1. An Overview of Bacterial Cell Structure Viro102: Bacteriophages & Phage Therapy 3 Credit hours NUST Centre of Virology & Immunology LECTURE 2:

2. Cell types Prokaryotic Cell Eukaryotic Cell Bacteria & Archaea animals, plants, fungi

3. Bacterial Cell Morphology

4. Bacterial Cell no membrane bound organelles  Capsule  Cell Wall  Cell membrane  Ribosome  Plasmids  Chromosome  Flagellum  Pilli

5. Capsule  Outer most covering of some bacteria  Well organized layer, not easily washed off  Helps to protect bacteria against oPhagocytosis, A capsule specific antibody may be required for phagocytosis to occur oBacteriophages oThey also exclude bacterial viruses and oHydrophobic toxic materials such as detergents

6. Capsule  Capsules also contain water which protects bacteria against desiccation  Allow bacteria to adhere to surfaces and cells.  Composed of polysaccharides  Bacillus Anthracis has protinaceous capsule composed of poly-D glutamic acid

7. Slime Layer

8. Glycocalyx

9. Bacterial Cell no membrane bound organelles  Capsule  Cell Wall  Cell membrane  Ribosome  Plasmids  Chromosome  Flagellum  Pilli

10. A comparison b/w G(+ve) & G(-ve)

11. GRAM POSITIVE GRAM NEGATIVE Cytoplasm Cytoplasm Lipoteichoic acid Peptidoglycan-teichoic acid Cytoplasmic membrane Inner (cytoplasmic) membrane Outer Membrane Lipopolysaccharide Porin Braun lipoprotein Periplasmic space

12. Classification of Bacteria  In 1884, Hans Christian Gram developed a staining technique for the identification of bacteria Gram staining  Based on this technique, Bacteria is characterized into oGram positive oGram negative  The difference is due to the cell wall differences in these bacteria

13. Gram Staining Technique

14. What happens in Gram staining?  Crystal Violet (purple) oPrimary stain; positive stain. oStains all cell walls purple.  Iodine oMordant (Any substance used to facilitate the fixing of a dye to a fibre). oCombines with CV to form an insoluble complex that gets trapped in thicker peptidoglycan layers.  Ethanol oDecolorizer. oCV complex washed out of Gram negative organisms because it cannot be trapped by outer layer.  Safranin (pink) oCounterstain. oStains all cells, but only the negative ones actually appear pink.

15. Prepare a thin smear of fresh bacteria Apply primary stain & then wash Apply mordant & then wash Apply ethanol Apply secondary stain & then wash Visualize

16. Results under a Microscope Yogurt: (Lactobacillus {G+} and Enterobacteria{G-})

17. Gram stain of cerebrospinal fluid showing B. anthracis Bacillus G +ve G -ve

18. Bacterial Cell No membrane bound organelles  Capsule  Cell Wall  Cell membrane  Ribosome  Plasmids  Chromosome  Flagellum  Pilli

19. Cell Membrane  Composition: oPhospholipid bilayer just like eukaryotic PM (Fluid Mosaic) oLipids are not homogeneously distributed in membrane oContain fatty acids with additional methyl hydroxyl or even cyclic groups oThe relative proportions of these fatty acids can be modulated by the bacterium to maintain the optimum fluidity of the membrane (e.g. following temperature change)

20. Cell Membrane  Functions: oActing as a permeability barrier for most molecules oServing as the location for the transport of molecules into the cell oFunction in energy production as the location about which a proton motive force is generated oAct as site for photosynthesis in Cyanobacteria

21. Bacterial Cell No membrane bound organelles  Capsule  Cell Wall  Cell membrane  Ribosome  Plasmids  Chromosome  Flagellum  Pilli

22. Ribosomes  Site of protein synthesis  Made up of protein and RNA  The 70S ribosome is made up of a 50S and 30S subunits  The 50S subunit contains the 23S and 5S rRNA  The 30S subunit contains the 16S rRNA

23. Bacterial Cell No membrane bound organelles  Capsule  Cell Wall  Cell membrane  Ribosome  Plasmids  Chromosome  Flagellum  Pilli

24. Bacterial Chromosome  Present in the cytoplasm  The bacterial chromosome is not packaged using histones to form chromatin as in eukaryotes but instead exists as a highly compact supercoiled structure, the precise nature of which remains unclear.  Most bacterial chromosomes are circular although some examples of linear chromosomes exist (e.g. Borrelia sp.)

25. Bacterial Cell No membrane bound organelles  Capsule  Cell Wall  Cell membrane  Ribosome  Plasmids  Chromosome  Flagellum  Pilli

26. Plasmids  Most bacteria also contain small independent pieces of DNA called plasmids that often encode for traits that are advantageous but not essential to their bacterial host

27. Bacterial Cell No membrane bound organelles  Capsule  Cell Wall  Cell membrane  Ribosome  Plasmids  Chromosome  Fimbriae  Pili  Flagellum

28. Fimbriae  Many prokaryotes have short, fine, hair like appendages that are thinner than flegella known as Fimbriae  Interchangeably term fimbraie and pili.  Cell may covered by 1000s of fimbraie, visible under electron microscope

29. Cont’d  Slender tubes composed of helical arrangement of protein subunits  3-10 nm in diameter  Several millimeter long  Usually facilitate the attachment of a bacterium to a surface (e.g. to form a biofilm) or to other cells (e.g. animal cells during pathogenesis)

30. Cont’d  Moreover help in motility  Pseudomonas aeroginosa, Neisseria gonorrhoeae, some strain of E.coli

31. Bacterial Cell No membrane bound organelles  Capsule  Cell Wall  Cell membrane  Ribosome  Plasmids  Chromosome  Fimbriae  Pili  Flagellum

32.  Hair like appendages  Different from fimbraie  Pili are larger  1-10 / cell  Genetically determined by conjugative plasmid  Phages specifically bind on sex pili, hence infect bacteria Pili

33.  During bacterial conjugation, a sex pilus emerging from one bacterium ensnares the recipient bacterium  This is direct contact merging the cytoplasms of two bacteria via a controlled pore which allows for the transfer of bacterial DNA from the bacteria with the pilus (donor) to the recipient bacteria  Usually the fertility factor is required to produce sex pili Pili

35. Bacterial Cell No membrane bound organelles  Capsule  Cell Wall  Cell membrane  Ribosome  Plasmids  Chromosome  Fimbriae  Pili  Flagellum

36. Flagellum  Flagella are whip like structures protruding from the bacterial cell wall and are responsible for bacterial motility  Composed of different proteins including flagellin  The arrangement of flagella about the bacterial cell is unique to the species observed. Common forms include:  Peritrichous Multiple flagella found at several locations on the cell.  Polar Single flagella found at one of the cell poles.  Lophotrichous A clump of flagella found at one cell pole.

37. Flagellum

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