The Study of Zooplankton and Invertebrates in the Biola Creek

Sam Hammer

Biology II Lab Section 5

Professor Billock

Friday, May 14th, 2015

The Department of Biological Sciences, Biola University, La Mirada, California

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Abstract

California is a destination from chaparral to predominate dessert that is rich in biological

diversity. Found abundantly in almost every region with different abiotic factors is the

zooplankton and invertebrates of the Biola Creek in the La Mirada community. The purpose of

this study was to figure out how abiotic factors affect the diversity of zooplankton and

invertebrates. By recording species that were retrieved at one location with different varying

abiotic factors, in order to better understanding which region (Deep or Shallow water)

zooplankton and invertebrates prefer to inhabit. The area of the creek where we took water

samples and data was behind the Horton dormitory, and it was the main location at which

samples were retrieved.

After further testing, diversity of zooplankton and invertebrates among the location

displayed a relationship with the changing abiotic factors. Each location displayed a different

level of diversity. The deep waters with and without algae displayed the highest level of diversity

and the shallow waters with and without algae displayed the least amount of diversity. Thus, the

results show that different abiotic factors ultimately lead to different levels of diversity in the

Biola Creek.

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Introduction

There are many micro and macro-invertebrates found abundantly in freshwater streams.

They play a major role in their habitat’s food web and are considered bio-indicators of the water

quality in which they inhabit (Camargo and Alvaro 2005). Species of these chaparral stream

macro-invertebrates have received little attention and are in need of more research (Camargo and

Alvaro 2005). Communities of zooplankton and invertebrates rely on abiotic factors for survival

and these factors effect there sustainability in maturation.

Different geographical locations may contain different abiotic factors that can influence

the diversity of the zooplankton and invertebrates in their communities. All Invertebrates are

going to have specific adaptations in response to their current geographical location, but a

different geographical location may lead to different traits and diversity (Armstrong 1964). This

means a species’ traits may show better adaptation to certain geographical locations as opposed

to other locations. A factor such as deep water, in which species that are adapted to inhabit

deeper water have greater diversity than species that are adapted to shallow water in a location

abundant with algal growth.

There are multiple factors that play a major role in the diversity of invertebrates. Water

temperature is a significant factor in an aquatic insect’s life cycle (Sefton 1972), affecting

successful development and can prevent them from inhabiting certain locations.

Surrounding vegetation (algae) can also be very crucial in determining diversity, by affecting

stream temperature and debris involvement. Invertebrate species rely heavily on the surrounding

vegetation for food and most are categorized as collectors and grazers (Walsh 2005). Species of

invertebrates categorized as collectors feed on fine particulate organic matter (FPOM) and will

thrive in an environment with plenty of FPOM while grazers tend to favor algae for food (Walsh

2005).Hammer 3 of 20

The purpose of this study was to document the zooplankton and invertebrate

communities of different water levels within the Biola Creek. In addition, a focus on planarian

and mayfly growth due to abiotic factors were observed, and a measured analysis of diversity in

algal regions was recorded. Several environmental variables were measured at the collection

site (Sensory observations, water chemical tests, water depth, temperatures, and stream

velocity) in order to better understand the abiotic factors which effect community structure and

the diversity of invertebrates.

Methods and Materials

Samples were taken at one geographical location in the Biola Creek.

Figure 1: Shallow Water Description (first time on experimental sight), Our shallow water was

to the left of the algae and in front of the big bush.

Figure 2: Deep Water Description (first time on experimental sight), Our Deepwater was to the

right of the algae and to the right of the big bush.

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Samples were also taken on the Friday between March 2nd- 27th between 10:30am-

2:30pm (Pacific Time Zone) at Biola Creek which sits at an elevation of about 59 meters above

sea level. A thermometer was used to record water temperature at the time of sampling. The

temperature and precipitation of all 27 days of March are listed in Graph 1.

Graph 1: There was low amounts of precipitation and a moderate temperature ranging in mid-

teens Celsius, and the high temperature ranged around mid-30 Celsius.

