Abiotic Factors Affecting Soil Invertebrates
Introduction
Over the next two weeks, we will be sampling soil organisms in different
microclimates on campus. In addition to enumerating the number and types of invertebrates
in these areas, we will also be obtaining measures of physical factors that might influence
the distribution and abundance of these organisms.
Invertebrates living in the soil and the organic litter above it are difficult to
sample. Most are small and many live in the interstices of soils, decomposing wood, and
leaves. Because of this, hand sorting of soil and litter samples under a dissecting microscope
is a very tedious and inefficient method of obtaining counts of soil and invertebrates. An
alternative is to use a technique that forces the invertebrates to leave the litter-soil
matrix on their own. We will be using just such a technique (known as the Tullgren funnel
method) to obtain counts of soil invertebrates in lab over the next two weeks. The Tullgren
funnel method employs heat and desiccation to drive organisms downward from the sample
through a funnel apparatus and into a vial of preservative.
The soil arthropod sampling procedure outlined in this exercise involves three main
steps: (1) collection of samples, (2) Tullgren funnel extraction, and (3) identification and
counting of the extracted organisms. We will focus on distribution of various types of soil
invertebrates across two microhabitats, the hillside by the Science Building and within the
forest.
First Week:
Collection of Samples
At each of the two sites (Hillside and Forest), each group will
sample one location. The procedure for collecting samples should be
designed to allow standardization of the surface area and depth of the
soil or leaf litter samples collected. For general ecological work, this
requirement can usually be satisfied through the use of some type of
cylindrical metal coring device. For sampling in porous soils or sands, a tin can with the top
and bottom removed may be satisfactory. To obtain the sample, force the edges of the can
into the soil or leaf litter. First, using a trowel and your hands, remove all soil and litter
from the top 1 centimeter. The sample should be removed from the corer with as little
disturbance as possible and placed in the first bag (to be used to extract invertebrates this
week, and to estimate soil particle size and organic content next week). Next remove a
small handful of soil from the area adjacent to where the previous sample was removed (to
be used to estimate soil moisture in lab). Be sure all bags are labeled with your group
number and site name (Hillside or Forest)!
Extraction Procedure
The Tullgren funnel employs electric lights to heat and dry the samples. The soil or
leaf litter material to be extracted should be wrapped in one layer of cheesecloth and
placed on the hardware-cloth screen above the funnel. The cheesecloth functions to reduce
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the amount of soil sifting down into the specimen vial as the sample dries. After placing the
soil sample in the funnel, place a jar with 80% ethanol under the funnel. Be sure all jars
are labeled with your group number and site name (Hillside or Forest)! Extraction
should be nearly complete after 36-48 hours.
Abiotic Factors
Soil moisture and organic content will be determined on the soil from the second bag
collected. The sample will be weighed on a balance, placed in a drying oven (100°C) and reweighed to estimate percent moisture content. Then the same sample will be placed in a
furnace (440°C) and then re-weighed to estimate percent organic matter (carbon in organic
molecules is converted to carbon dioxide gas at these temperatures)
Second Week:
Invertebrate Identification and Enumeration
In the second week of lab, each team needs to identify and count the invertebrates
collected from the Tullgren funnels. Animals may be identified and counted by placing the
sample in a flat dish and examining it with a dissecting microscope.
Although most animals can be identified to order or family with the aid of standard
reference books, we will be using a "natural taxonomy" approach. For most soil arthropod
groups, however, identification beyond this level is very difficult. The large number of
species and small body size of many groups mean that careful anatomical study is required
for complete identification. Furthermore, many of the individuals encountered in samples
are immature forms for which descriptions are generally unavailable. We will be using a
simplified approach that will allow us to obtain the needed data in the limited time we have
available. You will use a dichotomous key to place each individual invertebrate into the
appropriate category. A dichotomous key is organized into a series of couplets. Each
couplet will have a unique number associated with it and couplets are numbered sequentially.
Each couplet consists of two statements about the organism you are observing. If written
correctly, only one of the statements will be true. At the end of each statement you will
find one of two things, either a direction to precede to another couplet, or a category that
includes the organism you are observing. By following these directions, you should be able
to place each of your organisms into one of six categories. In this key, the categories we
will be using are in bold type.
Abiotic Factors The soil in the funnel will be used to estimate particle size distribution
by sieving samples. Procedures for these estimates will be discussed in class.
