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10500 BENTHIC MACROINVERTEBRATES*

10500 A. Introduction

1. Definition total number of individuals is divided into a reasonably balanced


distribution of species. Such communities respond to changing
Benthic macroinvertebrates are animals that inhabit the sedi- habitats and water quality by altering community structure (in-
ment or other substrates on the bottom of freshwater, estuarine, vertebrate abundance and composition). However, many habi-
and marine ecosystems. During their life cycles, these organisms tats— especially disturbed ones—may be dominated by a few
may construct attached cases, tubes, or nets that they live on or species.
in; roam freely over rocks, organic debris, and other substrates; The responses of macroinvertebrate communities to environ-
or burrow freely in substrates. According to historical defini- mental changes are useful in assessing the effects of municipal,
tions, macroinvertebrates are visible to the unaided eye and industrial, oil, and agricultural wastes, and of other land uses on
retained on a U.S. Standard No. 30 sieve (0.595- or 0.600-mm surface waters. Investigators have documented macroinverte-
openings).1 In actuality, however, some are difficult to see with- brate community responses to increased inorganic nutrients,
out magnification. increased organic loading, substrate alteration, and toxic chem-
The standard sieve for collecting freshwater, estuarine, and ical pollution. Inorganic nutrients and severe organic pollution
marine benthic macroinvertebrates is the U.S. Standard No. 30 usually reduce the variety of macroinvertebrates to only the most
sieve; however, some estuarine and marine programs use the tolerant ones [usually associated with low dissolved oxygen
U.S. Standard No. 50 sieve (0.300-mm openings) or the U.S. (DO) concentration] and increase their density. Sometimes se-
Standard No. 35 sieve (0.500-mm openings). For all aquatic vere organic pollution, siltation, or toxic chemical pollution may
assessment programs, the No. 30 sieve is recommended for reduce or even eliminate the entire macroinvertebrate commu-
collecting benthic fauna from freshwater, estuarine, and marine nity from an affected area. However, macroinvertebrate re-
habitats or from any water transport system. To accommodate sponses may be mediated by other environmental (biological,
some old historical databases (if the study’s data-quality objec- chemical, and physical) conditions.
tives permit), investigators might use a U.S. Standard No. 28 Assessing the effects of a pollution source generally involves
sieve (1.0-mm openings). To obtain a more representative sam- comparing macroinvertebrate communities and their physical
ple of the benthos (one that includes smaller forms, early life- habitats in polluted areas with those in adjacent unpolluted sites
stages, and other taxa of macroinvertebrates), they may use a (e.g., a gradient away from point sources of contamination). The
U.S. Standard No. 60 sieve (0.250-mm openings). procedure includes sampling and analyzing communities from
The standardization of bioassessment for species composition, different sites and subsequently determining whether and how
taxa richness, diversity, evenness, trophic levels, and major they differ. The basic information required in most structure
taxonomic spatial and temporal patterns may be enhanced sig- analyses cases is a count of individuals per taxon. Then, inves-
nificantly by the conventional use of a U.S. Standard No. 30 tigators can characterize and compare communities according to
sieve. community structure, density, diversity, metrics, pollution indi-
The major macroinvertebrates found in freshwater are flat- cators, or other analyses,1 including various statistical methods
worms, annelids, mollusks, crustaceans, and insects. The major (see 10500D). They also can estimate biomass and productiv-
macroinvertebrates found in estuarine and marine waters are ity.1– 4 Equally important is a characterization of DO concentra-
bryozoans, sponges, annelids, mollusks, roundworms, cnidarians tion, substrate, water depth, sediment type and grain size, total
(coelenterates), crustaceans, insects, and echinoderms. organic carbon (TOC), and other site- and situation-specific
characteristics.
2. Response to Environment While the following macroinvertebrate methods are tradition-
ally used to sample and quantify benthic invertebrate communi-
The species composition and density (numbers of individuals ties, other methods are also being evaluated in an effort to
per unit area) of macroinvertebrate communities in streams, develop and implement narrative biological criteria for surface
lakes, estuaries, and marine waters can be uniform from year to waters.5 Not discussed here are U.S. Environmental Protection
year in unperturbed environments. However, life-cycle dynamics Agency (EPA)-developed rapid bioassessment techniques6,7 and
produce variations in species composition and abundance either Environmental Monitoring and Assessment Program (EMAP)
temporally or spatially. protocols for field operations, as well as methods for sampling
Most aquatic habitats—particularly free-flowing streams and macroinvertebrates and assessing the ecological conditions of
waters with acceptable water quality and substrate conditions— wadeable streams.8,9
support diverse macroinvertebrate communities in which the
3. References

* Approved by Standard Methods Committee, 2010.


1. KLEMM, D.J., P.A. LEWIS, F. FULK & J.M. LAZORCHAK. 1990. Mac-
Joint Task Group: Donald J. Klemm (chair), Peter M. Chapman, Philip A. Lewis, roinvertebrate Field and Laboratory Methods for Evaluating the
Don W. Schloesser, William T. Thoeny. Biological Integrity of Surface Waters, EPA-600/4-90-030. Envi-

