Annual plant neighbourhoods

As a team, we are discussing the fine-scale grain of sampling for estimating annual-annual plant interactions in deserts. We are particularly interested in the Mojave Desert to examine pollinator-herbivore interactions with annuals that are mediated by the other immediately adjacent congeneric species. Here is a brief compilation of key papers examining this challenge.

scale matters, a plant’s eye view

Publications describing the fine-scale annual plant neighbourhood concept

Mack, R. N. and Harper, J. L. 1977. Interference in dune annuals: spatial pattern and neighbourhood effects. – Journal of Ecology 65: 345-363.

Holzapfel, C. and Mahall, B. E. 1999. Bidirectional facilitation and interference between shrubs and annuals in the Mojave desert. – Ecology 80: 1747-1761.

Schiffers, K. and Tielbörger, K. 2006. Ontogenetic Shifts in Interactions among Annual Plants. – Journal of Ecology 94: 336-341.

Lortie, C. J. and Turkington, R. 2008. Species-specific positive effects in an annual plant community. – Oikos 117: 1511-1521.

Emery, N. C., Stanton, M. L. and Rice, K. J. 2009. Factors driving distribution limits in an annual plant community. – New Phytologist 181: 734-747.

Luzuriaga, A. L., Sánchez, A. M., Maestre, F. T. and Escudero, A. 2012. Assemblage of a Semi-Arid Annual Plant Community: Abiotic and Biotic Filters Act Hierarchically. – PLOS ONE 7: e41270.

Underwood, N., Inouye, B. D. and Hambäck, P. A. 2014. A Conceptual Framework for Associational Effects: When Do Neighbors Matter and How Would We Know? – The Quarterly Review of Biology 89: 1-19.

Underwood, N., Hambäck, P. A. and Inouye, B. D. 2020. Pollinators, Herbivores, and Plant Neighborhood Effects. – The Quarterly Review of Biology 95: 37-57.

Personal vote

I am a fan of the 15cm scale for fine-scale but often sample with a 15cm ring nested within a second 30cm metal ring. I construct using wire.

A workflow for pollen identification.


The reproductive ecology of cactus is not well-studied. A small, side project of mine is to determine the pollinator guild of buckhorn cholla at Sunset Cove, Mojave Desert, and with which plant species, if any, it shares pollinators. The genera Opuntia and Cylindropuntia are known to be insect-pollinated, but I am curious which of the more than 659 species of bees in the Mojave Desert desert are pollinators.

As visitation does not necessarily lead to pollination, I removed the pollen loads from 22 bee visitors I caught during insitu observation periods. I also removed stigma from the cholla to quantify heterospecific pollen deposition i.e. evidence of pollinator sharing. Pollen ID is not easy task and so I have developed a workflow to make it more streamlined.


Prep a reference collection:

  1. Create a reference collection by removing pollen from the anthers of several flowers of every species blooming in the area. Store in ethanol.
  2. Mount and stain the pollen with fushcin jelly.
  3. Image each species of pollen grain at 3 magnifications. Measure the length and width of about 10 grains per species. I calibrated Lumenera’s Infinity Analyze software using a stage micrometer to make this really quick.
  4. Make a reference document to consult. I use a word doc where every page is a species. Add in the photos at several magnifications, the mean size and any notes.
Sample reference page for Echinocereus engelmanni (Hedgehog cactus)


To go through the stigma or bee pollen load samples, I use my Canon EOS 60D dslr with a 60mm macro lens pointed confocally into a light microsite at 100x. I used the remote shooting utility from Canon to control the camera with my computer and display the view onto a second monitor.

Home example of confocal setup
  1. I designate each coverslip on the slide as a zone and do 8 transects through each, counting the grains. Each line in my spreadsheet is a transect, each column is a species. I use 5 columns for buckhorn so I never have to count very high.
  2. I don’t count damaged grains, or grains in air bubbles.
  3. Each slide gets its own folder. I take photos of each heterospecific grain with the file name as the zone + transect + species, which is simple using the photo utility. Knowing where the grain is on the slide and what its surroundings are will be helpful if you need to find it again.
  4. The species can be tentative for now so don’t get too bogged down.
  5. Take photos of unknowns when first encountered and assign them morphospecies ID. I put these in a separate folder as a reference.
  6. Some species are easy to ID. Quite a few are not. The more grains you see the easier it is to spot the differences.
  7. To help ID, we can take a page from entomologists. Sort the photos by their tentative IDs, putting each species in a folder so they are visible all at once (do a bulk rename to append the folder name first). It is difficult to compare grains unless they are side by side, which isn’t realistic with one microscope.
  8. Sort until each folder contains identical grains, then assign them a species from the reference collection. Or assign them to a species group for species that are virtually identical (likely Asteraceae!). Assign any remaining to morphospecies. Update the datasheet with the corrections.  
Buckhorn cholla (larger) and silver cholla (smaller). Thankfully the most abundant grains are simple to differentiate.

Quick notes on plant harvesting

Ok, our competition trials never look that good.

useful paper: Designs for greenhouse studies of interactions between plants

Protocols

  1. Aboveground harvest: clip from soil surface.
  2. Belowground harvest: need to harvest entire plant at once by removing plants and roots from pot experiments (for instance) and gently washing to remove all soil but keeping roots and shoots intact.  Then, snip aboveground growth from below.
  3. Depending on level of replication and lowest possible independent sample unit, harvest one individual, one species, or all individuals of one species per pot into independent paper bags.
  4. Place in ovens at 68F for at least 2 days.
  5. Leave all plants in paper bags in oven until the moment you are ready to weigh.
  6. Remove from paper bags to weigh for small plants. Typically, I weigh to 3-4 decimal grams for small desert annual plants.
  7. Return plants to bag, do not return to oven, store in a paper box for a few weeks or until all data entered and checked.

#density series idea notes

Concepts

Do native species density trials at two scale – micro and mesoscale using pots and large plastic buckets. Mix of potting soil and sand best – 50:50.

Consider intra and interspecific competition series – with replacement, i.e. keep net densities per pot consistent.  Finally, perhaps consider competition with an exotic such as red brome.

Designs
with and without shade mimic in greenhouse
umbrella over buckets in mesocosm experiment
natives against brome in density series

Natives

a

b

c

then a*b, a*c, b*c

Or run each out in solo then against red brome.

 

Round 1.
a vs rb
b vs rb
c vs rb
at 1,2, 5, 10, 25 seeds etc or a simple with-replacement density series.
THEN, ready for it…. red brome from cali and red brome from Israel
Round 2. 
a+b vs rb
etc.

 

Scales