Fundamentals of Microbiology

EVA VLKOV
FANFR ground floor 29 (consulting hours: Tuesday 1 -3 p.m.) or lab 2 nd floor 230 vlkova@af.czu.cz

Fundamentals of Microbiology

12 lectures, FANFR 220 EXAM: written report on selected microbiological theme 10 minutes presentation on 5 th January 2012 written test

Literature to study

Prescott et al.: Microbiology, Wm. C. Brown Publishers, 1996 any other textbook of fundamentals of microbiology or general microbiology http://kmvd.agrobiologie.cz/index.php?obsah =skripta

Lectures topics

1. Introduction, history of microbiology 2. Viruses 3. Bacteria, prokaryotic cell structure and function 4. Fungi, eukaryotic cell structure and function 5. Microbial nutrition and growth 6. Control of microorganisms by physical and chemical agents 7. Metabolism and enzymes 8. Generation of energy, respiration 9. Fermentations, catabolism of carbohydrates and polymers, carbon cycle 10. Lipid and protein catabolism, biosynthesis 11. Nitrogen, phosphorus, and sulphur cycles 12. Microbial genetic

Written report
5 10 pages literature overview minimum 5 literature sources (textbooks, research papers, http://portal.isiknowledge.com, http://portal.isiknowledge.com)

Content:
introduction main part conclusions list of references (enclosures)

Fundamentals of Microbiology

24 hours of laboratory exercises FA N F R 2 2 0 C R E D I T: 2 written protocols on chosen laboratory t r a i n i n g t h e m e ( e . g . AT B t e s t , i d e n t i f i c a t i o n o f b a c t e r i a , w a t e r e x a m i n a t i o n ) , t i l l 1 5 th D e c e m b e r 2 0 11 ( c o n t e n t : p r i n c i p l e , m e t h o d o l o g y, r e s u l t s , c o n c l u s i o n s ) attendance on trainings

Microbiology

is defined as the science studying organisms or agents which cannot be seen by eye and must be examined by microscope (objects less than about 1 mm) its subjects are viruses, bacteria, many algae and fungi, and protozoa can be divided into virology, bacteriology, algeology, mycology etc. or on general, systematic, applied microbiology

Microorganisms

are necessary for the production of bread, cheese, beer, antibiotics, vaccines, vitamins, enzymes... they make possible the cycles of carbon, oxygen, nitrogen, sulphur.... they also caused many diseases

History of Microbiology
Fracastorius
medical doctor 1546 declared, that small nonvisible organisms exist and are responsible for diseases

History of Microbiology
1665 1670 microbiology as a science starts construction of first microscopes by Robert Hooke and Antony van Leeuwenhoek
described cocci and rods, blood cells and protozoa

Louis Pasteur (1822-1895)

discovered fermentations discovered anaerobic life sterilization, pasteuratization vaccination (smallpox) isolation and identification of causers of some infection diseases

Robert Koch (1843-1910)

solidifying of liquid cultivation media (gelatine, agar) construction of Petri dish new cultivation media colouring of bacteria by aniline colours isolation and identification of causers of cholera, anthrax and tuberculosis

Kochs postulates
the microorganism must be present in all ill objects and absent in all healthy objects the causer of disease must be isolated and grown in pure culture when the isolated microorganism is inoculated into healthy host, the same disease will developed the same microorganism must be isolated again from the host

Some important historical facts

1786 Muller first classification of microorganisms 1867 Lister antiseptic surgery 1884 Metchnikoff phagocytosis 1884 Gram Gram colouring 1892 Ivanowsky viruses 1907 Metchnikoff probiotics 1923 first edition of Bergeys manual 1928 Fleming penicillin 1937 procaryotes x eucaryotes 1953 Watson and Crick structure of DNA 1983 HIV identification 1984 PCR

Five kingdom system

1. MONERA = PROCARYOTAE SUBCELLULATA Vira viruses PROTOCELLULATA Bacteria bacteria Cyanophyta cyanophytes 2. PROTISTA eucaryotic cell - algae, protozoa, some simple fungi 3. FUNGI fungi and mashrooms 4. PLANTAE plants 5. ANIMALIA animals