1
BENTHIC MACROINVERTEBRATES (10500)/Introduction

ronmental Monitoring Systems Laboratory, U.S. Environmental tribution 1611. Appalachian Environ. Lab., Univ. Maryland, Frost-
Protection Agency, Cincinnati, Ohio. burg.
2. WETZEL, R.G. & G.E. LIKENS. 1991. Limnological Analyses, 2nd ed. LEONARD, P.M. & D.J. ORTH. 1986. Application and testing of an index
Springer-Verlag, New York, N.Y. of biotic integrity in small, coolwater streams. Trans. Amer. Fish.
3. PLANTE, C. & J.A. DOWNING. 1989. Production of freshwater inver- Soc. 115:401.
tebrate populations in lakes. Can. J. Fish. Aquat. Sci. 46:1489. WARWICK, R.M. 1986. A new method for determining pollution effects
4. PLANTE, C. & J.A. DOWNING. 1990. Empirical evidence for differ- on marine macrobenthic communities. Mar. Biol. 92:557.
ences among methods for calculating secondary production. J. N. HILSENHOFF, W.L. 1987. An improved biotic index of organic stream
Amer. Benthol. Soc. 9(1):9. pollution. Great Lakes Entomol. 20:31.
5. GIBSON, G.R., ed. 1996. Biological criteria. In Technical Guidance ROHM, C.M., J.W. GIESE & C.C. BENNETT. 1987. Evaluation of aquatic
for Streams and Small Rivers, EPA-822-B-96-001. Off. Science & ecoregion classification of streams in Arkansas. Freshwater Ecol.
Technology, U.S. Environmental Protection Agency, Washington, 4:127.
D.C. SHACKELFORD, B. 1988. Rapid Bioassessments of Lotic Macroinverte-
6. PLAFKIN, J.L., M.T. BARBOUR, K.D. PORTER, S.K. GROSS & R.M. brate Communities: Biocriteria Development. Arkansas Dept. Pol-
HUGHES. 1989. Rapid bioassessment protocols for use in streams and lution Control & Ecology, Little Rock.
rivers. In Benthic Macroinvertebrates and Fish, EPA-440/4-89-001. STEEDMAN, R.J. 1988. Modification and assessment of an index of biotic
Off. Water, U.S. Environmental Protection Agency, Washington, integrity to quantify stream quality in southern Ontario. Can. J.
D.C. Fish. Aquat. Sci. 45:492.
7. BARBOUR, M.T., J. GERRITSEN, B.D. SNYDER & J.B. STRIBLING. 1999. U.S. ENVIRONMENTAL PROTECTION AGENCY. 1988. Proc. 1st National
Rapid Bioassessment Protocols for Use in Streams and Wadeable Workshop on Biological Criteria, Lincolnwood, Ill., Dec. 2– 4,
Rivers: Periphyton, Benthic Macroinvertebrates, and Fish, EPA 1987, Rep. No. 905/9-89/003. U.S. Environmental Protection
841-B-99-002. Off. Water, U.S. Environmental Protection Agency, Agency, Chicago, Ill.
Washington, D.C. WHITTIER, T.R., R.M. HUGHES & D.P. LARSEN. 1988. Correspondence
8. LAZORCHAK, J.M., D.J. KLEMM & D.V. PECK. 1998. Environmental between ecoregions and spatial patterns in stream ecosystems in
Monitoring and Assessment Program-Surface Waters: Field Oper- Oregon. Can. J. Fish. Aquat. Sci. 45:1264.
ations and Methods for Measuring the Ecological Conditions of OHIO ENVIRONMENTAL PROTECTION AGENCY. 1989. Biological Criteria for
Wadeable Streams, EPA 620-R-94-004F. National Exposure Re- the Protection of Aquatic Life: Volume III. Standardized Biological
search Lab., U.S. Environmental Protection Agency, Cincinnati, Field Sampling and Laboratory Methods for Assessing Fish and
Ohio, and National Health & Environmental Effects Research Lab., Macroinvertebrate Communities. Ohio Environmental Protection
U.S. Environmental Protection Agency, Corvallis, Ore.
Agency, Div. Water Quality Monitoring & Assessment, Surface
9. BARBOUR, M.T. 1997. Perspectives from North America on the
Water Section, Columbus.
programmatic and scientific elements of an effective bioassessment.
REYNOLDSON, T.B., D.W. SCHLOESSER & B.A. MANNY. 1989. Develop-
Human Ecol. Risk Assess. 3:929.
ment of a benthic invertebrate objective for mesotrophic Great
Lakes waters. J. Great Lakes Res. 15:669.
4. Bibliography BURD, B.J., A. NEMEC & R.O. BRINKHURST. 1990. The development and
application of analytical methods in benthic marine infaunal stud-
HYNES, H.B.N. 1963. The Biology of Polluted Waters. Liverpool Univ.
Press, England. ies. Advan. Mar. Biol. 26:169.
MACAN, T.T. 1963. Freshwater Ecology. John Wiley & Sons, New York, NATIONAL RESEARCH COUNCIL. 1990. Managing Troubled Waters: The
N.Y. Role of Marine Environmental Monitoring. National Academy
RUTTNER, F. 1966. Fundamentals of Limnology. Univ. Toronto Press, Press, Washington, D.C.
Toronto, Ont. WESTON, D. 1990. Quantitative examination of macrobenthic changes
MACKENTHUN, K.M. & W.M. INGRAM. 1967. Biological Associated Prob- along an organic enrichment gradient. Mar. Ecol. Prog. Ser. 61:
lems in Freshwater Environments. Federal Water Pollution Control 233.
Admin., Washington, D.C. YOUNT, J.D. & G.J. NIEMI. 1990. Recovery of lotic communities and
HYNES, H.B.N. 1970. The Ecology of Running Waters. Univ. Toronto ecosystems from disturbance—a narrative review of case studies.
Press, Toronto, Ont. Environ. Mgmt. 14:547.
ODUM, E.P. 1971. Fundamentals of Ecology, 3rd ed. Saunders Publish- KARR, J.R. & B.L. KERANS. 1992. Components of biological integrity:
ing Co., Philadelphia, Pa. their definition and use in development of an invertebrate IBI. In
COULL, B.C., ed. 1977. Ecology of Marine Benthos. University of South T.P. Simon & W.S. David, eds. Environmental Indicators: Mea-
Carolina Press, Columbia. surement and Assessment Endpoints, Chapter 1, EPA 905-R-92-
PEARSON, T.H. & R. ROSENBERG. 1978. Macrobenthic succession in 003. U.S. Environmental Protection Agency, Chicago, Ill.
relation to organic enrichment and pollution of the marine environ- BLAKE, J.A. & A. LISSNER. 1993. Taxonomic atlas of the benthic fauna
ment. Oceanogr. Mar. Biol. Annu. Rev. 16:229. of the Santa Maria Basin and Western Santa Barbara Channel, Vol.
BROWER, J.R. & J.H. ZAR. 1984. Field and Laboratory Methods for 1. Santa Barbara Mus. Natural History, Santa Barbara, Calif.
General Ecology. Wm. C. Brown Publ., Dubuque, Iowa. CAIRNS, J., JR. 1993. A proposed framework for developing indicators of
RESH, V.H. & D.M. ROSENBERG. 1984. The Ecology of Aquatic Insects. ecosystem health. Hydrobiologia 263:1.
Praeger Publ., New York, N.Y. ROSENBERG, D.M. & V.H. RESH, eds. 1993. Freshwater Biomonitoring
WARD, J.V. 1984. Ecological perspectives in the management of aquatic and Benthic Macroinvertebrates. Chapman-Hall, New York, N.Y.
insects. In V.H. Resh & D.M. Rosenberg, eds. The Ecology of GURTZ, M.E. 1994. Design of biological components of the National
Aquatic Insects, p. 558. Praeger Scientific, New York, N.Y. Water-Quality Assessment (NAWQA) Program. In S.L. Loeb & A.
WIEDERHOLM, T. 1984. Responses of aquatic insects to environmental Spacie, eds. Biological Monitoring of Aquatic Systems. Lewis
pollution. In V.H. Resh & D.M. Rosenberg, eds. The Ecology of Publishers, Ann Arbor, Mich.
Aquatic Insects, p. 508. Praeger Scientific, New York, N.Y. DAVIS, W.S. & T.P. SIMON, eds. 1995. Biological Assessment and
CUMMINS, K.W. & M.A. WILZBACH. 1985. Field Procedures for Analysis Criteria: Tools for Water Resource Planning and Decision Making.
of Functional Feeding Groups of Stream Macroinvertebrates, Con- Lewis Publishers, Boca Raton, Fla.

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HAUER, F.R. & G.A. LAMBERTI, eds. 1996. Methods in Stream Ecology. Louis, Vol. IV, Part A. John Wiley & Sons, New York, N.Y.
Academic Press, New York, N.Y. PATRICK, R. 1998. The Mississippi River and Its Tributaries South of St.
KARR, J.R. 1996. Aquatic invertebrates: sentinels of watershed condi- Louis, Vol. IV, Part B. John Wiley & Sons, New York, N.Y.
tion. Wings 19(2):22. U.S. ENVIRONMENTAL PROTECTION AGENCY. 1998. Lake and Reservoir
PATRICK, R. 1996. Rivers of the United States: Vol. III: The Eastern Bioassessment and Biocriteria, Technical Guidance Document,
and Southeastern States. John Wiley & Sons, New York, N.Y. EPA 841-B-98-007. Off. Water, U.S. Environmental Protection
PEPPER, I.L., C.P. GERBA & M.L. BRUSSEAU. 1996. Pollution Science. Agency, Washington, D.C.
Academic Press, San Diego, Calif.
BATZER, D.P., R.B. RADER & S.A. WISSINGER. 1999. Invertebrates in
U.S. ENVIRONMENTAL PROTECTION AGENCY. 1996. Biological Criteria:
Freshwater Wetlands of North America: Ecology and Management.
Technical Guidance for Streams and Small Rivers, EPA 822-B-96-
John Wiley & Sons, New York, N.Y.
001. Off. Science & Technology, U.S. Environmental Protection
Agency, Washington, D.C. GRIFFITHS, R.W. 1999. BioMAP: Bioassessment of Water Quality. Cen-
BARBOUR, M.T. 1997. The re-invention of biological assessment in the tre for Environmental Training, Niagara College, Niagara-on-the-
U.S. Human Ecol. Risk Assess. 3:933. Lake, Ontario, Canada.
U.S. ENVIRONMENTAL PROTECTION AGENCY. 1997. Priorities for Ecological KARR, J.R. & E.W. CHU. 1999. Restoring Life in Running Waters. Island
Protection: An Initial List and Discussion Document for EPA, EPA Press, Washington, D.C.
600-S-97-002. Off. Research & Development, U.S. Environmental GERRITSEN, J., B. JESSUP, E.W. LEPPO & J. WHITE. 2000. Development of
Protection Agency, Washington, D.C. lake condition indexes (LCI) for Florida. Tetra Tech, Inc., Owings
MACBROOM, J.G. 1998. The River Book. Connecticut Dept. Environ- Mills, Md., Appendices A-1 to B-13. Florida Dept. Environmental
mental Protection, Hartford. Protection, Stormwater & Nonpoint Source Management Section,
PATRICK, R. 1998. The Mississippi River and Its Tributaries North of St. Tallahassee.

