Plant microbiome below ground

Root microbiome

3 main compartments

1. Rhizosphere – the soil tightly attached to the roots

2. Rhizoplane – microbes physically attached to the surface of the root

3. Endosphere – all the microbes that live in the internal compartments (within and between cells)

Microbial filtering

• Plant will select subsets of microbes which survive in its environment

• Factors that control the structure of the microbiome include:

  • edaphic factors (soil factors like pH)

  • rhizodeposits

  • host genome

  • host immune system

  • microbe-microbe interactions.

• These factors have varying degrees of importance from the rhizophere through to the endosphere

• Soil microbiome very diverse, only a small portion can survive in the rhizosphere, of those only a small portion can colonise the plant roots etc. With each level closer to the plant, you get increased selectivity of microorgansims

1. The rhizosphere 

• Hotspot of microbial activity

• Includes soil 2mm from the root surface

• Interface between the plant and the environment – lots of nutrient transfer

• Some plants can release up to 40% of their photosynthetic carbon into this space

• Plants are able to alter their rhizosphere depending on who they want to colonise their roots or what they need

• Difficult to define as its made up of lots of gradients, which makes it difficult to standardise across studies

  • E,g: as you move closer to the roots, you change oxygen concentration, the influence of different exudates present etc.

components of the rhizosphere: rhizodeposits

• Term which includes anything a plant does to alter its rhizosphere

• Includes:

  • Sloughed off root cap and border cells – acts as a lubricant to help cells move through

  • Mucilage

  • Exudates – carbons pumped out to feed microbes

  • Enzymes – pick up plants own nutrients

• Other forms of carbon exchange:

  • Volatiles – released when plants are attacked and used to signal to other plants that there are predators nearby

  • Symbioses

  • Cell death – carbon sequestered into soil

rhizodeposits: carbon excahnges 

Carbon exchanges

• Typically categorised in 2 broad groups of ways of releasing carbon

 Active

• Usually secretions

• High Mw

Passive

• Usually occurs via diffusion (can be passive or facilitated)

• Low molecular weight

Rhizodeposits: Exudates

Exudates are typically categorised into high molecular weight like mucilage and low Mw like the rest of them.

High molecular weight exudates - mucilage 

• High molecular weight and active secretion

• Functions:

  • Lubrication for a growing root

  • Reduces dessication

  • Soil aggregation – alters water and oxygen dynamics

  • Can help keep moisture gradient in drought

  • Certain types of roots house nitrogen fixers

  • Can help with microbial defence – traps pathogens

Low molecular weight exudates 

• Hundreds of low Mw carbon compounds

• High diversity

• Chemical fingerprint of the plant – use it figure out the species, lifestage, stress state etc.

• Generally grouped into organic acids, amino acids, proteins, sugars and phenolics (free and bound to lignins)

• Exudate function:

  • Nutrient acquisition

  • Antagonism/allelopathy – carbon compound that is toxic for other organisms

  • Microbial attraction

  • Allows for resource partitioning

• Each plant has a chemical fingerprint, things that influence this includes:

  • Breeding

  • Plant genotype

• Theories on why exudates exist: could be excess carbon but most likely has a role in shaping the symbioses

Physiological role of exudates

Exudates as a defence – some of the physiological roles of exudates

• Mucilage – physically traps pathogens

• Phytoalexins – compounds that plants release in response to infection

• Phytoanticipins – compounds that plants release all the time that act as antimicrobial peptides, such as benzoxanoids (nitrogen containing compounds)

• Phenolics – including flavonoids and phenylpropanoids

• Terpenoids

Microbial attraction – chemotaxis

• Exudates allows for chemotaxis to occur – recruitment of microorganisms through chemical gradients

• Bacteria localise to the root tips where the most exudates are

Exudates: phenolics

• Phenolics are antimicrobial but specialised slow growing microbes consume these phenolics – these microbes called oligotrophic

• Some phenolics, such as caffeic acid, are very inhibitory – each phenolic has a different role

• Some of them can cause an increase in respiration in microbes, some of them cause a decrease in respiration in microbes

Example of changes in exudate profile

• Correlation between sugars and phenolics

• In the emergent stage the root is flooded with sugars but as it grows, levels of sugars decrease as instead of forming the rhizosphere the plant is focused on maintaining the rhizosphere

how do we study exudates 

In the lab

• Put plant roots in a hydroponic solution and they release their exudates

• Can analyse these using mass spec, liquid chromatography

In the field

• Dig up the root of tress and place them in syringes containing solution

• Can now analyse solution using same techniques

function of the rhizosphere

• Physically structures the abiotic environment which alters the biotic environment.

• See a different chemical profile depending on the life stage of the plant – this environment that the plant generates controls which bacteria can survive – how they shape the rhizosphere.

Typical rhizosphere composition 

• Microbes enriched: r strategists which are copiotrophs (fast growing organisms with high turnover rate.

