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Title: Microbial Diversity
Description: 10 credit IBERS first year module, taught by Gareth Griffith. covers prokaryote structure, growth ecology, and disease, viruses, fungal diversity and technology, and fungal diseases.

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Tree of Life
26 January 2015

13:21

History
• Antonie van Leeuwenhoek (1650s)
○ Animalcules
• Louis Pasteur (1822-1895)
○ Microbe disease link
• Robert Koch (1843-1910)
○ Identified cholera
• Search for classification of microbes
○ Aristotle - plant or animal
○ Von Nägelo (1857) - Plant
○ 5 kingdoms
 Animal, plants, fungi, protists
 Monera (bacteria)
○ Chatton (1937) introduced the term prokaryote
○ Stanier (1961) proposed current definition
○ Murray (1968) proposed Prokaryotae

Eukaryotes vs
...
6mm
• Thiomargarita namibiensis 0
...

• E coli doubling time = 20 minutes
○ 1 cell --> 4
...
g
...

• Most organisms cannot survive
• Extremophiles
○ Mostly archaea
• Sporulation sometimes used

What do cells need?
• All cells require energy
○ Chemotrophs
 Organic
 Inorganic
○ Phototrophs
• All cells require carbon
○ Heterotrophs
 Organic compounds
○ Autotrophs
 CO2
Biogeochemical cycles
• The water cycle
• Carbon cycle
• Nitrogen cycle
○ Movement of nitrogen between atmosphere and organisms
○ Nitrogen fixing bacteria (N2 to ammonia)
○ Nitrifying bacteria (ammonia to nitrates/nitrites)
○ Denitrifying bacteria (reduction of nitrates)
○ Symbiotic bacteria in root nodules
• Sulphur cycle
• Phosphorous cycle
Microorganisms in soil
• Bacteria, fungi algae, viruses
• Bacteria make up 70%
• Affected by abiotic factors
○ pH, moisture, [O2]
• Role of soil bacteria
○ Nitrogen fixation
○ Decomposition of organic matter
 Carbon cycle

Microorganisms in air
Microbial Diversity Page 11

Microorganisms in air
• Too dry for growth
• Spores are carried in air
• Vegetative cells carried on dust/water droplets
• Variable type/number

Microorganisms in water
• Marine and freshwater
○ Photosynthesis - carbon fixations
○ Pathogens
 E coli
□ Detection
 Selective medium
 Differential medium to show presence

Microbial Diversity Page 12

Prokaryote Disease
13 February 2015

14:03

Bacteria and Humans
• Internal tissues
○ e
...
blood, brain, muscle
○ Free of micro-organisms
• Surface tissues
○ e
...
skin, mucous membranes
○ Colonised by micro-organisms
• Many of the bacteria on human skin are mutualistic
• Normal microbial fauna
○ Vary in number and identity
 Body part
 Age
 Diet
○ Beneficial
 Prevent infection
 Aid digestion/nutrition
• Clostridium difficile
○ 5% healthy guts
○ Overgrowth following treatment
○ Rehydration/antibiotic therapy
○ Recurrent cases
 Vancomycin 31%
 Faecal transplant 81%
○ Change to gut bacteria
Friendly bacteria
• Natural microbial flora
• Pre+probiotic products
○ Add more bacteria
○ Promote growth of bacteria
• Protect against pathogens
• Help nutrient absorption
○ Increase obesity?

Bacteria are all around us
• We inhale them
• We ingest them
• Non-pathogenic > pathogenic

Overcoming the barriers
• Skin
○ Entry through cut or wound
○ Vector, e
...
flea, mosquito
• Damage to barriers
○ Lung cilia - smoking
○ Stomach - antacids
○ Digestive system - antibiotics
○ Skin - hyaluronidase; hyaluronic acid holds tissues together
Bacterial stomach infections
Microbial Diversity Page 13

Bacterial stomach infections
• Peptic ulcer and chronic gastritis
○ Heliobacter pylori
 Lives in mucus layers
 Damages epithelium

Steps in disease process
• Contamination
• Infection
• Disease
Virulence factors
Structural or physiological
1
...
Capsule - protects organism from host defences
3
...
Toxins that can directly cause disease

