微生物學

1. 微生物學 許勝傑 博士 長庚大學 生物醫學系 助理教授 E-mail: schsu@mail.cgu.edu.tw 電話 (03)211-8800#3690

5. Chapter 1 The Evolution of Microorganisms and Microbiology CHAPTER GLOSSARY Archaea Bacteria Eukarya Fungi Genome Genomic analysis Koch’s postulates Microbiology Microorganism Prions Prokaryotic cells Protists Spontaneous generation Viroids Viruses Virusoids Figure/ Table Summary

6. What is microbiology? • study of organisms too small to be clearly seen by the unaided eye (i.e., microorganisms) • these organisms are relatively simple in their construction and lack highly differentiated cells and distinct tissues

7. The Importance of Microorganisms • most populous and diverse group of organisms • found everywhere on the planet • play a major role in recycling essential elements • source of nutrients and some carry out photosynthesis • benefit society by their production of food, beverages, antibiotics, and vitamins • some cause disease in plants and animals Microbes are estimated to contains 50% of biological carbon and 90% of biological nitrogen on Earth.

8. Fig. 1.1 Concept map showing the types of biological entities studied by microbiologists.

9. Members of the microbial world • prokaryoticcells lack a true membrane-delimited nucleus • eukaryoticcells have a membrane-enclosed nucleus, are more complex morphologically and are usually larger than prokaryotic cells Classification schemes • five kingdom scheme includes Monera原核界, Protista, Fungi, Animalia and Plantae with microbes placed in the first three kingdoms • three domain alternative, based on a comparison of ribosomal RNA, divides microorganisms into Bacteria(true bacteria),Archaeaand Eucarya (eucaryotes)

10. Discovery of Microorganisms • Antony van Leeuwenhoek (1632-1723) • first person to observe and describe microorganisms accurately

11. The Conflict over Spontaneous Generation • spontaneous generation • living organisms can develop from nonliving or decomposing matter • Francesco Redi (1626-1697) • disproved spontaneous generation for large animals • showed that maggots on decaying meat came from fly eggs

12. Louis Pasteur (1822-1895) • his experiments • placed nutrient solution in flasks • created flasks with long, curved necks • boiled the solutions • left flasks exposed to air • results: no growth of microorganisms

13. Koch’s postulates • The microorganism must be present in every case of the disease but absent from healthy individuals. • The suspected microorganism must be isolated and grown in a pure culture. • The same disease must result when the isolated microorganism is inoculated into a healthy host. • The same microorganism must be isolated again from the diseased host.

14. Earliest Molecules - RNA • original molecule must have fulfilled protein and hereditary function • ribozymes • RNA molecules that form peptide bonds • perform cellular work and replication • earliest cells may have been RNA surrounded by liposomes

15. Chapter 2 The Study of Microbial Structure: Microscopy and Specimen Preparation CHAPTER GLOSSARY Atomic force microscope Bright-field microscope Confocal scanning laser microscope (CSLM) Dark-field microscope Differential interference contrast (DIC) microscopy Differential staining Fixation Fluorescence microscopy Gram stain Negative staining Parfocal Phase-contrast microscope Refractive index Resolution Scanning electron microscope (SEM) Simple staining Transmission electron microscope (TEM)

16. Lenses • focus light rays at a specific place called the focal point • distance between center of lens and focal point is the focal length • strength of lens related to focal length • short focal length more magnification

17. The Light Microscope • bright-field microscope • dark-field microscope • phase-contrast microscope • fluorescence microscope

18. Microscope Resolution • ability of a lens to separate or distinguish small objects that are close together • wavelength of light used is major factor in resolution shorter wavelength  greater resolution

19. Fixation • preserves internal and external structures and fixes them in position • organisms usually killed and firmly attached to microscope slide • heat fixation – routine use with procaryotes • preserves overall morphology but not internal structures • chemical fixation – used with larger, more delicate organisms • protects fine cellular substructure and morphology Preparation and Staining of Specimens • increases visibility of specimen • accentuates specific morphological features • preserves specimens