10500 B. Sample Collection

1. General Considerations investigators adhere to the following basic rules, they can design
a sound survey.
Before conducting a benthic survey, determine specific data a. Always establish a reference station(s) outside the influ-
quality objectives and clearly define the information sought. ence of all wastewater discharges of concern (but in the same
Data quality objectives are qualitative and quantitative state- water body). Because most surveys are made to determine the
ments developed to specify the quality of data needed to support damage that pollution causes aquatic life, this will be the basis
specific decisions and conclusions about the information sought. for comparing biota in polluted and unpolluted areas. Preferably
Discussion with water chemists, hydrologists, limnologists, and establish at least two reference stations: one far from the effluent
individuals from other disciplines will be helpful. Ultimately, the discharge and the other near the discharge, but not subject to its
choice of methodology will depend on whether the habitat to be influence. (For example, if the discharge were in a river, one
studied is a stream, lake, reservoir, or marine area. For example, station would be far upstream of the discharge and the other
investigators only need a few sampling stations upstream and would be immediately downstream.) Whenever feasible, use
downstream of a discharge to determine whether the macroin- reference stations with physicochemical characteristics similar to
vertebrate community downstream is damaged. However, if the the receiving area’s substrate and overlying water.
objective is to delimit the extent of damage from a discharge or b. Locate a station immediately downstream of each dis-
series of discharges, they will need reference stations upstream charge or in the affected area in its immediate vicinity, as
of, downstream of, and bracketing all discharges. In marine appropriate.
waters, they may need to sample a nearby estuary. In open ocean c. If the discharge does not mix completely on entering the
waters, they may need to sample some distance from the dis- body of water, but instead channels along one side or disperses
charge point. in a specific direction, then locate stations in the left-bank
Characterize the physicochemical properties of faunal sam- (looking upstream), midchannel, and right-bank sections of the
pling-station substrate and overlying water. Measure such prop- stream; in concentric arcs in lakes and oceanic waters; or in any
erties as sediment size class distribution (sand, silt, and clay); other configuration that will meet study objectives.
organic content and toxic pollutant concentrations; temperature, d. Establish stations at various distances downstream from the
salinity, hardness, alkalinity, DO, total organic carbon (TOC), last discharge of concern to determine the linear extent of dam-
ammonia, sulfides, and nutrient (total and dissolved) concentra- age. In a marine environment, a nearby estuary may be sampled;
tions; biochemical oxygen demand; water depth; and velocity of in open ocean waters, samples may be taken in a nearby area
flowing streams. with comparable currents, depth, sediment characteristics, and
After gaining a thorough understanding of the water body’s salinity.
characteristics, select specific areas to be sampled. There is no e. To permit comparison of macroinvertebrate communities,
predetermined number of sampling stations that will be appro- be sure that all sampling stations are ecologically similar. For
priate for all situations. No water quality survey is routine, nor example, select stations with similar bottom substrate (e.g., sand,
can one be conducted totally on a “cookbook” basis. However, if gravel, rock, mud, organic content), depth, presence of riffles and

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BENTHIC MACROINVERTEBRATES (10500)/Sample Collection

pools, stream width, gradient, flow velocity, bank or shore cover, Collecting a representative statistical sample is difficult be-
salinity, hardness, TOC, nutrient and DO concentrations, and cause of variation in successive scientific samples. Without
wave exposure. knowing the sampling variability, investigators cannot know the
f. Collect samples for physical, toxicological (if applicable), degree to which the data truly represent the population. Make
and chemical analyses as close to biological sampling stations as replicate observations of a population if definitive statistical
possible to ensure correlation of findings; take such samples at inferences about the population will be made.2–11
the same time and from the same grab when possible. Collect Standardize sampling design to consider the following re-
substrate samples for physicochemical analyses from the upper quirements:
few centimeters, where most organisms live. a. Approximate the set of all samples that can be selected
g. Locate macroinvertebrate sampling stations in the best (i.e., separate the sampling universe into all possible samples).
physical habitat [areas not influenced by atypical conditions For example, if the location (site) containing the population has
(bridges, dams, etc.)]. an area of 1000 m2 and the sampling device samples an area of
h. Discharges in coastal areas may be subject to various 1 m2, then 1000 samples could be collected in the sampling
degrees of salt water intrusion (salt water wedge). Macroinver- universe.
tebrate populations may change drastically in such areas; docu- b. Assign each sample an equal probability of being selected.
ment and/or allow for this effect. Using the situation above, divide the area to be sampled into
i. When sampling in small, wadeable, first- to third-order 1000 discrete units.
streams, begin at the station farthest downstream and proceed c. Use a table of random numbers to select sites for sampling
upstream to minimize disruptions induced by the sampling itself. (i.e., sample randomly, not haphazardly).
This is unnecessary for non-wadeable streams and rivers. d. The sampling design outlined above is known as simple
For a long-term biological monitoring program, consider col- random sampling. When using this design, it is often advanta-
lecting macroinvertebrates at each station at least once during geous to determine the number of samples necessary for a certain
each of the annual seasons, though this may not always be level of precision:
necessary and would depend on the study design.1 More frequent
sampling may be necessary if effluents’ characteristics change or
spills occur. Make allowance for collections at night where
“drift” or night feeding organisms are of special concern. In
N ⫽ 冉 冊
t⫻s
D ⫻ x̄
2

general, the most critical period for macroinvertebrates in where:


streams is during periods of high temperature and low flow,
whereas in estuarine and marine environments it is the period of N ⫽ number of samples,
maximum stratification and poor vertical mixing. If available t ⫽ tabulated t value at 0.05 level with the degrees of
time and funds limit sampling frequency, make at least one freedom of preliminary survey (generally t ⬇ 2.0 at
survey during the critical time. larger sampling sizes),
s ⫽ sampling standard deviation of samples, known from a
2. Sampling Design preliminary survey,
D ⫽ required level of precision expressed as a decimal (0.30
Some terms have multiple meanings. In biology, for example, to 0.35 usually yields a statistically reliable estimate),
a population is a group of individuals that are all members of the and
same species or taxonomic group. In statistics, a population is x̄ ⫽ sample mean density of preliminary survey.
the entire set of values for the characteristic of interest in a whole
sampling universe. For example, researchers interested in deter- To estimate the number of samples needed, analysts first need
mining the mean density of worms on a lake bottom might take specific information (the mean and standard deviation) about the
ten grabs from lake sediments. The number of worms in each population to be sampled. Because this information is unknown
grab would be an observation, the density of worms would be the (because sampling has yet to occur), estimate the population’s
characteristic of interest, and the contents of each grab would be mean and standard deviation by one of three ways: conduct a
an experimental unit or sampling unit. The entire lake bottom pilot study, use results from an earlier or similar study, or make
would be the sampling universe and enough grabs to equate with educated estimates.3,12 For example, if investigators want to
the area of the entire lake bottom would be the population (of determine the mean chironomid density in relatively homoge-
units). neous lake sediments during summer, and they know that six
Similarly, the term sample has two often contradictory uses. In grabs taken the previous summer produced a mean density of
typical studies, observations usually are not made of all possible 4230 chironomids/m2 and a standard deviation of 1628 chirono-
sampling units; instead, observations are only made of a small mids/m2, then they can use these data to estimate their study’s
fraction of the total. Statistically, this set of observations is called mean chironomid density [⫾30%, with a 5% probability of error
a sample. In the example given above, the ten grabs collectively (␣ ⫽ 0.05)]. Using the formula given above,
would be a sample. However, in everyday language (and as used
in this book and most scientific publications), a sample is a
portion of the real world that has been selected for measurement
N⫽ 冉 2.5706 ⫻ 1628
0.30 ⫻ 4230
冊 2

(e.g., a water sample, plankton haul, or bottom grab). Therefore,


in the example above, each individual grab would be a sample (t ⫽ 2.5706 at a 5% probability of error and 5 degrees of
(i.e., “ten samples were taken”). freedom)

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BENTHIC MACROINVERTEBRATES (10500)/Sample Collection

Figure 10500:2. Ponar® grab.

Figure 10500:1. Petersen grab.