Main differences between microorganisms

criterion viruses
no
no

bacteria
procaryotic
yes

fungi
eucaryotic
yes

protozoa
eucaryotic
no

cell type
cell wall size

20-400 nm

1-2 (10) m

5-10 m

50-300 m

Differences between procaryotic and eucaryotic cell

criterion
nucleus membrane size usually >2 m mitochondria endoplasmic reticulum (ER) placing of ribosomes chromosomes procaryotic cell free in cytoplasmic matrix circular eucaryotic cell + + + +

on ER linear

Viruses

General properties of viruses

simple acellular organization virion = complete virus particle one or more molecules of DNA or RNA (absence of both in the same virion) enclosed in protein capside, sometimes other layers (carbohydrates, lipids, additional proteins) exist in two phases: extracellular and intracellular obligatory intracellular parasites (intracellular phase, replicating nucleic acid, induce host metabolism to synthesize virion components) inability to reproduce independently on the host cells few or no enzymes (extracellular phase) without own metabolism

Virion
size: 10 300 (400) nm nucleic acid in central part DNA or RNA, single- or double-stranded, linear or circular capsid = protein coat composed from capsomers morphological types of capsides: helical and icosahedral

Virion structure

Size and morphology of viruses

Virus taxonomy
lack of knowledge of viruses origin and evolution viruses are separated into large groups based on their host preferences: animal and plant viruses, bacteriophages divided into 73 families by: a) nucleic acid type b) nucleic acid strandedness c) presence or absence of an envelope

Bacteriophage structure

Bacteriophage adsorption

Animal virus entry into the host cell

Lysogenic and lytic cycle of bacteriophage

Lytic cycle of bacteriophage

Bacteria

Differences between procaryotic and eucaryotic cell

criterion
cell wall size usually >2 m mitochondria endoplasmic reticulum (ER) placing of ribosomes chromosomes procaryotic cell free in cytoplasmic matrix circular eucaryotic cell + + + +

on ER linear

Bacterial size and shape

cocci roughly spherical cells, 1 m rods (bacilli) rod shape, 1 x 2-4 (10) m actinomycetes filamentous bacteria, filaments are 1 m width and several mm long

Morphology of bactrerial cell

G+ cell wall structure

G- cell wall structure

Glycocalyx

o adherence, enzyme storage o polysaccharides filaments

bacterial cell

Pili and Fimbriae

o attachment to surfaces o sex-pili conjugation o protein filaments

Flagella

o movement, antigens

o o o o o

proteins (flagellin) monotricha lofotricha amfitricha peritricha

Plasma membrane
o o o o o o o contain proteins and phospholipids lipids forms bilayer, hydrophilic x hydrophobic ends thin structure, thick 5-10 nm peripheral x integral proteins selectively permeable barrier nutrient and water transport metabolic processes

Nutrient transport
o main role play cell wall and plasma membrane

Passive (simple) diffusion

o concentration gradient o no energy consumption o ions, glycerol, O2, CO2

Osmosis

Facilitated diffusion
o o o o carriers proteins permeases by concentration gradient no energy use limited in procaryotes, typical for eucaryotes

Active transport

o against concentration gradient o use of energy o membrane proteins

Group translocation

o against concentration gradient o use of energy o membrane proteins o molecules are modified during the transport o typical for eucaryotes

Mesosome

o internal membrane system o invagination of plasma membrane o function is unknown o involved in cell wall formation during division o chromosome replication o energy metabolism o artefacts generated during the fixation for electron microscopy

Cytoplasmic matrix
o o o o o protoplast inclusion bodies granules enzymes ribosomes (synthesizing proteins) o nucleoid (DNA and associated proteins)

Nucleoid

o o o o o

one circle chromosome single- or double-stranded DNA about 3 500 genes no membrane enzymes

DNA replication

Plasmids
o circle double-stranded DNA o can exist and replicate independently of the chromosome or may be integrated with it o are not required, genes that give advatages

Ribosomes

o protein synthesis o rRNA, mRNA, tRNA o proteins

Endospores

o o o o o

resistant dormant structure no reproduction Clostridium, Bacillus viable for over 500 years multilayered (central core, membrane, cortex, coat, exosporium) o low water content, metabolism and ribosome number reduction

Bacteria identification
Classification = arrangement of organisms into groups or taxa Identification process of determining that a particular isolate belongs to recognized taxon Nomenclature names for taxonomic groups Basic taxon = species (binomial system) Species = set of cells with the same characteristics Strain = set of cells which grew from one cell Classification systems: phylogenetic numerical taxonomy phenetic