Each site had its flow velocity measured by recording the amount of time it took a plastic float

to move a fixed distance.

Graph 2: The deep flowrate steadily increased than decreased again while the shallow flow rate

was an increase from 1st week and slowly decreased by 4th week.Hammer 5 of 20

Graph 3: The deep temperature stayed moderately constant from its drop its 1st week to a peak

at the 2nd week mark. The Shallow water temperature stayed consistently warm dropped by the

3rd week and increased majorly by the 4th week.

Sensory observations were recorded of the vegetation that was growing around the bank, on

top of the water, and the surrounding lawn area around the experiment region.

Sensory Observations first time on experimental sight:

1) The odor of the creek was not to smelly, in the mossy areas and algal areas it smelled a

little musty, or even a bit rotten grassy smell. I think this was caused by the algal growth

and the murky water of the creek and the organism’s digestive wastes.

2) The water color looks a little polluted as one may observe in our original pictures that

were dated on (March 6th 2015) the first day of the creek project. We happen to believe

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that some construction workers who have been in the area dumped paint into our creek.

That is why the water looks somewhat like watered down milk. There is no evidence of oil

hazards, but the paint hazard is sufficient enough.

3) The turbidity was very clear, the clear water was not very affected by the haziness of

divided particles that are suspended in the creek. The turbidity was definitely affected by

the paint that was poured into the river.

4) There was a major deposit of silt our 1st week because some construction workers who had

dumped paint deposit into the water. I believe the silt will brush away as the current

moves on.

To measure the depth, we extended a tape measurement into the deep and shallow areas of the

water in order to measure in length the height of the water in several areas and took an average.

We used a protractor and observed the angle from the bank in several areas and got an average of

the angle which was: θ= 150◦.

Figure 3: Diagram of angle of the river bank with respect to the riverbed.

During sampling, changes to the location were recorded. For classification, the

invertebrate quick guide table found in lab was used to determine the type and stage of

development the invertebrates were in. The width was measured at each sampling spot by using

a measuring tape to measure the distance from one bank of the stream to the other. One

measurement was recorded, at approximately 1.5 meters apart (from bank to bank). The length

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was about 1 meter long as well. Then the measuring tape was used to measure the depth of the

stream. One end was positioned on the stream floor and the measurement was taken at the

surface of the water.

Graph 4: The depth stayed constant for the shallow water region and increased by the 3rd week

for the deep water region.

Invertebrates were collected using a surber stream bottle sampler at the experiment site

location. The surber sampler has a square opening of .3048 meters by .3048 meters and an

attached net with a mesh opening of 1mm. Six samples were taken at the site once a week. Two

samples were of the water regions of the deep and shallow. Two other samples were of the

algae water by location (deep and shallow), and the last two samples were for chemical tests

(deep and shallow regions).

Table 1: Chemical Analysis of Deep and Shallow water.

Water SamplesShallow Water Weeks Chlorine (Total ppm) Chlorine (Free ppm) Hardness (grain/gal) Alkalninity (ppm) Copper ppm (mg/L) Sulfide ppm (mg/L) Ph Nitrate Nitrogen 60s ppm Nitrite Nitrogen 30s ppm

1 0 0 0.058823 240 0 0 8.4 10 0.152 0 0 0.058823 240 0.2 0 8.4 5 0.153 0 0 0.058823 240 0.5 0 8.4 2 0.34 0 0 0.058823 240 0.5 0 8.4 2 0.15

Water SamplesDeep Water Weeks Chlorine (Total ppm) Chlorine (Free ppm) Hardness (grain/gal) Alkalninity (ppm) Copper ppm (mg/L) Sulfide ppm (mg/L) Ph Nitrate Nitrogoen 60s ppm Nitrite Nitrogen 30s ppm

1 0 0 0.058823 240 0 0 8.4 10 0.32 0 0 0.058823 240 0 0 8.4 5 0.33 0 0 0.058823 240 0.2 0 8.4 5 0.154 0 0 0.058823 240 0.5 0 8.4 2 0.15

Chemical Data:

Chemical Data:

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There was a lot of debris and sticks, along with various insects, that would get stuck into

the back of the surber sampler. After each collection, the contents of the net were emptied

through a large plastic funnel into a 100ml plastic containers. Samples were then returned to the

lab where they were analyzed for the presence of invertebrates and zooplankton.