20. Dyes and Simple Staining • dyes • Ionizable dyes have charged groups • basic dyes have positive charges • acid dyes have negative charges • chromophore groups • chemical groups with conjugated double bonds • simple stains • a single stain is used • use can determine size, shape and arrangement of bacteria

21. Gram staining Differential Staining • divides microorganisms into groups based on their staining properties • divides bacteria into two groups based on differences in cell wall structure

22. Differential Staining Figure 2.19

23. Electron Microscopy • beams of electrons are used to produce images • wavelength of electron beam is much shorter than light, resulting in much higher resolution

25. Newer Techniques in Microscopy • confocal scanning laser (CLSM) microscopy and scanning probe microscopy • have extremely high resolution

26. Confocal Microscopy • laser beam used to illuminate a variety of planes in the specimen • computer compiles images created from each point to generate a 3-dimensional image • used extensively to observe biofilms

27. Scanning Probe Microscopy • atomic force microscope • sharp probe moves over surface of specimen at constant distance • up and down movement of probe as it maintains constant distance is detected and used to create image

28. Chapter 3 Bacteria and Archaea CHAPTER GLOSSARY Lipopolysaccharides (LPSs) Nucleoid Peptidoglycan Periplasmic space Plasmid Porin proteins Sex pili S-layer Spirillum螺旋菌屬 Spirochete螺旋體 Bacillus Capsule Cell envelope Chemotaxis Coccus Endospore Fimbriae Fluid mosaic model Gas vacuole Glycocalyx Inclusions

29. Common features of bacterial and archaeal cell structure • prokaryotes differ from eukaryotes in size and simplicity • most lack internal membrane systems • prokaryotes are divided into Bacteria and Archaea • Bacteria are divided into 2 groups based on their Gram stain reaction

30. Size, Shape, and Arrangement • cocci (s., coccus) – spheres • diplococci (s., diplococcus) – pairs • streptococci – chains • staphylococci – grape-like clusters • tetrads – 4 cocci in a square • sarcinae – cubic configuration of 8 cocci • bacilli (s., bacillus) – rods • coccobacilli – very short rods • vibrios – resemble rods, comma shaped • spirilla (s., spirillum) – rigid helices • Spirochetes 螺旋體– flexible helices • Mycelium 菌絲– network of long, multinucleate filaments • Pleomorphic多形性– organisms that are variable in shape • Archaea • pleomorphic, branched, flat, square, other unique shapes

31. Figure 3.2

32. largest – • 50 μm in • diameter • smallest – • 0.3 μm in • diameter

33. Size – Shape Relationship • important for nutrient uptake • surface to volume ratio (S/V) • small size may be protective mechanism from predation掠食

35. Bacterial Cell Envelope • Plasma membrane • Cell wall • Layers outside the cell wall Functions of the plasma membrane • encompasses the cytoplasm • selectively permeable barrier • interacts with external environment • receptors for detection of and response to chemicals in surroundings • transport systems • metabolic processes

36. Fluid Mosaic Model of Membrane Structure • Lipid bilayerin which proteins float Membrane proteins • peripheral proteins • loosely associated with the membrane and easily removed • integral proteins • embedded within the membrane and not easily removed

37. The asymmetry of most membrane lipids • polar ends • interact with water • hydrophilic • nonpolar ends • insoluble in water • hydrophobic Bacterial Membranes • differ from eukaryotes in lacking sterols • do contain hopanoids, sterol-like molecules • a highly organized, asymmetric system which is also flexible and dynamic

38. The Bacteria Cell Wall • rigid structure that lies just outside the plasma membrane Functions of cell wall • provides characteristic shape to cell • protects the cell from osmotic lysis • may also contribute to pathogenicity • very few procaryotes lack cell walls

