Exercise 4

Environmental Parameters as Limiting Factors

I. Introduction

The term limiting factor is defined as any factor or any environmental parameter
that when it is decreased or increased, absent or present, it affects the growth, metabolic
processes, abundance, and distribution of organisms in the natural environment.

The response of an organism to a parameter has a minimum, optimum, maximum

effect between which lies a range of gradient that is known as the zone of tolerance which
organisms are still capable of functioning and further that range known as the zone of
intolerance can cause death of the organism.

This exercise intends to investigate the responses of plant and animal species to
varying levels of environmental factors and to determine the factors’ limiting effects such
as: temperature on Aeges aegypti (mosquito) larvae, soil moisture on Oniscidea(sowbugs),
pH on Zea mays (corn) and Artemia (brine shrimps), and salinity on Zea mays (corn) and
Cornu aspersum (snails).

II. Materials and Methods

A. Temperature Effects

Nine beakers were filled with tap water and its temperatures were adjusted. Three
beakers were labelled as jar 1, another three as jar 2 and the rest three jars as jar 3 so that
every jar has three trials. All jar 1 served as the control set-up (room temperature) and its
water temperature is recorded. For all jar 2, the water temperatures were lowered to about
10-15 °C by adding ice water. For all jar 3, the water temperatures were increased to about
40-45 °C by adding freshly boiled water. Temperatures were measured by the use of
mercury thermometer and were maintained for 20 minutes.
 Two mosquito larvae were placed carefully into each jar for 20 minutes. Motility and
swimming movements were observed and noted. After 20 minute exposure, living larvae
were counted and % mortality for each particular temperature was determined.

B. Soil Moisture Gradient

Four oven-dried soil samples were weighed for about 500 grams each and were
placed in different quadrats in a box. The soil samples were labelled as sample 1, 2, 3, 4.
Sample 1 was left as it is, sample 2 was added with 10 ml of tap water, sample 3 was added
with 30 ml of tap water, and sample 4 was added with 60 ml of tap water.

20 sowbugs were placed at the center of the box. After 20 minutes, sowbugs found
in different quadrats were observed and counted.

Three trials were made but before doing another trial, the sowbugs were allowed to
recover from stress.

Chi-square test was used to measure the degree of correspondence of observed

values with the variables. Values expected on the basis of no differential response therefore
may be calculated by dividing the total number of individuals counted during the run by
the number of sections in the gradient. To compare the expected and observed numbers of
each section of the gradient, the value of chi-square (x2) was calculated by the following
formula:
(Observed - Expected)2
x2 = ∑ Expected

C. pH Effects

i. On Zea mays

Test solutions of pH 2, 4, 7, and 9 were prepared by adding hydrochloric acid and/or

sodium hydroxide to water. pH were measured by the use of pH meter.

Twelve petri-dishes with filter-paper at the bottom were prepared. Into each dish,
nine corn seeds were placed and moistened with the following solutions:
 1st set of 3 dishes – pH 2
2nd set of 3 dishes – pH 4
3rd set of 3 dishes – pH 7
4th set of 3 dishes – pH 9

After a week, seed germination and shoot length (cm) of the corn seeds were
counted and measured. The formula that is used to calculate the percentage germination of
each dish is:

% germination = number of germinated seeds x 100

total number of seeds

ii. On Artemia

Four culture tubes containing 100-150 brine shrimp eggs were filled each with ¾
solution of a particular pH namely, pH 2, pH 4, pH 7, and pH 9. Three trials were made

After a week, the number of living shrimps in every pH solution was counted using a
dissecting microscope.

D. Salinity Effects
i. On Zea mays

150 corn seeds were soaked overnight. Solutions with different salt content such as
0 ppt., 2.5 ppt., 5 ppt., 10 ppt., and 20 ppt., were prepared by dissolving a varying amounts
of salt in 1000 ml distilled water. No salt was added to yield 0 ppt., 2.5 grams salt was
dissolved to yield 2.5 ppt., 5 grams salt was dissolved to yield 5 ppt., 10 grams salt was
dissolved to yield 10 ppt., 20 grams salt was dissolved to yield 20 ppt.

Fifteen petri-dishes lined with filter paper at the bottom were placed with seven
soaked but ungerminated corn seeds. Each set of three were moistened with the
corresponding salt solutions.