Table 2: This table the average abiotic factors for each creek region are displayed.

Average Abiotic Factors Deep Water Shallow Water Temperature ( C ) 24.4 25.6

pH 7.8 7.8Water Speed (m/s) 0.2248 0.4668Water Depth (m) 0.2334895 0.101346Nitrate 60s ppm 5.5 4.75

A pre-made wet mount was used to analyze each bottle containing samples. The bottles

were emptied one at a time and a 500ml squeeze bottle filled with RO/DI was used to wash any

substrate left behind on the slide and to also provide liquid adhesion for the specimens on the

slide. Occasionally we would have to use protist slowing agent for faster moving invertebrates

to contain them on to the slide. Sharp forceps were used to remove mayfly species floating

around in the sample tray. A detailed search by the microscope was performed so as not to miss

any zooplankton, or aquatic invertebrates that could be in the sample.

Materials Used:

Field Equipment: Lab Equipment: Electronic Resources:

- Measuring Tape

- 100ml Plastic

containers (6)

- Surber stream bottle

sampler

- Protractor

- Microscope

- Sulfide Chemical

Test

- Kit

- Pipettes

- 500ml squeeze

- Microsoft Xcel

2013.

- Microsoft Word

2013.

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Apparatus

- Notebook

- Pencil

- Camera

bottle filled with

RO/DI

- Nitrate/ Nitrite

Chemical Strips

- Wet Mount Slides

- Protist detainment

solution

- Copper Chemical

Testing Strips

- 5 in 1 Chemical

Test strips

(Alkalinity,

Hardness, Free

Chlorine, Chlorine,

pH) Testing Strips.

Results

The samples were recorded and taken on site our location (Figure 1), and there were

several abiotic factors observed (Graph 1- Table 1). There was 1 sample of each water volume

that were taken and brought back to the lab for further review. In the standing water alone we

found numerous types of invertebrates and recorded there picture and found the phylum.

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Graph 5: This graph illustrates the total number of invertebrates found over four weeks in each

region.

In all the samples taken over the 4 week period the total invertebrate diversity came out to

be increasingly larger for the deep water than for the shallow water regions of the experiment

site.

Graph 6: This graph illustrates the total number of species found in algae from each region over

the 4 weeks. There was a clear preference for deep water algae, as supposed to shallow water

algae.

In all the samples taken over the 4 week period the total invertebrate diversity came out to

be increasingly larger for the deep algal water than for the shallow algal water. I chose to

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independently study two organisms specifically to observe how they would increase in their

diversity due to the abiotic factor changes over the 4 weeks and I found wonderful results

concluding that. I am lead to believe that the invertebrate habitat in the Biola Creek is abundant.

Graph 7: Over the 4 weeks of the experiment, the water temperatures fluctuated. The number of

mayflies steadily increased as the temperature increased. There was no effect to the species

numbers when the temperature hit its low point.

Graph 8: At the beginning of the experiment, the deep water nitrate levels were considerably

high. They decreased as the time went on. The planarian diversity steadily increased as the nitrate

levels decreased.

There exists a very diverse ecosystem, as I have observed, in the Biola Creek of La

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Mirada. The results (Graph 5-8) of this experiment prove two things in regards to my

hypothesis, they are: (1) Chemical and Physical (temperature mainly) abiotic factors are

important in the regulation of environment the invertebrates live in. Second, the placement of

invertebrates and what region they chose to inhabit is heavily dependent upon food web (algae

resource) and most of the inhabitants in the creek are arthropods (Graph 5). To restate my

hypothesis, found in the lab notebook, I hypothesized that more organismal invertebrates would

be present in the deep waters of our testing location and sub regions than found in the shallow

waters of the same testing location. These results support the basis in that the abiotic factors

influence the habitat of the observed invertebrates, through temperature, turbidity, algal growth

and physical changes that have occurred upon the creek. All these factors play a role in

benefiting the marine community, or dismantling it.

