Monday, June 18, 2012

Fear is in the soil

Leaf litter and other plant debris are broken down in the soil by fungi and microbes, which decompose the organic matter into molecules which they can use.  We know that all living matter is connected through complex ecosystem interactions.  Changes in predator populations affect the prey populations which may then cause changes to the plants they eat.  Changes in plant populations would alter the amount of decomposition of leaf litter, the soil microbes and the amount of carbon dioxide released by the microbes.  This interconnection between predator and soil decomposition is not unexpected and seems fairly logical.  However, as reported by Hawlena et al. in Science last week, there is another way in which predators can affect the belowground communities.

Fear changes molecular composition of prey
When predators are present in an ecosystem, the prey are afraid and stressed out.  The stress actually causes a slight change in the ratio of carbon to nitrogen (C:N) in the bodies of the prey.  This has been reported before, but let’s try to understand why this would happen.  There are two very important types of molecules that affect the C:N ratio.

1) Sugars/carbohydrates: sugars are our main source of food and energy.  They are made of carbon, oxygen and hydrogen, but not nitrogen.  

 2) Proteins: proteins have many functions in the cell, but are usually not used as a direct source of energy. Proteins are made up of amino acids, each of which has at least one nitrogen atom.

Therefore, the amount of carbon is most affected by concentrations of sugars and the amount of nitrogen is most affected by the presence of proteins.

When prey are afraid and on guard against attack, they consume more carbs, because that’s the best source of energy.  They make less protein, because that process takes up precious energy that is needed to escape predators.  They also breakdown their remaining proteins into glucose to fuel the cells.  In other words, the prey increase their ratio of sugar to protein, which means they will have an increased C:N ratio.

When the prey with altered C:N content die in the soil, will this affect the soil microbes’ ability to decompose other materials?

Scare the grasshoppers, harm the soil
The authors set out to test this question in various settings.  They did an experiment in the field and did a number of others in a pseudo field/lab setting, so they could control the variables.  They raised grasshoppers in the field with or without the risk of spider predation.  The spiders scared the grasshoppers (understandably) and raised the grasshopper C:N ratio by just 4%.  The spiders couldn’t actually kill the grasshoppers because their mouths had been glued shut, so the authors sacrificed the scared grasshoppers and the controls and took them back to the lab.  They added the grasshoppers to large natural soil samples to let them decompose.  Both types of grasshoppers decomposed at roughly the same rates (measured by the amount of CO2 released by the soil microbes). 

The authors then added grass litter to the soils which had previously broken down the grasshoppers.  Remember there was only a 4% change in C:N ratio of the scared grasshoppers.  This small change caused a threefold decrease in decomposition of the plant material!  Small changes to the amount of carbon and nitrogen that is entering the soil can have a major impact on how well the soil breaks down plant litter.  Wow!

It’s the protein
The nitrogen content in the soil is very important to the functioning of decomposers.  Nitrogen is used by the microbes to produce enzymes that catalyze the degradation of complex carbohydrates (found in high concentrations in plants).  Therefore, the high C:N ratio in the scared grasshoppers is most likely hurting the soil microbes by offering less nitrogen.  To test this idea, Hawlena et al. created “artificial grasshoppers”, basically a mass of chitin (the shell of insects), carbohydrates and proteins.  They could change the ratio of carbon to nitrogen and specifically the amount of protein.  The more protein they added to the artificial grasshopper, the more the grass was broken down later on and CO2 emitted.  In other words, the microbes need nitrogen from the proteins of the dead herbivores in order to decompose the plant litter.  Even a small change in nitrogen content can throw everything way out of whack.

A grasshopper in the presence of predators (like the spider) will make less proteins.  When the grasshopper dies, less nitrogen (from protein) will enter the soil ecosystem.  The decomposers will not be able to make key enzymes necessary to break down plants, so the levels of decomposition will drop.

So a predator in an ecosystem can have effects all the way down to the way the tiny microbes in the soil decompose organic matter.

Wednesday, June 13, 2012

This is why pre-meds have to learn about fungi

By now we all know there are tons of bacteria living in our guts (we are made up of 10x more bacteria than human cells!)  The bacteria help us digest food and make new molecules.  If our microbiome gets disrupted it can lead to serious digestive problems.  It has also been suggested that our bacteria help us fight infections by interacting with the immune system.  Bacteria aren’t the only microorganisms living inside us, though.  It turns out there is a diverse population of fungi living in our intestines, as reported by Iliev et al. in Science last week.

Fungus like mushrooms?
When we think of fungus, most people think about mushrooms.  I can almost guarantee you no one is growing mushrooms in their colons.  There are lots of other types of fungi, though, including single-celled creatures like Saccharomyces “yeast”.  Fungi are one phylogenetic group, so they have characteristics in common with each other, including many of their DNA sequences. 

In order to test for fungi in animal guts, the authors did PCR to test mice for the presence of a certain fungus-only DNA sequence.  They found fungus throughout the entire mouse gastrointestinal tract, with the highest density towards the end of the colon (or large intestine).  They also found fungi in rat, guinea pig, rabbit, pig, dog and human feces!  I’m glad humans weren’t left out of this fungal bounty.

The authors isolated the DNA from mouse feces and sequenced all the fungal DNA they could find.  They identified over 100 different known fungal species and more than 100 potentially new species of fungi!  Most of these fungi were found in low concentrations, except for Candida tropicalis which accounted for 65% of all the fecal fungi.  Candida tropicalis is an opportunistic pathogen, which only causes a problem when it grows out of control in people with suppressed immune systems.