After a week, the root and shoot length were measured in cm using a ruler in each
dish. The root/shoot ratio of the germinated seeds in each treatment was calculated.
 ii. On Cornu aspersum

Five jars were filled with corresponding salt solutions as described above. Into each
jar, five freshwater snails were placed.

After a week, the total number of dead snails in each jar was recorded.

III. Results and Discussion

The following tables and figures below represent the data gathered from the
experiments according to the exercise objectives which is to investigate the responses of
certain plant and animal species to varying levels of temperature, moisture, pH and salinity,
in order to came up with a determination of the limiting effects of such factors. This part
will elaborate more through table and chart interpretation of the gathered data.

Trials

Figure 1

Figure 2

Temperature (°C)
 In figure 1, there is no significant result in the data of number of living larvae vs.
temperature since all larvae survived even in the varying temperatures. It is because of the
limited time given to be observed which is 20 minutes but if it is about days, the larvae
population will become lowered because some may not be able to tolerate the harsh
environment which is when the temperature is lowered to 10-15°C or increased to 40-45°C.
The mortality percentage for each particular temperature was 0%.

Illustrated in figure 2, when the temperature is increased or decreased from 29°C

the motility of larvae became slower compared to its motility when it is in 29°C. The
exercise in figure 1 and 2 can be explained with Shelford’s law of tolerance which states
that the absence or poor performance of a species may be controlled by the qualitative or
quantitative deficiency or excess of any factor that approaches the limit of tolerance.

In the exercise, one of the factors used is temperature. The mosquito larvae has a
specific optimum range for temperature for them to survive and function normally which is
29°C in the exercise, minimum which is 10-15°C, and maximum which is 40-45°C.

Trials

Figure 3

water (mL)
 As we can see in figure 3, given three trials, sowbugs prefer soil added with 30ml
water than others, not too moist not too dry. Sowbugs naturally live in damp habitats such
as under rocks, dead leaves, flowerbeds or other objects on damp ground where they feed
on decaying plant matter or organic debris. Sowbugs are dry-land crustaceans, they
breathe through their gills, thus they need moisture in order to respire but not too moist so
that they can compensate properly with the environment by helping in the degradation of
organic matter.

This exercise shows two combined ideas; the limits of tolerance, and the factor
compensation. The optimum soil moisture for the sowbugs in the exercise, is when the soil
is added with 30ml tap water. This is the ideal moisture for sowbugs to survive and to
compensate.

Table 1

The chi-square value in the exercise is 7.4 and the degrees of freedom is 2. Since our
x2 statistic (7.4) exceeded the critical value for 0.025 probability level (7.30), the null
hypothesis should be rejected because the observed values and the expected values have a
highly significant difference in the number of sowbugs.
 Figure 4

Figure 5

Seed germination and seedling growth are influenced by many factors however in
the exercise the factor used was pH. The optimum pH range condition for a corn seed is
between 5.5-6.5. In the exercise all had germinated in pH 7 in all three trials and also
yielded the longest shoot lengths with an average of 78.55 cm followed by pH 9 (average=
57.33cm), 2 (average= 42.33cm), and 4 (average= 28.33cm). Shelford’s law of tolerance
can be applied here with corn having a wide range of tolerance in pH since it still grows
despite the varying pH used but the rate of growth is much slower when the pH is
increased or lowered from pH 7.

Figure 6

Brine shrimp are crustaceans that are classified in the phylum Arthropoda (the
largest phylum in the animal kingdom, which includes insects and other creatures with
jointed legs and exoskeletons). They live in inland bodies of saltwater with an ideal pH
conditions of around 8, but no lower than 5 and no higher than 10.

The reasons why it only hatched one brine shrimp in pH 9 is because of the set-up
and apparatus used. In order to successfully hatch and raise brine shrimp, the materials to
be used should be a wide-mouth quart jar instead of a test tube, the water to be used
should be salt water instead of fresh, number of brine shrimp eggs should be lowered about
50 rather than 150, and there should be constant air-pump and food supply after 1-3 days
when the eggs start hatching.
Figure 7

Figure 8

Figures 7 and 8 used the factor salinity to determine its effect in corn seed
germination and snail survival. We can deduce in the graph that as salt concentration
increases, the number of germinated seeds decrease down to 14% germination while the
number of dead snails increase up to 100% mortality. This is because of the concept of
osmosis and tonicity. Osmosis is the net movement of water molecules across a semi-
permeable membrane from a region of low solute concentration to a region of high solute
concentration (until equilibrium is reached) while the ability of an extracellular solution to
make water move into or out of a cell by osmosis is known as its tonicity.