Discussion

The results of our experiment proved our hypothesis to be true because there was way

more invertebrates that lived in the deep waters by themselves and with algae than the shallow

water with, or without algal growth (Graph 5-6). I expected to find this through my given

hypothesis and because abiotic factors do effect the environment and those that live in it. How

my results compared to those expected are that I theoretically thought that more organisms would

inhabit an increased volume of water biomass over a smaller volume of biomass. When there is

algae involved, a lot of small invertebrates like planarian, stoneflies, and crustaceans enjoy

phytoplankton and use its nutrients as a resource to live. As talked in previous lab sections,

invertebrates are found around algae, attached to algae, and swimming in algae. The most

unexpected result was trying to find zooplankton micro-organisms. Having established the same

logical reasoning in which I made my hypothesis, I figured there would be more invertebrates in

the deeper water regions than the shallower regions of the test site. There was not many micro-

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organisms recorded and found in our samples. In Graph 6 the results show a low population of

zooplankton (Epistylis) and in the different regions where samples were taken. It was

unexpected, but we found an organism in the creek that has not been analyzed before in the lab,

which was the protozoan called Epistylis. We as also found an unidentified organism which we

could later identify has a caddisfly. These explanations I believe are a result of increased nitrate

levels residing in the Biola Creek.

In an aquatic environment the most common ionic (reactive) forms of inorganic nitrogen

are ammonium (NH4+

), Nitrite (NO2-) and Nitrate (NO3

-). These ions may be present naturally in

aquatic ecosystems as result of atmospheric deposition, surface and ground water runoff

(Camargo and Alvaro et al. 2005). In consequence concentrations of nitrate in freshwater and

marine ecosystems usually are higher than those of ammonium and nitrite induced regions.

Nitrate however may be removed from water by aquatic plants like algae and bacteria which

assimilate thinking of it as a source of nitrogen (Camargo and Ward 1992). Due to these results

found in the San Gabriel watershed of Southern California one can refer to the methods Table 1

to see the high nitrate levels.

Graph 9: The deep water nitrate decreased from its high concentration consistently till the 4th

week, while the shallow water nitrates continues to decrease than leveled out by the 4th week.

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The reason why the Nitrate levels were so high the 1st week was because if you refer to Figure 1

and Figure 2 one can observe the turbidity and silt (as recorded in methods sensory observations

section 3 and 4) of the creek is very high. By analysis of the flow rate (Graph 2) it correlates to

Graph 9 in how the nitrate concentration decreased over the next 3 weeks post our first data

collection on site.

Camargo also states: the main toxic action of nitrate on aquatic invertebrates is due to the

conversion of the oxygen carrying pigment of invertebrates are incapable of carrying oxygen

anymore because the toxicity of nitrate (Camargo and Ward 1992). Invertebrates react to

increased levels of nitrate and the effects are just as serious as if elevated nitrate concentrations

in drinking water would have serious risk on humans. This is why I also believe that there was a

low result of our population of invertebrates over our 1.5m2 area because the waste deposit of silt

from the construction workers, but also because with the rise and fall of the water depth (Graph

4) over time, creates a environment where the pollution in the amount of water due to the rise

and fall of water level undergo toxic reactions that end up killing organisms. Nitrate toxicity to

aquatic invertebrate’s increases with increasing nitrate concentrations and exposure time.

Conversely nitrate toxicity decrease with increasing body size and water salinity (Camargo and

Alvaro et al., 2005). The solution to the problem of nitrate waste that is being dumped into the

creek is to increase the Salinity. Salinity is an important factor in determining many aspects of

the chemistry of natural waters and of biological processes within it, and is a thermodynamic

state variable that, along with temperature and pressure, governs physical characteristics of the

water inhabitants (Camargo and Ward 1992).