Before using each grab sampler, calibrate it for actual surface
area sampled.
N ⫽ 10.88 ⬇ 11 1) The Petersen grab (Figure 10500:1) is used for sampling
hard bottoms (e.g., sand, gravel, marl, and clay) in swift currents
Thus, it is estimated that 11 grabs will be necessary. and deep water.3 It is an iron, clam-type grab manufactured in
e. A simple random sampling design is useful when sampling various sizes that will sample an area between 0.06 and 0.09 m2.
relatively homogeneous areas. However, most taxa are not dis- It weighs approximately 13.7 kg, but may weigh as much as
tributed uniformly over water bottoms. Different habitats (sand, 31.8 kg when auxiliary weights are bolted to its sides. The extra
mud, gravel, or organic material) support different densities and weights make the grab stable in swift currents and provide more
species of organisms. In which case, a stratified random design cutting force in fibrous or firm bottom materials. Modify the
is more useful. sampler by adding end plates, by cutting large strips out of the
In a stratified random sampling design, a heterogeneous uni- top of each side, and by adding a hinged 30-mesh screen (as in
verse (different bottom substrates, current velocities, depths, the Ponar grab).14
temperatures, etc.) is divided into more homogeneous strata. To use the Petersen grab, set the hinged jaws and lower to the
Once the strata are defined, use simple random sampling within bottom slowly to avoid disturbing lighter bottom materials. Ease
each stratum. Stratified random sampling has two important rope tension to release the catch. As the grab is raised, the lever
advantages: it provides data on various subsets of a population system closes the jaws.
(e.g., density of benthic invertebrates in each sediment type), and 2) The Ponar grab (Figure 10500:2) is used increasingly in
it reduces variability because it deals with more homogeneous medium to deep rivers, lakes, reservoirs, and estuaries.15 It is
subpopulations, allowing for more accurate and precise popula- similar to the Petersen grab in size, weight, lever system, and
tion estimates. sample compartment, but has side plates and a screen on top of
The data needed to divide the population into various strata the sample compartment to prevent sample loss during closure.
usually is acquired via pre-study reconnaissance (a pilot study). With one set of weights, the standard 23- ⫻ 23-cm sampler
A systematic sampling design often is used in such pilot studies. weighs 20 kg. A 15- ⫻ 15-cm petite Ponar may be used. The
In a systematic-transect design, investigators conduct sampling large surface disturbance associated with a Ponar grab can be
at equal intervals along a number of transects in a habitat to reduced by installing hinged (rather than fixed) screen tops,
identify and locate existent strata.3,12 thereby reducing the pressure wave associated with the sam-
f. In descriptive studies, investigators should take at least pler’s descent. This sampler is best used for mud, sand, gravel,
three replicate sampling units per station.3,13 If statistical testing or small rocks with mud, but it can be used in all substrates
is planned, more replicates probably will be needed. except bedrock.
g. Standardize data acquisition and recording when practical. 3) The Van Veen grab (Figure 10500:3) is used to sample in
Use metric units. open marine waters and in large lakes. The sampler’s long arms
tend to act as stabilizers without disturbing water at the water–
3. Sampling Devices, Quantitative substrate interface. It is basically an improved version of the
Petersen grab for mud, gravel, pebble, and sand substrates. The
Quantitative and qualitative samplers have been designed to sampler is heavy; lower it from a boat or ship platform via
collect organisms from the bottom of different water bodies. The mechanical or hydraulic lifts.
most common quantitative sampling devices are the Petersen, 4) The Smith-McIntyre grab (Figure 10500:4) has the heavy
Ponar®,* and Ekman grabs and the Surber or square-foot stream steel construction of the Petersen, but its jaws are closed by
bottom sampler, all described below. strong coil springs.16 Its chief advantages are stability and easier
a. Grab samplers: control in rough water. Its bulk and heavy weight require oper-
ation from a large boat equipped with a winch. The 45.4-kg grab
* Registered trademark of Morris & Lee, Inc. d/b/a Wildlife Supply Co., Buffalo, can sample an area of 0.2 m2,17–19 but smaller models (0.1 m2 or
NY. 0.05 to 0.06 m2) are available.

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BENTHIC MACROINVERTEBRATES (10500)/Sample Collection

Figure 10500:3. Van Veen grab.

5) The Shipek®† grab (Figure 10500:5) is designed to take a


sample from virtually any substrate; samples have a surface area Figure 10500:5. Shipek grab.
of 0.04 m2 and are approximately 10 cm deep at the center.3
The sample compartment is composed of two concentric half
cylinders. When the grab touches bottom, inertia from a self- (23 cm or 30.5 cm tall) is available. To prevent sample overflow
contained weight releases a catch and helical springs rotate the and loss, place a Standard U.S. No. 30 sieve insert in the top of
inner half cylinder by 180°. The sample bucket may be disen- any Ekman grab sampler for deep sediments.
gaged from the upper semi-cylinder by releasing two retaining b. Riffle/run samplers:
latches. This grab is for special use in marine waters and large 1) Surber-type samplers (Figure 10500:7)20 consist of two
inland bodies of water (e.g., in compact substrates). brass frames— each 30.5 cm (1 ft) square— hinged together
6) The Ekman grab (Figure 10500:6) is only useful for sam- along one edge. When in use, the two frames are locked at right
pling mud, silt, muck, and sludge in water with little current.3 It angles, one frame marking off the area of substrate to be sam-
is difficult to use in areas with rocky or sandy bottoms or pled, and the other supporting a net to collect organisms washed
moderate macrophyte growth because small pebbles or grit or into it from the sample area.
macrophyte stems prevent proper jaw closure. The grab weighs
approximately 3.2 kg. The box-like part holding the sample has
spring-operated jaws on the bottom, which must be cocked
manually (exercise caution when cocking and handling the grab
because of possible injuries if jaws are tripped accidentally). At
the top of the grab are two hinged overlapping lids that are
partially held open during descent by water passing through the
sample compartment. These lids are held shut by water pressure
when the sampler is being retrieved. The grab is made in three
sizes—15 ⫻ 15 cm, 23 ⫻ 23 cm, and 30 ⫻ 30 cm— but the
smallest size is usually adequate. A taller model of this sampler

† Registered trademark of Morris & Lee, Inc. d/b/a Wildlife Supply Co., Buffalo,
NY.

Figure 10500:4. Smith-McIntyre grab. Figure 10500:6. Ekman grab.

6
BENTHIC MACROINVERTEBRATES (10500)/Sample Collection

Figure 10500:7. Surber or square-foot sampler.

The net usually is 69 cm long, and its first few centimeters and
its wings are constructed of heavier material (canvas, taffeta) to
increase durability. The rest of the net is a standard 30 mesh size
(595 to 600 ␮m). While a finer mesh might collect more of the
smaller invertebrates and young instars, it also will clog more
easily and resist the current more, possibly resulting in a loss of
organisms due to backwashing. This sampler is specific for
macrobenthos; many microcomponents of the benthos are not
collected.
Use this sampler in shallow, flowing water (no more than
30 cm deep). In deeper water, some organisms may flow over the
top of the sampler. Position sampler securely on the stream
bottom parallel to water flow, with the net portion downstream.
Take care not to disturb the substrate upstream of sampler. Leave
no gaps under the edges of the frame that would allow water to
wash under the net. Fill any gaps along the back edge of the
sampler by carefully shifting rocks and gravel along the outside Figure 10500:8. Phleger core sampler.
edge. When sampler is in place (it may be necessary to hold it in
place with one hand in a strong current), carefully turn over and
lightly hand-rub all rocks and large stones inside the frame to
synthetic sponge. Remove organisms that stick to foam and
dislodge organisms clinging to them. Examine each stone for
include in sample.
organisms, larval or pupal cases, etc., that may be clinging to it
2) Hess-type samplers are cylindrical with enclosed sides and
before discarding. Scrape attached algae, insect cases, etc., from
an open top. They function much like Surber-type samplers.3
the stones into the sampler net. Stir remaining gravel and sand
c. Core or cylindrical samplers: Use core or cylindrical sam-
with hands or a stick to a depth of 5 to 10 cm (depending on
plers to sample sediments in depth. Efficient use as surface
substrate) to dislodge bottom-dwelling organisms. It may be
samplers requires dense animal populations. Core samplers vary
necessary to hand-pick some mussels and snails that the current
from hand-pushed tubes to explosive-driven and automatic-
does not carry into the net.
surfacing models.3,21
Remove sample by inverting net into sample container. Care-
1) The Phleger corer (Figure 10500:8) is widely used and
fully examine net for small organisms clinging to it. Remove
operates via gravity.3 Styles and weights vary among manufac-
these—preferably with forceps to avoid damage—and include in
turers; some use interchangeable weights that allow variations
sample. Rinse sampler net after each use.
between 7.7 and 35.0 kg, while others use fixed weights weigh-
A common problem when using the Surber sampler is that
ing 41.0 kg or more. Core length will vary with substrate texture.
organisms wash under the bottom edge of the sampler. The
2) The KB®‡ core sampler (Figure 10500:9), or a modification
following modifications have been suggested for different sub-
known as the Kajak–Brinkhurst corer, may be useful in obtaining
strates:
estimates of the standing stock of benthic macroinvertebrates
• For loose gravel—Extend bottom edge of Surber frame to 5
inhabiting soft sediments.22
or more cm so frame can be inserted deeper into substrate. This
3) Box core samplers23–27 are used to sample soft substrate in
method works well in soft substrates (e.g., sand and gravel),
large rivers, lakes, and estuaries. They are available in several
where the current causes substrate shifting.
sizes, can sample a variety of sediments, and are used in marine
• For coarse gravel and rock—Add serrated extension to the
waters and in the Great Lakes3,28 to collect benthic macrofauna.
back edge of frame to secure it and reduce washing from under
The sampler may be deployed from ships or platforms, but
this edge. This method is helpful in hard gravel and rock sub-
diver-collected cores are preferred.
strates, where sinking the entire frame is impossible.
• For gravel and bedrock—Add a 5-cm band of flexible ma-
terial to the bottom edge of sampler to create a seal in rocky, ‡ Registered trademark of Morris & Lee, Inc. d/b/a Wildlife Supply Co., Buffalo,
uneven substrates. Make band of foam rubber or fine-textured NY.

