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:...
TRANSCRIPT
2
3
4
5
Chapter 1 The Evolution of Microorganisms and Microbiology
CHAPTER GLOSSARYArchaeaBacteriaEukaryaFungiGenomeGenomic analysisKoch’s postulatesMicrobiologyMicroorganismPrionsProkaryotic cellsProtistsSpontaneous generationViroidsVirusesVirusoids
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
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
The Importance of Microorganisms
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 prokaryotic cells lack a true membrane-delimited nucleus eukaryotic cells 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), Archaea and Eucarya (eucaryotes)
10
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
11
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.
12
Chapter 2
The Study of Microbial Structure: Microscopy and Specimen PreparationCHAPTER GLOSSARY
Atomic force microscopeBright-field microscopeConfocal scanning laser microscope (CSLM)Dark-field microscopeDifferential interference contrast (DIC) microscopyDifferential stainingFixationFluorescence microscopyGram stainNegative stainingParfocal
Phase-contrast microscopeRefractive indexResolutionScanning electron microscope (SEM)Simple stainingTransmission electron microscope (TEM)
13
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
14
Gram staining divides bacteria into two groups based on differences in cell wall
structure
Differential Staining divides microorganisms into groups based on their staining properties
15
Figure 2.19
Differential Staining
16
Chapter 3Bacteria and Archaea
CHAPTER GLOSSARY
BacillusCapsuleCell envelopeChemotaxisCoccusEndosporeFimbriaeFluid mosaic modelGas vacuoleGlycocalyxInclusions
Lipopolysaccharides (LPSs)NucleoidPeptidoglycanPeriplasmic spacePlasmidPorin proteinsSex piliS-layerSpirillum 螺旋菌屬Spirochete 螺旋體
17
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
Common features of bacterial and archaeal cell structure
18
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
19
Figure 3.2
20
•largest – 50 μm indiameter
• smallest – 0.3 μm in diameter
21
Size – Shape Relationship
important for nutrient uptake surface to volume ratio (S/V) small size may be protective mechanism from predation 掠食
S= 4πr2
V= 4/3 πr3
S/V = 3/r
22
23
Bacterial Cell Envelope
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
Plasma membrane
Cell wall
Layers outside the cell wall
24
Fluid Mosaic Model of Membrane Structure Lipid bilayer in 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
25
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
26
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
27
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 structureM: peptidoglycan or murein layerP: periplasmic spacePM: plasma membrane
28
Peptidoglycan ( 肽聚糖 )(Murein, 胞壁質 ) Structure
Diaminoacids present in peptidoglycan
meshlike polymer of identical subunits forming long strandstwo 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
L-lysine meso-diaminopimelic acid
29
peptidoglycan strands have a helical shape
peptidoglycan chains are crosslinked by peptides for strength
G(-)
G(+)
30
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
Isolated gram+ cell wall
teichoic acids• polymers of glycerol or ribitol ( 核糖醇 ) joined by phosphate groups
31
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
32
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
33
Lipopolysaccharides (LPSs) consists of three parts
lipid A core polysaccharide O side chain (O antigen)
Importance of LPS may contribute to attachment to surfaces and biofilm formation creates a permeability barrier contributes to negative charge on cell surface (core polysaccharide) protection from host defenses (O antigen) helps stabilize outer membrane structure (lipid A) can act as an endotoxin (lipid A)
34
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
35
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
36
lysozyme breaks 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
The Cell Wall and Osmotic Protection osmotic lysis
can occur when cells are in hypotonic solutions movement of water into cell causes swelling and lysis due to
osmotic pressure cell wall protects against osmotic lysis
37
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
38