Part of my experiment with the diversity of invertebrates was observing two organisms

Mayflies (Order: Ephemeroptera) and Planarian (Genus: Dugesia) and what their presence is in

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the creek due to the abiotic factors amidst the pollution and contamination going on. After seeing

the first day how polluted the creek looked I wanted to test what factors may affect the presence

of these invertebrates over just general toxicity of the creek water. The Dugesia is a common

member of stream communities (Armstrong, 1964). Local populations may consist of individuals

reproducing both asexually and sexually as individuals, or may consist exclusively of asexually

reproducing individuals only. Armstrong found that the pond populations he studied experienced

varying food supply, Nitrate concentration and temperatures, which ultimately affected both

reproduction and growth of Dugesia. As observed in Graph 8 Planarian diversity each week

increased due to the three, or one of three factors. Walsh in his research found that deforestation,

particularly in the riparian zone, is often identified as an important driver of urban impacts to

streams, with lowland urban development often resulting in restructuring, or loss of riparian

vegetation (Walsh 2005). Because of this problem streams cannot receive the nutrients it needs

for its inhabitants, that’s why our results showed that most invertebrates were found in deep water

with algae present (Graph 5-6). The Biola Creek is located in the middle of a residential urban

city. Walsh’s study found that the presence of urban pipelines affected concentrations of diatoms,

macro-invertebrates, and fishes that were associated with the urban density gradient, but were less

strongly affected by the presence of riparian forest. Because of Walsh’s the reason in which the

Planarian increased was because of decreased Nitrate levels, and weather patterns that were

present during times of the experiment.

Sefton found that Planaria is able to complete its life-cycle over a range of temperature

from 3.50 C (possibly 1.50 C) to 20.00 C (Sefton and Reynoldson 1972). It was evident from his

data that the temperatures of British lakes are well within the tolerance limits for the persistence

and breeding of the species. The temperatures during the testing time over the weeks were in the

low 20’s C (Graph 3) for the deep algal water (Graph 5-6) concentrated regions which consisted Hammer 16 of 20

of mostly all the planarian recorded in week 3 and 4 (Graph 5-6 ). The data proves that Planarian

(Dugesia) depend upon the temperature and nutrients of the algae to grow and reproduce (since

they live in an urban environment).

One the most consistent data sets (Table 1) along with free chlorine and chlorine is the pH

level of the creek water from all 4 weeks of collection. Mayflies (and their Larvae) Studies have

shown that in mayfly assemblages, Petrin found smaller body size, greater reproductive output,

faster life cycles and a larger proportion of gathering collectors and scrapers with increasing pH

in Mayfly populated areas of streams (Petrin 2011). This study correlates with my data in that

with the consistency reading of 8.4 neutral pH level there was a substantial amount of Mayfly

population being observed all 4 weeks (Graph 5-6).

Figure 4: Conceptual diagram illustrating the hypothesized changes in the mean trait levels along

the pH gradient.

I hypothesized faster life cycles (voltinism), higher fertility and smaller body size with increasing

pH (Petrin 2011). This does show how some scientist have hypothesized that pH composition and

its influence on invertebrates, but Petrin stated: it is still unclear why populations may be Hammer 17 of 20

impaired by low pH under some circumstances, but not under others. Concurrently, little is

known about how pH affects the composition of species traits that may control ecological

processes. Another hypothesis I have to the population of Mayflies in a given area is predicted by

the presence of predators surrounding the inhabiting location of Mayflies. Peckarsky predicted

also that Mayfly Larvae incubation is also effected by temperatures and the presence of predators

(Stoneflies). Peckarsky found that temperature and avoidance of Maylfly Larvae to Stoneflies is

proportional (Peckarsky, 1996). This solidifies the idea that prey may adjust their behavior to the

presence, or activity of predators over the short, and desire to move to warmer waters where

fertility is abundant (Graph 7). For example, many prey modify their behavior when faced with a

series of acute predator threats that vary in intensity (Peckarsky 1996). Therefore the reason in

which the population of Mayflies recorded are more abundant in the deep water of the stream

alone (Graph 6) is theoretically related to the idea that it is better to avoid Stoneflies predators

that want to eat them and move to warmer water for incubation of Larvae. By observing Graph 6

there were not as many Stoneflies present in the deep water region of the experiment site. Thus

proving the theoretical idea that population increase is dependent upon the abiotic factor of

predators that inhibit one region over another.