7
BENTHIC MACROINVERTEBRATES (10500)/Sample Collection

Figure 10500:10. Wilding or stovepipe sampler.

d. Drift samplers: Drift samplers, usually in the form of nets


(Figure 10500:11), are anchored in flowing water to capture
macroinvertebrates that have migrated or been dislodged from
the bottom substrates into the current. Drift organisms are im-
portant to the stream ecosystem because they are prey for fish
and should be considered in the study of fish populations. Drift
Figure 10500:9. KB corer. organisms respond to pollutional stresses (e.g., spills) by in-
creased drift from an affected area so drift is important in
water-quality investigations, especially of spills of toxic materi-
The KC Box Corer frame is made of 60 ⫻ 60 ⫻ 6 mm, als. Drift also is a factor in recolonizing denuded areas and
sandblasted, hot galvanized stainless steel square tubes.27 The 30 contributes to recovery of disturbed streams.
⫻ 20 cm or 32 ⫻ 32 cm sampler tube is made of 3-mm Use nets with a 929-cm2 upstream opening and mesh equiv-
electropolished stainless steel. The sampling surface area is alent to U.S. Standard No. 30 screen (595-␮m pore size). After
either 600 or 100 cm2, and it samples a depth of 20 to 40 cm. placing the net in the water, frequently remove organisms and
Preferably use a box coring device with a rectangular corer debris to prevent clogging and subsequent diversion of water at
whose cutting arm can seal the sample before retracting from the the net opening. Use replicate samples, as appropriate, to meet
bottom. To sample enough individuals and taxa, and integrate study objectives. Set drift-net samples for any specified time
the patchy distribution of benthic fauna, use a sampler with a (usually 1 to 3 h) but use the same time for each station.
surface area of at least 100 cm2 and a sediment depth of at least Sampling between dusk and 1 a.m. is optimal.
20 cm. A box corer that can sample deeper sediment may be The total quantity (numbers or biomass) of organisms drifting
needed to collect deep-burrowing infauna. For sandy sediments, past a given station is the best measure of drift intensity. Report
it may be necessary to substitute a grab sampler to adequately data in terms of numbers or biomass/m3.31–33
penetrate sediment and collect samples. Visually inspect each
sample to ensure that an undisturbed, adequate amount of sample 4. Sampling Devices, Qualitative
is collected.
4) The Wilding or stovepipe sampler (Figure 10500:10)29 –30 is When sampling qualitatively, search for organisms in as many
made in various sizes and with many modifications.3 The Wild- habitats as possible.34 Collect samples by any method that will
ing sampler is made from any tubular material (e.g., 60- to 75-cm capture representative species.
sections of 30-cm-diam stovepipe30 or 75-cm sections of 30-cm-
diam aluminum irrigation pipe fitted with handles). The Maine
Department of Environmental Pollution uses a 5-gal bucket with
the bottom removed.
The sampler is pressed into the substrate and its contents are
agitated. It is especially useful for quantitatively sampling a
bottom with dense, vascular plant growth. It may be used to
sample vegetation, mud–water interface sediment, or most shal-
low stream substrates. However, large volumes of vegetation,
when sampled in this way, may require a great deal of time for
laboratory processing. Figure 10500:11. Drift net sampler.

8
BENTHIC MACROINVERTEBRATES (10500)/Sample Collection

a. Dip, kick nets are the most versatile collection devices for
shallow, flowing water and for lake shorelines. When combined
with a standardized kicking technique,35 these nets are appropri-
ate for quantitatively sampling macroinvertebrates.36
b. Tow nets, dredges, or trawls range from simple sled-
mounted nets to complicated devices with teeth that dig into the
bottom. Some models feature special apparatus to hold the net
open during towing and to close it during descent and retrieval.
Available styles have been discussed elsewhere.21,37,38

5. Sampling Devices, Artificial Substrate Samplers

Artificial substrate samplers are devices of standard compo-


sition and configuration placed in water for a predetermined
exposure period to be colonized by macroinvertebrate commu-
nities. Because many of the physical variables encountered in
bottom sampling are minimized (e.g., depth, light penetration,
temperature differences, and species substrate preferences), ar-
tificial substrate sampling complements other types of sampling.
Like natural submerged substrates (e.g., logs and pilings), arti-
ficial substrates are colonized primarily by immature aquatic
insects, crustaceans, coelenterates, bryozoans, and to some ex-
tent worms, gastropods, and mollusks. In lotic systems, the
organisms that colonize artificial substrates are primarily drift
organisms (e.g., immature insects and eggs) carried by water
currents. Placement conditions should be similar so the numbers
and kinds of organisms reflect the capacity to support aquatic
life.
Position artificial substrates in the euphotic zone (0.3 m) for
maximum abundance and diversity of macroinvertebrates.13 Op-
timum time for substrate colonization is 6 weeks in most waters.
For uniformity of depth, suspend sampler from floats on a Figure 10500:12. Hester–Dendy artificial substrate unit.
3.2-mm steel cable. If vandalism is a problem, use subsurface
floats or place sampler near the bottom. Regardless of installa-
tion technique, use uniform procedures. the top 9 plates by one spacer. Separate Plate 10 by two spacers,
At shallow water stations (less than 1.2 m deep), install Plates 11 and 12 by three spacers, and Plates 13 and 14 by four
samplers so they are midway in the water column at low flow. spacers. The sampler is approximately 14 cm long and 7.5 cm in
For samplers installed in July, when water depth is about 1.2 m diameter, has an exposed surface area of approximately 1300 cm2,
and the August average low flow is 0.6 m, install 0.3 m above the and weighs about 0.45 kg. Do not reuse samplers exposed to oils
bottom. Take care not to let samplers touch the bottom or they and chemicals that may inhibit colonization. Because it is cylin-
may become covered with silt, thereby increasing the sampling drical, the sampler fits inside a wide-mouth container for ship-
error. In shallow streams with sheet rock bottoms, secure artifi- ping and storage. The sampler is inexpensive, compact, and
cial substrates to 0.95-cm steel rods driven into the substrate or lightweight.13,40,41
secure to rods mounted on low, flat, rectangular blocks. Another type of modified Hester–Dendy, multiple-plate arti-
Before removing samples from water, it may be necessary to ficial substrate sampler is constructed of 0.3-cm tempered hard-
enclose them in an oversized plastic bag (double wrapping) that board cut into 7.6-cm square plates and 2.5-cm square spacers.34
is tightly sealed to prevent possible organism loss or else remove Eight plates and twelve spacers are used for each sampler. The
them via a large dip net (mesh equivalent to a U.S. Standard plates and spacers are placed on a 1/4-in. (0.64-cm) eyebolt so
No. 30 sieve). Disassemble sampler and brush it in a pan of water there are three single spaces, three double spaces, and one triple
in the field or add preservative to the bag containing the intact space between plates. The sampler’s total surface area, excluding
sampler, and disassemble and brush it later in the laboratory. the eyebolt, is 939 cm2 (0.9 m2). Generally, five samplers are
Although many styles of artificial substrate samplers have used and placed in streams tied to a concrete construction block
been tested,39 the basket sampler13 and the Fullner40 modifica- as anchor. This prevents samplers from coming into contact with
tion of the Hester–Dendy41 multiplate sampler are widely used. natural substrates.
a. Multiple-plate (modified Hester–Dendy) sampler (Figure b. The basket sampler13 (Figure 10500:13) is a cylindrical
10500:12) is constructed of 0.3-cm-thick tempered hardboard “barbecue” basket 28 cm long and 17.8 cm in diameter, filled
with 7.5-cm round plates and 2.5-cm round spacers with center- with approximately thirty 5.1-cm-diam rocks or rocklike mate-
drilled holes. The plates are separated by spacers on a 0.63-cm- rial weighing 7.7 kg. A hinged side door allows access to the
diam eyebolt, held in place by a nut at the top and bottom. In contents. The sampler provides an estimated 0.24 m2 of surface
each sampler, 14 large plates and 24 spacers are used. Separate area for colonization. The factors governing proper installation

9
BENTHIC MACROINVERTEBRATES (10500)/Sample Collection

specific sampling sites, but a SCUBA diver is required to collect


samples.45 More accurately located sampling sites and the ability
to collect a large number of replicate samples may outweigh the
disadvantage of using a diver. Suction samplers have been used
widely in sampling marine environments, but they have obvious
depth limitations.