Three terms—hypotonic, isotonic, and hypertonic—are used to compare the

osmolarity of a cell to the osmolarity of the extracellular fluid around it. In the case of corn
seeds and snails, as the salt concentration is increased, the water inside their cell tends to
move out faster to the region of higher salt concentration which is outside of the cell hence,
it will cause in the shrinking of plant and animal cells due to water loss. We know that
water is very essential in survival and growth of organisms and when water is absent it is
expected that organisms will not be able to grow or survive and will eventually die.

IV. Conclusion

Based on the observations and results presented, the following conclusions

are drawn:

In order to grow and survive, every individual organism of a species population,

need certain elements or abiotic factors. Out of these naturally occurring elements, some
are very essential and needed in a large amount whereas others are needed in very small
amounts. This is the law of limiting factors which is also incorporated in the law of
minimum that when an essential factor is in its least supply it will tend to be the limiting
one.

Every factor, there is a limit, there is a range in which an organism can tolerate.
There is a lower and an upper limit in the range of environmental factor between which it
functions efficiently and organisms may have a wide range or a narrow range of tolerance.
(Shelford’s law of tolerance)

Organisms are not actually living at the optimum range (as determined
experimentally) with regard to a particular physical factor. In such cases some other factor
or factors are found to have greater importance.

V. Answers to Questions
 1. How does temperature affect living organisms? Discuss the cellular and
physiological basis of temperature effect on organisms.

Temperature has long been recognized as an important environmental factor in

both terrestrial and aquatic ecosystems in observed to its key role over biological activity
such as development, growth and reproduction of organisms. Mainly temperature
influences enzymatic reactions through hormonal and nervous control to digestion, from
respiration and osmoregulation to all aspects of an organism’s performance and behaviour.

 Temperature and cell:

The minimum and maximum temperatures have lethal effects on the cells and their
components. If too cold, cell proteins may be destroyed as ice forms, or as water is lost and
electrolytes become concentrated in the cells; heat coagulates proteins (Lewis and Taylor,
1967).

Most of metabolic activities of microbes, plants and animals are regulated by varied
kinds of enzymes and enzymes in turn are influenced by temperature, consequently
increase in temperature, upto a certain limit, brings about increased enzymatic activity,
resulting in an increased rate of metabolism.

 Temperature and physiology:

The study of physiological ecology begins with an understanding of the influence of

ambient temperatures. Organisms have evolved a multitude of mechanisms for regulating
the temperature of their bodies in order to flourish in their particular environment. A high
metabolic rate helps supply heat. An efficient circulatory system conducts and distributes
heat around the body. Hibernation enables animals to survive long periods of cold.

2. Explain the Hoff’s rule. Is this applicable in all environments? Why?

  Temperature as an Environmental Factor

Within a limited range of temperature, the rate of chemical (biochemical) reactions

double with every 10°C rise in temperature. This is known as Van’t Hoff’s rule. Thus
temperature exerts a profound influence on the physiological activities of organisms. So yes,
it all applies in all environments because every physiological function has temperature
limits-an optimum, minimum, and maximum temperature. Organisms differ in their
tolerance limit to extreme temperatures.

3. What do eury- and steno- prefixes imply in ecology? How do organisms vary
in their range of tolerance?

An organism with a narrow tolerance range is said to be "steno.." for that factor (e.g.
stenothermal or stenohaline). An organism with a wide tolerance range is said to
be "eury.." (e.g. eurythermal). Organisms can be steno with respect to one factor and eury
with respect to a different one. These tolerance ranges are not independent. An organism
near the tolerance limits for one factor will probably be under stress, so its ability to
tolerate other factors will be reduced. The tolerance ranges for environmental factors
partly define the organism's niche. If salinity tolerance is graphed against temperature
tolerance, a niche area is defined.