Functions of capsules, slime layers, and S-layers protection from host defenses (e.g., phagocytosis) protection from harsh environmental conditions (e.g., desiccation 乾燥 ) 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
39
Glycocalyx ( 醣外被 ) network of polysaccharides extending from the surface of the
cell a capsule or slime layer composed of polysaccharides can also
be referred to as a glycocalyx
40
Archaeal membranes composed of unique lipids
isoprene units (five carbon, branched) ether linkages rather than ester linkages to glycerol
some have a monolayer structure instead of a bilayer structure
41
Archaeal Cell Walls Differ from Bacterial Cell Walls
lack peptidoglycan most common cell wall is S layer may have protein sheath external to S layer S layer may be outside membrane and separated by
pseudomurein pseudomurein may be outermost layer – similar to gram-
positive microorganisms
42
Archaeal cell walls lack peptidoglycan cell wall varies from species to species but usually consists of
complex heteropolysaccharides Methanogens have walls containing pseudomurein
Cell envelopes of Archaea
pseudomurein
43
Bacterial and Archaeal Cytoplasmic Structures
Cytoskeleton Intracytoplasmic membranes Inclusions Ribosomes Nucleoid and plasmids
44
Cytoplasmic Matrix
substance in which nucleoid, ribosomes and inclusion bodies are suspended
lacks organelles bound by unit membranes composed largely of water is a major part of the protoplasm (the plasma membrane and
everything within)
45
The Procaryotic Cytoskeleton homologs of all 3 eucaryotic cytoskeletal elements have recently
been identified in Bacteria and one has been found in Archaea functions include roles in cell division, protein localization and
determination of cell shape
46
FtsZ – many bacteria and archaea forms ring during septum formation in cell division
MreB – many rods, some archaea maintains shape by positioning peptidoglycan synthesis machinery
CreS – rare, maintains curve shape
Tubulin
Actin
Intermediate filament
47
Intracytoplasmic Membranes
plasma membrane infoldings observed in many photosynthetic bacteria
analogous to thylakoids of chloroplasts reactions centers for ATP formation
observed in many bacteria with high respiratory activity
anammoxosome in Planctomycetes organelle – site of anaerobic ammonia oxidation
48
Inclusions granules of organic or inorganic material that are stockpiled by the
cell for future use some are enclosed by a single-layered membrane
membranes vary in composition some made of proteins; others contain lipids
Organic inclusion bodies glycogen
polymer of glucose units poly-β-hydroxybutyrate (PHB)
polymers of β-hydroxybutyrate cyanophycin granules
large polypeptides containing about equal quantities of arginine and aspartic acid
carboxysomes contain the enzyme ribulose-1,5,-bisphosphate carboxylase
(Rubisco), enzyme used for CO2 fixation
49
gas vacuoles found in cyanobacteria and some other aquatic procaryotes provide buoyancy aggregates of hollow cylindrical structures called gas vesicles
50
Inorganic inclusion bodies polyphosphate granules
also called volutin granules and metachromatic granules linear polymers of phosphates
sulfur granules Magnetosomes Fe3O4
found in aquatic bacteria magnetite particles for orientation in Earth’s magnetic field cytoskeletal protein MamK helps form magnetosome chain
51
Ribosomes complex structures consisting of protein and RNA sites of protein synthesis smaller than eucaryotic ribosomes
procaryotic ribosomes 70S eucaryotic ribosomes 80S
S = Svedburg unit
52
•bacterial and archaeal ribosomal RNA
•16S small subunit
•23S and 5S in large subunit
•archaea has additional 5.8S (also seen in eukaryotic large subunit)
•proteins vary
•archaea more similar to eukarya than to bacteria
53
The Nucleoid ( 類核體 ) irregularly shaped region location of chromosome
usually 1/cell not membrane-bound
Procaryotic chromosomes are located in the nucleoid, an area in the cytoplasm
54
The procaryotic chromosome
a closed circular, double-stranded DNA molecule looped and coiled extensively nucleoid proteins probably aid in folding
nucleoid proteins differ from histones
Plasmids usually small, closed circular DNA molecules exist and replicate independently of chromosome have relatively few genes present genes on plasmids are not essential to host but may confer
selective advantage (e.g., drug resistance)
