AFB Practical #1

Question 1

What species of mycobacteria are nucleic acid probes available for?

Answer 1

• M. tuberculosis complex
• M. avium complex
• M. gordonae
• M. kansasii

Question 2

List 3 characeristics of Nocardia sp.

Answer 2

• Often appear powdery white due to production of aerial hyphae
• Branching GPR
• Partially acid fast

Question 3

Nocardia sp.

• Modified acid-fast stain
• Kinyoun acid-fast stain

Answer 3

• Modified acid-fast stain positive (red)

- Kinyoun acid-fast stain negative (blue)

Question 4

What is a confirmation test for M. tuberculosis and M. bovis?

Answer 4

Nitrate test

• M. tuberculosis (+)
• M. bovis (-)

Question 5

What reagent must be added to a negative nitrate test? Why?

Answer 5

Addition of zinc dust confirms nitrogen gas

Question 6

Niacin accumulation test is used to differentiate b/w which two mycobacterial species?

Answer 6

• M. tuberculosis (+)

- M. bovis (-)

Question 7

Name 4 characteristics of M. mucogenicum

Answer 7

• Rapid grower
• Partially acid-fast
• Nonchromogen
• Wet/mucoid looking colonies

Question 8

How is Kinyoun stain interpreted?

Answer 8

• Pos: red

- Neg: blue/purple

Question 9

Known as the “cold” carbolfuschin-based stain

Answer 9

Kinyoun

Question 10

The phenomenon known as cording is observed in which mycobacterium species?

Answer 10

M. tuberculosis (virulent factor)

Question 11

Name 3 characteristics of M. marinum

Answer 11

• Grows at 28C
• Cutaneous infection associated w/ fish tanks or salt water
• Photochromogen

Question 12

What type of stain is the auramine-rhodamine (AR) stain?

Answer 12

Fluorochrome stain

Question 13

What are the advantages fo the AR stain?

Answer 13

• Most sensitive stain
• Stains both viable and nonviable organisms
• Performed directly on specimens

Question 14

What are the disadvantages of the AR stain?

Answer 14

Won’t stain rapid growers

Question 15

What reagents are used in the N-acetyl-L-cysteine method of decontamination?

Answer 15

• N-acetyl-L-cysteine
• NaOH
• Sodium citrate
• Neutralizing buffer
• Phenol red

Question 16

N-acetyl-L-cysteine

- Purpose of N-acetyl-L-cysteine

Answer 16

Works as a mucolytic agent

Question 17

N-acetyl-L-cysteine

- Function of NaOH

Answer 17

Functions as a bacterial decontaminant

Question 18

N-acetyl-L-cysteine

- Function of sodium citrate

Answer 18

Binds to heavy metals in the specimen

Question 19

N-acetyl-L-cysteine

- Function of neutralizing buffer

Answer 19

Stabilizes the pH and stops the decontamination process

Question 20

N-acetyl-L-cysteine

- Function of phenol red

Answer 20

Present in the digestion solution to monitor pH

Question 21

Incubation requirements for MAC w/o crystal violet

Answer 21

Ambient air @ 28-30°C for 7 days

Question 22

Which two mycobacterial species is the sodium chloride tolerance test useful in identifying? Results?

Answer 22

• M. abscessus (+)

- M. chelonae (-)

Question 23

Incubation requirements for the Tween 80 hydrolysis test

Answer 23

Ambient air @ 35-37°C for 10 days

Question 24

How is the Tween 80 hydrolysis test interpreted?

Answer 24

• Pos: pink/red

- Neg: orange/no color change

Question 25

What is HPLC and how do we use it in the lab?

Answer 25

High performance liquid chromatography; provides an ID based on mycolic acid pattern, analysis of mycolic acids contained within mycobacterial cell walls

Question 26

How do mycobacteria typically appear in gram stains?

Answer 26

Beaded or ghosted GPR (can look similar to strep)

Question 27

Name 3 characteristics of M. gordonae

Answer 27

- Found in tap water - Scotochromogen - Non-pathogenic

Question 28

Name 3 characteristics of M. tuberculosis

Answer 28

- Rough and buff - Slow grower - Nonchromagen - Must confirm ID using biochemical methods

Question 29

Which mycobacteria species deomonstrates candycane striations?

Answer 29

M. kansasii

Question 30

Requires heme to grow, optimal temperature is 30C generally isolated from immunocomposed individualsq

Answer 30

M. haemophilum

Question 31

What is the MB bottle (Myco F Lytic) used for?

Answer 31

Liquid medium used for blood, put into Bactec Fx, has heme (CHOC) in media to break apart RBCs to release organism

Question 32

What is the MP bottle (MGIT) used for?

Answer 32

Liquid medium used for all specimen excepts blood; put into MGIT instrument, clear if uninooulated

Question 33

Name 2 characteristics of M. kansasii

Answer 33

- "Candy cane" striations | - Photochromogen

Question 34

What is the 7H10 slant used for?

Answer 34

All purpose, enriched for AFB, clear agar, if from a superficial source a second slant is set up and incubated at 30°C

Question 35

Purpose of the choclate slant?

Answer 35

Used for specimens rich in heme (BM, blood), incubated at 30C

Question 36

Purpose of LJ slant?

Answer 36

Lowenstein-Jensen, used in biochemical testing

Question 37

Function of the 7H10 plate?

Answer 37

Used when isolation of colonies is desired

Question 38

What PPE is used in the AFB lab

Answer 38

- Gloves - Gown - Mask (N95 or better) - Foot covers

Question 39

What unique characteristic is seen in M. szulgai?

Answer 39

Colonies are either smooth or rough

Question 40

What is he media used in the 5% NaCl tolerance test?

Answer 40

LJ slant with 5% NaCl

Question 41

What are the incubation requirements for the 5% NaCl tolerance test?

Answer 41

Incubated at 35-37C in CO2 up to 4 weeks