Stomatal Responses to Water

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Stomatal Responses to Water Status in Cottonwood Trees!

Eric Hansen and Lauren Huntington!

Biology 2023, Fall 2014!

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Abstract!

A study was done to test the responses of leaf stomata density to apparent drought

conditions. A stomata density count was taken from leaves of a Populus deltoides in Casper,

Wyoming where the tree grows alone in the center of a cement parking lot. Other counts were

taken from the same species along the North Platte River just a few miles away from the

parking lot tree. Previous studies show that stomata density actually increases in areas of mild

drought, but decreases in areas of severe drought. The following paper discusses how the

Populus deltoides’ stomata responds to the drought-like conditions of a parking lot in

comparison to the trees that have direct access to a water source. The drought-like conditions

of the parking lot appear to be severe enough to cause a decrease in number of stomata per

leaf. !

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Introduction!

We know that water is crucial to living things, especially plants. So imagine a plant that does

not have any direct access to a water source. It does not grow along the shores of a river or a

lake. It is not thriving in somebody’s backyard where it can take advantage of grassy, moist

ground. No, this plant is stationed right in the middle of a cement parking lot. This plant is a

Populus deltoides, or an eastern cottonwood tree and it lives an interesting, urban lifestyle.

There is no visible soil surrounding this tree. The entire base of it is engulfed by mounds of

cement. Yet, it seems to be thriving in this environment.!

This unusual (for Wyoming) situation brings about many questions. One question that arises

is how does this large cottonwood tree thrive in such a dry environment? We chose to

investigate how the lack of water this tree is exposed to might affect how many stomata are

present on its leaves compared to a tree of the same species that lives right on the shores of a

water source: the North Platte River. !

Stomata are minute aperture structures on plants found typically on the outer leaf skin layer,

also known as the epidermis. They consist of two specialized cells, called guard cells that

surround a tiny pore called a stoma. Their main function is to allow gases such as carbon

dioxide, water vapor and oxygen to move rapidly into and out of the leaf.1 We hypothesize that

the stomata density in the lonely, urban parking lot tree will be less than that of the same

species that has direct access to a water source. Due the restricted environment, the parking lot

tree will have less stomata because of its need to conserve the little water it receives. The tree

along the river will have a more dense stomata count because it doesn’t have to worry about the

water loss that occurs when stomata open and close due to its accessible water supply. !

In one study, moderate water deficits had positive effects on stomatal number, but more

severe deficits led to a reduction. The present results indicate that high flexibilities in stomatal

density and guard cell size will change in response to water status. 2

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Several reports have shown that the stomatal density and its index increase with water

stress, but the number of stomata per leaf decreases. With decreasing precipitation, stomatal

density also increases, whereas plant height, density, and leaf area decrease. An increase in

stomatal density was observed under moderate drought, but a decrease occurred with drought

severity. 3 !

Water stress can be defined as reduced water availability; either by water scarcity (drought)

or osmotic stress (high salt concentrations) or water logging; too much water. Water stress may

reduce photosynthesis, respiration and ion uptake, change the metabolic and growth patterns in

the plant and in severe cases result in plant death. In nature water stress is common either for

long or short periods of time, depending on the local climate. Most plants therefore have some

adaptation or response to enhance the growth and survival rate during water stress and

subsequent recovery. 4!

As a plant is exposed to moderate drought, a stomatal aperture may be decreased through

sensing physical or chemical signals such as hydraulic press and Abscisic acid. A decline of the

stomatal conductance may limit the net photosynthetic rate and water transpiration with

progressive water stress. 5!

The research seems to back up the hypotheses only if the drought conditions are extreme

enough. Experimental research was done to test the hypothesis. !

Materials and Methods!

! As stated above, we began our experiment with specimen collection of the leaves of

Cottonwood trees with different environmental conditions. Our controls were the trees located

within a 50ft vicinity of the North Platte River in Casper, Wyoming. We chose the Platte River

Parkway as our locale to gather the leaves for this portion of the research experiment (figure 2.)