An example of taxonomic ranks and names

Rank Kingdom Phytum Class Order Example Procaryotae Actinobacteria Actinobacteria Bifidobacteridae

Family
Genus Species Subspecies

Bifidobacteriales
Bifidobacterium Bifidobacterium animalis Bifidobacterium animalis ssp. lactis

Morphological characteristics

o o o o o

cell shape cell size staining behavior cilia and flagella endospore shape and location

Cultivation characteristics

o morphology of colonies o colour of colonies o growth in liquid media

Physiological characteristics
o o o o o o growth temperature optimum and range osmotic tolerance oxygen relationships pH optimum and growth rang salt requirements and tolerance sensitivity to metabolic inhibitors and antibiotics

Metabolic characteristics
o o o o o o carbon and nitrogen sources energy sources fermentation products mechanisms of energy conversion secondary metabolites formed storage inclusions

Molecular characteristics

o o o o o

comparison of proteins nucleic acid base composition G+C content nucleid acid hybridization nucleic acid sequencing polymerase chain reaction

Micromycetes
Yeasts and Moulds

Micromycetes - introduction
eucaryotic, filamentous or holocarpic haploid, except zygotes hyphae with or without cross walls (septa) diameter of hyphae 5-10 m, length from m to hundreds mm mycelium network of hyphae cell wall with chitin or cellulose mono-, di-, polycaryotic structure

Kingdom: Mycota (Fungi)

Division: Eumycota (True fungi) Subdivision: Zygomycotina Genus: Mucor, Rhizopus Subdivision: Ascomycotina Genus: Aspergillus, Penicillium,

Saccharomyces
Subdivision: Deuteromycotina (Fungi imperfecti) Genus: Candida, Monilia

Micromycetes
Yeasts and Moulds

Scheme of yeast cell

Life cycle of yeast Saccharomyces cerevisiae

Reproduction of yeasts: buding(vegetative) ascospores (sexual spores)

Cell-division cycle in a yeast Saccharomyces cerevisiae

Buding and Pseudomycelia

Important yeast genera

Saccharomyces ethanol fermentation, vitamin
production, bakery yeasts, food spoilage (honey, jam, ketchup)

Kluyveromyces milk products (kefir), feed

mixtures, spoilage of cheeses

Candida some are pathogenic, food spoilage

(beer, milk products, meet, vegetable), feed mixtures, cocoa bean fermentation

Torulopsis feed mixtures

Methylene blue staining

this method distinguish live (colourless) and dead (coloured) cell Saccharomyces cerevisiae a drop of yeast is placed in the centre of a slide one drop of methylene blue is added cover glass examine with dry objectives (10x, 45x)

Moulds
Nonseptated hyphae Septated hyphae

Fructification organs Sporangiophore Connidiophore

Reproduction of Moulds

Vegetative disintegration of mycelium

Sexual spores zygospores, ascospores Asexual spores fructification organs - sporangiophore - conidiophore

Asexual spores of moulds

Mucor

Penicillium

Zygomycota

nonseptated mycelia sexual zygospres, asexual sporangiospores saprophytic soil, fruits, food spoilage, decomposition of polysaccharides (pectin) Rhizopus, Mucor

Life cycle of Zygomycota

Ascomycota

sexual ascospores, asexual conidiospores moulds with septated mycelia sporogenic yeasts Penicillium, Aspergillus, Candida, Saccharomyces

Life cycle of Ascomycota

Deuteromycota (Fungi imperfecti)

conidial stages of Ascomycota sexual reproduction has not been primarily observed two names for one genera one genera placed into both groups Ascomycota and Deuteromycota food spoilage, plant pathogens, decomposition of organic matter

Penicilin antibiotic, P. notatum, Sir Alexander Flemming 1929 P. expansum patulin P. roquefortii and P. camembertii P. marneffei pathogenic for immunodeficient people soil, decomposition of organic matter food spoilage bread, vegetable

Aspergilus flavus 1961 aflatoxins (mycotoxins) food spoilage vegetable, meat, dried meat, crops, jams production of enzymes (amylases, lipases and pectinases), soya sauce Asp. niger production of citric acid

