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Some Differential Characteristics of Procaryotes and Eukaryotes CHARACTERSISTIC CYTOLOGICAL FEATURES nucleoplasm (genophore, nucleotide separated from the cytoplasm by a unti-membrance system of(nuclear membrane) size of smallest dimension of cells (width or diameter): usually 0.2-2.0 um usually >2.0 um mitochondria present chloroplast present in phototroph vacuoles, if present, enclosed by unit membrane gas vacuoles presents golgi apparatus present lysosmes present microtubular systems present endoplasmic reticulum present ribosome location dispersed in the cytoplasm attached to an endoplasmic reticulum cytoplasmic streaming, pseudopodial movement, endocytosis, and exocytosis either nucleoplasm or cytoplasm flagella, if present diameter: 0.01-0.02um ca. 0.2um in cross section, have a characteristic " 9+2" arrangement of microtubules endospores present ANTIBOITIC SUSCEPTIBILITY Susceptible to: Penicillin, streptomycin, or other antibiotics specific for prokaryotes cycloheximde or other antibiotics specific for eukaryotes FEATURES BASED ON CHEMICAL ANALYSIS poly B hydroxbutyrate present(as a storage compound in cytoplasmic inclusions) teichoic acids present (in cell walls) polyunsaturated fatty acids possibly present (in membranes). branched-chain iso-or anteiso-fatty acids and cyclopropane fatty acids present(in membranes) PROCARYOTES EUCARYOTES - +b D -d + D + + + + + D D D + + D + = + + D - - D - D D D - D Rare Common Rare Common - CHARACTERSISTIC sterols present (in membranes) diaminopimelic acid present (in cell walls) muramic acid present (in cell walls) peptidoglycan (containing muramic acid) present in cell walls NUTTRITION nutrients acquired by cells as soluble small molecules; to serve as sources of nutrients, particulate matter or large molecules must first be hydrolyzed to small molecules by enzymes external to the plasma membrane METABOLIC FEATURES respiratory and photosynthetic functions and associated pigments and enzymes (e.g. chlorophylls, Cytochrome), if present, are associated with the plasma membrane or invaginations thereof chemolithotrophic type of metabolism occurs (inorganic compounds can be used as elctr4on donors by organisms that derive energy from chemical compounds) ability to fix N2 ability to carry out anoxygenic photosynthesis ENZYMES FEATURES type of super oxide dismutase: Cu-Zn type Mn and/or Fe type REPRODUCTIVE FEATURES cell division occurs by mitosis, and a micro tubular (spindle) system is present meiosis occurs mechanisms of gener transfer and recombination, if they occur, involve gametogenesis and zygote formation MOLECULAR BIOLIGICAL PROPTERTIES number of chromosomes present per nucleiod chromosomes circular chromosomes linear sedimentation constant of ribosome's 70S 80S sedimentation constants of ribosomal RNA 16S, 23S, 5S 18S, 28S, 5.85S, 5S PROCARYOTES EUCARYOTES -1 Common Dg h D h D - + D +1 - D - D + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + CHARACTERSISTIC MOLECULAR BIOLOGICAL PROPERTIES (CONTINUED) first amino acid to initiate a polypeptide chain during protein synthesis: Methionin N-Formymethionin messenger-RNA binding site at AUCACCUCC at 3 end of 16S or 18S ribosomal RNA PROCARYOTES EUCARYOTES - - +b - +b - a. Symbols: +, positive;-, negative; D, differs among organisms. b. A few bacteria (e.g., certain treponemes, mycoplasmas, haemobartonella) may have a width as small as 0.1 um; a few bacteria (e.g., Achromatium, Macromonas) may have a width greater than 10 um. c. Gas vacuoles are not bounded by a unit membrane. The vesicles composing the vacuoles can be caused to collapse by the sudden application of hydrostatic pressure- a feature essential to their identification. d. However, certain intracellular fibrils that may be microtubules have been reported in Spiroplasma, certain spirochetes, the cyanobacterium Anabaena, and in bacterial L forms... e. Bacterial ednospores are usually resistant to heat treatment of 80C or more for 10 min. however, some types of endospores may be killed by this heat treatment and may require testing at lower temperatures. f. Except in membranes of most mycoplasmas. g. Present in virtually all gram-negative eubacteria and in many gram-positive bacteria. h. Present in walled eubacteria except Chlamydia; absent in archaeiobacteria i. However in cyanoabacteria there may be j. With a few rare exceptions, such as certain phtobacteria. k. Except in mitochondria, in which the Mn type occurs. l. Except in mitochondria and chloroplast, which have 70S ribosomes. CHAPTER IV THE FOUR MAJOR CATEGORIES OF BACTERIA The four major (on a phenotypic basis) categories of bacteria are described briefly below, followed by tables of characteristics useful for differentiating some of these categories. 