It was afternoon on a sunny day. Temperatures were mild and there was little wind. We collected

small branches with approximately 6 leaves on each. The branches were approximately 195 cm

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off the ground. The trees that were collected from were approximately 75-85 cm in

circumference. !

The tree in the parking lot stands in the neighborhood of Paradise Valley (figure 1.) The

specimens from this tree were collected earlier in the same day and were gathered from a

branch within standing reach (approximately 195 cm). The urban tree was much larger in size

with its main trunk splitting into four separate trunks of 60-65 cm in circumference each. !

! All samples were taken to the laboratory that afternoon. After only a few hours, the

samples had begun to lose their flexibility and had become rigid. We quickly took our casts of

the stomata by applying clear nail polish to the undersides of the leaves and pulling the casts off

with clear packing tape. We chose the 4 best leaves from each tree to create our casts. !

! A week later we began our counts to gather the data for stomata density. Using a

microscope set to 400x magnification we counted the number of stomata in each .1385mm^2

window. For the River trees we counted the number of stomates in 4 separate windows of 3

different samples. The Paradise valley tree samples were collected from 3 separate windows of

4 sample leaves. !

! Our measurement techniques weren’t as specific as they could have been. More specific

and accurate measurements could improve the experiment design. The height of the leaves

were based upon the height of the person conducting the experiment and what was within

reach. We also had no way of telling how tall the trees were in comparison to each other. The

measurement of the circumference of each trunk was based upon the wing span of the person

doing the experiment as we did not have access to a cloth tape measure to ensure accuracy. !

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!Fig. 1 !

This map shows the location of the tree in Paradise Valley, Casper, Wyoming. Note the parking

lot surrounding the tree.!

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Fig. 2!

This map shows the location of the trees located along the Platte River in Casper, Wyoming.!

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Results!

Fig.3!

This graph shows our specific stomata counts per mm^2 for the tree located in Paradise Valley.!

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!Fig.4 !

This graph shows our specific stomata counts per mm^2 for the trees along the North Platte

River.!

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Fig. 5!

This line graph shows a comparison between the stomata counts of the Paradise Valley Tree

alongside the stomata counts of the Platte River Trees.!

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Fig. 6!

This graph shows the comparison in stomata density means between the tree located in

Paradise Valley and the trees located along the North Platte River.!

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! Our results indicate that our data supports our original hypothesis. The stomata density

of the trees located along the Platte River show an increase over that of the tree surrounded by

cement located in the parking lot in Paradise Valley. !

! The mean stomata density of the river trees averaged to 140.7942 stomata per mm^2.

(see fig. 6) The paradise valley tree showed a much lower mean number of 105.2089 stomata

per mm^2. (See fig. 6) !

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! Each tree showed highly comparable numbers in stomata density counts conducted on

the 12 samples, with a small degree of variance. The Paradise Valley tree was consistently

lower in stomata counts. (See fig. 3) The Platte River trees had consistently higher numbers in

stomata density counts of the 12 samples we collected. (See fig. 4) These results are visibly

apparent on the graph showing the comparison of stomata counts. (See fig. 5) Our p-value for

our data collected was 0.0001345. This result was highly significant which leads us to believe

there is a very slight chance that our hypothesis is incorrect.

Discussion!

Plants need water to survive. That’s a fact. By this logic, the amount of water a plant has

access to should affect it in one way or another. A study was done to compare stomata density

on an interesting Populus deltoides (eastern cottonwood) that grows in the middle of a cement

parking lot. Surrounded by cement, it is assumed this tree does not receive much water in the

dry environment of Casper, Wyoming. When precipitation does occur, there is not much, if any,

soil around the tree to absorb the water. !

We hypothesized that this situation would cause this tree to have less stomata than that of a

tree near the North Platte River where it would have direct access to a water source.!

According to a Tansley Review on the influence of environmental factors on stomatal

development, changes in the environment modulate the developmental and patterning

pathways to determine stomata frequency. This means that the two different collection sites with

different environmental factors should lead to a different stomata density. 6!

In one study that was done on leaf stomatal density and its relationship to water status, it

was found that moderate drought conditions actually caused an increase in stomata density, but

the more severe drought conditions caused a decrease in stomata density. 2!