7. References
Figure 10500:13. Basket sampler.
1. ALDEN, R.W., III, S.B. WEISBERG, J.A. RANASINGHE & D.M. DAUER.
1997. Optimizing temporal sampling strategies for benthic environ-
and collection are the same as those described for the multiplate mental monitoring programs. Mar. Pollut. Bull. 34:913.
sampler. Some investigators prefer using the basket because 2. SNEDECOR, G.W. & W.G. COCHRAN. 1967. Statistical Methods. Iowa
natural substrate materials are used for colonization. State Univ. Press, Ames.
c. Marsh net sampler (Figure 10500:14) is used for sampling 3. KLEMM, D.J., P.A. LEWIS, F. FULK & J.M. LAZORCHAK. 1990. Mac-
macroinvertebrates in estuarine and marine environments.42 It roinvertebrate Field and Laboratory Methods for Evaluating the
can be used in different habitats (e.g., marsh, beach, tidal creek, Biological Integrity of Surface Waters, EPA-600/4-90-030. Envi-
and tidal flat) of estuarine and marine intertidal zones to depths ronmental Monitoring Systems Laboratory, U.S. Environmental
Protection Agency, Cincinnati, Ohio.
of 3 m. The metal frame is constructed of No. 22 galvanized
4. GREEN, R.H. 1979. Sampling Design and Statistical Methods for
sheet metal and 1/4-in. (6-mm) welding rods. A 0.5-m plankton Environmental Biologists. John Wiley & Sons, New York, N.Y.
net of nylon monofilament screen is laced to the posterior end of 5. ZAR, J.H. 1984. Biostatistical Analysis, 2nd ed. Prentice-Hall,
the frame. The net has a bayonet-type cod end for easy removal. Englewood Cliffs, N.J.
The mesh size of the plankton net and cod end is about 1 mm 6. UNDERWOOD, A.J. & C.H. PETERSON. 1988. Towards an ecological
(bar measure). The frame and net weigh 5 kg. The collecting framework for investigating pollution. Mar. Ecol. Progr. Ser.
procedures are the same in all intertidal zone habitats. The net is 46:227.
placed at one end of the sampling area, and 30 m of rope is paid 7. MANTEL, N. 1987. The detection of disease clustering and a gener-
out in an arc to prevent the operator from disturbing the sampling alized regression approach. Cancer Res. 27:200.
site. The net is then retrieved by hand at a rate of about 0.3 m/s. 8. LEGENDRE, P. & M.J. FORTIN. 1989. Spatial patterns and ecological
analysis. Vegetation 80:107.
Advantages are that the sampling distance does not have to be
9. GREEN, R.H., J.M. BOYD & J.S. MACDONALD. 1993. Relating sets of
measured before taking the sample, the net can be towed at a variables in environmental studies: the sediment quality triad as a
constant speed, and samples also can be taken over soft mud paradigm. Environmetrics 4:439.
bottoms.42 10. UNDERWOOD, A.J. 1994. On beyond BACI: sampling designs that
might reliably detect environmental disturbances. Ecol. Appl. 4:3.
6. Suction Samplers 11. LANDIS, W.G., G.B. MATTHEWS, R.A. MATTHEWS & A. SARGENT.
1994. Application of multivariate techniques to endpoint determi-
Suction samplers are widely used to collect benthic macroin- nation, selection and evaluation in ecological risk assessment. En-
vertebrate samples.43,44 These samplers can be placed directly on viron. Toxicol. Chem. 13:1917.

Figure 10500:14. Marsh net sampler.

10
BENTHIC MACROINVERTEBRATES (10500)/Sample Collection

12. GAUGUSH, R.F. 1987. Sampling Design for Reservoir Water Quality Biological Field Assessment of Ohio Surface Waters & Volume III,
Investigations, Instruction Rep. E-87-1. Waterways Experiment Sta- Standardized Biological Field Sampling and Laboratory Methods
tion, Vicksburg, Miss. for Assessing Fish and Macroinvertebrates Communities. Ohio En-
13. MASON, W.T., JR., C.I. WEBER, P.A. LEWIS & E.C. JULIAN. 1973. vironmental Protection Agency, Div. Water Quality Monitoring &
Factors affecting the performance of basket and multiplate macro- Assessment, Surface Water Section, Columbus.
invertebrate samplers. Freshwater Biol. 3:409. 35. FROST, S., A. HUN & W. KERSHAW. 1971. Evaluation of a kicking
14. WEBER, C.I., ed. 1973. Biological Field and Laboratory Methods for technique for sampling stream bottom fauna. Can. J. Zool. 49:167.
Measuring the Quality of Surface Waters and Effluents, EPA-670/ 36. CROSSMAN, J.S., J. CAIRNS, JR. & R.L. KAESLER. 1973. Aquatic
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Ponar grab sampler. In Standard Guide for Collection, Storage, icological Testing and Selection of Samplers Used to Collect Ben-
Characterization, and Manipulation of Sediments for Toxicological thic Invertebrates, 2010 Annual Book of Standards, Vol. 11.06:
Testing and Selection of Samplers Used to Collect Benthic Inver- Biological Effects and Environmental Fate; Biotechnology. [Annex
tebrates, 2010 Annual Book of Standards, Vol. 11.06: Biological A1 in ASTM E1391-03 (Reapproved 2008)]. American Soc. Test-
Effects and Environmental Fate; Biotechnology. [Annex A1 in ing & Materials, W. Conshohocken, Pa.
ASTM E1391-03 (Reapproved 2008)]. American Soc. Testing & AMERICAN SOCIETY for TESTING and MATERIALS. 2010. ASTM D 4401-84:
Materials, W. Conshohocken, Pa. Standard practice for collecting benthic macroinvertebrates with
AMERICAN SOCIETY for TESTING and MATERIALS. 2010. ASTM D 4343-84: Petersen grab sampler. In Standard Guide for Collection, Storage,
Standard practice for collecting benthic macroinvertebrates with Characterization, and Manipulation of Sediments for Toxicological
Ekman grab sampler. In Standard Guide for Collection, Storage, Testing and Selection of Samplers Used to Collect Benthic Inver-
Characterization, and Manipulation of Sediments for Toxicological tebrates, 2010 Annual Book of Standards, Vol. 11.06: Biological
Testing and Selection of Samplers Used to Collect Benthic Inver- Effects and Environmental Fate; Biotechnology. [Annex A1 in
tebrates, 2010 Annual Book of Standards, Vol. 11.06: Biological ASTM E1391-03 (Reapproved 2008)]. American Soc. Testing &
Effects and Environmental Fate; Biotechnology. [Annex A1 in Materials, W. Conshohocken, Pa.
ASTM E1391-03 (Reapproved 2008)]. American Soc. Testing & AMERICAN SOCIETY for TESTING and MATERIALS. 2010. ASTM D 4407-84:
Materials, W. Conshohocken, Pa. Standard practice for collecting benthic macroinvertebrates with
AMERICAN SOCIETY for TESTING and MATERIALS. 2010. ASTM D 4344-84: orange peel grab sampler. In Standard Guide for Collection, Stor-
Standard practice for collecting benthic macroinvertebrates with age, Characterization, and Manipulation of Sediments for Toxico-
Smith–McIntyre grab sampler. In Standard Guide for Collection, logical Testing and Selection of Samplers Used to Collect Benthic
Storage, Characterization, and Manipulation of Sediments for Tox- Invertebrates, 2010 Annual Book of Standards, Vol. 11.06: Biolog-
icological Testing and Selection of Samplers Used to Collect Ben- ical Effects and Environmental Fate; Biotechnology. [Annex A1 in
thic Invertebrates, 2010 Annual Book of Standards, Vol. 11.06: ASTM E1391-03 (Reapproved 2008)]. American Soc. Testing &
Biological Effects and Environmental Fate; Biotechnology. [Annex Materials, W. Conshohocken, Pa.
A1 in ASTM E1391-03 (Reapproved 2008)]. American Soc. Test- AMERICAN SOCIETY for TESTING and MATERIALS. 2010. ASTM D 4556-85:
ing & Materials, W. Conshohocken, Pa. Standard guide for selecting stream-net sampling devices for col-
AMERICAN SOCIETY for TESTING and MATERIALS. 2010. ASTM D 4345-84: lecting benthic macroinvertebrates. In Standard Guide for Collec-
Standard practice for collecting benthic macroinvertebrates with tion, Storage, Characterization, and Manipulation of Sediments for
Van Veen grab sampler. In Standard Guide for Collection, Storage, Toxicological Testing and Selection of Samplers Used to Collect
Characterization, and Manipulation of Sediments for Toxicological Benthic Invertebrates, 2010 Annual Book of Standards, Vol. 11.06:
Testing and Selection of Samplers Used to Collect Benthic Inver- Biological Effects and Environmental Fate; Biotechnology. [Annex
tebrates, 2010 Annual Book of Standards, Vol. 11.06: Biological A1 in ASTM E1391-03 (Reapproved 2008)]. American Soc. Test-
Effects and Environmental Fate; Biotechnology. [Annex A1 in ing & Materials, W. Conshohocken, Pa.
ASTM E1391-03 (Reapproved 2008)]. American Soc. Testing & AMERICAN SOCIETY for TESTING and MATERIALS. 2010. ASTM D 4557-85:
Materials, W. Conshohocken, Pa. Standard practice for collecting benthic macroinvertebrates with
AMERICAN SOCIETY for TESTING and MATERIALS. 2010. ASTM D 4346-84: Surber and related type samplers. In Standard Guide for Collection,
Standard practice for collecting benthic macroinvertebrates with Storage, Characterization, and Manipulation of Sediments for Tox-
Okean 50 grab sampler. In Standard Guide for Collection, Storage, icological Testing and Selection of Samplers Used to Collect Ben-
Characterization, and Manipulation of Sediments for Toxicological thic Invertebrates, 2010 Annual Book of Standards, Vol. 11.06:
Testing and Selection of Samplers Used to Collect Benthic Inver- Biological Effects and Environmental Fate; Biotechnology. [Annex
tebrates, 2010 Annual Book of Standards, Vol. 11.06: Biological A1 in ASTM E1391-03 (Reapproved 2008)]. American Soc. Test-
Effects and Environmental Fate; Biotechnology. [Annex A1 in ing & Materials, W. Conshohocken, Pa.
ASTM E1391-03 (Reapproved 2008)]. American Soc. Testing & AMERICAN SOCIETY for TESTING and MATERIALS. 2010. ASTM D 4558-85:
Materials, W. Conshohocken, Pa. Standard practice for collecting benthic macroinvertebrates with