Blacks, wet walls of food factories

Tokay wines vegetable pathogen

Microbial nutrition requirements

Bacterial cell composition
Elements: C 50%, N 8%, H 8%, O 30%, P 3%, S 1%, K, Mg, Ca, Na, Cl (all 0,4%), Fe, Cu, Mn, Co, I, F, Zn Matter: water 80%, dry matter 20% (proteins 60%, DNA 3%, RNA 16%, polysaccharides 3%, lipids 15%, other compounds 3%)

Requirements
macroelements: C, N, H, O, S, P, K, Ca, Mg, Fe microelements: Mn, Zn, Co, Mo, Cu, ...... biosynthesis and source of energy (chemothrophs)

Carbon, Hydrogen and Oxygen

often satisfied together the skeleton of organic molecules (saccharides, peptides, lipids) CO2 or organic carbon (source of energy, are reduced and donate electrons to other molecules saprophytes parasites

Nitrogen

amino acids, proteins, enzymes, purines, pyrimidines (DNA, RNA) organic amino acids, peptides (pepton, trypton, yeast extract) inorganic NH4+, NO3, N2 (reduction to NH4+)

Phosphorus

nucleic acids, phospholipids, ATP, ...... inorganic soluble forms of phosphate (H2PO4-, HPO42-, PO43-) organic phosphatase, nucleic acids, phospholipids

Sulphur

amino acids (methionine, cysteine) vitamins (biotin, thiamin) inorganic usually preferable, SO42organic S amino acids

Microbial growth
usually the growth of microbial population is studied, not the growth of individual cell changes in the population during cultivation time describes growth curve batch and continual cultivation in bacteria, each cell separates into two new cells in every generation

The Growth Curve in a Closed System

Measurement of microbial growth

direct counting cultivation turbidity measurement

Lag phase
cells do not divide preparation for multiplication ribosomes, enzymes synthesis number of cells can decrease the length depends on the age of inoculum and medium quality

Exponential phase
maximum growth and division rate the shortest generation time the highest consumption of nutrients the highest production of primary products determination of growth characeteristics (matematical rules are valit at that phase)

Stationary phase
the total number of viable cells is constant balance between cell division and cell death nutrient limitation accumulation of metabolites maximum production of secondary metabolites

Death phase
decreasing number of live cells nutrient deprivation toxic metabolites small number of cells resists in anabiosis

Mathematical characteristics of the growth

lag phase duration generation time (time for each cell division, time necessary for cell number doubling in population)
Generation time (h) 0.20 (12) 0.35 (21)

Microorganism Streptococcus thermophilus Escherichia coli

Bacillus subtilis
Clostridium botulinum Mycobacterium tuberculosis Saccharomyces cerevisiae Protozoa

0.43 (26)
0.58 (35) 12 2 10

Mathematical characteristics of the growth

specific growth rate (number of generation per time unit, amount of biomass generated per time unit, amount of cells...)
Microorganism specific growth rate (h-1) 3,47 1,98 1,61 1,19 0,06 0,35 0,07

Streptococcus thermophilus Escherichia coli Bacillus subtilis Clostridium botulinum Mycobacterium tuberculosis Saccharomyces cerevisiae
Protozoa

Continuous cultivation
growth in open system population in exponential phase chemostats regulation by constant chemical composition of the media turbidostat constant cell number

Factors influencing growth

(Staphylococcus aureus) (Halobacterium)

(Lactobacillus) (Escherichia) (proteolytic bacteria)

(Chlamydomonas nivalis) (Pseudomonas florescens) (E. coli) (Thermus aquaticus) (Sulfolobus, Pyrococcus)

obligate anaerobe: Micrococcus luteus, Pseudomonas, fungi facultative anaerobe: E. coli, Enterococcus, Saccharomyces aerotolerant anaerobe: Streptococcus pyogenes obligate anaerobe: Clostridium, Bacteroides, Bifidobacteria microarophile: Campylobacter, Lactobacillus

Influence of nutrient
Growth of Bifidobacterium animalis on media with glucose (Glu) or raffinose (Raf)

Influence of nutrient (diauxic growth)

