol 4. counterstain with safranin

identify steps of acid fast staining procedure in order

smear and heat fix 1. carbolfuchsin 2. rinse h2o 3. decolorize acid alcohol 4. rinse h2o 5. counterstain methylene blue 6. rinse h2o

Micrococcus luteus

spheres

differential stain

stains specific morphological structures - cell wall and cell membrane

Serratia marcescens

*This microbe forms gram-negative short rods Gram-Negative Bacilli, Gram negative motile rod, lives in soil and water. commonly found in bathrooms and is pink and slimy in appearance. Often infects aids patients, burn victims, and is resistant to antibiotics.

Klebsiella pneumoniae

...

What stain is used for a negative stain?

...

How many major layers make up the cell walls of gram-positive bacteria? gram-negative bacteria?

1 2

with acid-fast staining, identify appearance of cell at each step based on cell wall type

1. acid fast red and non acid fast red 2. acid fast red and non acid fast colorless 3. acid fast red and non acid fast blue

with gram stain identify appearance of cell at each step based on cell-wall type

1. both purple 2. both purple 3 gram neg colorless, gram pos purple 4. gram neg pink or red an gram pos purple

purpose of each step in acid fast stain

1. carbolfuchsin penetrates cell wall 2. acid alcohol strips stain from all non acid fast bacteria 3. methylene blue stains non acid fast bacteria blue

identify steps of the spore staining procedure

1. papertowel on stain and place over steam 2. flood paper with malachite green 3. decolorize with distilled water 4 counterstain with safranin

with the spore staining, identify purpose of each step

1. papertowel reduces evaporation of smear 2. malachite green stains endspore 3. distilled water decolorizes 4 safranin colors resting cell

purpose of each step of gram staining

1.Primary stain (crystal violet) will stain all bacteria purple 2. Mordant (grams iodine) will combine inside cell with crystal violet and form insoluble crystal violet iodine complex in gram positive cells 3. decolorizing agent (ethyl alcohol) washes out primary stain of other bacteria except gram positive 4. counterstain (safranin) stains bacteria again. gram negative wll be pink or red and gram positive still purple

It is best to use cultures for Gram staining within what time period?

24 hours

What does Gm-/Gm+ do help us determine?

Appropriate antibiotic treatments

Which bacteria has a glycocalyx comprised of polypeptides?

Bacillus anthraces

Which 2 bacterial species produce endospores?

Bacillus spp. Clostridium spp.

Stalked

Bacterial strains of this shape can be engineered to efficiently take up toxins present at low concentrations in water sources and are being researched for bioremediation applications.

What kind of dye is Crystal Violet? Cationic or Anionic?

Cationic

Cocci

Cocci are also broken down into more descriptive terms - diplo (pairs), staphylo (clusters), and strepto (chains)

Sarcina

Coccus arrangement arising from division in four planes.

Diplococcus

Coccus arrangement arising from division in one plane.

Tetrad

Coccus arrangement arising from division in two planes.

What does CFU stand for and what does it mean?

Colony Forming Units; the number of colonies grown during incubation

What did we use as the primary stain?

Crystal Violet

Cell color results

Crystal Violet (primary stain) Gram's Iodine (mordant) Acetone-alcohol (decolorizer) Safranin (counterstain)

Name the four staining reagents.

Crystal Violet, Iodine, Ethanol, Safranin

serial dilution

Dilution of a substance several times by the same amount each time

Bacillus

Electron Micrograph

Diplococcus

Electron Micrograph

Spirochete

Electron Micrograph

Vibrio

Electron Micrograph

Highly dehydrated resting cells produced in environmental stress.

Endospores

What did we use as the decolorizing agent?

Ethanol

Food at risk for food poisoning by Staphlyococcus?

Foods at highest risk of contamination with Staphylococcus aureus and subsequent toxin production are those that are made by hand and require no cooking. Some examples of foods that have caused staphylococcal food poisoning are sliced meat, puddings, some pastries and sandwiches.

What does the Crystal Violet stain do within the cell wall?

Forms a purple complex

Gram stain for streptococcus pneumoniae would be

Gram +

Escherchia coli

Gram negative Pink Flagella Rods

Bacillus

Gram negative Pink rods

Staphylococcus epidermidis

Gram negative spheres

Neisseria sicca

Gram negative spheres or cocci

Bacillus cereus

Gram positive

Staphylococcus arrangement of cocci

Gram positive Purple spheres

Corynebacterium xerosis

Gram positive rods

What are the two groups of bacteria and their colors?

Gram-positive and Gram-negative bacteria; purple and pink, respectively

Why are gram-positive bacteria more susceptible to penicillin?

Gram-positive bacteria possess more target sites because over 90% of their tetrapeptides are cross-linked

Who invented the gram stain?

Hans Christian Gram

Why is gram staining important?

Identifies unknown bacteria

What did we use as the mordant?

Iodine

What is the purpose of the mordant?

Iodine complexes the crystal violet to the peptidoglycan layer

What is the purpose of the mordant, Iodine?

Keeps the Crystal Violet from being removed from the Gram-positive bacteria

What is the 3rd amino acid of the cell wall tetrapeptide for gram-positive bacteria? gram-negative bacteria?

