Forest canopy fixes nitrogen shortage

The two billion hectares of forest canopy remaining on our planet are ideal habitat for nitrogen fixing microorganisms that can convert N2 to ammonia.

StantonCanopy

View of the forest canopy at the research site. Credit: D. Stanton.

The forest canopy tends to be nutrient-poor because there is no access to nutrients that accumulate in the soils on the forest floor, and rainfall can leach away any nutrients that do accumulate in the canopy from atmospheric deposition. So if you are a microbe, and you want to enjoy the view from the canopy, it is to your advantage to be able to fix atmospheric nitrogen so you can build essential molecules such as proteins and ATP.

As I mentioned in a previous post (Nitrogen continues to confound convention) both phosphorus (P) and molybdenum (Mo) are essential nutrients for biological nitrogen fixation.  Daniel Stanton and his colleagues hypothesized that nitrogen fixation in the canopy might be limited by the availability of P and Mo, so they designed a series of experiments to explore the role of these nutrients at the San Lorenzo Canopy Crane in San Lorenzo National Park in the Republic of Panama.  The crane provides about 1 ha of canopy access to non-acrophobic ecologists.

Stantoncrane

The crane at the research site: Credit: D. Stanton.

In one experiment, Stanton and his colleagues filled thin nylon stockings with vermiculite to form 40 cm long cylinders of 4 cm diameter.  Each cylinder was then soaked in either pure water (control), a molybdenum (Mo) compound, a phosphorus (P) compound, or a combination of Mo and P,  thus establishing four treatments. They attached each of these stockings to five different trees and allowed them to reside in the canopy for six months, to be colonized by microorganisms.

Stantonstockings

Nylon stockings treated with nutrients (or untreated controls) and affixed to branches in the canopy. Credit: D. Stanton.

The researchers measured the rate of nitrogen fixation by cutting a 50 cm2 rectangle from the area of densest growth on each stocking, and incubating it (along with the colonizing microorganisms) in a closed bottle that they had inoculated with heavy nitrogen (15N).  They then measured how much 15N the colonizers took up during a 12 hr incubation period.

Stantonfixationlab

Samples incubating for 12 hr to measure the rate of nitrogen fixation. Credit: D. Stanton.

The most common colonizers were nitrogen fixing filamentous cyanobacteria. These cyanobacteria fixed nitrogen at a somewhat (but not statistically significant) higher rate with Mo addition and at a much higher rate with P addition, and even more so with Mo + P addition.

StantonFig1A

Nitrogen fixation rates for each experimental treatment. C = control.  Note that the y-axis is logarithmic, so these differences in fixation rates are substantial.  Non-overlapping lowercase letters above the bars indicate significant differences between the means.

Nitrogen fixation is complex and costly.  Part of the complexity arises because nitrogenase, the enzyme that catalyzes the reaction, cannot tolerate oxygen.  To deal with this problem, cyanobacteria have evolved heterocysts, which are specialized anaerobic cells where nitrogen fixation occurs.  How does nutrient addition influence heterocyst abundance and function?

There are actually two aspects to this story.  One finding is that Mo addition had no effect on heterocyst abundance, while P addition had a pronounced effect.

StantonFig1B

Heterocyst frequency for each experimental treatment.

A second aspect is that Mo addition had a pronounced effect on the efficiency of nitrogen fixation.  For one analysis the researchers compared the nitrogen fixation rate per heterocyst for the phosphorus addition treatments either without or with Mo addition (in other words, they compared the P added treatment to the Mo + P treatment). Nitrogen fixation rates were much higher in the Mo + P treatments.  So while Mo does not increase heterocyst abundance, it does dramatically increase heterocyst fixation efficiency.

StantonFig2B

Quantity of N fixed per heterocyst per day in relation to absence (left bar) or presence (right bar) of Mo.  P was added for both treatments.  Dark horizontal lines are the median values, quartile range is represented by top and bottom of each box, and the whiskers represent the range of values for each treatment.

Phosphorus acts by markedly increasing the overall cyanobacterial growth.  It increases the amount of cyanobacteria that colonizes the canopy and also increases heterocyst density per filament. In contrast molybdenum’s effect is more nuanced as it increases the efficiency of the nitrogen fixation reaction without having any (obvious) effect on cyanobacterial structure.

How do these findings influence our understanding of tropical forests in the western hemisphere?  It turns out that episodes of nutrient addition actually happen in nature, courtesy of vast plumes of nutrient-rich rock-derived dust that periodically blow over the Atlantic Ocean from the Sahara desert in western Africa. Preliminary estimates by Stanton and his colleagues indicate that nutrient enrichment from these dust plumes is sufficient to  profoundly increase the rates of nitrogen fixation in tropical forests.  This may require us to reconsider our understanding of how nitrogen cycles within and between ecosystems.

note: the paper that describes this research is from the journal Ecology. The reference is Stanton, D. E., S. A. Batterman, J. C. Von Fischer, and L. O. Hedin. 2019. Rapid nitrogen fixation by canopy microbiome in tropical forest determined by both phosphorus and molybdenum. Ecology 100(9):e02795. 10.1002/ecy.2795. Thanks to the Ecological Society of America for allowing me to use figures from the paper. Copyright © 2019 by the Ecological Society of America. All rights reserved.

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