Growth of E. coli on media with glucose (glukzy) and sorbitol

Antimicrobial substances

normal growth

time of substance addition

bacterio- (microbi-) static substance

bacterio- (microbi-) cide substances

Metabolism, Carbon Cycle

Sources of energy for microorganism

ATP adenosine triphosphate

o energy carrier o ATP breaks down to ADP and orthoposphate (Pi), energy is available for work o energy from photosynthesis, respiration and fermentation is accumulated in ATP

The cells energy cycle

Catabolism

Anabolism

Enzymes
o specific proteins catalyzing metabolic reactions o increase the rates of reactions o formation of the enzymesubstrate complex o its conversion to products o cleavage of substrate

Allosteric regulation
o modulator binds to regulatory site o changes enzyme conformation o alteration in the shape of the active site o possibility to bind the substrate

A general diagram of aerobic catabolism in a chemoorganoheterotroph

Glycolysis
o glucose degradation to pyruvate o (Embden-Meyerhof pathway) o cytoplasmic matrix

Carbon cycle in the environment

Catabolism of Carbohydrates

polymers (starch, cellulose) hydrolysis or phosphorolysis monomers (glu, man, fru, gal) glycolysis

pyruvate
fermentation respiration

Fermentations
o 2 ATP from glucose o anaerobic processes o electrons from organic molecules are donated to organic electron acceptors o products of fermentations are simpler organic molecules

Alcoholic fermentation
o glycolysis pyruvate decarboxylation (acetaldehyde) reduction to ethanol o C6H12O6 2CH3CH2OH + 2CO2 (2ATP)

Alcoholic fermentation
Saccharomyces o bakery, alcoholic beverages, vitamins o anaerobic fermentation x aerobic respiration o mezophiles o mono- or disaccharides Candida Zymomonas (bacteria)

Lactic acid fermentation

o glycolysis pyruvate + H+ lactic acid

homolactic fermentation: o main product is lactic acid (> 90 %)

heterolactic fermentation: o about 50% lactic acid, acetic acid, ethanol, CO2, other organic acids

Lactic acid bacteria (LAB)

o o o o

anaerobic, facultative anaerobic, microaerophilic mesophilic, thermophilic acidotolerant mono or disaccharides, organic N, vitamins

Lactic acid bacteria (LAB)

Homolactic fermenters Lactococcus milk, milk products, Lc. lactis, Lc. cremoris Streptococcus yogurt, Sc. thermophilus Enterococcus intestinal tract, probiotic, indicators of faecal
pollution, E. faecium, E. fecalis

Pediococcus milk, silage, fermented meat products Lactobacillus: homo as well as heterolactic
Lb. deslbrueckii ssp. bulgaricus yogurt Lb. acidophilus milk, intestinal tract, probiotic Lb. plantarum plants, silage

Lactic acid bacteria (LAB)

Heterolactic fermenters
Lactobacillus Lb. fermentum, Lb. brevis silage, souerkraut, plants Leuconostoc milk, milk products (production of polysaccharides), silage

Fermentation by bifidobacteria
sugars (mono or oligosaccharides) 60% acetic acid + 40% lactic acids

Bifidobacterium
o o o o o anaerobes, mezophilic intestinal tract, probiotics B. bifidum B. animalis B. longum

Propionic acid fermentation

monosaccharide or lactic acid pyruvate propionate + acetic acid + CO2 + H2O

Propionibacterium
o facultative anaerobe, mezophilic

skin (P. acne), Emmenthal chesses (flavour and holes),

intestinal tract

Butyric acid fermentation

sugars (mono-, oligo- or polysaccharides) pyruvate butyric acid + acetic acid + CO2 + H2 + alcohols or other organic acids

Clostridium
o o o o o anaerobic, mezophilic, G+, sporeforming intestinal tract, soil, food spoilage C. perfringens C. butyricum C. botulinum

Methanogenesis
o acetic acid or CO2 + H2 CO2 + CH4 o methanogenic bacteria (Methanobacterium, Methanococcus, Methanobrevibacter) o biogas production o greenhouse gas

Mixed acid fermentation

pyruvate ethanol + acetic acid + lactic acid + succinic + formic acid + CO2 + H2

Escherichia
o facultative anaerobic, mezophilic, G-, enterobacteria o intestinal tract, probiotic, indicators of faecal pollution, pathogenic o E. coli

Mixed acid fermentation

Salmonella
o facultative anaerobic, mezophilic, G-, enterobacteria o intestinal tract, zoonotic infection (transferred between animals and human), food o S. enterica