Shelford’s Law of Tolerance

Every environmental factor varies through a wider range of intensity, which any
organism can tolerate characteristically. Each individual has a lower and an upper limit in
the range of environmental factor between which it functions efficiently. For any one factor,
different organism finds optimal conditions for existence. So the minimum quantity /
condition for any factor in the list of an organisms requirement is called its threshold level.
It may be any chemical or physical factor such as light, temperature, moisture or phosphate
etc. Above threshold the rate of function increases more or less rapidly until a maximum is
reached beyond which there is usually a decline in the rate of process either because of
deleterious effect produced, or interference of some other factor.

4. What is the significance of soil moisture to soil dwelling organisms?

 The amount of water present in a soil sample is called the moisture content.
Moisture is very important and its amount needs to be in careful balance—not too dry and
not too wet—for organisms to thrive. Specifically, the soil’s moisture content needs to
match the needs of the plants, animals and other organisms living in the habitat. Most often
soil with evenly balanced moisture is a haven for plants and small, soil-dwelling animals for
there is more food, and great shelter for them.

5. Discuss the physiological effects of pH changes on living organisms. How do

results here relate to the problem of acid rain?
 Effect on Flora

According to National Geographic, acid rain is particularly harmful to trees and

other plants. Acid rain adds aluminum to the soil and destroys important nutrients. As a
result, trees and plants are less able to absorb the ground water they need for growth.
Additionally, acid rain generally damages plant health, making them less resistant to insect
damage and disease. Acidic water similarly affects aquatic plant life, destroying important
food sources.

 Effects on Fish

Acidic water robs fish and other aquatic species of sodium in the blood and oxygen
in the tissues. Additionally, it affects the functioning of fish gills. Some species tolerate
acidic water better than others. Even if the acidity does not kill fish, the additional stress
can stunt growth and make them less able to compete for food. Acidic water also poisons
fish eggs, as they will not hatch if water pH is too low. The U.S. Environmental Protection
Agency has found that most eggs will not hatch in water with a pH level of 5.0 or less.

 Effects on Microorganisms

In addition to the direct effects on fish, acid water also destroys ecosystems by
killing organisms lower on the food chain. For example, the Pennsylvania Department of
Environmental Protection found that mayflies are particularly vulnerable to acidic water,
because it lowers sodium in the blood. Consequently, species feeding on mayflies will leave
an affected area, or starve.

6. Provide the physiological explanation of the effect of salinity on fresh water

organisms as well as certain plant species.
 The Effect of Salinity on Plants

Salts in the soil water may inhibit plant growth for two reasons. First, the presence
of salt in the soil solution reduces the ability of the plant to take up water, and this leads to
reductions in the growth rate. This is referred to as the osmotic or water-deficit effect of
salinity. Second, if excessive amounts of salt enter the plant in the transpiration stream
there will be injury to cells in the transpiring leaves and this may cause further reductions
in growth. This is called the salt-specific or ion-excess effect of salinity (Greenway and
Munns, 1980). As salinity is often caused by rising water tables, it can be accompanied by
waterlogging. Waterlogging itself inhibits plant growth and also reduces the ability of the
roots to exclude salt, thus increasing the uptake rate of salt and its accumulation in shoots.

 The Effect of Salinity on Animals and Environment

Changing salinity is a master factor in the distribution of both marine and estuarine
species and is limiting to freshwater organisms; hence salinity is fundamental in modifying
aquatic ecosystem assemblage structure and functioning. The effects of changing salinity
on the ecology of different habitats is driven ultimately by the underlying physiology and
tolerances of organisms and their ability to cope with salinity fluctuations on both long and
short time scales. Estuarine species are often euryhaline, adapted to tolerate fluctuating
salinity, whereas many marine species are stenohaline and limited by their narrow range of
physiological tolerance. For estuarine species, lowered salinities may be a subsidy, i.e.
benefiting the organisms by reducing competition, whereas for non-tolerant species they
are a stressor. Salinities at the margins or outside the tolerance range of particular species
will prevent their occurrence, change their behaviour, or limit reproduction and
germination, reducing their fitness for survival in that environment. Salinity can act
synergistically or antagonistically with other environmental stressors, producing a
dynamic and changing environment.
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