Cultivation and Isolation of Bacteria

What are the different types of stains?

- Gram stain

- Ziehl-Neelsen/Kinyoun/Modified

- Fluorochrome

- Acridine Orange

Gram positive pathogens stain ______.

purple

What is the order of a gram stain procedure?

- Fixation

- Crystal violet

- Wash

- Iodine treatment

- Wash

- Decolorization

- Wash

- Counter stain (safranin)

- Wash

- Blot dry

Ziehl-Neelsen/Kinyoun stain is used for ______.

mycobacteria

Ziehl-Neelsen is associated with the use of ______ temperatures, while Kinyoun is associated with ______ temperatures.

hot, cold

The primary stain used in Ziehl-Neelsen is ______.

Carbol-Fuschsin

The decolorizer used in Ziehl-Neelsen is ______.

3% HCl in ethanol

______ is used as the counter stain in Ziehl-Neelsen.

Methylene blue

Modified Z-N/Kinyoun is used for partially ______ organisms.

acid-fast

______ is the primary stain and ______ is the counter stain used in Modified Z-N/Kinyoun.

Carbol-Fuchsin, methylene blue

______ is the decolorizer used in Modified Z-N/Kinyoun.

1% H2SO4

Fluorochrome staining is used for ______.

mycobacteria

Fluorochrome is the primary stain used on all specimens sent for mycobacterial ______.

culture

In Fluorochrome staining, ______ is used as the primary stain.

Auramine-Rhodamine

In Fluorochrome staining, ______ is used as the counterstain.

K permanganate

What is the decolorizing solution for Fluorochrome staining?

0.5% HCl in 70% EtOH

What are the 4 different types of plating media?

- All-purpose/non-selective

- Enriched

- Selective

- Selective/Differential

All-purpose/non-selective media supports the growth of most ______ bacteria.

non-fastidious

All-purpose/non-selective is a ______ agar.

nutrient

Blood agar is a type of ______ agar consisting of Trypticase Soy Agar with 5% sheep red blood cells.

enriched

With blood agar, an MLS should be acutely aware of ______ after colony formation.

hemolysis

What are the types of hemolysis that can present on blood agar?

- Alpha

- Beta

- Gamma

Alpha hemolysis produces a ______ discoloration surrounding the colonies.

greenish

This greenish discoloration is due to ______ hemolysis of the RBCs in the blood agar.

incomplete

Bacteria that produce alpha hemolysis on blood agar are always ______ negative.

catalase

Catalase negative bacteria causes an accumulation of ______, causing alpha hemolysis coloration.

H2O2

Beta hemolysis on blood agar produces a ______ zone around the colonies.

clear/colorless

This clear zone is caused by ______ hemolysis of RBCs on the blood agar plate.

complete

Organisms that produce a beta hemolytic zone on blood agar are always ______ positive.

hemolysin

Gamma hemolysis means there is ______ hemolysis/discoloration.

no

Chocolate agar is a type of enriched media that consists of RBCs that are ______ before they are mixed into the agar.

lysed

Chocolate agar supports the growth of ______ organisms such as Neisseria and Haemophilus.

fastidious

What is the purpose of selective agar?

Inhibit the growth of some bacteria and promote the growth of others

Through what mechanisms can selective agar function through?

- Nutrient manipulation

- pH

- Osmotic adjustments

- Oxygen tension

- Dyes

- Antimicrobial agents

What are the three types of selective agar?

- Phenylethyl alcohol (PEA)

- Columbia Nalidixic Acid (CNA)

- Thayer-Martin

Selective/Differential agar ______ between the bacteria that does grow.

differentiates

What are the types of selective/differential agar?

- MacConkey

- Hektoen

- Mannitol Salt Agar

MacConkey agar is selective for ______.

gram negative rods

MacConkey agar differentiates between bacteria that is/is not ______.

lactose fermenting

Colonies that are the same color as the MacConkey agar plate are _______.

non-lactose fermenters

Colonies that are pink against the MacConkey agar plate are ______.

lactose fermenters

Hektoen agar is selective for ______.

gram negative rods

Hektoen agar differentiates between bacteria that are _______ fermenters.

lactose/sucrose

Colonies that appear green on Hektoen agar are _______ fermenters.

non-sucrose, non-lactose

If these green colonies have a black center, they are ______ positive.

H2S

Colonies that appear orange on Hektoen agar are _______ fermenters.

lactose and sucrose

Mannitol Salt Agar plating that yields a yellow color means the corresponding bacteria _______.

ferment mannitol

Mannitol Salt Agar plating that yields no change in color (stays red) means the corresponding bacteria _______.

don't ferment mannitol

Mannitol Salt Agar is ______ selective, along with anything that can grow in high ______ levels.

staph, salt

Enrichment broth is used for ______ the number of organisms present in a sample.

increasing

Transport media is used to maintain organism ______.

viability

What are some of the transport medias used in microbiology?

- Cary-Blair

- Amies

- Stuart

What are the two types of media inoculation techniques?

- Dilution streak

- Quantitative cultures

Dilution Streak Plating Technique

Quantitative Culture Plating Technique

What is the optimal temperature used for media incubation?

35C (+/-2C)

For aerobic bacteria, what should be the atmospheric compound levels, from highest to lowest?

N2 --> O2 --> CO2

For microaerophilic bacteria, what should be the atmospheric compound levels, from highest to lowest?

N2 --> CO2 --> O2

For anaerobic bacteria, what should be the atmospheric compound levels, from highest to lowest?

N2 --> H2 --> CO2

The optimal incubation time for media is ______.

24 hours

What components are observed when looking at colony morphology?

- Hemolysis

- Size

- Margin

- Elevation

- Density

- Color

- Consistency

- Odor