Download BacteriaDiversityMDS_07_v2

Lecture 21: Bacterial diversity
and Microbial Ecology
Dr Mike Dyall-Smith
Haloarchaea Research Lab., Lab 3.07
[email protected]
Ref: Prescott, Harley & Klein, 6th ed., parts of
chapters 21-24 (refer to these notes).
Also p590-1 (lichens)
Main Topics
• That, along with the Archaea, the Bacteria are
very widespread in nature
• That the Domain Bacteria contains many,
different groups, with considerable metabolic
diversity
• How microbial diversity is studied
Microbial habitats
•Archaea and Bacteria are found wherever there is:
• Water
• Energy source
• C, N, P, S, etc.
• Within physicochemical limits (, pH, salt,...)
Ecological characteristics of bacteria
• live almost anywhere there is liquid water
• occur in large numbers
• Most bacterial cells are relatively small
• Species diversity is very large (and growing)
• Most of the ~35 phyla are poorly understood
• Can be studied to some extent without cultivation
Ecological characteristics of bacteria
• Most of the 10
30
bacterial cells are relatively small
soil bacteria
0.3 × 0.5 µm
marine bacteria
0.3 × 1 µm
Escherichia coli
1 × 3 µm
Epulopiscium*
50 × 600 µm
Thiomargarita*
750 µm
* exceptions
Microbial habitats
•Within physicochemical limits:
• TEMP: –10 to 113 
(20 - 40 )
• pH: 0 to 11 (3 - 5 units for any one species)
• [NaCl]: 0 to 6 M (~saturation) depending on
species
• Others: Oxygen (toxicity), pressure, radiation
Phylogeny of Bacteria (using 16S rRNA)
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At least 35 phyla
Groupings and relationships are
very informative: e.g.
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Gram positives cluster together
in Firmicutes
Cyanobacteria: photosynthetic
Spirochaetes: helical cells;
motility by axial filaments
placement of a new isolate into
a phylogenetic grouping can be
highly predictive
Phylogeny of Bacteria (using 16S rRNA)
•
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Most of the studied bacteria
belong to just 4 phyla - Proteobacteria
- Bacteroidetes
- Firmicutes
- Actinobacteria
Some phyla have no cultured
representatives (white wedges)
These are detected by 16S rRNA
sequences directly from natural
samples, but cannot be grown in
pure culture in the laboratory
How to study microbial diversity and ecology
•
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Cultivation dependent - ideal, but has problems!
Cultivation independent:
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Sequence information - eg. 16S rRNA sequences,
genome sequences
rRNA targeted probes, eg. FISH
(Fluorescent In Situ Hybridization)
Allows a visual inspection of phylogenetic groups of
cells in a natural sample
Cultivation dependent
• Pure cultures are the basis of the traditional
way of studying bacteria
• Usually only 1% of cells in a natural sample
will form colonies on plates
• Different bacteria have different abilities to be
cultured; from easy to difficult
• Known examples that cannot be cultured
Bacteria: examples that have not yet
been cultured
• Mycobacterium leprae (leprosy)
• Treponema pallidum (syphilis)
• Epulopiscium fishelsoni
• All members of the TM7 phylum
(a major lineage of Bacteria)
Mycobacterium leprae
Treponema pallidum
Epulopiscium
Cultivation independent: Sequence data
• 16S rRNA sequences, specific genes, mRNAs,
whole genome sequences, metagenomes
• Discovered many new groups of Bacteria
- but physiologies yet unknown
• Can use sequence information to directly
visualise specific bacteria in situ (in their natural
state)
Fluorescent In Situ Hybridization (FISH) ...
Cultivation independent: Sequence data
• 16S rRNA sequences, specific genes, mRNAs,
whole genome sequences, metagenomes
• HOW ?