1. gram negative Eubacteria that have cell walls these are prokaryotes that have a complex (gram-negative type) cell-wall profile consisting of an outer membrane and an inner, thin peptidoglycan layer which contains muramic acid and is present in all but a few organisms that have lost this portion of wall) and a variable complement of other components outside or between these layers. They usually stain gram negative. Cell shapes may be spheres, ovals, straight or curved rods, helices, or filaments; some of these forms may be sheathed or capsulated. Reproduction is by binary fission, but some groups show budding, and a rare group (pleurocasples) shows multiple fission. Fruiting bodies and myxospores may be formed by the mycobacterium. Swimming motility, gliding motility, and nonmotility are complying observed. Members of the division may be phototrophic or no phototrophic (both lithotrophic and heterotrophic) bacteria and include aerobic, anaerobic, facultative) anaerobic, and microaerophilic species some members are obligate intracellular parasites. II gram positive eubacteria that have cell walls profile of the gram positive type; reaction with grams' stain generally, but not always, is positive. Cells may be spheres, rods, or filaments the rods and filaments may be none branching, but many show true branching. Cellular reproduction is generally by binary fission; some produces spores as restings forms endospores or spores on hyphae). They are not photosynthetic; generally, they are chemosynthetic heterotroph and include aerobic, anaerobic, facultative anaerobic, and Microaerophile species. The members of this division include simple asporogenous and asporogenous bacteria, as well as the actinomycetes and their relatives. IIIEubacteria lacking cell walls There are prokaryotes that lack cell walls (commonly called the mycoplasmas and including the class Mollicutes) and do not synthesize the precursors of peptidoglycan. They are enclosed by a unit membrane. The cells are highly Pleomorphic and range in size from large deformable vesicles to very small (0.2um), filterable elements. Filamentous forms with branching projections are common. Reproduction may be by budding, fragmentation and or binary fission some groups who a degree of regularity of form due to the placing of internal structures. Usually, they are nonmotile, but some species show a form of gliding motility. No resting forms are known. Cells stain gram negative. Most require complex media for growth (high-osmotic-pressure surroundings) and tend to penetrate the surface of solid media forming characteristic friend egg colonies. The organisms resemble the naked L-forms that can be generated from many species of bacteria (notably gram positive eubacterial) but differ in that the mycoplasmas are unable to revert and make cell wall. Most species are further distinguished by requiring both cholesterol and long-chain fatty acids for growth; unesterified cholesterol is a unique component of the membranes of both sterol requiring and nonrequiring species if present in the medium. The guanine plus cytosine content of ribosomal RNA is 4348 mol% 9lower than the 50-54 mol% of walled gram negative and gram positive eubacteria); the guanine plus cytosine content of the DNA is also comparatively low, 23-46 mol% and the genome size of the mycoplasmas is less than that the other prokaryotes at 0.5-1.0 x 10 Daltons. The mycoplasmas may be saprophytic, parasitic, or pathogenic, and the pathogens cause disease of animal’s plants, and tissue cultures. IV Archaeiobacteria The arch bacteria are predominantly terrestrial and aquatic microbes, occurring in anaerobic hyper saline or hydrothermally and geothermally heated environment; also, some occur symbionts in animal’s digestive tracts. They consist of aerobes anaerobes, and facultative anaerobes that grow chemolithoauthotrophically, orgnaorrophically, Archaeobacteria may be mesophiles or thermophiles, with some species growing even above 100C A unique biochemical feature of archaeiobacteria is the presence of glycerol isopranyl ether lipids. The lack of murein (peptidoglycan-containing muramic acid) in cell walls makes archaeiobacteria insensitive to B lactam antibiotics. The common arm of the RNAs contains pseudouridineor 1-methylpseudourindine instead of ribothymidine. The sequences of 5S, 16S, and 23S RNAs are very different from the corresponding ones in eubacteria and eucaryotes. Archaeiobacteria share some molecular features with eukaryotes: (a ) the elongation factor 2(EF-2) contains the amino acid diphthamide and is therefore ADP-ribosylable by diphtheria toxin, (b) amino acid sequences of the ribosomal A protein exhibit sequence homologies with the corresponding eukaryotic (L-7/L-12) protein, (c) the methionyl initiator RNA terminates with the base paid AU (f) the DNA dependent RNA polymerases re multi component enzymes and are insensitive to the antibiotics rifampicin and streptolydigin (g) like the DNA polymerases of