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Soil Invertebrate Lab Data Sheet
Group #____________________
Location ___________________
Number per taxa:
Insect Larvae
Unsegmented Worm
Earthworm (segmented worm)
Springtail
Myriapod
Spider
Mite
Other insects
Other
Soil particle size:
Weight of coarse sieve w/o
sample (g)
Weight of fine sieve w/o
sample (g)
Weight of bottom pan w/o
sample (g)
Weight of coarse sieve w/
sample (g)
Weight of fine sieve w/
sample (g)
Weight of bottom pan w/
sample (g)
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Simplified Key to Soil Invertebrates
1) This couplet addresses the general shape of the organism.
a) Shape is generally worm-like, without legs or with 6 or fewer leg-like appendages
(See Figures 1, 3, 4, 5)………………………….…………… Go to 2
b) Shape is bug-like with hard exoskeletion, with at least 6 leg-like appendages (Fig. 2,
6, 7) ………………………………………………………………………… Go to 4
2) This couplet requires identification of the presence or absence of appendages.
a) No recognizable appendages (Fig. 3, 4) …………………………………………………… Go to 3
b) Possesses recognizable appendages, or head capsule (Fig. 1, 5) ……… Insect Larvae
3) This couplet requires identification of segmentation.
a) Lacks visible segmentation (Fig. 3) ………………………………… Unsegmented Worm
b) Has visible segmentation (Fig. 4) …………………………………… Earthworm
4) How many legs?
a) Six legs (Fig. 6)……………………………………
b) More than six legs (Fig. 2, 7) …………
Go to 5
Go to 6
5) How many body and antennal segments?
a) Nine or fewer body segments behind the head; antenna with four segments; hinged
tail appendage often visible (Fig 8)………………………… Springtail
b) Otherwise ……………………………………………………………… Other insect
6) How many legs, again?
c) Eight legs (Fig. 2) ……………………………………Go to 7
d) More than eight legs (Fig. 7) ……………… Myriapod
7) Considers thorax and abdomen.
a) Thorax and abdomen separated by constricted “waist” ………………………… Spider
b) Thorax and abdomen fused into one (Fig. 2) …………………………………………… Mite
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Figure 3
Figure 2
Figure 1
Figure 5
Figure 4
Figure 8
Figure 7
Figure 6
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What to turn in for this Lab
Prepare and turn in a Results section and a Discussion section
Results section. The Results section should always include a written summary of important
trends in your data tables and graphs.
Downloaded Excel data set and calculate based on the data from all groups:
 Total number of individuals and total number of taxa found at each sampling point.
 Mean and standard deviation for total number of individuals and total number of
taxa at each site.
 Mean and standard deviation for each physical parameter at each site.
In this lab, you must choose how to best present your data in graph form to address the
comparison of sites (graphs may be done either by hand on graph paper or by computer).
There are two general strategies you may wish to pursue. First, plot parameters among
sites in bar graphs for selected biotic parameters (for example relative proportion of taxa
at each site) and abiotic parameters (e.g. soil moisture), and compare and discuss trends. A
second approach might be to plot the individual biotic and abiotic parameters against one
another in a scatter graph.
Discussion section. In the Discussion, you will describe how the reported results support
your final conclusion. Use the literature provided at the end of this handout The Ecology of
Soil Invertebrates, to support your conclusions. As part of the discussion, write a few
paragraphs that address the following questions:
 If there are differences among sites, explain what may have caused the
differences observed? Use the information on “The Ecology of Soil
Invertebrates” provided or other sources to support your explanation.
 If there are no differences among sites (or differences are not as expected),
explain why might this be (discuss other possible alternative explanations,
experimental design flaws, or incorrect assumptions)?
 How confident are you about your conclusions? How might you further test
these explanations? What might be the relevance of your work?
Don't be afraid to speculate in this section (but make sure it is clear to the reader what is
speculation and what is not). No single scientific study is definitive. Thoughtful speculation
is a key ingredient in the scientific process!
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Random Number Table:
1 092725 012157 827052 297980 625608 964134
2 104460 007903 484595 868313 274221 367181
3 676071 388003 266711 323324 044463 762803
4 881878 862385 203886 261061 096674 811548
5 534500 336348 086585 241740 581286 008435
6 094276 615776 242112 985859 075388 082003
7 333848 513630 474798 841425 331001 542740
8 847886 629263 596457 589243 576797 800957
9 942495 695172 523982 264961 771016 118797
10 450553 679145 324036 715835 963418 533048
11 024670 615375 717260 171144 340939 208712
12 932959 205554 113225 704406 263818 633643
13 039831 202271 212602 089507 469224 639594
14 988302 547676 746372 209684 217507 290574
15 175854 538509 553171 532143 360606 597321
16 495315 444204 428810 387020 053680 418895
17 753091 146845 416561 310332 393625 548843
18 953724 288220 149970 386262 386069 597807
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The Ecology of Soil Invertebrates
The soil is a radically different environment for life than the ones on and above the
ground; yet the essential requirements do not differ. Like organisms that live outside the
soil, life in the soil requires living space, oxygen, food, and water. Without the presence and
intense activity of living organisms, soil development could not proceed. Soil inhabitants
from bacteria and fungi to earthworms convert inert mineral matter into a living system.
Soil possesses several outstanding characteristics as a medium for life. It is stable
structurally and chemically. The atmosphere remains saturated or nearly so, until soil
moisture drops below a critical point. Soil affords a refuge from extremes in temperature,
wind, light, and dryness. These conditions allow soil fauna to make easy adjustments to
unfavorable conditions. On the other hand, soil hampers the movement of animals. Except
to such channeling species as earth worms, soil pore space is important. It determines the
living space, humidity, and gaseous conditions of the soil environment.