CandidaImage source

Fungal receptor
Our immune systems are trained to seek out and destroy foreign invaders like bacteria, viruses and fungi.  How does the immune system recognize fungi?  They have a cell wall (human cells do not) that has the molecule B-1,3,-glucans (a sugar polymer).  Macrophages in our immune system express a receptor called Dectin-1, which can recognize B-1,3-glucans and initiate an immune attack.  Since we have potentially pathogenic fungi living in our guts, our immune systems are probably always keeping them in check, so they don’t grow out of control and make us sick.  In fact, some of the diseases that cause inflammatory bowel disorders might be caused by fungi, and not bacteria, inducing an inflammatory response. 

To test this idea, the authors chemically induced colitis (inflammation of the colon) in mice (I’ll spare you a photo).  These mice were found to have circulating antibodies against fungal proteins, which implies that fungi have something to do with the pathology of colitis.  The authors wondered what would happen to mice which do not have the Dectin-1 receptor.  Since they can’t detect fungi, their immune systems would not be able to mount any attack.  Sure enough, the mutant mice had much worse colitis symptoms.  They had way more C. tropicalis (and other pathogenic fungi) in their guts than the wildtype mice which also had colitis.  In other words, missing the fungal receptor didn’t necessarily cause colitis, but it made the symptoms much worse. 

To further prove that colitis is aggravated by the native fungal population, the authors gave the mutant mice an antifungal drug during the induced colitis session.  These mice were much healthier than their siblings who did not receive the drug.  As long as something, an antifungal drug or the functioning immune system, can control fungal growth, then colitis is much milder.

Relevance to human disease
The inability to control fungi in the gut leads to more severe colitis in mice.  What about in humans?  Could a similar mutation be responsible for human colitis?  The authors focused on patients who have ulcerative colitis.  This comes in various forms; 30% of the patients have a severe form that does not respond to medical therapy.  They sequenced the gene for the fungal receptor and found that patients with the severe form were more likely to have a particular gene sequence compared to patients with a more mild type of colitis.  Though this doesn’t prove anything, it is in line with the idea that fungal growth can aggravate the inflamed colon.  Perhaps this sequence difference in the severe patients prevents their fungal receptor from working at full efficiency.

To summarize all this, we have fungus living in our intestinal tract.  Some of these fungi are totally harmless and I bet they help us with some physiological function (yet to be determined).  Other native fungi, including C. tropicalis, are potentially pathogenic, but are controlled by the immune system normally.  If the immune system cannot recognize fungal cells because the fungal receptor is defective, then the fungi will begin to grow out of control, which can upset the natural order in our guts.  Inflammatory bowel diseases will become more severe when the fungi are able to grow unchecked by the immune system. 

I have to say I don’t like the idea of a bunch of pathogenic fungi living in my intestines and just waiting for my immune system to let down its guard.  Stay vigilant macrophages!

Check out this excellent article by Carl Zimmer in the NY Times about the human microbiome.

Wednesday, June 6, 2012

What makes a bloody butcher taste so damn good?

Ever wonder what makes a good tomato taste so great?  Well, so do the plant geneticists trying to produce the “better” tomato.  The amount of sugar has a lot to do with it, but what about that tomato smell?  Our perception of taste is enhanced by how food smells before we put it in our mouths and as we chew it.  The chemicals that produce the tomato aroma are called volatiles.  The food industry assumed that the volatiles that are found in the highest concentrations are the ones that make a tomato a tomato, and these should be the targets of genetic manipulation.  A paper by Tieman et al, which appeared this week in Current Biology, challenges this thinking by systematically investigating what combination of chemicals found naturally in tomatoes makes a delicious tomato.

Mass produced tomatoes are relatively homogenous, so the authors decided to examine the chemical composition of 152 heirloom varieties.  I think it’s worth noting the names of a few of these varieties: Bloody Butcher, Giant Oxheart, Crimson Sprinter, Tasti-Lee, Mr. Stripey, and Mexico Midget.  

Bloody Butcher variety of heirloom tomato. (Credit: Totally Tomatoes)

After figuring out the concentration of chemicals in the tomatoes, the researchers asked consumers to rate the flavor of the tomato varieties.  Surprisingly, a number of generic supermarket tomatoes scored quite high.  There was no simple pattern of chemicals that defined a good tomato.  As you would expect, the flavor profile of a tomato is quite complex, but the authors were able to pull out some new interesting information from their analysis.

1) The volatiles that are the most concentrated in tomatoes do not necessarily correlate with perceived flavor intensity.  In other words, some of the odors that are in the highest concentrations are not associated with flavor intensity.  Take them or leave them, either way the consumer wouldn’t notice.  The authors proved this by testing the flavor of mutant tomatoes that cannot enzymatically produce some of the volatiles that are normally found in high concentrations.  There was no difference in preference between the mutants and normal tomatoes.

2) Some of the volatiles contributed to the perception of sweetness.  In particular, an odor called geranial was positively correlated with sweetness.  To investigate this further, they used a mutant tomato that could not make geranial but still had the same amount of sugars and acids.  Consumers rated these mutants as being less sweet even though the sugar:acid ratio was exactly the same as the normal tomato.  Think about that for a minute... a smell increased the sweetness of a food.  We could replace excess sugars in processed foods with geranial to lower the calories without affecting the overall taste of the food!

So what makes a bloody butcher tomato taste so good?  High levels of geranial and other volatiles that trick your taste perception into thinking you’ve bitten into a slice of heaven.