•Take sample, extract DNA (or RNA)
•a) PCR amplify 16S rRNA genes
•clone individual genes, sequence
•b) Sequence DNA directly (metagenomics)
- usually difficult to reconstruct individual
microbial genomes as too many species
FISH - Fluorescent In Situ Hybridisation
- short DNA sequence
- complementary to rRNA
- specific sequence (eg. to genus)
- fluorescent tag attached
rRNA
• Permeabilize cells so that
the DNA probe can enter
Allow it to find its matching
sequence on rRNA
FISH - Fluorescent In Situ Hybridisation
View cells (in situ) under
fluorescent microscope, and
see what cells fluoresce,
showing they have bound the
probe
• Fluorescent DNA probe will bind to rRNA in the
cells only if it exactly matches complementary
sequence of rRNA target region
• Many different coloured fluors, so can do
simultaneous probes for different genera,
families....
FISH - Fluorescent In Situ Hybridisation
Growth in Laboratory media versus
natural conditions
Oligotrophy is the rule in nature
• Most of the biosphere has low available
nutrients (or at least one limiting nutrient)
• oligotrophy (‘small feeding’) is growth at
low nutrient concentrations
Dissolved organic C
NATURE
LAB.
coastal waters
soil
M9 minimal
0.1 - 1 mg/l
5 - 20 mg/l
800 mg/l
Surface area to Volume ratio
• Small cell size is a way to cope with low
substrate availability. It increases the surface
area to volume ratio.
• Substrate uptake is via cell
membrane proteins.
Increasing SA/Vol improves
the ability to supply
nutrients to the cytoplasmic
volume
natural microbial
populations
• Large numbers of small cells
• Nutrient levels usually very low
• Population size controlled and limited by
nutrient availability
• Low growth rate (as nutrients removed
rapidly), and just matches the death rate
• Energy mainly used for cell maintenance
Microcystis bloom in Matilda Bay, Swan-Canning Estuary, Western Australia.
Photo by Tom Rose (WA Waters and Rivers Commission)
an un-natural cyanobacterial bloom due to excessive nutrients (pollution)
Phylum: Cyanobacteria
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Largest and most diverse group of
photosynthetic bacteria (24 genera)
carry out oxygenic photosynthesis:
similar to eukaryotes. Fix CO2
Cells contain thylakoid membranes
Significant proportion of marine
plankton (and marine microbial food
web)
Phylum: Cyanobacteria
Cells contain thylakoid membranes
Phylum: Cyanobacteria
• photolithoautotroph: energy from
•
light, inorganic electron source,
carbon from CO2
many filamentous forms possess
heterocysts, where nitrogen
fixation occurs
heterocyst
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Typical gram -ve cell wall structure
diverse modes of reproduction
some show gliding motility
Can form symbiotic relationships with fungi = lichens.
•Lichens: an association between
two partners: an ascomycete
(fungus) and a cyanobacteria (or
alga).
•Partnership forms when both are
nutritionally deprived
•Cyanobacteria provide organic
compounds via photosynthesis,
and can fix nitrogen
•Fungus provides protection, water
retention, extracts minerals and
nutrients from substrate
Summary
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16S rRNA gene sequence comparisons allow a
phylogenetic framework to be discerned. Useful for
taxonomy, ecological and evolutionary research
Domain Bacteria has 35 phyla, and are species rich
Great metabolic and genetic diversity within phyla
Many phyla are poorly studied because no
members have yet been cultivated
Despite this, useful information can be obtained
using cultivation independent methods (e.g. 16S
rRNA sequences, genome sequences, FISH)
One example given, phylum Cyanobacteria
Summary
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50 - 90% of all biomass on Earth is microbial
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oligotrophy is the most common state in nature
bacteria widespread, only limited by: free water,
energy source, components of biomass, and where
biomolecules can be stable
cell size is small in order to increase the surface
area to volume ratio, hence improving the ability to
take up nutrients at low concentration
•final note: the vast majority of bacteria are not
pathogens. They work for us, in the environment
•PICTURE CREDITS
•Treponema pallidum:
http://www.brooksidepress.org/Products/OperationalMedicine/DATA/operation
almed/Manuals/GMOManual/clinical/Dermatology/Treponema%20pallidum50
0.jpg
•Mycobacterium leprae: www.wissenschaft-online.de
•Cyanobacterial bloom in Swan river:
http://www.ozestuaries.org/indicators/econ_cons_algal_blooms.jsp Photo by
Tom Rose (WA Waters and Rivers Commission)
•Others from Prescott, Harley and Klein.