Nitrogen nurses

Alfred Lord Tennyson puzzled over the conflict between love as a foundation of Christianity, and the apparent violence of the natural world.

Who trusted God was love indeed

And love Creation’s final law

Tho’ nature, red in tooth and claw

With ravine, shriek’d against his creed

The good poet would be relieved to learn that modern ecologists have uncovered a softer, gentler side of the natural world – facilitative interactions, in which one species (the facilitator) helps out a second species. In many, but not all, cases, the second species also helps out the first species. Ecologists describe these mutually-beneficial interactions as mutualisms. As an example, Mimosa luisana is a mutualist with Rhizobium bacteria, providing the bacteria with root nodules to live in and carbohydrates as an energy source, while receiving ammonia (NH3) that the bacteria fix (convert) from atmospheric N2. A second type of mutualism, a mycorrhizal association, is a very common facilitative interaction between plants and fungi, which grow alongside or within the plant roots. In many mycorrhizal associations, the plant provides carbohydrates to the fungi, which import and share nutrients and water.

Mimosa plant

Mimosa luisana. Credit: Leticia Soriano Flores, algunos derechos reservados (CC BY-NC)

Alicia Montesinos-Navarro and her colleagues, and researchers before them, noticed that in arid and semi-arid environments, plant-plant facilitation was most common between two plant species that were structurally and functionally very distinct, and that tended to be very distantly related to each other. In particular, M. luisana tends to associate with many different species of plants, including many cacti that look nothing like it, and are very distantly related. M. luisana is called a nurse plant, because other species tend to grow under its branches, which shade the soil and reduce water loss from evaporation. Recent work by Montesinos-Navarro and her colleagues showed another benefit of nursing – some plants receive nitrogen from these nurse plants via the network of mycorrhizal fungi.

Traditionally, ecologists have argued that associations between distantly-related plants occur because the plants have very different ecological niches, using different resources in different ways, so they are not competing with each other. Montesinos-Navarro and her colleagues argue that a second process might be important in this and other systems. Close relatives of M. luisana might tend to have high nitrogen levels and thus not benefit from nitrogen transfer from the nurse plant, while more distantly-related plants might tend to have lower nitrogen levels and thus benefit from any nitrogen arriving from M. luisana. They explored this hypothesis in the semi-arid Valley of Zapotitlan in the state of Puebla, Mexico.

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Study site dominated by the columnar cactus Neobuxbaimia tetezo, Credit: Alicia Montesinos-Navarro.

Measuring nitrogen transfer from the nurse plant to the recipient is not the world’s easiest task. Fortunately there is a rare form or isotope of nitrogen, 15N, which can be distinguished from the more common 14N. The researchers soaked M. luisana leaves in urea that was made up of primarily 15N, and the leaves took up the urea. Consequently, any exported nitrogen would contain a disproportionately high concentration of 15N, resulting in high 15N levels in the recipient plant. They then measured 15N levels in 14 different species of plants that used M. luisana as their nurse. The researchers were able to test two hypotheses. First, they could see whether close relatives to M. luisana tended to have higher N-levels than more distantly related species. Second they could see whether distant relatives tended to receive more nitrogen from nurse plants than did close relatives.

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Mimosa luisana branch taking up 15N-labeled urea. Credit: Alicia Montesinos-Navarro.

The graph below summarizes the results. The y-axis measures how much the 15N level in the facilitated species increased by the end of the experiment (15 days). The x-axis measures the evolutionary relationship between M. luisana and the facilitated species – more precisely how long ago the two species shared a common ancestor. Lastly, the size of the dot measures the initial difference in leaf N-levels between M. luisana and the facilitated plant.

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Influence of evolutionary relationship between M. luisana and the facilitated species (x- axis) and nitrogen gradient – the initial difference in nitrogen levels between the two species (size of dots) on the amount of nitrogen imported by the facilitated species.

Several trends are evident. First, close relatives of M. luisana tended to have similar leaf nitrogen values to M. luisana (medium sized dots), while distant relatives tended to have much less nitrogen than M. luisana (largest dots). Second, the most distant relatives tended to have the greatest increase in their 15N levels, which indicates that they received the greatest nitrogen export from their nurses.

One question is how the nitrogen is transported. Montesinos-Navarro and her colleagues describe how they treated soil with a fungicide, presumably killing the mycorrhizae, which resulted in a substantial reduction in nitrogen transport. This suggests that the mycorrhizal network is important for nitrogen transport. But more pressing is what do the nurse plants get out of the relationship. The researchers suggest that the recipient plants may provide M. luisana with either water or phosphorus, both of which may be in short supply in arid environments.

