Intro Flashcards
Why study nutrient cycling + prokaryotes?
studying prokaryotes allows us to classify organisms at a fundamental level and provide an understanding of life that evolved prior to organisms with nuclei
What was Carl Woese’s main discovery when he studied 16s rRNA?
He determined that prokaryotes are not monophyletic as was previously thought, but instead, exist as 2 different groups with different sequences of nucleic acids and rRNA
(EU) bacteria and (Archae) bacteria = bacteria and archaea
What environments do Archaea typically live in?
extreme environments typically with high sulphur content
What are the points of view that biological systems can be defined by?
community structure
trophic dynamic levels (population)
energy transfer and nutrient cycling
Describe how community structure can be used as a point of view to describe complex biological systems
the system is defined by the primary producers and the plant community (ex. trees)
Describe how trophic dynamic levels (population) can be used as a point of view to describe complex biological systems
the system is defined by the animal populations and their interactions
Describe how energy transfer and nutrient cycling can be used as a point of view to describe complex biological systems
the system is defined by the prokaryotes that function and have functioned in the past
Why is studying prokaryotes important for understanding eukaryotes?
the current paradigm is that eukaryotic cells evolved from a symbiosis of prokaryotic cells billions of years ago that retained their ancestral rRNA
What key nutrients do bacteria and archaea play an essential role in transforming? How?
organic carbon, nitrogen, sulfur
they are key regulators of ecosystem productivity
How are trophic interactions linked to nutrient cycling?
primary producers produce different phytochemicals which influence trophic interactions and ecosystem nutrient dynamics (ex. lignin, cellulose, tannins, terpenes)
the phytochemicals will influence the herbivorous populations which will influence the carnivorous populations and the herbivorous community will limit the type of and abundance of plants
the phytochemicals will influence the nutrient dynamics which in turn will influence which plants grow and how well they grow
there’s 3 feedback loops:
- between the phytochemicals produced by plants and trophic interactions
- between the phytochemicals produced by plants and ecosystem nutrient dynamics
- between trophic interactions an ecosystem nutrient dynamics which includes the other 2 loops
How are spatial and temporal variations linked by feedback loops?
there’s different layers on the phytochemical landscape (ie., the way the phytochemicals influence other aspects of the ecosystem) which include:
nutrient availability
phytochemistry
herbivore activity
predation pressure
What kind of qualitative and quantitative methods are needed to develop and provide evidence for the spatial and temporal links to the feedback loops on the phytochemical landscape?
- observation and identification of members of the ecosystem (plants, animals, bacteria)
- quantitative data collection
- field sampling and statistical analysis using bio, chemical, and physical methods
- uni- and multivariate
- correlation/regression - experimenting in the lab and in the field
- statistical theory of decision
- use random block design
- mathematical distribution (ex., Guassian, Lognormal, Poisson)
What are the practical reasons for studying nutrient cycling + prokaryotes?
agricultural reasons:
- N2 fixation
- nutrient cycling
- animal husbandry
- gut bacteria in the animal gut microbiome
food industry:
- food preservation (heat, cold, radiation, chemicals)
- fermented foods
- food additives (monosodium glutamate, citric acid, yeast)
disease:
- identifying new diseases
- treatments, cures, and prevention`
energy/environmental reasons:
- biofuels
- fermentation
- bioremediation
- microbial mining
biotechnology:
- GMOs
- pharmaceuticals (ex. insulin)
- gene therapy
Where are prokaryotes?
everywhere! in all biomes
What are biomes?
a biogeographical unit within which a biological community exists in response to a similar regional climate
ex. temperate rainforest, steppe, grassland
Who was the first to establish the major biogeoclimatic zones in BC?
Krajina
How can the communities within a biome vary? explain why
multiple communities, aside from the dominant one, can develop on specific habitats within a biome and this can include different plants and bacteria
this occurs because:
- water moisture levels can vary depending on the slope (low at the top of the slope, high at the bottom of the slope = different habitats
define xeric
when moisture levels are low
define hydric
when moisture levels are high
Explain how Mt Doug is a good representation of multiple communities existing in different habitats related to elevation gradient
there are 3 communities on Mt Doug
at the bottom, where moisture levels are high and nutrients are abundant: Western Red Cedar dominates
in the middle, where moisture and nutrient levels are intermediate: Douglas-fir dominates
at the top, where moisture levels and nutrient levels are low: Garry Oak dominates
What conditions do Douglas-fir grow best in?
high nitrate (NO3-) rather than ammonia (NH4+)
What did Krajini suggest as an explanation for how Douglas-fir has come to dominate the forest ecosystem on Mt Doug? What other factor is being considered now as well?
nitrification of ammonia to nitrate
now being more considered is the role of mycorrhiza
What is the scientific name for Douglas-fir?
