1. Differentiation of Bacteria by Cell Wall Composition

2. The cell wall is the outer most layer of the cell. In many cases the cell wall comes in direct contact with the environment. Function  Protection of the cell  Maintains the shapes of the cell  Maintains the osmotic integrity of the cell

3.  Provide structure and shape and protect cell from osmotic forces  Assist some cells in attaching to other cells or in eluding antimicrobial drugs  Not present in animal cells, so can target cell wall of bacteria with antibiotics  Bacteria and archaea have different cell wall chemistry

5. Consider the three morphological distinctions of bacteria. Which of these shapes would need to be flexible in order to exhibit the shape of the bacterium?

6. Bacteria are classified into two groups based on the composition of their cell walls. The two groups are known as gram positive and gram negative.

7. ◦ Most have cell wall composed of peptidoglycan ◦ Peptidoglycan is composed of sugars, NAG, and NAM ◦ Chains of NAG and NAM attached to other chains by tetrapeptide crossbridges  Bridges may be covalently bonded to one another  Bridges may be held together by short connecting chains of amino acids

8. Gram-positive cell walls  Relatively thick layer of peptidoglycan  Contain unique polyalcohols called teichoic acids  Some covalently linked to lipids, forming lipoteichoic acids that anchor peptidoglycan to cell membrane  Retain crystal violet dye in Gram staining procedure; so appear purple

9. The gram positive cell wall is referred to as a peptidoglycan layer and is composed of alternating subunits of two carbohydrates: n-acetyl muramic acid and n-acetyl glucosamine.

10. The gram positive cell wall is referred to as a peptidoglycan layer and is composed of alternating subunits of two carbohydrates: n-acetyl muramic acid (NAM) and n-acetyl glucosamine (NAG).

11. ◦ Gram-negative cell walls  Have only a thin layer of peptidoglycan  Bilayer membrane outside the peptidoglycan contains phospholipids, proteins, and lipopolysaccharide (LPS)  May be impediment to the treatment of disease  Appear pink following Gram staining procedure

12. The gram negative cell wall consists of two distinct layers. The outer most layer is referred as the outer membrane and is composed of lipopolysaccarhides (LPS). The second layer a peptidoglycan layer that is homologous to a gram positive cell wall.

14. Why does the gram stain work? How is the differences of cell wall types used to distinguish the different bacterial types?

15. Scenario You have been give a slide with a mixture of gram positive (coccus) and gram negative bacteria (spirochete). Think about the gram stain procedure as it relates to the cell wall differences. What would the end result of each step of the gram stain be for each of the bacteria types?

16. Gram Staining Procedure ReagentFunction Results Crystal VioletPrimary Stain IodineMordant Acetone-AlcoholDecolorizer SafraninCounter Stain

18. Why does the gram stain work? Because of the differences of cell wall composition.

19. The gram positive cell wall is referred to as a peptidoglycan layer and is composed of alternating subunits of two carbohydrates: n-acetyl muramic acid and n-acetyl glucosamine. The gram negative cell wall consists of two distinct layers. The outer most layer is referred as the outer membrane and is composed of lipopolysaccarhides (LPS). The second layer a peptidoglycan layer that is homologous to a gram positive cell wall.

21.  Archaeal Cell Walls ◦ Do not have peptidoglycan ◦ Contains variety of specialized polysaccharides and proteins ◦ Gram-positive archaea stain purple ◦ Gram-negative archaea stain pink