Direct actions of bacteria
• Adherence or attachments
○ Adhesins
 Proteins or glycoproteins
 On pili and capsules
• Colonisation
○ Growth of microorganisms on epithelial surfaces
• Invasion
○ Ability to invade and growth in host tissues
Bacterial tissues
• Any substance poisonous to other organisms
○ Exotoxins - soluble substances secreted into host tissues
○ Endotoxin - part of the cell wall, released into the host tissues by gram negative bacteria

Stages in the course of an infectious disease

Microbial Diversity Page 14

Microbial Diversity Page 15

Tuberculosis
13 February 2015

14:20

• Mycobacterium tuberculosis
○ Straight or slightly curved rids
○ Slow-growing
○ Stain acid-fast, because of mycolic acid in cell walls
• Robert Koch 1882
• Very slow-growing in culture
• Lung disease
○ Tubercle
• Ancient disease
• Massive global health problem
○ 1/3 of world population
• Acquired by inhalation
○ Respiratory secretions or dry sputum
• Organism multiply slowly inside white blood cells
○ WBCs rupture
○ Further cells infected
○ Acute inflammatory response
○ Fluid produced
○ Pneumonia-like symptoms

Resurgence
• HIV infection
• Diabetes, malnutrition, alcoholism, IV drug use
• Drug resistance strains
○ Isoniazid and rifampificin

Mycobacteria
• M leprae - leprosy
• M bovis - bovine tuberculosis
• M marinum - TB in fish, skin lesions in humans

Microbial Diversity Page 16

Viruses
16 February 2015

17:08

• No independent metabolism
• Require elements of host metabolism for replication
• Helical capsids
○ Multiple, identical protein subunits
○ Spiral staircase arrangement
○ Coats genome
• Naked vs
...
Mycelial Growth
• Yeast
○ Non-polar growth
○ Growth in liquid
○ Mobile
○ Lower SA:V
• Mycelium
○ Polar growth
○ Growth at interface
○ Sessile
○ Elongated cells

Multicellularity
• Unicellular - yeasts
• Multicellular
○ Determinate
 Strict body plan
 Genetically hardwired development
○ Indeterminate
 Developmentally plastic
 Responsive to environment
 Modular?

The Hypha
• Apical vesicular complex AVC



○ Spitzenkorper
• Cell wall
○ Chitin microfibrils
 N-acetyl glucosamine subunits
○ Glucans
 Amorphous branched glucose polymer
Microbial Diversity Page 26

 Amorphous branched glucose polymer
○ Hydrophobins
 Water resistant
○ Melanin
○ High tensile strength
○ Expansion



• Septa
○ Valves controlling flow within mycelium
○ Regulate passage of organelles, cytoplasm, nuclei
○ Modified in growth of cell-wall

Hyphal Branching
• Growth rate and diameter
○ Primary > secondary > tertiary
• Hyphal growth unit = total length/number of apices

Tropic responses
• Chemotropism
○ O2
○ Nutrients
○ pH
• Phototropism
• Changes in
○ Apex direction
○ Branch frequency
○ Branch angle

Mycelium
• Pressurised hydrodynamic system
• Septa control cytoplasm flow
• Turgor pressure from osmotic water uptake

Multihyphal structures
• Mycelial cords
○ Aggregated hyphae
○ Foraging
○ Translocation of water
 Dry rot
• Rhizomorphs
• Sclerotia
• Fruit bodies

Microbial Diversity Page 27

Rumen Protozoa
09 March 2015

17:30

• Rumen in ruminants allows a diet of fibrous material
• 5 litre rumen in sheep
...