Pseudotsuga menziesii
What is the scientific name for Western Red Cedar?
Thuja plicata
What is the scientific name for Garry Oak?
Quercus garryana
Explain the relationship between Monotropa uniflora, Thuja plicata and Mahonia nervosa (Oregon grape), and Russula sp. fungi
Monotropa uniflora look like fungi but are actually plants that have lost the gene that codes for chlorophyll so they appear white (hence, the name ‘ghost plant’)
this plant parasitizes Russula sp. fungi
Russula sp., infect the roots of WRC and provide the tree with N and P and receive carbon from the tree in return (ie., mycorrhizal relationship)
because of the parasitism, the Monotropa uniflora steals the carbon from the Russula provided by the tree and the N and P from the Russula
Why are soil bacteria difficult to study?
the scale is sooo small
bacteria are the roughly the same size as clay particles (1-5 microns)
What are the biogeoclimatic zones on Vancouver Island?
Alpine tundra
mountain hemlock
coastal douglas-fir
coastal western hemlock
What’s the Ecological Reserve Act?
a Land Act approved in 1971 to set aside ecological reserves that people do not have access to
reserve sites are meant to be representatives of the biogeoclimatic zones in BC
Which province in Canada was the first to set aside land parcels with representative ecosystems?
BC for both terrestrial and marine ecosystems
Where are some nearby ecological reserves?
10 Mile Point and Oak Bay Island in the Saanich Inlet include both terrestrial and marine ecosystems
Why is it important to reserve land in regards to the ecology of bacteria?
some reasons include:
- only specific bacteria can fix nitrogen into forms that are usable for plants (agriculture and forestry reasons)
- bacteria can be used in wastewater treatment
- bacteria can degrade anthropogenic chemicals like oil and pesticides
- bacteria and archaea have key roles in biogeochemical cycles in soil and aquatic ecosystems (including the transformation of C, N, S, and P)
What forms of nitrogen are essential for plant growth? What forms of nitrogen are available in the terrestrial environment?
Mineral forms (ammonia, nitrate) are essential to plant growth but these are limited
Nitrogen (N2) is most common form of nitrogen in the environment
How do bacteria contribute to soil fertility and plant growth?
Specific bacteria can fix/convert N2 into ammonia - these are called diazotrophs
What are diazotrophs?
bacteria that can convert N2 into ammonia (NH3)
What are the key forms of nitrogen in the nitrogen cycle?
ammonium (NH4+)
nitrate (NO3-)
nitrite (NO2-)
Who suggested that nitrogen was a limiting factor in plant growth?
Justus Liebig of the University of Giessen
Who won the Nobel Prize in atmospheric chemistry for discovering the relationship of N2O to the ozone degradation in 1970?
Paul Crutzen
Where is the biggest reservoir of nitrogen?
the atmosphere
Which form of nitrogen is the most common key form?
ammonium (NH4+)
In the nitrogen cycle, what catalyzes the transformation between different forms (ie., what are the arrows)? give an example
enzymes coded by genes in specific bacteria
ex. Nitrosomonas
Which form of carbon is most commonly used by bacteria in combination with ammonium for energy?
CO2
What angle is the carbon cycle conceptualized from?
based on CO2
What does all organic matter eventually become?
CO2 and methane
Describe the change of atmospheric methane historically?
pre-industrial: methane levels in the atmosphere were stable
massive increase = indicates instability = there’s either too much production and not enough consumption or both
How is the level of atmospheric methane studied?
gas chromatography
How can we study the historical conditions of atmospheric methane?
ice cores from Antarctica are extracted and preserved gas bubbles from these are analyzed
the deeper in the ice core, the older the information
Describe methanotrophs
prokaryotes that use methane (CH4) as their carbon energy source
What are the 2 types of methanotrophs? Describe them and the conditions they exist in
MOB Type I: membrane is bundled in the middle of the cell
- typically in high methane, high Nitrogen conditions
MOB Type II: membranes layered around the cell wall
- typically in low methane, low nitrogen conditions
Describe the nitrogen, methane, and oxygen levels in a lake profile
surface:
oxygen 100%
low methane
lower NH4
bottom:
very low oxygen
very high methane
slightly lower NH4 than methane
Given the oxygen, methane, and ammonium concentrations within a lake profile, where would you expect to find MOB type 1 and type 2?