curing is the loss of a plasmid classification of plasmids based on mode of existence, spread, and
function
55
Camphor 樟腦Toluene 甲苯
56
Functionprotection, attachment to surfaces, horizontal gene transfer, cell movement
External Structures
57
Pili and Fimbriae
fimbriae (s., fimbria) short, thin, hairlike, proteinaceous appendages
up to 1,000/cell mediate attachment to surfaces some (type IV fimbriae) required for twitching motility or
gliding motility that occurs in some bacteria sex pili (s., pilus)
similar to fimbriae except longer, thicker, and less numerous (1-10/cell)
required for mating
58
Patterns of Flagella Distribution monotrichous – one flagellum polar flagellum – flagellum at end of cell amphitrichous – one flagellum at each end of cell lophotrichous – cluster of flagella at one or both ends peritrichous – spread over entire surface of cell
extends from cell surface to the tip hollow, rigid cylinder composed of the protein flagellin some procaryotes have a sheath around filament
filament
59
Flagellar Ultrastructure
The Hook and Basal Body hook
links filament to basal body basal body
series of rings that drive flagellar motor
60
Flagellar Synthesis an example of self-assembly complex process involving many genes and gene products new molecules of flagellin are transported through the hollow
filament growth is from tip, not base
61
Differences of Archaeal Flagella flagella thinner more than one type of flagellin protein flagellum are not hollow hook and basal body difficult to distinguish more related to Type IV secretions systems growth occurs at the base, not the end
62
MotilityFlagellar movementSpirochete motilityTwitching motilityGliding motility
Bacteria and Archaea have directed movement
chemotaxis
move toward chemical attractants such as nutrients,
away from harmful substances
move in response to temperature, light, oxygen, osmotic
pressure, and gravity
63
Bacterial Flagellar Movement flagellum rotates like a propeller
very rapid rotation up to 1100 revolutions/sec in general, counterclockwise (CCW) rotation causes forward motion (run) in general, clockwise rotation (CW) disrupts run causing cell to stop and tumble ( 打滾 )
64
Mechanism of Flagellar Movement flagellum is 2 part motor producing torque ( 項鍊 , 轉矩 ) rotor ( 轉子 )
C (FliG protein) ring and MS ring turn and interact with stator stator ( 發電機的定子；定片 )- Mot A and Mot B proteins
form channel through plasma membrane protons move through Mot A and Mot B channels and produce energy through proton motive force torque powers rotation of the basal body and filament
65
Spirochete ( 螺旋體 ) Motility
multiple flagella form axial fibril which winds around the cell
flagella remain in periplasmic space inside outer sheath
corkscrew shape exhibits flexing and spinning movements
66
Twitching ( 抽動 ) and Gliding ( 滑行 ) Motility
may involve Type IV pili and slime twitching
pili at ends of cell short, intermittent, jerky motionscells are in contact with each other and surface
gliding smooth movements
67
Chemotaxis ( 化學趨化作用 ) movement towards a chemical attractant or away from a chemical
repellent concentrations of chemical attractants and chemical repellents
detected by chemoreceptors on surfaces of cells
68
Chemotaxis Towards Attractant
in presence of attractant (b) tumbling frequency is reduced and runs in direction of attractant are longer
69
The Bacterial Endospore formed by some bacteria dormant resistant to numerous environmental conditions
heat radiation chemicals desiccation 乾燥
Endospore Structure
spore surrounded by thin covering called exosporium
thick layers of protein form the spore coat
cortex 皮層 , beneath the coat, thick peptidoglycan
core has nucleoid and ribosomes
70
What makes an endospore so resistant?
calcium (complexed with dipicolinic acid) small, acid-soluble, DNA-binding proteins (SASPs) dehydrated core spore coat and exosporium protect
71
Sporogenesis Also called endospore formation or sporulation normally commences when growth ceases because of lack of
nutrients complex multistage process
72
Germination-Transformation of dormant spores into active vegetative cells
activation prepares spores for germination often results from treatments like heating
germination spore swelling rupture of absorption of spore coat loss of resistance increased metabolic activity
outgrowth emergence of vegetative cell