ANSWERS

1. List the 3 basic shapes of bacteria and give an example of each group.
   • Cocci - spheres.
   • Bacilli - rods.
   • Spirilla - spirals/helixes.
2. What are curved bacilli known as?
   Vibrios.
3. What are variable shaped bacteria called?
   Pleomorphic.
4. Describe the following: diplococci, streptococci, staphylococci.
   • Diplococci - pairs of cocci.
   • Streptococci - chains of cocci.
   • Staphylococci - clusters of cocci.
5. List the components of a generalised bacterial cell and briefly describe the function of each.
   • Cell wall (made of peptidoglycan/murein) - maintains cell shape, prevents cell from bursting and provides protection from damage.
   • Capsule/slime layer - protection, aids adherence, aids virulence, prevents phagocytosis, may serve as a food reserve.
   • Cell membrane - acts as a diffusion gradient and controls the passage of substances into and out of the cell, site of enzyme activity.
   • Meosomes (folds of the cell membrane) - concerned with respiration and cell division.
   • Cytoplasm - site of synthesis processes.
   • Bacterial chromosome - contains DNA which carries the hereditary information of the cell.
   • Plasmids - involved in conjugation; extra pieces of DNA which can replicate independently from the chromosome.
   • Flagellae - concerned with motility of the cell.
   • Pili - aid adherence, sex pili are involved in conjugation.
6. How do antibacterial drugs work?
   They target the bacterial cell wall (which is chemically unlike any other structure found in animal cells). This means that bacteria may be attacked and destroyed without harming the host. Once the wall is damaged, the cell takes up water, swells and bursts. Penicillin prevents the synthesis of the cell wall particularly in Gram positive bacteria.
7. Gram positive bacteria may produce harmful microbial exotoxins. Give 3 examples and state the disease caused by each.
   • Clostridium tetani - tetanus.
   • Clostridium botulinum - botulism.
   • Staphylococcus aureus - food poisoning.
8. List the essential requirements for bacterial growth.
   • Nutrient supply.
   • Optimum temperature for growth (usually body temperature).
   • Correct pH (usually 7-7.4).
   • Water.
   • Correct gaseous environment.
9. What are facultative anaerobes?
   Organisms that grow aerobically when oxygen is available but can also function anaerobically.
10. What is the process by which bacteria reproduce called?
    Binary fission (asexual reproduction).
11. Briefly describe the process by which bacteria reproduce.
    • Bacterial cell reaches a certain size.
    • Chromosome replicates to form 2 identical chromosomes.
    • Cell enlarges, chromosomes are separated and cell membrane grow inwards at the centre.
    • Cell wall grows inwards at the centre forming a septum.
    • Cell divides forming 2 daughter cells which may separate completely or remain attached (forming chains or clusters - see Q3).
12. What is meant by conjugation?
    The passage of DNA from one bacterial cell (the donor cell) to another (the recipient).
13. Briefly describe conjugation.
    • The donor and recipient cells are pulled together by the sex pilus.
    • Pilus retracts bringing the 2 cells very close to one another.
    • Cell membranes fuse to form a channel between the 2 cells.
    • Plasmid replicates and 1 strand passes through the channel to the recipient.
    • The 2 cells separate - the recipient is now a donor cell since it contains the plasmid.
14. What is a bacterial endospore?
    A dormant form of bacteria that can survive in unfavourable conditions. Endospores are formed when the growing cells are deprived of some factor e.g. inadequate nutrition).
15. Name 2 genera in which spore formation is common.
    • Bacillus.
    • Clostridium.
      Bacterial spores are extremely resistant and can survive extremes of temperature and pH and exposure to some disinfectants.
16. The identification of bacteria may be assisted by the use of microbial staining procedures. List 2 simple stains suitable for staining bacteria.
    • New methylene blue - shows the presence and morphology of bacteria.
    • Polychromatic methylene blue - used to stain anthrax bacilli in blood smears and to demonstrate McFadyean's reaction.
17. What is meant by a differential stain?
    A stain which allows differentiation between more than one type of bacteria. Gram's stain is an example which is used to differentiate intact, morphologically similar bacteria into 2 groups based on cell colour after staining.
18. List the 4 chemicals used in a Gram's stain.
    • 0.5% Aqueous crystal violet.
    • Lugol's iodine.
    • Methylated spirit.
    • Dilute carbol fuchsin (or saffranine).
19. Of the chemicals listed in Q17, which is the mordant?
    Lugol's iodine.
20. What is the difference between Lugol's and Gram's iodine?
    Lugol's is more concentrated than Gram's and is used to obtain darker colourisation.
21. What colour do Gram positive bacteria stain?
    Deep blue/purple.
22. What colour do Gram negative bacteria stain?
    Red.
23. What is the name of the stain used to detect acid fast bacilli such as mycobacteria (tuberculosis and Johne's disease)?
    Ziehl Neelsen stain.
24. Why are bacterial smears heat fixed?
    • To prevent the sample from being washed off.
    • To kill the bacteria but preserve cell morphology.
25. What is the function of a culture medium?
    Provision of an appropriate nutrient material in order to cultivate bacteria.
26. List 3 examples of simple (basal) culture media.
    • Nutrient broth.
    • Nutrient agar.
    • Peptone water.
27. What is meant by enriched culture media?
    A media containing a supplement in order to promote the growth of more fastidious bacteria which will not grow in a simple medium.
28. Give an example of enriched culture medium.
    Chocolate agar (containing ruptured red blood cells); suitable for the cultivation of Neisseria species.
29. What is the difference between enriched and enrichment culture media?
    Enriched media are supplemented culture media designed simply to promote the growth of certain bacteria, whilst enrichment media are only used when a required species of bacteria is present in a mixed sample in very small numbers. It favours the growth of the wanted species, enabling it to become dominant.
30. What are selective media?
    Media designed to inhibit the growth of certain bacteria while not affecting the growth of others.
31. Give 2 examples of selective media.
    • MacConkey's broth - inhibits non-enteric bacteria.
    • Deoxycholate-citrate agar - inhibits many non-enteric bacteria but most strains of Salmonella will form colonies on it.
32. MacConkey's agar, in addition to being an example of a selective culture medium, is also a differential medium. What do you understand by this term?
    Differential media are used to distinguish between different species of bacteria on the same agar plate.
33. What is the optimum temperature for the cultivation of campylobacter?
    40 Degrees C.
34. At what temperature should unused agar plates be stored at and why?
    2-6 Degrees C. A cool temperature prevents evaporation of the culture medium, and plates may be stored in the refrigerator for up to 3 weeks and inhibits the growth of pathogens.
35. Why are agar plates incubated upside-down?
    In order to prevent condensation from pooling on the medium.
36. What is the correct method for the disposal of culture plates?
    Autoclave and put into a yellow clinical waste sack with biohazard seal ready for incineration.
37. Name 2 proprietary media suitable for performing sensitivity testing.
    • Mueller-Hinton.
    • Sentest.
38. Briefly describe the use of biochemical media.
    These are used to distinguish between different species of bacteria by detecting differences in their biochemical reactions with the media. Differentiation may be determined by various factors including the ability of bacteria to ferment certain sugars, and the production of enzymes.
39. For what purpose are transport media employed?
    Transport media provide a suitable environment for the storage of bacterial samples prior to laboratory culture. They do not support growth, but ensure the survival of any organisms present until the material can be examined.