Lab Exam #1(Microbiology

scientific method

1.Observation 2.Hypothesis 3. Experimentation 4. Data collection 5.Analysis and conclusion

Coarse adjustment knob

adjust the height of objective lens

Fine focus adjustment knob

(the distance between the objective lens and the specimen

Substage condenser

brightness and control contrast

Resolution

is wavelength of light and design of condenser

How can resolution be maximized?

Condenser being kept at high can maximize the resolution

parafocal

Specimens to remain in focus when objectives are switched

Cyanobacteria (Anabaena)

o   oxidize hydrogen sulfide to sulfur, found in hot springs and stagnant water

Amoeba

shapeless,amorphous form

Paramecium

cilia for locomotion and water-regulating vacuoles

Euglena

unicellular, eukaryotic

Characteristics of fungi

Cell walls, multicellular molds (eukaryotic-true nucleus , heterotrophic, saprophytic(feeding on dead decaying matter, grows fuzzy on plates)

3 general shapes of bacteria

1. Coccus

2. Bacillus

3. Spirillum

steps for preparing a successful smear

1.     The bacteria must be evenly and thinly dispersed

2.     Let air dry completely

3.     The bacteria needs to be attached to the slide

    heat fix bacteria

Bacteria becomes heat fixed when organisms are killed and can attach them to the slides being used

why we stain bacteria

easier to view when it is stained

the difference between a positive & negative stain

Positive stain adhere to the cell surface and will be colored with the stain

Negative stain determines cell size and arrangement

Simple stain

stain use only one stain and basic/acidic staining

Differential stain

uses more than one stain and examples of this acid-fast staining, endospore stain

Gram positive cell walls

have a thick peptidoglycan cell wall

Gram negative cell walls

have a thin peptidoglycan cell wall

Steps of gram stain-

1.      After performing a smear prep on the slide, saturate the smear with crystal violet for 20 seconds.

2.     Rinse with water completely.   

3.     Saturate the smear with iodine for 1 minute.

4.     Rinse with water completely.   

5.     Decolorize with Gram decolorizer (75% ethanol + 25% acetone) for 10 seconds; if you leave the decolorizer on too long, it will remove the crystal violet out of a gram positive cell as well!!

6.     Rinse with water completely.   

7.     Counterstain with safranin for 1 minute.

8.     Rinse with water completely.   

9.     Carefully blot the slide dry with bibulous paper.

10.  Observe the slide using a light microscope, draw under oil immersion.

Gram negative bacteria

be stained the reddish-pink color of safranin

Gram positive bacteria

be stained the purplish color of crystal violet

Staphylococcus epidermidis

spherical cocci,gram positive

Escherichia coli

rod-shaped-bacillus Gram negative

Bacillus subtilis

(rod shaped-bacillus,gram positive

Mycobacterium smegmatis

bacillus,gram positive

What is the purpose of steam in acid -fast stain?

Loosens mycolic acid and secures carbol fuchsin

name & function of the acid found in cell wall of acid-fast bacteria

Mycolic acid  and it resists the uptake of simple dyes such as the ones in a gram stain.

Acid-fast organism at end of stain

resist de-colorization by acid alcohol

Not acid-fast organism

decolorized by acid alcohol and are counterstained with methylene blue

Mycobacterium smegmatis

bacillus(shape),is acid fast bacteria

Staphylococcus epidermidis

o   spherical cocci(shape), NOT a acid fast bacteria

Two pathogenic acid fast bacteria

1.     Mycobacterium tuberculosis

2.     Mycobacterium leprae

Steps and reagments in Endospore stain

The purpose of the endospore in some bacteria?

Endospore allow bacteria to survive without nutrients for very long periods of time and can resist harsh environmental conditions that are exposure to heat and radiation

·      Under what conditions do endospores form?

Endospores form when a bacteria encounter harsh environmental conditions or nutrient depletion.

Endospore

allow bacteria to survive nutrients for long period of time

Vegetative stage

the organism can return to the metabolically active stage

What is the shape of Bacillus subtilis and does it form endospore?

(rod-shaped) bacterium and doesn’t form endospores

·      Give an example of at least two pathogenic endospore forming bacteria

Clostridium tetani and Clostridium botulinum

Aspetic technique

minimizes the possibility of introducing contamination.; Prevents contamination of microbial cultures

Disinfection

meaning that the number of living microbes particularly those potentially hazardous to humans has been greatly reduced by application of a chemical or chemical mixture

Sterilization

:- (completely free of living microbes

Pure culture

microbes are genetically identical to one another and free of other species or strains

Mixed culture

comprised of two/more distinct species or strains in a shared environment

·      the procedure for performing a broth or slant transfer

Aseptic technique is used for broth or slant transfer

The streak method and why we do this?

Streak method dilutes a population bacteria to an agar surface that separates individual cells. Helps create pure cultures

What do we invert media (plates) for incubation?

·      This helps prevent condensation droplets from falling onto the agar surface.

Selective

o   : allow growth of only certain types of organism

Example: Phenyl ethyl alcohol agar (PEA): selective for gram +, inhibits gram –

Differential

o   which allow growth of closely-related organisms that will appear visibly different on the media due to their growth patterns

Example: Blood agar and TSI

Enriched:

o   Contains additional nutrients to enhance growth Example: blood agar

Phenyl ethyl alcohol plate (PEA)

o   selective for gram + and inhibits gram;-alcohol dissolves outer membrane of gram negative;selective media

Blood agar

contains 5% sheep blood;cultivation of fastidious bacteria such as Streptococcus, by observation of hemolysis;differential media

Triple sugar Iron Agar (TSI)

o   fermentation and production of gas;- fermentation using indicator phenol red; differential media; differentiates gram-enteric

Mac Conkey agar (MAC)

o   contain crystal violet; contain lactose and neutral red selective and differential; differentiates for lactose fermenters

Mannitol salt agar (MSA

o   contain 8-10% NaCl and contain mannitol and phenol red; selective and differential; differentiates for mannitol fermenters

Eosin Methylene Blue agar (EMB)

selective/differential 2 dyes are toxic(methylene blue and eosin); lactose fermentation produces acids; selects for gram – bacteria, differentiates lactose fermentation.