Type I will be at the bottom of the lake (high methane, high N)
Type II will be closer to the surface (low methane, low N)
Explain and give an example of how bacteria can be important in biomining and biotechnology
Discovering new bacteria growing in extreme conditions has led to new industrial enzymes
ex. Thermus aquaticus was found in a thermal pool in Yellowstone (Thomas Brock) and since, the taq polymerase used for PCR was extracted from this prokaryote
ex. thermostable amylases from thermophilic organisms have been found to be important in the food industry
How has the definition of ecology been updated since our understanding of prokaryotes has improved?
ecology is now defined as the study of the distribution, abundance, and DIVERSITY of living organisms (Bell, 2001)
What is the definition of the ecology of prokaryotes?
the study of the distribution, abundance, and diversity of prokaryotes in nature
What makes microbial ecology such a large field of study?
there are so many types of interactions
parasitism, predation, commensalism, amensalism
diversity of organisms with different physiologies
most scientists in this field have focused on transformation of C, N or S
What are prokaryotes?
organisms lacking membrane-bound nuclei and organelles
these organisms cannot be seen without the help of microscopes
Who first discovered the existence of microorganisms?
Antonie van Leuwenhoek (1676) with the first rudimentary microscope
What are some milestones in the study of microbial ecology?
1676: discovery of microorganisms with the first developed microscopes (Leuwenhoek)
1850-1900: classical techniques and methods of studying developed by founders of microbiology (Pasteur, Koch, Winogradsky, Beijerinck)
- cultures on solid media, etc
1900-1970: research mainly in labs and defined by habitat (ex. soil microbiology, marine microbiology, aquatic microbiology)
1960-1990: microbial ecology becomes considered a critical science for ecosystem ecology and environmental sciences
1990-onward: molecular genetic techniques developed for understanding community structure and function (analytical chemistry and bacterial activity)
- ex. liquid and gas chromatography, mass spectrometry
- ex. PCR, tRFLP, RT-PCR, allows the identification in situ
What are the 6 characteristics of prokaryotes?
self-feeding (metabolism)
self-replicating (growth)
differentiation
chemical signaling (communication)
movement
evolution
Describe the metabolism/self-feeding characteristic of prokaryotes
the cell is an open system
metabolism includes:
- chemical uptake from environment
- transformation of chemicals within the cell
- elimination of waste products back into the environment
Describe the self-reproduction/growth characteristic of prokaryotes
chemicals taken up from the environment are transformed into new cells by the direction of pre-existing cells
mitotic replication to produce identical daughter cells
What’s a major difference between replication in bacteria vs archaea?
Archaea have histones
Bacteria do not
Describe Class I reactions that occur within prokaryotes regarding self-feeding
Class I: glucose is taken up from the environment and transformed via cellular respiration
Describe Class II reactions that occur within prokaryotes regarding self-feeding
Class II: NH4+, PO4^3-, SO4^2- are taken up from the environment and combine with carbon to produce hexosamines, nucleotides, and amino acids (respectively)
Describe Class III reactions that occur within prokaryotes regarding self-feeding
Class III: products from the Class II reactions (NH4+, PO4^3-, SO4^2) are converted into peptidoglycans, RNA and DNA, and proteins
What happens to the products of the Class III reactions in regards to self-feeding?
peptidoglycans function in the cell wall
proteins are involved in a myriad of functions, but mainly they are in the cell membrane and ribosomes
RNA has ribosomal functions
DNA has chromosomal functions
Describe the differentiation characteristic of prokaryotes
new cell structures can form with specific functions, for example, spores
generally these occur as a part of the cell life cycle (different structures have specific functions at certain life stages)
Describe the communication/chemical signaling characteristic of prokaryotes
communication in prokaryotes occurs mainly by taking up and releasing chemicals
Describe the movement characteristic of prokaryotes
most prokaryotes have flagella and are capable of self-propulsion
movement is key for searching for optimal environments for growth
NOT ALL have this characteristic
Describe the evolution characteristic of prokaryotes
evolution occurs in prokaryotes to adapt new biological properties
How has the evolution of prokaryotes differed to eukaryotes?