12
BENTHIC MACROINVERTEBRATES (10500)/Sample Processing and Analysis

drift nets. In Standard Guide for Collection, Storage, Characteriza- Book of Standards, Vol. 11.06: Biological Effects and Environmental
tion, and Manipulation of Sediments for Toxicological Testing and Fate; Biotechnology. [Annex A1 in ASTM E1391-03 (Reapproved
Selection of Samplers Used to Collect Benthic Invertebrates, 2010 2008)]. American Soc. Testing & Materials, W. Conshohocken, Pa.
Annual Book of Standards, Vol. 11.06: Biological Effects and AMERICAN SOCIETY for TESTING and MATERIALS. 2010. ASTM E 1469-92:
Environmental Fate; Biotechnology. [Annex A1 in ASTM Standard practice for collecting benthic macroinvertebrates with
E1391-03 (Reapproved 2008)]. American Soc. Testing & Materials, multiplate sampler. In Standard Guide for Collection, Storage,
W. Conshohocken, Pa. Characterization, and Manipulation of Sediments for Toxicological
AMERICAN SOCIETY for TESTING and MATERIALS. 2010. ASTM E 1468-92: Testing and Selection of Samplers Used to Collect Benthic Inver-
Standard practice for collecting benthic macroinvertebrates with basket tebrates, 2010 Annual Book of Standards, Vol. 11.06: Biological
sampler. In Standard Guide for Collection, Storage, Characterization, Effects and Environmental Fate; Biotechnology. [Annex A1 in
and Manipulation of Sediments for Toxicological Testing and Selec- ASTM E1391-03 (Reapproved 2008)]. American Soc. Testing &
tion of Samplers Used to Collect Benthic Invertebrates, 2010 Annual Materials, W. Conshohocken, Pa.

10500 C. Sample Processing and Analysis

1. Sample Processing tissue proteins to retain characteristics of the soft body (e.g.,
segmented worms) form.8,9
After collecting a benthic sample, pour the slurry gradually For qualitative samples, place rocks, sticks, and other objects
into a sieve bucket. Gently wash slurry over screen to prevent in a white pan partially filled with water. Many animals will float
damaging or losing specimens. Slurries that clog the screen free from these objects and can be removed with forceps.
require removal of screened material. A series of one or two Assign identification numbers either in the field or at the
coarser screens (e.g., 1-cm and 0.5-cm mesh) will hold back laboratory and transcribe information from the labels to a per-
larger materials (e.g., leaves, sticks, shells, and gravel) while manent ledger. The ledger provides a convenient reference in
permitting organisms and smaller materials to pass through to identifying the number of samples collected at various places,
the bottom sieve. Carefully check rocks, sticks, shells, and other time of sampling, and water characteristics.
objects for attached or burrowed organisms before discarding. A Preserve and store in 70% ethanol organisms taken in the field
soft-bristled toothbrush may be used to remove attached inver- or from artificial substrates and sieved with a U.S. Standard
tebrates from rocks, sticks, and similar objects. No. 30 sieve. For special studies and to retain anatomical form
and structures, fix soft-bodied organisms first with 5 to 10%
Wash residual material on the screen into a container. A
buffered formalin. (CAUTION: For health and safety reasons,
cheesecloth bag is useful because it does not restrict the quantity
always take care when using 5 to 10% buffered formalin, or
of wash water. Label containers with a collection code but do not
avoid using it to fix or preserve organisms in the field or in
affix labels to lids. Similar labels can be written with pencil or the laboratory. Never discard fixatives or preservatives into
indelible ink on high-rag-content paper and placed in the con- the environment.)
tainer. Record label code on a field sheet that describes location,
date, type of sample, collector’s name, and other pertinent in-
formation. 2. Sorting and Identification
Use laboratory elutriation devices,1,2 as appropriate, to reduce
time required to sort benthic organisms from samples containing Whether organisms are sorted in the field or the laboratory,
large amounts of silt, mud, or clay. Wash screened material into follow consistent procedures. Before processing a sample, trans-
a container and fix the contents in a solution of 10% buffered fer information from the label to a data sheet that provides space
formalin or 70% ethanol.3– 6 If ethanol is used, do not fill more for scientific names and number of individuals. Place sample
directly in a shallow white tray with water for sorting. To
than one-half the container with screened material. Preserve and
facilitate sorting organisms from detritus, the organisms may be
store animals with calcareous shells or exoskeletons (mussels,
stained with rose bengal (200 mg/L or enough to achieve a light
snails, crayfish, and ostracods) in 70% ethanol.6,7
pink color) in the formalin or ethanol preservative for at least
Some macroinvertebrates (soft-bodied animals) are identified 24 h.10 (NOTE: Excessive staining may prevent specific identifi-
more easily if they are relaxed to prevent constriction during cation of some specimens.) Examine entire sample and separate
preservation. Common relaxants include carbonated water (soda organisms unless they occur in very large numbers. If a sub-
water) or carbon dioxide added to water. Other relaxants include sample is sorted, take care that rare forms are not excluded. As
aqueous solutions of 70% ethyl alcohol, 2% nicotine sulfate, organisms are picked from the sample, sort under a scanning lens
propylene phenoxetol, or 5 to 10% solutions of either chlorotone, or stereoscopic microscope, separate them into different taxo-
chloral hydrate, or magnesium sulfate added gradually to water nomic categories, identify to the lowest taxonomic level to meet
containing the soft-bodied animals until the degree of relaxation data quality objectives, and record on the data sheet. Place
sought is reached. Narcotize organisms before fixing them. Ide- animals in separate vials according to category and fill vials with
ally, fix annelid specimens (oligochaetes) in 5 to 10% buffered 70% ethanol. Inside vials, place labels containing sample track-
formalin before preserving them in 70 to 80% ethanol (NOTE: ing number, date collected, sampling location, and names of
Alcohol is not a satisfactory tissue fixative). Fixation stabilizes organisms.

