1、Introduction of Microbiology, Bacterial Structure,Pin Lin (凌 斌), Ph.D. Departg ment of Microbiology 6th edition) 2. 醫用微生物學 (商惠芳 審閱, 6th edition),課程要點 (Outline),Introduction of Medical MicrobiologyBacterial ClassificationBacterial Structure,History of Microbiology,In 1674 Dutuch biologist Leeuwenhoek
2、 discovered a world of tiny “animalcules” (microbes) by microscope. In 1840 German Pathologist Friedrich Henle proposed “Germ Theory” for proving microorganisms causing diseases. 3. Robert Koch & Louis Pasteur confirmed this theory in late 1870 -1880s. 4. A. Fleming discovered that the mold Penicill
3、ium prevented the multiplication of staphyloocci.= The first antibiotic, Penicillin, was identified.,History of Microbiology-II,In 1946, American microbiologist John Enders develop the virus culture for vaccine development.,Four Groups of Microbes,(Prokaryotic),(eukaryotic),(eukaryotic),研究微生物 了解它們所造
4、成的疾病發展出控制它們的方法,細菌分類-I (Bacterial Classification),細菌以外形特徵分類 (Phenotypic classification): 顯微形態學 (Microscopic morphology) 巨觀形態學 (Macroscopic morphology) 生物分型 (Biotyping) 血清分型 (Serotyping) 抗生素圖譜 (Antibiogram patterns) 噬菌體分型 (Phage typing),細菌分類-II (Bacterial Classification),細菌以成份分類 (Analytic):細胞壁脂肪酸分析 (C
5、ell wall fatty-acid analysis) 全細胞脂質分析 (Whole cell lipid analysis) 全細胞蛋白質分析 (Whole cell protein analysis) 酵素電泳分型 (Multifocus locus enzyme electrophoresis),細菌以基因型分類 (Genotypic):鳥糞螵呤和胞嘧啶比率分析 (Guanine plus cytosine ratio) DNA 雜交法 (DNA hybridization) 核酸序列分析法 (Nucleic acid analysis) 質體分析法 (Plasmid analysi
6、s) 染色體DNA片段分析法 (Chromosomal DNA fragment analysis),細菌分類-III (Bacterial Classification),Differences Among Prokaryotes: Bacteria have different shapes. Coccus: spherical bacterium staphylococcus; grapelike clusters, diplococcus; two cells together Rod-shaped bacterium: BacillusEscherichia coli : bacil
7、lus. Spirillum: Snakelike treponeme some bacteria (螺旋菌屬),原核細胞 (Prokaryote)的特徵,真核細胞 (Eukaryote)的特徵,Eukaryote vs. Prokaryote,Bacterial Ultra-structure,Gram-positive vs. Gram-negative bacteria,1. Gram-positive vs Gram-negative bacteria:- Similar Internal structures- Different External structures. 2. Th
8、e cytoplasm of the bacteria contains - DNA chromosome, mRNA, ribosomes, proteins, and metabolites. 3. The bacterial chromosome - A single copy (haploid) and double-stranded circle in a discrete area known as the nucleoid. - No histones,Cytoplasmic Structures-I,4. Plasmids (質體) : - Smaller, circular,
9、 extrachromosomal DNAs - Most commonly found in gram-negative bacteria- Not essential for cellular survival - Provide a selective advantage: many confer resistance to one or moreantibiotics.,Cytoplasmic Structures-II,The cytoplasmic membrane - A “lipid bilayer structure” similar to that of the eukar
10、yotic membranes - Contains no steroids (e.g., cholesterol); mycoplasmas are the exception. Involves in electron transport and energy production, which are normally achieved in mitochondria in eukaryotes.,Cytoplasmic Membrane-I,Contains transport proteins = exchange metabolites ion pumps = a membrane
11、 potentialMesosome- A coiled cytoplasmic membrane - Acts as an anchor to bind and pull apart daughter chromosomes during cell division.,Cytoplasmic Membrane-II,Bacterial Cytoplasmic Membrane,ATP production machinery,1. The structure components and functions of the cell wall distinguish gram-positive
12、 from gram-negative bacteria. (A). Gram positive bacteria:(1) Peptidoglycan (murein, mucopeptide)(2) Teichoic acid（胞壁酸) & Lipoteichoic acid(3) Polysaccharides,Cell Wall,革蘭氏陽性菌細胞壁 (Gram-positive bacterial cell wall),Essential for the structure, for replication, and for survival in the hostile conditi
13、ons.Interfere with phagocytosis and has pyrogenic activity (induces fever). Degraded by lysozyme, an enzyme in human tears and mucus,Functions of Peptidoglycan,1. Water-soluble polymers, containing ribitol or glycerol residues joined through phosphodiester linkages.2. Constitute major surface Ag of
