Microbiology Lab Quiz #1

Ubiquitous (describing microbes)

present or existing everywhere

Microorganisms/Microbes

Living creatures that are too small to see with the naked eye

Contamination

unintentional exposure to microbes

inoculation

intentional exposure to microbes

Culture

controlled growth of microbes in a lab

Fomite

object or surface that can harbor and transfer microbes

incubate

develop, grow

Pellicle

bacteria layer on top of incubated broth

scum/oily contact lens appearance

Sediment

bacteria layer at bottom of incubated broth

sand/salt appearance

Turbidity

cloudiness

in broth, turbidity indicates growth of microbes

Fastidious

picky eater

Morphology

shape

Descriptive texture terms (of bacteria)

moist

dry

dull

shiny

mucoid

Descriptive edge terms (of bacteria)

smooth

wavy

rough

curled

filamentous (root-like)

Motility

ability to move

bacterial movement can be tracked with TCC deeps (semi-solid media)

What is the classification for SabDex media?

Selective media

What is the purpose of SabDex media?

Grow certain microbes and prevents others

What are some unique traits of SabDex media?

Low pH

High sugar

Grows fungi

What is the classification for NA media?

Standard media

What is the purpose for NA media?

Basic nutrients to grow microbes

What are the unique traits for NA media?

None

What is the classification for TSA media?

Enriched media

What is the purpose for TSA media?

Extra nutrients to help fastidious bacteria grow

What are some unique traits of TSA media?

Good at growing bacteria from human skin

3 general arrangements of bacteria

Bacilli

Cocci

Spirrila

Types of bacilli

bacillus (1 rod shape)

diplobacilli (2 rods connected end to end)

streptobacilli (3+ rods connected end to end)

palisades (rods leaning on each other side by side, like books)

streptobacilli

palisade bacilli

Types of cocci

coccus (single sphere)

diplococci (2 spheres connected)

streptococci (3+ spheres connected)

staphylococci (grape cluster)

tetrad (4 spheres in square formation)

streptococci

staphylococci

tetrad cocci

Types of spirilla

vibrio (comma)

spirillum

spirochete

vibrio

spirillium

spirochete

milliliter (mL)

1/1,000 L

microliter (µL)

1/1,000,000 L

P100 micropipette

10-100 µL

... 0.01-0.1 mL

P1000 micropipette

100-1,000 µL

...0.1-1 mL

Agar

Gelatin-type material derived from seaweed

Which physical form of media has 1.5% (the most) agar?

Solid media (slants and plates)

Which physical form of media has 0% (the least) agar?

Liquid media (broth)

Which physical form of media has 0.5-0.7% agar?

Semi-solid media (deep)

What is semi-solid media used for?

Test for motility or O2 requirement

Needle used to stab line of bacteria down center of media

What is solid slant media used for?

transport and storage

What is solid plate media used for?

to quantify or isolate

What is liquid media used for?

culturing large quantities of bacteria

What is a loop used for?

Inoculating liquid media and solid media

Making smears ("loop-full" of solid growth on a "loop-full" of water)

Aseptic inoculation techniques

Flame needle/loop from back to tip

Work within range of Bunsen burner

Hold tubes at sideways-ish to prevent air contamination

Flame tubes after opening and before closing

Clamshell plates

Simple staining

use a single stain to observe bacteria

ex: Direct stain, Negative stain, Capsule stain

Differential staining

use two stains to distinguish bacterial cells based on their cell wall structure

ex: Gram stain, Acid-Fast stain, Endospore stain

Mordant

something used to make dye penetrate cell wall and stick

ex: iodine, heat

What 2 types of staining did we talk about NOT heat fixing in lab?

Negative simple stains and Capsule staining

What is the charge of the outside of a bacterial cell?

-

What is the charge of the surface of a glass slide?

+

What is the charge of an acidic dye?

-

What is the charge of a basic dye?

+

Disadvantage of Direct simple stain?

Heat fixing causes some shape and size distortion

Advantage of Negative simple stain?

View true morphology and size of bacteria

Obligate aerobe

grows only in O2 presence

Deep results: top-only growth

Obligate anaerobe

grows only in O2 absence

Deep results: bottom-only growth

Facultative anaerobe

grows with or without O2

Deep results: throughout growth

Gram staining

distinguishes bacteria on thickness of peptidoglycan

Gram staining process

Smear bacteria on plate

Air dry

Heat fix

1st stain

Mordant (iodine)

Decolorizer (Alcohol, medium strength)

2nd stain

Gram staining results

thick peptidoglycan positive: purple

thick peptidoglycan negative: pink

Acid fast staining

distinguishes bacteria on presence of waxy mycolic acid in cell wall

Acid-Fast staining process

Smear bacteria on plate

Air dry

Heat fix

1st stain

Mordant (heat 8 mins)

Decolorizer (Acid alcohol, strong)

2nd stain

Acid-Fast staining results

mycolic acid positive: red

mycolic acid negative: blue

Mycobacterium

a large family of bacteria that has mycolic acid in cell wall

often found in soil and water

Endospore

A thick-walled protective spore that forms inside a bacterial cell and resists harsh conditions.

Endospore staining

distinguishes bacteria from endospores (bacteria "eggs")

not all bacteria make endospores

Endospore staining process

Smear bacteria on plate

Air dry

Heat fix

1st stain

Mordant (heat 8 mins)

Decolorizer (Water, weak)

2nd stain

Endospore staining results

endospores: green

vegetative bacteria: red

*there will be more endospores in old cultures of endospore-creating bacteria than in young cultures

Capsule staining

Used to observe the microbial CAPSULE, a protective glycocalyx layer surrounding the cells of some bacteria and fungi

Mix of negative stain and direct stain

Capsule staining process

Smear bacteria on plate

Air dry

Stain

Rinse with 20% copper sulfate

Capsule staining results

capsule positive: halo around stained bacteria

capsule negative: slime glycocalyx will not show halo

Possible causes of false positive for Gram stain

-too much bacteria on plate

-too weak or too little decolorizer used

-mordant left on too long

-wrong secondary stain used

Possible causes of false negative for Gram stain

-wrong primary stain used

-mordant not on long enough

-too strong or too much decolorizer used