eukaryotes are not inhibited by aphidicolin or burylphenyl dGTP, and (h) protein synthesis is inhibited by anisomycin but not by chloramphenical Autotrophic archaeiobacteria do not assimilate carbon dioxide via the Calvin cycle, in Methoanobacterium CO is fixed via an acetyl CoA pathway, whereas in acidianus and thermoproteus autotrophic CO is fixed via a reductive tricarboxlylic acid pathway. Fixation of N ahs been demonstrated by some methnogens. Gram stain results may be positive or negative within the same order because of very different types of cell envelopes. Gram positive species posses pscudomurein, methanochondroitin, and heteropolysaccharide cell walls gram negative cells have (glycol) protein ride cell walls, gram negative cells have the cells may have a diversity of shapes including spherical spiral plate or rod shaped unicellular and multicellular forms in filaments or aggregates also occur the diameter of anindividual cell may be 0.1-15 um, and the length of the filaments can be up to 200um. Multiplication is by binary fission, budding, constriction, fragmentation or unknown mechanism colors of cell masses may be red, purple, pink, orange brown, yellow green greenish black, gray, and white. The major groups of archaobacteria include (a) the methanogenic archaeiobacteria, (b) the archaeiobacteria sulfate reducers, (c) the extremely Halphilic archaeiobacteria, (d) the cell wall less archaeiobacteria, and (e) the extremely thermophilic metabolizes. CHARACTERSISTIC GENERAL MORPHOLOGIC AND METABOLIC FEATURES Strict anaerobes that form methane as the predominant metabolic end product from H2CO2, format, acetate, methanol, methylamine or H2 methanol. Cells exhibit a blue green epifluroescence when excited at 420mm Strict anaerobes that form H2S from sulfate by dissmilatory sulfate reduction. Extremely Thermophilic (growth up to 92 C) exhibit blue green epifluroescence when excited at 420mm Cells stain gram negative or gram positive and are aerobic or facultative anaerobic Chemoorganotroph. Rods and regular to highly irregular cells occur; cells require a high concentration of NaCl (1.5 More above) Neutrophilic or Alkaliphilic. Mesophilic or slightly Thermophilic (up to 55C) some species contain the red- purple photoactive pigment bacteriorhodopsin and are able to use light for ATP synthesis Thermoacidophilic, aerobic, facultative anaerobic or strictly anaerobic gram negative rods, filaments, or cocci. Optima growth temperature between 70C and 105C acidophilus and Neutrophilic autotrophic or heterotrophic most species are sulfur metabolizes PROCARYOTES ARCHAEOBACTERIA - - +b +b - - - - CELL WALLS(IF PRESENT) contain muramic acid ANTIBIOTIC SUSCEPTIBILITY susceptible to penicillin or other antibiotics that inhibit synthesis of muramic acid containing peptidoglycan LIPIDS membrane phospholipids consist of long chain alcohol(phytanois) that are either linked to glycerol to form C20 diphytanyl glycerol diethers or C40 dibiphytanyl digilycerol tetraethers long chain aliphatic fatty acids that are ester linked toglycerol pathway used in formation of lipids: mevalonate pathway malonate pathway MOLECULAR BIOLOGICAL FEATIURES ribothymine is present in the common arm of the RNA pseudouridine or 1methylpseudourindine is present in the common arm of the RNA CHARACTERSISTIC First amino acid to intiate a polypeptide chain during protein synthesis. Methionin N-F formylmethiorine aminoacyl stem of the initiator IRNA terminates with the base pair AU protein synthesis by ribosomes inhibited by anisomycin kanamycin chloramphenicol ADPRibosylation of the peptide elongation factor EF-2 is inhibited by diphtheria toxin Elongation factor 2(EF-2) contains the amino acid diphthamide. Some RNA genes contain introns. DNA-dependent RNA polymerases are multicomponent enzymes inhibited by rifampicin and streptolydigin replicating DNA polymerases are inhibited by aphidicolin or butylphenyl d GTP Symbols+, positive,-, negative D differs among organisms - - +b - +b - - - - - D -d D -d PROCARYOTES ARCHAEOBACTERIA - - +b - +b - D - D - - - -d - -d - Some of the characteristics differtiating eubacterial Major form eubacterial major coategory CHARACTERSISTIC MAJOR MAJOR CATEGORY 1 CATEGORYII CYTOLOGICAL FEATURES gram staining reaction an outer membrane is present in the cell wall in addition to the plasma cytoplasm membrane +b +b acid fast staining endospores present filamentous growth with hyphae that show true branching LOCOMOTION Gliding motility occurs. D D CHEMCIAL FEATURES percentage of the dry weight of the cell wall that is represent by lipid teichoic or lipteichoic acids present lipopolysaccharide (LPS) occurs ( in the outer membrane of the cell wall) -d -d 2-keto-3-deoxyoctonate (KDO) present percentage of the dry weight of the cell wall is represented by peptidoglycan mycolic acids present phosphatidylinositol mannosides present + + METABOLIC