14、those gram-positive species = Bacterial Serotyping3. Promote attachment to other bacteria as well as to specific receptors on mammalian cell surfaces (adherence).Important factors in virulence, initiate endotoxic-like activities.,Teichoic & Lipoteichoic acid,Backbone: - N-acetylglucosamine & N-acety
15、lmuramic acid- The backbone is the same in all bacterial species.Tetrapeptide side chain attach to N-Acetylmuramic acid.,Peptidoglycan Synthesis,肽聚糖合成-I (Peptidoglycan Synthesis-I),Peptidoglycan A major component of cell wall Forms a “Meshlike layer” consisting: a polysaccharide polymer cross-linked
16、 by Peptide bonds Cross-linking reaction is mediated by:- Transpeptidases- DD-carboxypeptidases- Targets of Penicillin,肽聚糖合成-II (Peptidoglycan Synthesis-II),胜肽聚糖合成-III (Peptidoglycan synthesis-III),革蘭氏染色法 (Gram stain) Gram stain is a powerful, easy test that allows clinicians to distinguish between
17、the two major classes of bacteria and to initiate therapy. Bacteria heat-fixed stained with Crystal violet this stain is precipitated with Grams iodine washing with the acetone- or alcohol-based decolorizer A counterstain, safranin, red Gram-positive bacteria, Purple, the stain gets trapped in a thi
18、ck, cross-linked, meshlike structure.,革蘭氏染色法 (Gram stain),革蘭氏陰性菌細胞壁 (Gram-negative bacterial cell wall),More complex than gram-positive cell walls.Consists three major parts. (1) Outer membrane（外膜)- -Unique(2) Periplasmic space（細胞質外腔）(3) Cytoplasmic membrane3. Major Components - Lipopolysaccharide (
19、LPS) (Endotoxin)- Lipoprotein,革蘭氏陰性菌細胞壁 (Gram-negative bacterial cell wall),Unique to Gram-negative bacteria.- An “asymmetric bilayer” structure - different from any other biologic membrane in the structure of the outer leaflet of the membrane. Maintains the bacterial structure a permeability barrie
20、r to large molecules (e.g., lysozyme) and hydrophobic molecules. 3. Provides protection from adverse environmental conditions such as the digestive system of the host (important for Enterobacteriaceae organisms).,Gram (-) bacteria: Outer membrane,5. The outer membrane is held together by divalent ca
21、tion陽離子 (Mg+2 and Ca+2) linkages between phosphates on LPS molecules and hydrophobic interactions between the LPS and proteins. 6. These interactions produce a stiff (硬的), strong membrane that can be disrupted by antibiotics (e.g., polymyxin) or by the removal of Mg+2 and Ca+2 ions (using ion chelat
22、or, eg. EDTA).,Gram (-) bacteria: Outer membrane,Lipopolysaccharide (LPS) (Endotoxin),1. O antigen 2. Core polysaccharide 3. Lipid A-active component of LPS,1. Induce innate immune response 2. Activate macrophage to secrete cytokines like IL-1, IL-6 & TNF-a,1. The outer membrane is connected to the
23、cytoplasmic membrane at adhesion sites and is tied to the peptidoglycan by lipoprotein2. The lipoprotein is covalently attached to the peptidoglycan and is anchored in the outer membrane.,Lipoprotein,Table 2-4,1. Capsules 夾膜a. Some bacteria are closely surrounded by loose polysaccharide or protein l
24、ayers called capsules b. Capsules and slimes are unnecessary for the growth of bacteria but are important for survival in the host.c. The capsule is poorly antigenic and antiphagocytic and is a major virulence factor (e.g., Streptococcus pneumoniae). d. Bacillus anthracis炭疽桿菌: polypeptide,External S
25、tructures,Ropelike (繩索式) propellers composed of helically coiled protein subunits (flagellin) - Anchored in the bacterial membranes through hook and basal body structures.- Driven by membrane potential.2. Flagella provide motility for bacteria, allowing the cell to swim (chemotaxis) toward food and
26、away from poisons. 3. Express Antigenic & strain determinants.4. Four types of arrangementa. Monotrichous: single polar flagellum b. Amphitrichous: flagella at both poles.c. Lophotrichous: tuft of polar flagellad. Peritrichous: Flagella distributed over the entire cell.,Flagella 鞭毛,1. Pili are hairl