prokaryotes have evolved to streamline their genome and get rid of redundancies
What are some basic differences between prokaryotes and eukaryotes?
prokaryotes:
- lack membrane bound organelles and nucleus
- single, circular, haploid chromosomes
- mitotic cell division
- have plasmids as extra-chromosomal DNA
- ribosomes are 70S
- extremely diverse physiology
- types: bacteria, archaea
eukaryotes:
- membrane-bound organelles and nuclei
- several, linear, diploid (some exceptions) chromosomes
- meiotic and mitotic cell division
- no extra-chromosomal DNA
- 80S ribosomes
- physiology limited to oxygenic respiration or oxygenic photosynthesis
- types: Protozoa, fungus, algae
What are the effects of prokaryotes being so small?
rapid diffusion of solutes through cytoplasm
high and diverse metabolism
essential role in nutrient cycling
What are the 2 domains of prokaryotes? How was this determined?
Bacteria and Archaea
Woese used 16S rRNA as a molecular marker to determine that methanogens are not Bacteria, but Archaea
What are the 3 main points of view that be used to describe complex terrestrial and aquatic ecosystems?
community structure: primary producers are used to describe the community (ex. Douglas-fir vs western Red Cedar)
trophic levels/interactions: how energy is transferred through a system
nutrient/biogeochemical cycling
What is the main link between trophic interactions and nutrient cycling?
using the phytochemical landscape as a demonstration
plant type and growth depends on the nutrients available in the soil (depends on the prokaryotes, nutrient cycling, degradation of organic material etc.)
the plants produce and release chemicals (ex. tannins, terpenes) which can affect the herbivorous community which will affect the carnivorous community
what is the link between nutrient cycling and atmospheric trace gases and global warming?
the post-industrial levels of harmful trace gases in the atmosphere are significantly higher than pre-industrial
this suggests there’s an imbalance between the amount that prokaryotes are consuming and releasing
if prokaryotes are not consuming enough methane for example, or producing too much, there will be an imbalanced level in the atmosphere
From the Wibbly Wobbly Circle of Life: how many people were hired to work at ICI in Billingham in 1960? What for?
20,000 people hired to work in the largest nitrogen fixation plant (to produce fertilizer) in the world
From the Wibbly Wobbly Circle of Life: What chemical processed was used in the nitrogen fixation plant at ICI?
Haber-Bosch
How much of the world’s agricultural yield would be lost without the Haber-Bosch process?
30-50%
How much of the fixed nitrogen from the plant is lost to the environment?
90%
What are the 2 major consequences of the loss of fixed nitrogen?
reduction of biodiversity
climate change
What are the 3 components of fertilizer?
ammonia
phosphate
potassium (potash)
What is the driver of fertilizer prices?
fossil fuel prices
How do farmers in poorer countries manage with the increasing costs of fertilizer?
crop rotation to reduce fertilizer use
What makes the concept of the Anthropocene more concrete?
the nitrogen cycle
Who was John Lawes?
He was the Founder of the Economist and Founder of the Rothamsted experiment
What did Lawes do at his estate?
he tested how fertilizer effects the growth of wheat to get rid of tariffs on wheat by the British ‘Corn Laws’
Who was Justus Liebig? How is he connected to Lawes?
a German agricultural chemist
discovered the law of the minimum which Lawes had demonstrated at Rothamsted
Explain why nitrogen is essential for life?
nitrogen is an essential component in proteins and nucleic acids which are both key for growth
What type of catalyst can some bacteria use that fixes nitrogen like the plant in Billingham?
Nitrogenase: an enzyme protein
How have these bacteria acquired Nitrogenase?
evolution through series of gene duplication, mutation, and natural selection
Why did Laws suggest that wheat crops should be followed by legumes? Was this a new agricultural innovation?
legumes have the nitrogen-fixing bacteria that contain the nitrogenase enzyme to fix atmospheric nitrogen and maintain plant-available forms of nitrogen in the soil
no, Lawes did not discover this, crop rotation has been a thing since 79 AD
What’s guano? How has it been used in Chile?
bat and bird feces
in Chilean islands, accumulated feces was collected and used as fertilizer
What was the ‘second agricultural revolution’ according to Francis Thompson?
chemical fertilizers
How much nitrogen is fixed by humans every year?