39. Bacterial Cell walls • bacteria are divided into two major groups based on the response to gram-stain procedure. • gram-positive bacteria stain purple • gram-negative bacteria stain pink • staining reaction due to cell wall structure

40. Peptidoglycan (肽聚糖)(Murein, 胞壁質) Structure • meshlike polymer of identical subunits forming long strands • two alternating sugars • N-acetylglucosamine (NAG) (N-乙酰葡萄糖胺) • N- acetylmuramic acid (NAM) (N-乙酰胞壁酸) • alternating D- and L- amino acids Diaminoacids present in peptidoglycan

41. peptidoglycan strands have a helical shape • peptidoglycan chains are crosslinked by peptides for strength G(-) G(+)

42. Gram-Positive Cell Walls • composed primarily of peptidoglycan • may also contain large amounts of teichoic acids (negatively charged) • help maintain cell envelop • protect from environmental substances • may bind to host cells • some gram-positive bacteria have layer of proteins on surface of peptidoglycan • teichoic acids • polymers of glycerol or ribitol (核糖醇) joined by phosphate groups Isolated gram+ cell wall

43. Periplasmic Space of Gram + bacteria • lies between plasma membrane and cell wall and is smaller than that of Gram - bacteria • periplasm has relatively few proteins • enzymes secreted by Gram + bacteria are called exoenzymes • aid in degradation of large nutrients

44. Gram-Negative Cell Walls • consist of a thin layer of peptidoglycan surrounded by an outer membrane • outer membrane composed of lipids, lipoproteins, and lipopolysaccharide (LPS) • no teichoic acids • peptidoglycan is ~5-10% of wall weight • periplasmic space differs from that in Gram + cells • may constitute 20-40% of cell volume

45. Lipopolysaccharides (LPSs) • consists of three parts • lipid A • core polysaccharide • O side chain (O antigen) Importance of LPS • may contribute to attachmentto surfaces and biofilm formation • creates apermeability barrier • contributes to negative charge on cell surface (core polysaccharide) • protection from host defenses (O antigen) • helps stabilizeouter membrane structure (lipid A) • can act as an endotoxin(lipid A)

46. Other Characteristics of the Outer Membrane • more permeable than plasma membrane due to presence of porin proteins and transporter proteins • porin proteins form channels through which small molecules (600-700 daltons) can pass

47. The Mechanism of Gram Staining • thought to involve shrinkage of the pores of the peptidoglycan layer of gram-positive cells • constriction prevents loss of crystal violet during decolorization step • thinner peptidoglycan layer and larger pores of gram-negative bacteria does not prevent loss of crystal violet

48. The Cell Wall and Osmotic Protection • osmotic lysis • can occur when cells are in hypotonicsolutions • movement of water into cell causes swelling and lysis due to osmotic pressure • cell wall protects against osmotic lysis • lysozymebreaks the bond between N-acetyl glucosamine and N-acetylmuramic acid • penicillin inhibits peptidoglycan synthesis • If cells are treated with either of the above they will lyse if they are in a hypotonic solution • protoplast – cell completely lacking cell wall • spheroplast – cell with some cell wall remaining

49. Capsules, Slime Layers, and S-Layers • capsules • usually composed of polysaccharides • well organized and not easily removed from cell • slime layers (黏液層) • similar to capsules except diffuse, unorganized and easily removed • slime may aid in motility • S-layers • regularly structured layers of protein or glycoprotein • In bacteria the S layer is external to the cell wall • common among Archaea, where they may be the only structure outside the plasma membrane S-layers

50. Functions of capsules, slime layers, and S-layers • protection from host defenses (e.g., phagocytosis) • protection from harsh environmental conditions (e.g., desiccation) • attachment to surfaces More functions… • protection from viral infection or predation by bacteria • protection from chemicals in environment (e.g., detergents) • facilitate motility of gliding bacteria • protection against osmotic stress