109 bacteria, 105 bacteria
Large and complex populations
Essential
Imbalance can lead to severe illness
Major contribution to GHG
○ 500-600L methane/day

Energy supply to ruminants
• Volatile Fatty Acids - 70%
• Microbial Cells - 50%
• Digestible unfermented food - 20%
Anaerobic Fungi
• First isolated 1975
• Zoospores attach to feed (mistaken for protozoa)
• ~30 species, <105 cells/ml

Rumen Bacteria
• 5 groups
○ Free living in liquid
○ Loosely associated with food particles
○ Firmly adhered to food particles
○ Associated with rumen epithelium
○ Attached to protozoa and fungi
• Feed-adhering groups
○ 75% population
○ 90% endonuclease activity
○ 70% of amylase activity
○ 75% protease activity
• Adhesion
○ Large multicompound complexes
○ Filamentous extracellular material pili-protein complex
○ Carbohydrate epitopes of bacteria glycocalyx
○ Enzyme binding domains
Protozoa
• Flagellates, ciliates
• Reduce flow of microbial protein from rumen
• Control
○ Saponin - reduces populations
 Inhibitors used to prevent breakdown
• Uncertain how many genera by morphology - adaptations vary in different organisms and
condition
○ Molecular phylogeny - 18S RNA gene

Microbial Diversity Page 28

Decomposition
13 March 2015

11:50

Overall Photosynthesis = Respiratopn

• Dead matter is a major carbon pool
○ Oil
○ Peat
○ Coal
○ Leaf litter
○ Wood
○ Soil organic matter
• Food webs are complex and intertwined
○ Decomposer organisms are ultimately decomposed
Rate of Decomposition
• Resource quality
○ Carbon:nitrogen
• Environmental factors
○ Temperature
○ Water
• 95% decomposition
○ Rainforest - 6 months
○ Tundra - 100 years
• Given as half life/95%
○ Trace amounts can persist for a long time
Measuring decomposition
• Litter bags
○ 7mm mesh (earthworms can get in)
1mm mesh (no macrofauna)
Microbial Diversity Page 29

○ 1mm mesh (no macrofauna)
• Cotton strips and CO2 measures
○ Cotton strips lose tensile strength
○ Infrared Gas Analysis for CO2
• Up to 6km of hyphae/gram of soil
Biocides
• DDT, streptomycin, benomyl (kills fungi)
• Greatly reduce litter repiration

Soil animals
• Detritivores
○ Earthworms
○ Nematodes
○ Springtails
○ Woodlice
• Low biomass, high numbers
• Contribute <10% to soil respiration
• Main effect is physical
○ Comminution - chewing up of litter
○ Increase surface area

Nutrients
• Nitrogen
○ Often limiting
○ Required for protein
• Phosphorous
○ Low availability - insoluble

Fungal Nutrition
• Turgor pressure and tissue softening allow penetration of substrates
○ Cellulases
○ Ligninases
○ Proteases
○ Lipases
○ Phosphatases
○ nucleases
• Oxygen exchange in hypha
• Translocation of nutrients

Composition of plant litter
• Cellulose
○ 20-45%
• Lignins
○ 5-30%
• Lignocellulose mainly degraded by fungi

Microbial Diversity Page 30

Fungal Disease
16 March 2015

12:15

• Biotrophy
○ Living host tissues
• Necrotrophy
○ litter
• Saprotrophy
○ Humus
• Candida albicans
○ Candidiasis
• Aspergillus fumigatus
○ Aspergillosis
• Penicillium marneffil
○ In Far East
• Epidermophyton floccosum
○ Athlete's Foot (Tinea pedis)
• Geomyces destructans
○ Kills bats, infection during hybernation
• Dutch Elm Disease - Ophiosotma ulmi
○ 1920s and 70s epidemics
○ Diseased logs
○ Bar beetle vector
○ Breeding, sanitation and fungicides so far ineffective
• Late blight - Phytophthora infestans
○ Infects potato, tomato, relatives
• Often transmitted via airborne fungal spores
○ Allergens

Microbial Diversity Page 31

Mutualism
13 April 2015

13:07

Anton de Berg, Miles Joseph Berkeley
Mutualism benefits all involved

• Root endophytes
○ Role unknown
• Lichen
• Mycorrhizas
○ Fungus root
○ Decomposition benefits plant
○ SA:V increased
 Roots and narrow hyphae
○ Exoenzymes
Ectomycorrhizas
• Colonise outer root tissues
• Change root morphology
Fungal Haustoria

Most animals have mutualistic associations with microbes
• Lignocellulose energy
• Cellulolytic gut microbes
Termites
• Major agents of tropical wood decay
• Fungus gardens - Old World Termites
○ Chimneys to maintain temperature
• Gut microbes - New World Termites
○ Bigger termites as a result