FEATURES energy derived by the oxidation of inorganic iron, sulfur or nitrogen compounds ENZYMIC FEATURES D D citrate synthesis: inhibited by reduced nicotinamide adenine dinulceotide (NADH) molecular weight ca 250, 000 D D ca 100,000 succinate thiokinases, molecular weight of 70,000-75,000 D D 140,000-150,000 a. b. c. d. e. f. g. Symbols: +, positive;-, negative, D, differs among organisms. B the staining reaction may not be conclusive. A few bacteria that have a gram positive type of cell wall profile and whose wall contain teichoic acids may stain gram negative, probably because of the thinness of the wall. For example, see the genus butyrivibrio. The staining reaction may not be conclusive. A few bacteria that have a gram-negative type of cell wall profile and whose walls contain lipopolysaccharide may stain gram positive for example, see the genus xanthobacter. An exception may be the genus coxeilla. Acid fast staining occurs in the genus mycobacterium and in some nocardia species. Endospores occur in the general bacillus, clostridium desulfotomaculum sporosarcina, and thermoactinomyces Except for mycobacterium corynebacteruim nocardia, and other genera whose walls contain mycolic acids. h. LPS consists of lipid A (a B-linked d-glucosamine disaccharide to which phosphate residues are linked at positions 1 and 4 and fatty heteropolysaccharide) and Oantigens9side chains that are polysaccharide composed of repeating unties) i. In many but not all gram negative bacteria, the core polysaccharide contains KDO which, if present, can serve as an indictor of the presence of LPS. j. Mycolic acids occur in corynebacterium nocardia, mycobacterium bacterionema, faenia, and rhodocccus. k. For example in bacteroides l. Known exception include acetobcter, thermos, and cyanoabacteria m. One know exception is thermos., Cytochrome one of various inron-prophyrins that can undergo reversible oxidation reduction and that serves as an electron carrier in an electron transport chain. Endospore a thick-walled spore formed in a bacterial cell. It is very resistant to being killed by heat and various other chemical and physical agents. Endospores are best verified by demonstrating that cultures survive a temperature of 70-80C for 10 min. Facultative anaerobe an organism that can grow well both in the absence of oxygen and in the presence of a level of oxygen equivalent to that in an air atmosphere (21%oxygen). Some are capable of throwing aerobically by respiring with oxygen and of growing be fermentation. Others have a strictly fermentative type of metabolism and respire with oxygen. anaerobically not Fermentation an energy yielding metabolic process in which electrons derived from an organic substrate are used ultimately to reduce an organic electron acceptor that is made by the cell itself. Neither an electron transport chain nor an exogenous terminal electron acceptor is involved. (Contrast with respiration) Flavin an enzymes that contains as tightly bound prosthetic groups either flavin mononucleotide (FMN) or flavin adenine dinucleotide (FAD) Fruiting body a specialized sore-producing structure that is composed of slime and bacterial cells, is often brightly colored, and may be visible to the naked eye. Formed by myxobacteria. Gas vacuole a cavity in the cytoplasm that contains gas of composition similar to that of the surrounding atmosphere. The boundary of the cavity is membrane consisting of protein subunits. Gas vacuoles appear as bring areas within a cell by passé contrast microscopy and are identified by their ability to collapse when the cells are subject to a sudden increase in hydrostatic pressure. Gliding motility a type of movement across surface that is exhibited by some bacteria devoid of flagella and incapable of swimming motility Gram variable some gram positive cell occur in an otherwise gram negative pure culture. Halphilic growing best at high Na C concentrations. Heterotroph growing in three dimensions and having one or more turns; corksxerw-shaped Hyphae the individual filaments or threads of mycelium Meanquinoes vitamins K. 2-methyl-3-all trans polyprenol-1, 4-naphatroquinones possessing side chains varying in length from C5 C65. Meanquinoes function as electron carriers Mesophilic growing best at a moderate temperature range (25-40C) Methanogenic producing methane Microaerophile an organism that is capable of oxygen dependent growth but cannot grow in the presence of a level of oxygen equivalent to that present in an air atmosphere (21%oxygen). Oxygen dependent growth occurs only at low oxygen level. In addition to being able to respire with oxygen, some Microaerophile may be capable of respiring anaerobically with electron acceptors other than oxygen. Mutualism a type of interaction in which two or more organisms living together benefit each other. Mycelium a mass or network of interwoven filaments (hyphae).