Field studies were conducted by Ahn to assess the coastal water quality impact of storm

water runoff from the Santa Ana River (located in Orange County), which drains a large urban

watershed located in southern California. Storm water runoff from the river leads to very poor

surf zone water quality (Ahn et al. 2005). The impact of storm water runoff on coastal water

quality is of particular concern in arid regions like southern California because, on an annual

basis, a large percentage 99.9% of the surface water runoff and associated pollution flows into the

ocean during a few storms in the winter (Walsh 2005). These studies have shown that pollutants

and contamination are flowing through the rivers and creeks of Southern California exposing Hammer 18 of 20

several of millions of invertebrates that live in the habitats to toxic wastes and increased nitrate

levels. As the movement to restore urban streams grows, urban stream ecologists will be

challenged to identify the primary reasons of contamination, the best actions to attain goals for

restoration are through community efforts (Ahn et al. 2005). Further challenges involve engaging

the human communities of urban areas to achieve a shared understanding of what is achievable

and desirable to communities for their local streams. Urban streams attribute to life, but are

limited by wastes, such as mowed grass riparian zones, or paved streamside paths, may have

amenity values for some urban communities. Sometimes, value placed in such altered, unnatural

environments can be a product of people not missing what they never had, and stream ecologists

can play a role in educating communities on how streams that more closely resemble natural

conditions are often more desirable. However, for such education of urban communities to be

effective, restoration actions and attainable restoration needs to take place for invertebrates to

thrive.

Acknowledgments

I would like to thank Dr. Billock for her assistance, time, and effort in helping me

understand, identify, and analyze my samples for this project. I also want to thank Elizabeth

Curran and Cheyenne Carrel for their guidance, service and support during this project. Also

thanks to the Biological Sciences Department for their resources they let me use. Finally, thanks

to the other students in the class that I got to share this experience with.

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Works Cited

Ahn, Jong Ho, et al. 2005. Coastal water quality impact of stormwater runoff from an urban

watershed in southern California. Environmental Science & Technology 39.16: 5940-5953.

Armstrong, Joseph T.1964 "The population dynamics of the planarian, Dugesia tigrina.

Ecology): 361-365.

Camargo, Julio A., Alvaro Alonso, and Annabella Salamanca. 2005. "Nitrate toxicity to aquatic

animals: a review with new data for freshwater invertebrates.” Chemosphere 58.9:1255-

1267.

Camargo, J. A., and J. V. Ward. 1992. Short-term toxicity of sodium nitrate (NaNO 3) to non-

target freshwater invertebrates. Chemosphere. 24.1: 23-28.

Peckarsky, Barbara L. 1996. "Alternative predator avoidance syndromes of stream-dwelling

mayfly larvae." Ecology: 1888-1905.

Petrin, Zlatko. 2011. Species traits predict assembly of mayfly and stonefly communities along

pH gradients. Oecologia 167.2: 513-524.     

Reyes, Ismael. "The Effect of Geographical Location on Mayfly Diversity in Costa Rica." pdf.

Sefton, A. D., and T. B. Reynoldson. 1972. The effect of temperature and water chemistry on the

life-cycle of Planaria torva (Müller) (Turbellaria: Tricladida).The Journal of Animal Ecology

487-494.

Walsh, Christopher J., et al. 2005. "The urban stream syndrome: current knowledge and the

search for a cure." Journal of the North American Benthological Society24.3: 706-723.

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