• depleted microbes: nitrogen fixing bacteria

• enriched bacteria:

  • Bacteroidetes

  • actinobacteria

  • proteobacteria

• depleted bacteria

  • nitrospira

  • acidobacteria

  • chloroflexi 

• Common functions:

  • denitrification

  • nitrogen fixation

  • urolysis

  • chitinolysis

• decreased functions:

  • nitrification

  • respiration of sulphur compounds

2. the rhizoplane

• Tightly attached microbes to the surface of the root itself

• Selective as microbes have to be able to survive and colonise the root morphology —> limitation of space forces microbes to compete 

• Now we have difficulties occurring with the plant immune system and the presence of:

  • PRRS

  • PAMPs

  • Flagellin proteins – lots of bacteria have flagella but not all flagellin proteins will set off the plants immune system, quite a high degree of sensitivity

3. root endosphere 

• Internal environment of the root itself, made up of endophytes

• Problems associated with the plant immune system

• Contamination makes it difficult to study – usually when you want to study the endosphere crush up the root and do 16SRNA amplification using PCR to identify colonisers. But chloroplasts and mitochondria also carry 16SRNA genes which contaminate your sample

Internal microbes

• Can colonise intercellular and intracellular spaces

• Require specialised genes

  • Usually have an enrichment of polymer-degrading enzymes (cellulases and pectinases for example)

  • Also often have secretion systems

Plant associated taxa in the endosphere 

• Lots of commensals

• Plant growth promoting rhizobacteria (PGPR) – umbrella term for microbes that help the host grow

• Mycorrhizal fungi

• Pathogens

plant-microbiome interactions 

Microbiomes have a lot of control around plant health physiology, can:

• Alter root structure

• Alter plant immune responses

• Impact plant nutrition

• Reduce abiotic stress

• Alter plat nutrient status

Examples:

• Auxin (indole 3 acetic acid) – microbes produce or degrade auxin to influence growth pathways

• Jasmonic acid and salycilic acid – growth hormones important for tsress signalling and immunity. Microbes can inhibit the production or produce more of it.

• Ethylene – important for stress responses in drought. ACC oxidase or ACC deaminase can increase or decrease ethylene levels.

• Giberellins – growth hormones which can be produced by some microbes

• Nutrient availability of phosphorous, nitrogen, potassium and iron

Specific example: example of pseudomonas syringae manipulating signaling

• Produces phytotoxin coronatine (mimics jasmonic acid) which supresses siacylic acid defence signalling

• By manipulating salicylic acid pathway means that it can manipulate the host immune system to be less effective

• Does this by stiulating auxin production and using a T3SS promotes lateral root formation, which allows the bacteria to invade more easily.

collecting the microbiome

• To extract the rhizosphere – dig up the roots, shake the roots and vortex the solution

• Rhizoplane – requires sonication of the clean roots which helps the helps the bacteria attached to the roots detach 

• Root endosphere – use the whole sonicated, cleaned root and crush it up to extract the DNA. Can then vortex this to create a pellet

plant symbioses

Not just one interaction, involves 3 different types of interaction:

• Mutualism – both organisms behefit

• Commensalism – one benefits, the other is unaffected

• Parasitism – one organism benefits, the other is negatively affected

most common plant mututalisms are 

1. mycorrhizal fungi 

2. legume forming bacteria 

1. mycorrhizal fungi

• Invade and create arbuscules inside the plant which allows for nutrient exchange

• Mutualistic relationship with plants

• Extension of the root system – allow a greater SA to reach nutrients

• Exchange P and N for carbon

• Can improve plant stress tolerance as well as water access

• 99% of all plants form this relationship – plants that don’t do this are abnormal

• Evolved around 400-500mya

• Vital for this territorialisation of plants

• 4-20% of the photosynthetic carbon is transferred to the fungus in the form of sugars and lipids, in exchange the fungi exchange phosphorus and nitrogen 

mycorrhizal groups

• Endomycorhhizal fungi

  • Arbuscular mycorrhizal fungi

  • Ericoidmycorrhizal fungi

  • Orchid mycorrhizal fungi

• Ectomycorrhizal fungi

• Ectendomycorhizal fungi

  • Arbutoid mycorrhizal fungi

• Endomycorhhizal fungi

  • Arbuscular mycorrhizal fungi

  • Ericoidmycorrhizal fungi

  • Orchid mycorrhizal fungi

• Key interaction structures are arbuscules inside the cells of the roots

• 80% of plants have this type of fungi

• Vital for territorialisation alongside other fungi, mucoromycota

• Obligate – fungus cannot get carbon from anywhere else – requires carbon from the pant

• Invade the cell but plant cell wraps a membrane around the arbuscules so they don’t enter the cytosplasm

• Endomycorhhizal fungi

  • Arbuscular mycorrhizal fungi

  • Ericoidmycorrhizal fungi

  • Orchid mycorrhizal fungi

• Primarily ascomycota and some basidiomycota

• Associated with Ericaceae species (blueberries)

• Acidic and infertile soils

• Produce coils inside of the cells

• Some of them can also form ectomycorrhizal associations

• Can still pick up carbon themselves due to degradation genes

• obligate interaction

• Endomycorhhizal fungi

  • Arbuscular mycorrhizal fungi

  • Ericoidmycorrhizal fungi

  • Orchid mycorrhizal fungi

• Primarily restricted to the Basidiomycota (specifically Agaricomyces)

• Associated with orchids – orchids are partially mycoheterotrophic which means that it relies on the fungus to feed it in certain parts of its lifecycle. For an orchid seed to germinate it must associate with this fungi.