150 million tonnes/year
What is the natural process for how fixed nitrogen returns to the atmosphere?
bacterial denitrification
How much of the world’s cereal production does nitrogen oxide cost?
3-16%
What 3 things does Wim de Vries (Wageningen, Netherland) attribute biodiversity loss to?
habitat destruction, climate change, nitrogen deposition
How does the Wibbly Wobbly article suggest bacterial nitrogen fixation can help solve issues caused by fertilizers in the environment?
coat seeds with nitrogen-fixing bacteria
What is a rhizome? Is it related to nitrogen-fixing roots?
no, rhizomes are a type of root but not specifically nitrogen-fixing
nitrogen-fixation occurs in root nodules of legumes in which bacteria occur
When the paper was written, what was the accepted belief about Crenarchaeota?
they were only found in thermophilic marine environments
what was the main finding of the research in the Crenarchaeota paper?
they are more diverse than originally thought
they are found in mesophilic freshwater sediment
what was the key molecule analyzed in the Crenarchaeota paper? why was it selected?
SSU/16s rRNA gene (DNA that codes for the RNA gene)
easier to extract DNA and use PCR than with RNA
the information is still relevant because the 16s rRNA / SSU rRNA is universal and highly conserved domains involved in protein synthesis
What are the main steps in the Crenarchaeota paper?
- collect samples at 2 lakes
- extract DNA
- amplify 16S rRNA gene using PCR and gel electrophoresis
- clone amplified genes into plasmid bBluescript KS+
- analyze clone library using RFLP
- sequence DNA of clones from each genetic group
- use pairwise distance coefficient and phylogenetic inferences to analyze sequence
In the Crenarchaeota paper, why did they sample lake sediment?
lakes were in proximity to the lab and aquatic sediment resembles marine sediment
What did the researchers in the Crenarchaeota paper need to do successful PCRs?
they needed:
target DNA from sediment
dNTP
MgCl2
primers (forward and reverse)
thermostable DNA polymerase (Taq polymerase)
thermocycler
controls (positive and negative)
How many primers were used and why were they chosen? Crenarchaeota paper
3 forward primers - 4Fa, 89F, 542F
1 reverse primer - 1492 RPL was chosen because there’s 1492 nucleotides in the DNA sequence
4Fa — 89F ——– 542F ——- 1492R —
4Fa - 1492R is the full size of the 16s rRNA
4a will amplify Archaea
89F targets the marine Crenarchaeota
542F targets for Archaea in general
Did the researchers from the Crenarchaeota do a PCR for each sample? why?
no, they did 2 consecutive PCRs with different primers (Nested PCR)
because it will increase the number of 16s rRNA copies of Archaea
How did they ensure their PCR was successful and not a result of contamination? Crenarchaeota
they used sterile distilled water
why did they need to clone the amplified 16s rRNA genes? how did they clone (what vector did they use)?
clone because sequences are different in different species regardless if the gene is the same
how:
insert each amplicon into a plasmid (pBluescript KS+) containing multiple cloning sites in the lacZ gene
the lacZ gene can change XGal into a blue colour
if lacZ gene is disrupted, there will be no colour change = white colonies are the ones with a successful amplicon insert
Why did the Crenarchaeota researchers do RFLP? would this method still be used today?
RFLP was used to search the clone library for genetic diversity and find groups with matching sequences
not really used today because sequencing is fast and cheaper than it used to be
which sampling site was the sequence retrieved from? Crenarchaeota
Lake Griffy
how many nucleotides have been sequenced from the Lake Griffy sample? Crenarchaeota
1322
Did the researchers sequence the entire 16s rRNA gene from the Lake Griffy sample? how can they tell if it was complete or partial? Crenarchaeota
no, it was a partial sequence
look for start and stop codons in the sequence to determine if partial or complete
Was the sequence from the Lake Griffy sample DNA or RNA? Crenarchaeota
DNA - it was for a gene
What is the name of the Lake Griffy clone and what group does it belong to? Crenarchaeota
pGrfA4 from group 1 (17 sequences) which is different from group 2 and 3 which have 14 and 1 sequence