This study indicates that we need to look beyond traditional niche theory, and may need to  dig deeper to understand the structure of plant communities, and how facilitative interactions can explain the coexistence of very distantly related plants.

note: the paper that describes this research is from the journal Ecology. The reference is MontesinosNavarro, A., Verdú, M., Querejeta, J. I., & ValienteBanuet, A. (2017). Nurse plants transfer more nitrogen to distantly related species. Ecology, 98(5), 1300-1310. Thanks to the Ecological Society of America for allowing me to use figures from the paper. Copyright © 2017 by the Ecological Society of America. All rights reserved.

Cottonwood genes and spider hostels

Back in my working days, people that I met would sometimes ask me what I did for my research. I usually told them that I studied spider sex, which, while true, was a bit misleading, as my interests were actually slightly broader. But studying spider sex was a good fit for my disposition, because, more than anybody I know, I can stare at something for a very long time and not get bored. And when spiders have sex, there can be very long periods, when, to our eyes, nothing is going on. As it turns out there is a great deal of pheromonal communication going on, and considerable vibrational activity as well, but it is not that easy for humans (even abnormally patient ones) to detect.

Taphrina

A spider and her three egg cases within a web she has built in a Taphrina blister. Credit Matt Barbour

The point is that I have a very soft spot in my heart for spiders, and was delighted to learn about an indirect process that provided a comfortable home for needy spiders. Heather Slinn got interested in her project while an undergraduate summer intern. Her colleague, Matt Barbour, pointed out that when he flipped over blistered black cottonwood leaves (Populus trichocarpa) he often found a spider hanging out in there. This observation led to her to study the relationship between cottonwood trees, cottonwood genetics, pathogens that make leaf blisters and spider occupancy rates.

Taphrina fungi form cup-like blisters in the leaves of Populous trees. But these trees vary in how susceptible they are to leaf blisters. The researchers wanted to answer three questions about this relationship. First, do spiders prefer to live in leaf-blisters as opposed to unblistered leaves? Second, are differences in tree susceptibility to Taphrina a result of genetic differences between the trees? And third can differences in Taphrina-resistance account for differences in spider density?

One of the keys to the experiment was establishing a garden with distinct clones of trees of known genetic makeup (genotypes). Slinn and her colleagues studied five different genotypes, with approximately 40 trees per genotype. They dutifully watered them throughout the summer, and then sampled up to 30 leaves from each tree for blister density, blister size, and spider residency, using a modified shop vac to suck up all of the spiders.

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Black Cottonwood garden. Credit Matt Barbour

The researchers discovered that blistered leaves were 35 times more likely to have a spider and/or spider web than were unblistered leaves. Clearly spiders found blistered leaves to be highly attractive homes.

But there were pronounced differences among the five genotypes in blister density, blister size, spider density and the probability that a spider was occupying a blister. Graph A shows that genotypes 1 and 3 (G1 and G3) had the lowest mean density of blisters (about 2 or 3 per meter of plant), while G4 averaged about 20 blisters per meter. Although G3 had relatively few blisters, it did boast the largest blisters (see graph B). The researchers concluded that blister density and size were both under genetic control, but not linked to each other.

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Mean (A) blister density (number per meter of plant) and (B) blister size, for the five tree genotypes.

But how did blister density and size influence spider residency? G4, the genotype with the greatest number of blisters per plant, also had the greatest number of spiders (graph C). But on a per blister basis, we can see that the two genotypes with the largest blisters (G2 and G3 – see graph B) also had the highest probability of housing a spider in their blisters (graph D). So when spiders make decisions about where to live, both size and number are important.

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Mean (C) spider density and (D) probability a blister hosts a spider, for the five tree genotypes.

It is unclear why the spiders are attracted to blisters.  Some spider species have been shown to be attracted to structural complexity, because that provides more or better attachment points for web strands and egg cases.  Depressed blisters may also give protection from abiotic factors such as wind and precipitation.

Like most good studies, this research raises a host of related questions. Why is there so much genetic variation in Taphrina-resistance within this tree species? Slinn suggests that there may be tradeoff whereby investment into Taphrina-resistance might compromise a plant’s ability to invest in other functions such as cold-resistance, or rapid growth and/or high reproductive rates. A second question is how does spider presence influence other species – for example does hosting a spider reduce the number of herbivorous insects that might attack the tree? A third issue raised by the authors is that plants infected with Taphrina may be weaker and more susceptible to herbivores. In that case, the spiders may be present in blisters because they are attracted to the herbivores that are eating the leaves. And finally, herbivores eating the leaves may transmit other diseases affecting our forests, such as dutch elm disease, chestnut blight and white pine blister rust. Unraveling this complex chain of events will keep researchers busy for many years.

note: the paper that describes this research is from the journal Ecology. The reference is Slinn, H. L., Barbour, M. A., Crawford, K. M., Rodriguez‐Cabal, M. A., & Crutsinger, G. M. (2016). Genetic variation in resistance to leaf fungus indirectly affects spider density. Ecology 98(3): 875-881. Thanks to the Ecological Society of America for allowing me to use figures from the paper. Copyright © 2017 by the Ecological Society of America. All rights reserved.