Microbial Diversity Page 32

Lichens
17 April 2015

12:28

Fungal Mutualisms
• Mycorrhizas
○ Photosynthetic partner generally larger than the fungal partner
• Lichen
○ Photosynthetic partner is microscopic
○ Similarities with marine corals
• Geosiphon
○ Intracellular bladders contain cyanobacteria
○ Artuscular mycorrhizas with higher plants
• Anton de Barry
• Beatrix Potter
• Tundra and deserts dominated by lichens

Lichen symbiosis
• Mutualistic association between fungus and phototroph
• Fungus is dominant partner
• Algae/cyanobacteria in association with fungi
• Predominate in inhospitable habitats
• Growth rates range from 0
...
Basidiomycetes
• Lichens are polyphyletic
• Latin name describes the fungus
• Some lichen fungi can have >1 photobiont partner
Photobionts
• 25 genera of unicellular and filamentous chlorophyta (green algae)
• 15 genera of cyanobacteria
• Most photobionts can be free living
• 90% of lichen species involve chlorophyta photobionts
• Unicellular algae are most common
• Cyanobacteria photobionts (10%) are mainly Nostoc or Scytonema species
• Cyanobacteria can fix nitrogen

Dissociation/Resynthesis
• Mycobiont and photobiont can exist separately or reform a symbiotic relationship

Microbial Diversity Page 33



Lichen Morphology

Lichen Dispersal
• Apothecia (with asci)
○ Dispersal of fungal partner only
• Diaspores
○ Fragmentation propagules
○ Contain algal cells surrounded by fungus
○ Maintain the partnership
Thallus Ultrastructure








Algal cells protected by fungal cortex
Lower cortex acquires substrate nutrients
Heteromerous - distinctly layered thallus
Homoiomerous - Less structured
Hydrophobins maintain air channels
Haustorial penetration of photobiont allows nutrient transfer

Microbial Diversity Page 34

Somatic incompatibility
• Non-self-recognition - zone lines
• Intra- and inter-specific competition
• Protection of territory
• Successional processes
• Lichenicolous lichens - photobiont takeover
• Parasitic fungi
Lichens and air pollution
• Inhabit exposed habitats
• Sensitive to airborne pollution
• SO2 - acidification
• Nox pollution - affects competitive interactions
Lichens and rock weathering
• Organic acids are secreted to release nutrients
• Important in rock weathering
Lichens in medicine/biotech
• Lichen metabolites used in traditional and modern medicine
• Litmus
• Traditional pigments
• Ethanolic fermentation

Microbial Diversity Page 35

Eukaryotic Algae - Phytoplankton
25 April 2015

11:41

• Algae is no longer a taxonomic term
○ “…unless purely artificial limits are drawn the designation algae
must include holophytic (photosynthetic via chl
...
g
...
” Fritsch (1935)
...

 2 unequal, both with hairs - cryptophyta
 2 unequal, 1 girdle and 1 trailing - pyrrhophyta
 1 hairy, one row - euglenophyta
• Cell ultrastructure
○ Pro-eukaryote
○ Plastid, particularly thylakoid arrangement









Single thylakoid, with girdling - Rhodophyta
Paired, no girdling - Cryptophyta
3-pky, no girdling - diatoms, G/B algae
3-ply, girdling - Dinoflagellates, haptophytes
Never single stacked, no girdling - Green algae,
euglenids
Plastid envelope

□ Double membrane envelope - chlorophyta,
rhodophyta
□ Envelope +1 additional membrane - euglenophyta
and pyrrhophyta
□ Envelope plus CER or nucleus - chrysophyta
○ Nucleus, nuclear envelope, details or mitosis and meiosis,
cytokinesis and chromosomes
Classification of "Algae"
• Phylum - chrysophyta
○ Class - bacillarophyceae/diatoms
 Chlorophylls a+c
 Accessory pigments - fucoxanthin
 Cell wall - silica
 Pennate and centric diatoms
 Raphid diatoms unique sliding movement
○ Class - Prymnesiophyceae/haptophytes
 Chlorophylls a+c
 Accessory pigments - fucoxanthin
 Cell wall - cellulose/CaCO3
 Haptonema - specialise organelle
 Coccolithphorids calcium carbonate scales
○ Class chrysophyceae/golden brown algae
 Chlorophylls a+c
 Accessory pigments - fucoxanthin, beta carotene
 Cell wall - silica scales, chitin, cellulose rods
 Majority freshwater species
 2 perpendicular flagella
 Eye spot
 Photoreceptor
• Phylum - Pyrrhophyta
Microbial Diversity Page 37

