Question

MICROBIOLOGY:

Exercise 3-7 Acid-Fast Stain (Kinyoun Method TRUE FALSE 1. The Acid-Fast stain is a differential stain. 2. The counter stain for the Kinyoun Acid-Fast method is 3. The decolorizing agent for the Kinyoun Acid-Fast method is . The Kinyoun method requires heat for at least 5 minutes . An Acid-Fast microbe is 6. The Acid-Fast method is used primarily to identify microbes in the genus TRUE FALSE in color 7. What is the other type of acid-fast staining procedure? 8. The NON-acid-fast microbe is usually stained what color? 9. The same antibiotics that are used to kill Gram-positive organisms are used to kill acid-fast organisms TRUE FALSE 10. Why are acid-fast organisms so hard to Gram stain? Exercise 3-8 Capsule Stain (Two methods) 11. A smear for the capsule requires heat-fixing 12. The basic stain (Maneval's) is applied and remains for five minutes. 13. What is the purpose of the Congo red stain? 14. What is the purpose of Maneval's stain? 15. Nigrosin can be used in place of Congo red. 16. This stain is both a structural and differential stain. 17. Identify 3 species of bacteria that may have capsules. TRUE FALSE TRUE FALSE TRUE FALSE TRUE FALSE 18. Sheep serum or egg albumin is sometimes used in preparing bacterial smears instead of water Why? (see pg. 217/201)

Answer 1

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1) acid-fast staining is differential: TRUE

2) counterstain for Kinyoun stain: Methylene Blue

3) Decolorizing stain for Kinyoun stain: 1% Acid alcohol

4) Kinyoun stain requires heat for at least 5 min: FALSE (requires no heat)

5) An acid fast microbe is Mycobacterium tuberculosis, M. Leprae, nocardia, rhodococcus, legionella

6) Staining used primarily for the genus: Mycobacterium

7) Other type of acid-fast staining: Ziehl Neelson acid-fast staining

8) Non acid fast microbe stain with the counterstain Methylene Blue: light blue

9) Drugs usually used for gram positive bacteria like erythromycin and acid fast bacteria like rifampicin are effective for the other type because they target common processes like transcription and translation but to a lesser extent. In practice specific antibiotics like penicillin that targets peptidoglycan in gram positive bacteria and isoniazid that targets mycolic acid formation in Mycobacteria are used. So the answer is no they are not. A gram positive antibiotic will usually target peptidoglycan and an acid fast bacteria will target the lipids in the cell walls.

10) acidfast bacteria are hard to gram staining because their cell walls are nearly impermeable to most stains. The cell walls are composed of mycolic acids, fatty acids, waxes and complex lipids. So a stain that is lipid soluble can only penetrate the cell wall. Harsher conditions like heat also have to used. These resist staining by acid stains. Once stained, they resist decolourization and do not let the acid stain out. Hence they are acid fast.

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11) FALSE. Heat fixing will destroy protein capsules and will also dry the cell which will result in shrinkage of the capsule.

12) TRUE. Flood the slide and pour off excess.

13) Congo red stains the background.

14) the purpose is to make the capsule visible by staining the background and the cell so the capsule is colorless making it visible. This is why it is called a negative staining method.

15) nigrosin can be used instead of Congo red: TRUE

16) TRUE

17) Streptococcus pneumoniae, Klebsiella pneumoniae, Neisseria meningitidis.

18) The electrostatic nature of the protein in sheep serum or albumin allows the bacteria to attach to the glass very well if water doesn't work.

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19) Rinsing agent is WATER: to decolourize the cell which is weakly stained by malachite green.

20) TRUE: Structural because it makes a particular structure visible and differential because it uses more than one chemical stain.

21) The primary stain is MALACHITE GREEN. The counterstain is safranin.

22) vegetative cell stains PINK.

23) FALSE. In this staining, heat acts as the mordant. The smear should be steaming for 3-5 minutes.

24) TRUE. Due to the fumes from the evaporation.

25) Clostridium perfringes, C. tetani, C. botulinum, Bacillus cereus, B. anthracis

26)

Subterminal 3) endospore between the middle and end of the cell

Terminal 1) endospore is in the middle of the cell

Central 2) endospore is at the end of the cell

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27) A special mordant is used to increase the flagellar thickness and make them visible: TRUE. Flagella are 10-50 nm making it difficult to see them under a light microscope.

28)

d. Amphitrichous 3) Flagella at both ends of the bacterium

e. Peritrichous 1) Flagellar covering the entire surface of the bacterium

f. Monotrichous 2) A single flagellum found on one end of the bacterium

g. Lophotrichous 4) Tufts of flagella at one end of the bacterium

29) Harsh procedures in flagellar staining such as Fixation and dehydration kill the bacteria hence live samples cannot be visualized.