L-Lysine meso Diaminopimetic acid

Gm- cell walls have a higher _____ content than Gm+ cell walls?

Lipid

Gram-positive cell walls contain only one...?

Membrane

Gram-negative cell walls contain two...? Which are? What's in the middle?

Membranes; inner and outer; peptidoglycan

What are the bacterial equivalents of histones?

Mg2+ polyamines

What 2 genera contain acid-fast bacteria?

Mycobacterium Nocardia

Name the 2 alternating polysaccharide chains that make up the cell wall.

NAG (N-acetylglucosamine) NAM (N-acetylmuramic acid)

What does the Decolorizing Solution, Ethanol, do to Gram-positive bacteria?

Nothing, because of the thick cell walls

The area in the prokaryote cytoplasm where DNA aggregates.

Nucleoid

Observe the Stained Smear

Observe morphology Rod, coccus, or coccobacillus Growth pattern (pairs, clusters, chains)

The bacterial cell wall is composed of what? Where is it located?

Peptidoglycan; on top of the plasma membrane

Circular extrachromosomal DNA that are self-replicating.

Plasmids

What did we use as the counter-stain?

Safranin

Stalked

Shape improves bacteria's ability to take up molecules from its environment.

What are the two staining techniques?

Simple and differential

Which bacteria possess endoflagella?

Spirochetes (Leptospira sp.)

Why are Staphylococcus toxic?

Staphylococcus is salt tolerant and can grow in salty foods like ham. As the germ multiplies in food, it produces toxins that can cause illness. Staphylococcal toxins are resistant to heat and cannot be destroyed by cooking.

Bacteria react different based on what on their cells?

Surface composition

What does it mean to aliquot something?

Take a smaller volume of a sample

Why Gram stain?

The Gram stain reaction and the bacterial morphology provide clues to the identity of the bacteria and are used to determine what tests are done to confirm this identity (GN pic)

What is the purpose of safrin in gram staining

The alcohol removes the cell membrane from gram negative so the safrinin dies it pink or red and the gram postive will remain puple.

Square

The bacteria are the most numerous type of cell found in salt lakes and are believed to be responsible for their characteristic red colour.

What happens to the cell wall if they are old or are stressed by heat or cold?

The cell walls become thin

Why do gram-negative bacteria stain red/pink?

The crystal violet and iodine sit on the surface of the cytoplasmic membrane. The decolorizer strips out the crystal violet-iodine complex because the peptidoglycan layer is so thin. The counter-stain can then sit on the cytoplasmic membrane. 7

Explain why gram-positive bacteria stain purple.

The crystal violet passes through the peptidoglycan layer because it is hydrated. It sits above the cytoplasmic membrane; it does not enter the internal contents of the cell. The iodine also passes through the peptidoglycan layer and forms a complex with the crystal violet. The organic solvent dehydrates the peptidoglycan, causing it to shrink and trap the crystal violet-iodine complex. The safrinin cannot pass through the peptidoglycan and does not reach the cytoplasmic membrane. 4

How is Staphylococcus food poisoning caused?

The most common way for food to be contaminated with Staphylococcus is through contact with food workers who carry the bacteria or through contaminated milk and cheeses.

Square

Though the discovery was made decades ago, scientists found it extremely difficult to grow these bacteria in a lab to be able to study them. In 2004 they succeeded!

What is the purpose of Safranin to Gram-negative bacteria?

Turns it pink/red

What does the Decolorizing Solution, Ethanol, do to Gram-negative bacteria? Why?

Washes out the purple complex. Cell walls are too thin

Endospore Stain

a dye is forced by heat into resistant survival cells called spores and endospores. it is designed to distinguish between spores and the cells that make them - the so called vegetative cells. -green

Staphylococcus

a usually pathogenic bacterium that occurs in grapelike clusters of cocci

motility

ability to move spontaneously

What is the purpose of a negative stain?

allows the shape or morphology to be seen because they appear white or clear with a dark background

Capsule stain

an example of a negative stain that colors the background, allowing the capsule to stand out as a halo around an organism

Extensive invaginations of cytoplasmic membrane in bacteria.

mesosomes

Which acids give acid-fast bacteria a waxy appearance?

mycolic acids

Name 3 of 5 components of the periplasmic space.

nutrient transport proteins nutrient acquisition enzymes (proteases) detoxifying enzymes (beta-lactamases) membrane derived oligosaccharides (MDO) osmoprotectants

What is the principle component of the cell wall?

peptidoglycan

Endoflagella are enclosed in the _____.

periplasmic space

What is the name of the space between the inner and outer membranes of a gram-negative bacteria cell wall?

periplasmic space

Mesosomes are mainly seen in gram-_____ bacteria.

positive

heat fixing

proceess of affixing bacteria in a smear to a glass slide in preparation for staining

what is the purpose of heat fixing a slide

to adhere the bacteria to the slide.

axial filaments

type of flagella that originate from both ends of the cell and wrap around the cell body

Bacterial cells capable of growth.

vegetative bacterial cells

Rhodospirillum rubeum

vibrio

can you tell e. coli from staph a.

yes, E. coli is G- and Staph a is G+