Ddgghce

• Phylum - Pyrrhophyta
○ Class - Dinophyceae/dinoflagellates
 Chlorophylls a+b, a+c
 Accessory pigments - phycobilins, beta carotene
 Cell wall - cellulose plates either beneath membrane or
naked
 Condensed mesokaryotic chromosomes
□ Dinomitosis
 Special organelles - pusule, ocellus, nematocyst, trichocyst,
muciferous bodies
• Phylum - cryptophyta
○ Class - cyrophyceae/cryptomonads
 Chlorophylls a+c
 Accessory pigments - phycobilins
 Cell wall - naked
□ Protein plates associated with underside of cell
membrane
 Ejectosomes - trichocysts
 Well suited to low light levels - cryosphere
• Phylum - chlorophyta
○ Class - chlorophyceae/green algae
 Chlorophylls a+b
 Accessory pigments - beta carotene, lutein
 Cell wall - cellulose/glycoproteins
 Classification disputed
□ Variety of morphologies
 Unicellular
 Colonial
 Filamentous
 Multicellular
• Phylum - Euglenozoa
○ Class - englenoidea/euglenids
 Chlorophylls a+b
 Accessory pigments - beta carotene, neoxanthin
 Cell surface - unique pellicle
□ Proteinaceous strips under plasma membrane
 Metaboly
 Chloroplasts with three membranes
Reproduction
• Bacillarophyceae (Diatoms)
○ Cell division and reduction
 Asexual binary fission
 Progressive size reduction
 Finite
○ Auxospore formation (sexual)
 Isogamous (M/F identical)
 Oogamous (distinct)
 Spores can survive for up to 20 years
• Prymnesiophyceae (haptophytes)
○ Hymenomonas carterae
 Apistonema stage - 21+-1 chromosomes
 Hymenomonas stage - 41+- 1 chromosomes
• Chrysophyceae (Golden-brown algae)
○ Form stratospores
○ Silica deposition vesicle formation
• Pyrrhophyta (dinoflagellates)
Isogamous (different M/F gametes)
Microbial Diversity Page 38

○ Isogamous (different M/F gametes)
○ Gymnodinium sp
...

 Sexual - anisogamous
 Mostly haploid life cycle
• Chlorophyta (green algae)
○ Asexual reproduction
 Fragmentation
 Zoosporogenesis
 Aplanospores
 Autospores
○ Sexual
 Isogamous
 Anisogamous
 Oogamous
• Euglenids
○ Meiosis has not been observed
○ Mesokaryotic chromosomes (never uncondense)
○ Cyst formation

Toxins
• Harmful Algal Bloom conditions
○ Stable water column
○ Near water surface
○ Elevated pH
○ Wind driven upwelling
○ Spores can be formed
• Dinoflagellate toxins
○ DSP - diarrhetic shellfish poisoning
 Dinophysis sp
...

 Brevetoxins
○ PSP - paralytic shellfish poisoning
 Several species
 Gonyautoxins
 Saxitoxins
○ CFP - ciguatera fish toxins
 Gambierdiscus toxicus
 Maitotoxin
 Ciguatoxin
• Diatom toxins
○ ASP - Amnesic shellfish poisoning
 Pseudoniteschia sp
...
g
...

• Mutualistic
○ Endosymbionts of animals
e
...
Symbiodinium sp
...
g
...

• Parasitic association
○ Blastodinium in copepods
 Significant economic losses in copepods
• Mutualistic association
○ Zooxanthellae
 Symbiodinium sp
Title: Microbial Diversity
Description: 10 credit IBERS first year module, taught by Gareth Griffith. covers prokaryote structure, growth ecology, and disease, viruses, fungal diversity and technology, and fungal diseases.