Proteus
o facultative anaerobic, mezophilic, G-, enterobacteria o intestinal tract, urinary infections

Butanediol fermentation
pyruvate acetoin butanediol + ethanol + acids

Enterobacter, Serratia, Erwinia

o facultative anaerobic, mezophilic, G-, enterobacteria o intestinal tract

Catabolism of Carbohydrates

polymers (starch, cellulose) hydrolysis or phosphorolysis monomers (glu, man, fru, gal) glycolysis

pyruvate
fermentation respiration

Aerobic respiration
pyruvate acetyl-CoA tricarboxylic acid cycle electon transport and oxidative phosphorylation CO2 + H2O o o o o o 38 ATP from glucose aerobic process aerobic bacteria most of fungi electrons from organic molecules are donated to molecular O2 by way of an electron transport chain

Incomplete aerobic respiration

organic C-substrate simpler organic molecule + (CO2) + H2O + energy o acetic fermentation o citric acid fermentation

Acetic fermentation
ethanol + O2 acetic acid + H2O + energy (6 ATP)

Acetobacter
o aerobic, G- rods, acidophils o vinegar o spoilage of alcoholic beverages

Citric acid fermentation

saccharides + O2 citric acid + H2O + energy

Aspergillus niger
o ascomycetes o citric acid production (preservation of food and beverages, aroma)

Anaerobic respiration
o aerobic electrons from organic molecules are donated to molecular O2 by way of an electron transport chain o anaerobic electron transport chains that can operate with inorganic electron acceptors other than O2 o electron acceptor is an oxidized inorganic molecule o nitrates (reduction, denitrification), sulphate (desulfuration), CO2 (methanogens), ....

Catabolism of polysaccharides

o hydrolysis o secreting of hydrolytic enzymes o assimilation of smaller molecules

Disaccharide cleavage

Catabolism of polysaccharides
Anaerobic condition Aerobic condition

CO2 + H2O2

Cellulose cleavage
o structure polymer of glucose in plants

cellulases

Aerobic condition
o soil, mineralisation of organic matter o complex of cellulolytic bacteria: Cytophaga, Sporocytophaga, Cellulomonas o actinomycetes, micromycetes

Anaerobic condition
o o o o butyric fermentation Clostridium waterlogged soil, bottoms of rivers and lakes mulch, compost, sludge digestion, production of biogas

Cellulose cleavage in ruminants

o Fibrobacter o Butyrivibrio o Bacteroides o Ruminococcus o Clostridium o anaerobic moulds o in monogastric animals large intestine

Strarch cleavage
o reserve polymer of glucose

amylases

Aerobic condition
o moulds - Apergillus, Rhizopus o bacteria - Bacillus, Pseudomonas, Streptomyces o mineralization in soil, amylolytic enzymes, malt production, synthetic sugars

Anaerobic condition
o butyric fermentation o Clostridium o intestinal tract, waterlogged soil

Pectin cleavage
pectinases
o polymer of galacturonic acid, plant cell walls and tissues

Aerobic condition
o o Bacillus, Mucor, Alternaria plant pathogens, soil

Anaerobic condition
o butyric fermentation, except of galcturonic acid o Clostridium o soil, intestinal tract, sludge digestion, production of biogas

Lipid catabolism
o energy source o lipids = triacylglycerols (esters of glycerol and fatty acids) o hydrolyzed by lipases o glycerol is catabolised in the glycolysis o fatty acids are oxidized in the -oxidation pathway acetyl-CoA fed into the TCA cycle or used in biosynthesis

Protein and amino acid catabolism

o source of carbon and energy o some bacteria and fungi o pathogenic, food spoilage, soil microorganisms aerobic o Bacillus, Pseudomonas, Proteus, moulds o products: NH3, CO2 anaerobic o Clostridium o products: NH3, CO2, H2S, scatol, indol, mercaptans, organic acids,...