This was again proven in another study where an increase in stomatal density was

observed under moderate drought, but a decrease occurred with drought severity. !

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Our results did, in fact, show a lower stomata density count in the parking lot tree (see figure

6). According to the other studies done, this would mean this tree is under severe drought

conditions compared to other trees along a river. !

As shown in figure 5, the stomata density for the counts on the tree near the river were

higher, on average, then the counts in the parking lot tree. The graph shows counts on 12

different spots for each tree. Although some points on the graph overlap, the tree with the direct

water access had a higher density on 8 of the 12 counts. The tree near the river also had a

higher mean density than that of the parking lot tree as shown in figure 6. The mean density

tells us that, on average, the tree near the river has a higher stomata density than that of the

tree in the parking lot. This information supports our hypothesis. !

Using the information gathered in our study and from what other studies say, we can accept

our hypothesis and say that the drought conditions in the parking lot are severe enough to

cause the leaves of the eastern cottonwood to have less stomata than that of the same species

with direct access to water along the river. Our p-value of 0.0001345 tells us that our findings

are highly significant, and there is a very small chance our hypothesis is incorrect. !

Future Directions!

The future of our study rests on a few different factors that could steer where our tests lead

next. Accounting for mistakes in our methods, we could do a more thorough stomata density

count. To further test our hypothesis, we could conduct precipitation tests for both locations. To

continue our research we could also conduct transpiration experiments on both our control and

our variable. If the Paradise Valley tree actually receives much less precipitation, these tests

could help support our original hypothesis.!

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Appendix

!Density numbers were figured by dividing the window size of 0.1385mm^2 by the number of

stomata counted in each.!

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PV Tree Platte River Trees

Stomata Number 15=108.303 Density Stomata Number 20=144.404 Density

Stomata Number 13=93.863 Density Stomata Number 21=151.625 Density

Stomata Number 14=101.083 Density Stomata Number 22=158.845 Density

Stomata Number 14=101.083 Density Stomata Number 25=180.505 Density

Stomata Number 15=108.303 Density Stomata Number 13=93.863 Density

Stomata Number 13=93.863 Density Stomata Number 17=122.744 Density

Stomata Number 16=115.523 Density Stomata Number 15=108.303 Density

Stomata Number 17=122.744 Density Stomata Number 16=115.523 Density

Stomata Number 15=108.303 Density Stomata Number 23=166.065 Density

Stomata Number 17=122.744 Density Stomata Number 19=137.184 Density

Stomata Number 10=72.202 Density Stomata Number 23=166.065 Density

Stomata Number 13=93.863 Density Stomata Number 20=144.404 Density

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!Cited References!

1. Swarthout, D. (2012). Stomata. Retrieved from http://www.eoearth.org/view/article/156262 !

2. Zhenzhu X. and Guangsheng Z. Responses of leaf stomatal density to water status and its

relationship with photosynthesis in a grass [Internet]. 22 July, 2008. Journal of Experimental

Botony. Available from http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2529243/!

3. Daszkowska-Golec, A. and Szarejko, I. Open or Close the Gate – Stomata Action Under the

Control of Phytohormones in Drought Stress Conditions [Internet]. 13 May, 2013. Frontiers

in Plant Science. Available from http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3652521/!

4. Arve, LE, Torre, S, Olsen, JE and Tanino, KK. Stomatal Responses to Drought Stress and Air

Humidity In: Shanker, A, editor. Abiotic Stress in Plants - Mechanisms and Adaptations

[Internet]. 22 Sept., 2011. Available from http://www.intechopen.com/books/abiotic-stress-in-

plants-mechanisms-and-adaptations/stomatal-responses-to-drought-stress-and-air-humidity!

5. Zhenzhu X., Guangsheng Z., and Hideyuki S. Plant responses to drought and rewatering

[Internet]. June 2010. Plant Signaling and Behavior. Available from http://

www.ncbi.nlm.nih.gov/pmc/articles/PMC3001553/!

6. Casson, S. and Gray JE. Influence of Environmental Factors on Stomatal Development. 19

Oct. 2007. Tansley Review.

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