27、ike structures on the outside of bacteria; they are composed of protein subunits (pilin).Fimbriae can be morphologically distinguished from flagella because they are smaller in diameter (3 to 8 nm versus 15 to 20 nm) and usually are not coiled in structure.They may be as long as 15 to 20 m, or many
28、times the length of the cell. 4. Fimbriae promote adherence to other bacteria or to the host (alternative names are adhesins, lectins凝集素, evasins逃避素, and aggressins攻擊素).,Fimbriae (pili): Latin for “fringe”,5. As an adherence factor (adhesin黏附素), flmbriae are an important virulence factor for E. coli
29、 colonization and infection of the urinary tract, for Neisseria gonorrhoeae and other bacteria. 6. The tips of the fimbriae may contain proteins (lectins) that bind to specific sugars (e.g., mannose).7. F pili (sex pili) promote the transfer of large segments of bacterial chromosomes between bacteri
30、a. These pili are encoded by plasmid (F).,Fimbriae (pili): Latin for “fringe”,1. Some gram-positive bacteria, but never gram-negative such as : Bacillus & Clostridium 梭狀菌屬 2. Under harsh (惡劣的) environmental conditions, such as the loss of a nutritional requirement, these bacteria can convert from a
31、vegetative state （生長狀態）to a dormant state（冬眠）, or spore. 3. The location of the spore within a cell is a characteristic of the bacteria and can assist in identification of the bacterium.,Spores (芽胞)-I,4. Dehydrated, multishelled structure that protects and allows the bacteria to exist in “suspended
32、animation ”. 5. It contains (a) a complete copy of the chromosome;(b) the bare minimum concentrations of essential proteins and ribosomes; (c) High concentration of Ca2+ chelate of DPA (Ca-DPA, dipicolinic acid)(砒啶甲酸) . = DPA appears to be important in spore core dehydration and concomitant spore he
33、at resistance.6. The structure of the spore protects the genomic DNA from desiccation, intense heat, radiation, and attack by most enzymes and chemical agents.,Spores (芽胞)-II,7. Depletion of specific nutrients (e.g., alanine) from the growth medium triggers a cascade of genetic events (comparable to
34、 differentiation) leading to the production of spore.8. Spore mRNA are transcribed and other mRNA are turned off. Dipicolinic acid(DPA) is produced.9. Spore structure:Core: one copy of DNA and cytoplasmic contentsInner membrane and Spore wallCortex: peptidoglycan layerCoat: Keratine-like protein whi
35、ch protect the spore.Exosporium:外胞壁,Spores (芽胞)-III,Spore structure: Core: one copy of DNA cytoplasmic contents Inner membrane Spore wall Cortex: peptidoglycan layer Coat: Keratine-like protein which protect the spore. Exosporium:外胞壁,Thank You & The End,10. Germination:The germination of spores into
36、 the vegetative is stimulated by disruption of the outer coat by stress, pH, heat, or another stressor and requires water and a triggering nutrient (e.g., alanine). 11. The process takes about 90 minutes. 12. Once the germination process has begun, the spore will take up water, swell, shed its coats
37、 (脫除外套) , and produce one new vegetative cell identical to the original vegetative cell, thus completing the entire cycle. 13. Once germination has begun and the spore coat has been compromised (變弱), the spore is weakened and can be inactivated like other bacteria.,Another factor implicated in spore
38、 resistance properties and germination is the small molecule pyridine-2,6-dicarboxylic acid (dipicolinic acid DPA). The Ca2+ chelate of DPA (Ca-DPA) is a major constituent of the dormant spore core, accounting for approximately 10% of total spore dry weight (14, 15). The operon encoding the A and B
39、subunits of DPA synthetase (called spoVFAB or dpaAB) is expressed as part of the E K regulon in the mother cell compartment (1). DPA is synthesized in the mother cell and subsequently transported into the forespore by a currently unknown mechanism (1). DPA appears to be important in spore core dehydration and concomitant spore heat resistance, as spores of B. subtilis mutants lacking DPA due to null mutations in dpaAB have a lower core wet density and are sensitive to wet heat (19).,