Protein and amino acid catabolism

o proteases hydrolysis of proteins to amino acids transported to the cell and catabolised o deamination removal of the amino group from AA organic acids o transamination amino group is transferred to an keto acid acceptor o convertion of organic acids to pyruvate, acetyl-CoA, TCA cycle intermediate o nitrogen is excreted as ammonium

Anabolism - biosynthesis

The cells energy cycle

Catabolism

Anabolism

The Construction of Cells

heterotrophs autotrophs

The Organization of Anabolism

o biosynthetic products (in blue) are derived from intermediates o CO2 fixation (in red)

Fixation of CO2
o only autothrops high need for energy: o photosynthesis o oxidation of reduced inorganic molecules incorporation of CO2: o Calvin cycle o reductive pentose phosphate cycle carboxysomes: o site for CO2 fixation o thiobacilly o cyanobacteria o nitrifying bacteria

Photosynthesis

Synthesis of Saccharides
gluconeogenesis: o reversed glycolytic pathway o pyruvate synthesis, glucose or fructose, converted to other sugars synthesis of polysaccharides:

Synthesis of proteins

o amino acids joined by peptide bonds o different proteins have different amino acid sequences and structure amino acids o amination o transamination

Synthesis of lipids

fatty acids: o straight chained or branched o fatty acid synthetase

glycerol: o arises from the reduction of the glycolytic intermediates phospholipids

Nitrogen cycle

Ammonification

Ammonification
o mineralization, mainly in soil o proteins, amino acids, nucleic acids, urea, chitin, uric acid, peptidoglycan, etc. NH3, NH4+ The use of NH4+ o source of N for biosynthesis o source of energy through nitrification o volatisation (outflow into the atmosphere) o sorption on soil complex o leaching into underground and surface water

Nitrification

Nitrification
o o o o aerobic! oxidation of ammonium in two steps chemolitothrophic autotrophs source of energy for nitrifying bacteria (source of C usually CO2)

Nitritation NH4+ + O2 NO2- + 2H+ + H2O + E Nitrosomonas, Nitrosococcus, Nitrosolobus Nitratation NO2- + O2 NO3- + E Nitrobacter, Nitrococcus

Significance of Nitrification
o o o o o source of energy for nitrifying bacteria NO3- is main source of N for plants NO3- is used in denitrification NO3- is not stable losses of N by leaching limitation for use water as drinking eutrophisation of surface water developing of algae o undesirable oxidation of NH4+ in farmyard mature ( N losses by leaching)

Denitrification

Denitrification
o NO3- + H+ N2 + H2O + E + O2 o (NO3- NO2- NO N2O N2) o respiration chain in which O from nitrates is an acceptor of H+ o source of energy for denitrifying bacteria o anaerobic o Pseudomonas, Paracoccus, Propionibacterium, Thiobacillus o losses of N from soil o removing of NO3- from drinking water

Assimilatory nitrate reduction

Assimilatory nitrate reduction

o o o o

NO3- + H+ NH4+ + H2O + E anerobic respiration O2 from NO3source of energy and N for synthesis of amino acids and proteins

N2 Fixation

N2 Fixation

o o o o

returning N2 from athmosphere to soil nitrogenases high need for energy anaerobic or aerobic proces

o N = N HN=NH H2N-NH2 2 NH3 2 NH4+ AK o N2 + 8e- + 16 ATP + 10 H+ 2NH3 + H2 + 16ADP + 16 Pi

N2 Fixation - Diazotrophic bacteria

o soil free living bacteria: Clostridium, Azotobacter o associative bacteria: Azospirillum (G- curved rods) o symbiotic bacteria: Frankia (G+ filamentous), Rhizobium, Bradyrhizobium (G- rods)
Frankia Bradyrhizobium

N2 Fixation - Diazotrophic bacteria

Clostridium
o G+ sporeforming anaerobic rods o acidic waterlogged soils o fixation 5 kg/ha/year o only in absence of other sources

N2 Fixation - Diazotrophic bacteria

Azotobacter
o G- nonsporeforming aerobic rods (nonfixing stage) x cocci (fixing stage) o neutral, high quality soils

o fixation 5-10 kg/ha/year o only in absence of other sources

N2 Fixation - Diazotrophic bacteria

Rhizobium
o G- nonsporeforming aerobic pleomorphic rods o straight in soil and media, branched in root nodules = bacteroids (shape Y) o fixation only in the roots if the content of N in soil is low (50-150 kg/ha/year) o symbiosis with Fabaceae o plants give sugars, rhizobia N o to improve fixation seeds could be inoculated before sowing

Sulphur cycle