Only a part of the upper soil layer is available to most soil animals as living space.
Spaces within the surface litter, cavities walled off by soil aggregates, pore spaces between
individual soil particles, root channels, and fissures all are potential habitats. Most soil
animals are limited to pore spaces and cavities larger than themselves.
Water in the pore spaces is essential. The majority of soil organisms are active only
in water. Soil water is usually present as a thin film coating the surfaces of soil particles.
This film contains, among other things, bacteria, unicellular algae, protozoa, rotifers, and
nematodes. The thickness and shape of the water film restrict the movement of most of
these soil organisms. Many small species and immature stages of larger centipedes and
millipedes are immobilized by a film of water and are unable to overcome the surface
tension imprisoning them. Some soil animals, such as millipedes and centipedes, are highly
susceptible to desiccation and avoid it by burrowing deeper.
When water fills pore spaces after heavy rains, conditions are disastrous for some
soil inhabitants. If earthworms cannot evade flooding by digging deeper, they come to the
surface, where they often die from ultraviolet radiation and desiccation or are eaten.
A diversity of life occupies these habitats. The number of species of bacteria,
fungi, protists, and representatives of nearly every invertebrate phylum found in the soil is
enormous. A soil zoologist found 110 species of beetles, 229 species of mites, and 46
species of snails and slugs in the soil of an Austrian deciduous beech forest.
Dominant among the soil organisms are bacteria, fungi, protozoans, and nematodes.
Flagellated protozoans range from 100,000 to 1,000,000, amoebas from 50,000 to 500,000,
and ciliates up to 1000 per gram of soil. Nematodes, an unsegmented worm, occur in the
millions per square meter of soil. These organisms obtain their nourishment from the roots
of living plants and from dead organic matter. Some protozoans and free-living nematodes
feed selectively on bacteria and fungi.
Living within the pore spaces of the soil are the most abundant and widely
distributed of all forest soil animals, the mites (Acarina) and springtails (Collembola).
Together they make up over 80 percent of the animals in the soil. Flattened, they wiggle,
squeeze, and digest their way through tiny caverns in the soil. They feed on fungi or search
for prey in the dark interstices and pores of the organic mass.
The more numerous of the two, both in species and numbers, are the mites, tiny
eight-legged arthropods from 0.1 to 2.0 mm in size. The most common mites in the soil and
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litter are the Orbatei. They live mostly on fungal hyphae that attack dead vegetation as
well as on the sugars produced by the digestion of cellulose found in conifer needles.
Collembolae are the most widely distributed of all insects. Their common name,
springtail, describes the remarkable springing organ at the posterior end, which enables
them to leap great distances for their size. The springtails are small, from 0.3 to 1.0 mm in
size. They consume decomposing plant material, largely for the fungal hyphae they contain.
Prominent among the larger soil fauna are the earthworms (Lumbricidae). Burrowing
through the soil, they ingest soil and fresh litter and egest both mixed with intestinal
secretions. Earthworms defecate aggregated castings on or near the surface of the soil or
as a semiliquid in intersoil spaces along the burrow. These aggregates produce a more open
structure in heavy soil and bind light soil together. In this manner earthworms improve the
soil environment for other organisms.
Feeding on the surface litter are Myriapod millipedes. They eat leaves, particularly
those somewhat decomposed by fungi. Lacking the enzymes necessary for the breakdown of
cellulose, millipedes live on the fungi contained within the litter. The millipedes' chief
contribution is the mechanical breakdown of litter, making it more vulnerable to microbial
attack, especially by saprophytic fungi. Accompanying the millipedes are snails and slugs.
Among the soil invertebrates they possess the widest range of enzymes to hydrolyze
cellulose and other plant polysaccharides, possibly even the highly indigestible lignins.
Not to be ignored are termites (Isoptera), white wingless, social insects. Except
for some dipteran and beetle larvae, termites are the only larger soil inhabitants that can
break down the cellulose of wood. They do so with the aid of symbiotic protozoans living in
their gut. Termites dominate the tropical soil fauna. In the tropics termites are
responsible for the rapid removal of wood, dry grass, and other materials from the soil
surface. In constructing their huge and complex mounds, termites move considerable
amounts of soil. Detrital-feeding organisms support predators. Small arthropods are the
principal prey of spiders, beetles, pseudoscorpions, predaceous mites, and centipedes.
Protozoans, rotifers, myxobacteria, and nematodes feed on bacteria and algae. Various
predaceous fungi live on bacteria-feeders and algae-feeders.
Cox, G. W. 1996. Laboratory Manual of General Ecology, 7 th edition. McGraw-Hill Higher
Education, Boston, MA
Smith, Robert Leo and Thomas M. Smith. 1998. Elements of Ecology. Addison Wesley
Longman, Inc. Menlo Park, CA
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