• Produce coils as their interaction structure

• Retain more degradation genes than ectomycorrhizal fungi

coils

• are the interaction structures for ericoid and orchid mycorrhizal fungi

• Orchid seeds may absorb carbon from hyphae, forming coils inside the cell

• Coils are degrade and release the nutrients into the host

stimulating this interaction

• When plants are phosphorous starved, they produce strigalactones

• Fungal spores sense the striga lactones using lipochitooligosaccaride mycorrhizal factor (Myc factor) which cause them to germinate and colonise the host

• Causes this calcium spike known as common symbiosis pathway

• Fungus invade into the root and cause the production of arbuscules

• So obligate that the fungi don’t even produce its own lipids for growth (E.g: beta monoacylglycerol is a lipid required for fungal growth but can’t be produced by the fungi)

Ectomycorrizal fungi

• Not restricted to one taxonomic group, can be ascomycota or basidiomycota

• Does not penetrate into the cell itself – intercellularly

• Particularly associate with temperate trees due to nitrogen acquisition – as plants move further north theres less of a phosporous depletion and more depleted in nitrogen

• Key features are mantle and Hartig net

  • Mantle is a fungus layer covering the entire surface of the root. Exudates and water have to pass through this mantle

  • Hartig net is the fungus that forms between cells

• Still have the genes and the enzymes to degrade soil organic matter so are not obligate

• Can produce fruiting bodies – mushrooms

Ectendomycotizal fungi: Arbutoid

• Basidiomycota

• Associate with a subset of plant species within the Ericaceae species

• Form this mantle but can also go inside the cells (aspects of both)

Other groups: Monotropoid

• Basidiomycota

• Also associate with the Ericaceae

• Associate with plants without chlorophyll

• Key characteristics include the Hartig net and fungal peg

• No penetration into the cytoplasm

• Mycoheterotrophy – as these plants don’t photosynthesise, they acquire carbon from the fungi

comparing interaction structures

root traits and mycorrhizal fungi

• Nice relationship between root traits and microbiome associations

• Roots that are thicker, tend to be associated with more mycorrhizal fungi

• Other trees make really thin roots which are less reliant on the mycorrizal fungi due to the increased surface area – also means that they’re less invested in their symbiotic partner

• Some tree species tend to invest more into a partner, some tend to be more independent

plant parasitism

Mycoheterotrophy

• fungi which obtain nutrition from other plants (e.g: achlorophyllous fungi)

Striga

• Parasitic plant which responds to striga lactones and colonises it and takes the carbon

Could consider mycorrhizal fungi in some instances to be parasites

• Where you have a less co-operative fungal partner, costs more carbon to get the nutrients

• Not parasitism in the classical sense because one of the partners not being harmed

• Important when we use these fungi to stimulate growth as we want to use the fungal partner which will be the most co-operative

2. root nodule symbiosis (legume forming bacteria) 

• Nitrogen fixing root nodule symbiosis (rhizobia)

• Paraphyletic: alphaproteobacteria and betaproteobacteria

• Ancient relationship

• Nodules vital for nitrogen acquisition

• Rhizobia fixes nitrogen and sends it back in exchange for carbon and safety.

stimulating the rhizobial partenership

• Stimulated by flavonoids

• Lipochitooligosaccharides (Nod factors) are released and recognised by Nod factor receptor 1 (NFR1) and NFR5

• Common symbiosis pathway causes calcium spikes

• Rhizobium associated with a speciifc area of the root hair and Nod signalling causes localised calcium spiking. This causes the growth area to curl towards the attached bacteria.

• These continuously repeated, localized shifts in growth direction—driven by the shifting calcium gradient—cause the root hair to grow around the bacterial colony, eventually leading to the formation of a tight curl that entraps the bacteria

• This mechanism is also what allows the plant to form a structure called an infection thread which facilitates the entry of the nitrogen fixing bacteria into the root

ecosystem services - the bigger picture of plant symbioses

• Forests that are full of arbuscular-associated plants produce higher quality litter

  • More nitrogen in litter

  • Breaks down faster

• Results in overall changed ecosystem function in the forest

• Form and quality of the exudates in the rhizosphere imacts global carbon budgets

• Mycorrizal fungi can pull carbon from multiple plants at one time (proved by radiolabelled isotopes)

Applied potential 

• Use microorganisms to change the nutrient cycling dynamic of that environment

• Phosphorous is a limiting nutrient

• So much phosphorous is fixed or washed away from the soil and so only a small portion of it is available to the plant host

• We can leverage microbial activity to make phosphorous more bioavailable

• Can use ectomycorrhizal fungi to understand which tree species will thrive when trying to restore certain forest areas.