Bacteriology Lab Final

What technique is designed to keep the working environment as clean as possible by sterilizing equipment in contact with microorganisms?

Aseptic technique

What is the loop used for?

Transferring cultures to other mediums

Prepare bacterial smears

Streak plates

What is the straight wire used for?

Stab cultures

Pick single colonies from plate

What is the order of the stains in the gram staining technique?

1. Primary stain

2. Decolorizing agent

3. Counterstain

What is the primary stain in gram staining?

Crystal violet

What is the mordant, and what is it used for?

Gram’s iodine

Intensifies the colour of the stain

What is the decolorizing agent in gram staining and how does it differ in gram negative and gram positive organisms?

50% ethanol 50% acetone

Protein stain difficult to remove in gram-positive. Lipid concentration in outer layers of gram-negative easily dissolved by alcohol.

What is the counterstain in gram stain?

Safranin

Stains decolorized cells red.

What’s an example of a gram-negative bacteria?

E. coli

What is an example of a gram-positive bacteria?

Staphylococcus

What is Acid-Fast stain (Ziehl-Neelsen Method) used for?

Mycobacterium and Nocaridia

Why is the acid-fast stain useful for staining mycobacterium?

The thick waxy wall makes it difficult to remove the penetrated stain

What is the primary stain in acid-fast stain?

Carbol fuchsin (red)

How is penetration of Carbol fuchsin enhanced?

Application of heat

What is counterstain in acid-fast stain?

Methylene Blue

What stain is this?

Gram stain

What stain is this?

Acid-fast Stain

What hemolysis does Staphylococcus aureus display on blood agar?

Beta-hemolytic

Also forms a yellow film around colonies

What does staph aureus do to MSA?

Color change from red to yellow

What result does streptococcus produce in the catalase test?

Negative

What result does staphylococcus aureus produce in the catalase test?

Positive

What is the principle behind the catalase test?

The bacterium contains catalase that will break down H₂O₂ into H₂O and O₂

What is the coagulase test used for?

To differentiate between Staph aureus (positive) from coagulase negative staphylococcus

What test is seen here, and what organism could it be?

Tube coagulase test

Positive: Staph aureus

1. what test is seen here?

2. what organism is the middle streak

3. what organism gives a positive result? (arrow head)

1. CAMP test

2. Staph aureus

3. Streptococcus agalactiae

Which organisms are CAMP positive?

Listeria spp and Rhodococcus equi

How does the CAMP test work?

Interaction of the beta-hemolysin from staph aureus with a protein produced by group B streptococci causes synergistic hemolysis

Which organisms are catalase positive?

• Nocardia asteroids

• Rhodococcus equi

• Listeria monocytogenes

• Bacillus spp

What’s an example of a catalase negative organism?

Trueperella pyogenes

What is the media and what organism is present?

Listeria monocytogenes in motility media

1. What species is this?

2. Is it gram positive or gram negative

1. Bacillus spp.

2. gram positive

1. what organism is seen here?

2. Is it gram-positive or gram negative?

3. What is it’s shape?

1. Listeria monocytogenes

2. gram positive

3. rod-shaped

1. What organism is seen?

2. Is it gram-negative or positive?

3. What is the shape?

1. Rhodococcus equi

2. gram-positive

3. Coccobacilli

What organism is displayed on this plate?

Bacillus spp.

What organism is displayed on this plate?

Listeria monocytogenes

What organism is displayed on this plate?

Rhodococcus equi

1. What is the principle of the oxidase test?

2. What is an organism that is oxidase positive?

1. Cytochrome oxidase enzyme oxidizes the reagent to a purple end product

2. Pseudomonas aeruginosa

1. What occurs when the phenol indicator has a red slant and a red butt?

2. What organisms would cause this?

1. glucose and lactose non-fermentor

2. pseudomonas

1. What occurs when there is a red slant and yellow butt?

2. What organism/s could cause this?

1. glucose fermentation only

2. Shigella, Serratia

1. What occurs when there is a yellow slant, yellow butt and gas?

2. What organism/s can cause this?

1. glucose and lactose fermentation, and gas production

2. Escherichia, Klebsiella, Enterobacter

1. what occurs when there is a yellow slant and butt, gas, and a black colour?

2. What organism/s can cause this?

1. glucose + lactose fermentation, gas production, H₂S production

2. ?

1. What occurs when there is a red slant, yellow but, gas and black coloration

2. What organism/s can cause this?

1. Glucose fermenter, gas producing, H₂S producing

2. Salmonella, Proteus

1. What does a positive urease test look like?

2. What is the principal behind it?

3. What organism produces a positive result?

4. What organism produces a delayed-positive result? (weak pink coloration)

1. Production of a bright pink color on the slant that may extent to butt

2. Reaction from interaction of metabolic waste indicating the presence of enzyme (urease) required to break down carbohydrate.

3. Proteus spp.

4. Klebsiella pneumoniae

What organism produces a urease negative result?

Escherichia coli

What is the SIM medium used for?

Differentiation of microorganisms on the bases of hydrogen sulfide production, indole production and motility

1. What is motility indicated by?

2. What organism displays positive motility?

1. turbidity of the medium or growth extending from inoculating stab line

2. Salmonella spp

   (also produces H₂S)

1. What is the positive test for Indole production?

2. What organism produces this result

1. Red color after the addition of Kovac’s reagent

2. Shigella spp.

1. What test medium is this?

2. What’s the principle behind it

1. Simmons citrate agar

2. organisms that can utilize citrate contain permeases which results in the formation of CO₂ reacting with the medium to produce sodium carbonate, which raises the pH

1. What is the positive citrate test?

2. What organism causes this?

1. increase pH causes green colour to change to blue

2. Klebsiella pneumoniae, Citrobacter spp, Enterobacter aerogenes

1. What is EMB used for?

2. What is the result of lactose fermenters?

3. What is the result of non-lactose fermenters?

1. Isolation and differentiation of gram-negative bacilli

2. Blue-black with or without a green metallic sheen (E. coli)

3. Colourless and translucent

1. What is MacConkey Agar used for?

2. What is a positive test?

3. What organism gives a positive test?

1. Differentiation of lactose fermenting and non-lactose fermenting

2. Pink colonies

3. E. coli, Klebsiella