13
BENTHIC MACROINVERTEBRATES (10500)/Sample Processing and Analysis

Identify animals in each vial using stereoscopic and compound BOUSFIELD, E.L. 1973. Shallow-Water Gammaridean Amphipoda of
microscopes (whichever is needed) and available experience and New England. Cornell University Press, Ithaca, N.Y.
resources. Identify organisms to species level if possible. Addi- DAY, J.H. 1973. New Polychaeta from Beaufort, with a key to all species
tional sources of information on laboratory techniques are avail- recorded from North Carolina, U.S. Circ. No. 375. National Oce-
able, as well as identification guides and taxonomic keys of anic Atmospheric Admin., National Marine Fisheries Serv., Wash-
macroinvertebrates (see 10500C.4 and Section 10900). ington, D.C.
WATLING, L. & D. MAURER. 1973. Guide to the Macroscopic Estuarine
and Marine Invertebrates of the Delaware Bay Region, Delaware
3. References Bay Rep. Ser. Vol. 5, p. 178. Univ. Delaware, Newark.
WILLIAMS, A.B. 1974. Marine flora and fauna of the northeastern United
1. WORSWICK, J.M. & M.T. BARBOUR. 1974. An elutriation apparatus
States. Crustacean: Decapoda, U.S. Circ. No. 389. National Oceanic
for macroinvertebrates. Limnol. Oceanogr. 19:538.
Atmospheric Admin., National Marine Fisheries Serv., Washing-
2. LAUFF, G.H., K.W. CUMMINS, C.H. ERIKSON & M. PARKER. 1961. A
method for sorting bottom fauna samples by elutriation. Limnol. ton, D.C.
Oceanogr. 6:462. FOX, R.S. & K.H. BYNUM. 1975. The amphipod crustaceans of North
3. EDMONDSON, W.T., ed. 1959. Ward and Whipple’s Freshwater Bi- Carolina estuarine waters. Chesapeake Sci. 16:223.
ology, 2nd ed. John Wiley & Sons, New York, N.Y. MORRIS, P.A. 1975. A Field Guide to Shells of the Atlantic and Gulf
4. COOK, D.G. & R.O. BRINKHURST. 1973. Marine Flora and Fauna of Coasts and the West Indies. Houghton Mifflin Co., Boston, Mass.
the Northeastern United States, Annelida: Oligochaeta, NOAA SMITH, R.I. & J.T. CARLTON, eds. 1975. Light’s Manual: Intertidal
Tech. Rep. NMFS CIRC-374. U.S. Dept. Commerce, National Invertebrates of the Central California Coast, 3rd ed. University of
Oceanic Atmospheric Admin., National Marine Fisheries Serv., California Press, Berkeley.
Seattle, Wash. BLAXTER, J.H.S., S.F.S. RUSSELL & S.M. YONGE. 1980. The species of
5. KLEMM, D.J. 1982. Leeches (Annelida: Hirudinea) of North Amer- mysids and key to genera. Advan. Mar. Biol. 18:7.
ica, EPA-600/3-82-025. Environmental Monitoring & Support Lab., BUTLER, T.H. 1980. Shrimps of the Pacific Coast of Canada. Can. Bull.
U.S. Environmental Protection Agency, Cincinnati, Ohio. Fish. Aquat. Sci. 202:1.
6. PENNAK, R.W. 1989. Freshwater Invertebrates of the United SIEG, J. & R.N. WINN. 1981. The Tanaidae (Crustacea: Tanaidacea) of
States—Protozoa to Mollusca, 3rd ed. John Wiley & Sons, Inc., California, with a key to the world genera. Proc. Biol. Soc. Wash.
New York, N.Y. 94(2):315.
7. BURCH, J.B. 1972. Freshwater Sphaeriacean Clams (Mollusca: Pele- HEARD, R.W. 1982. Guide to Common Tidal Marsh Invertebrates of the
cypoda) of North America. U.S. Environmental Protection Agency, Northeastern Gulf of Mexico, MASGP-79-004. Mississippi–
Cincinnati, Ohio. Alabama Sea Grant Consortium.
8. KLEMM, D.J., ed. 1985. A Guide to the Freshwater Annelida PRICE, W.W. 1982. Key to the shallow water Mysidacea of the Texas
(Polychaeta, Naidid and Tubificid Oligochaeta, and Hirudinea) of coast with notes on their ecology. Hydrobiologia 93:9.
North America. Kendall/Hunt Publ. Co., Dubuque, Iowa. WRONA, F.J., J.M. CULP & R.W. DAVIES. 1982. Macroinvertebrate sub-
9. KATHMAN, R.D. & R.O. BRINKHURST. 1998. Guide to the Freshwater sampling: a simplified apparatus and approach. Can. J. Fish. Aquat.
Oligochaetes of North America. Aquatic Resources Center, College Sci. 39:1051.
Grove, Tenn. WILLIAMS, A.B. 1984. Shrimp, lobsters, and crabs of the Atlantic Coast
10. MASON, W.T., JR. & P.P. YEVICH. 1967. The use of phloxine B and of the Eastern United States, Maine to Florida. Smithsonian Insti-
rose bengal stains to facilitate sorting benthic samples. Trans. Amer. tution Press, Washington, D.C.
Microsc. Soc. 86:221. BRINKHURST, R.O. 1986. Guide to the Freshwater Aquatic Microdrile
Oligochaetes of North America, Canadian Spec. Publ. Fisheries &
4. Bibliography Aquatic Science 84. Dept. Fisheries & Oceans, Ottawa, Ont.
PENNAK, R.W. 1989. Fresh-Water Invertebrates of the United States.
PETTIBONE, M.H. 1963. Marine polychaete worms of the New England Protozoa to Mollusca, 3rd ed. John Wiley & Sons, Inc., New York,
region. I. Families Aphroditidae through Trochochaetidae. U.S. N.Y.
Nat. Mus. Bull. 227:1. VECCHIONE, M., C.F.E. ROPER & M.J. SWEENEY. 1989. Marine Flora and
SMITH, R.I., ed. 1964. Keys to marine invertebrates of the Woods Hole
Fauna of the Eastern United States. Mollusca: Cephalopoda. Na-
Region, Contrib. No. 11. Systematics-Ecology Program, Marine
tional Marine Fisheries Serv., National Systematics Lab., Washing-
Biological Lab., Woods Hole, Mass.
ton D.C.
MCCAIN, J.C. 1968. The Caprellidae (Crustacea: Amphipoda) of the
KLEMM, D.J., P.A. LEWIS, F. FULK & J.M. LAZORCHAK. 1990. Macroin-
Western North Atlantic, Bull. 278. Smithsonian Institute, Washing-
vertebrate Field and Laboratory Methods for Evaluating the Bio-
ton, D.C.
SCHULTZ, G.A. 1969. How to Know the Marine Isopod Crustaceans. logical Integrity of Surface Waters, EPA-600/4-90-030. Environ-
Wm. C. Brown Company Publ., Dubuque, Iowa. mental Monitoring Systems Lab., U.S. Environmental Protection
FOSTER, N.M. 1971. Spionidae (Polychaete) of the Gulf of Mexico and Agency, Cincinnati, Ohio.
the Caribbean Sea. Studies on the Fauna of Curacao and Other PACKARSKY, B.L., P.R. FRAISSINET, M.A. PENTON & D.J. CONKLIN, JR.
Caribbean Islands. Vitg. Natuurw. Studkring Suriname 63, 36(129):1. 1990. Freshwater Macroinvertebrates of Northeastern North Amer-
GOSNER, K.L. 1971. Guide to Identification of Marine and Estuarine ica. Cornell University Press, Ithaca, N.Y.
Invertebrates. Cape Hatteras to the Bay of Fundy. Wiley-Inter- KLEMM, D.J. 1991. Taxonomy and pollution ecology of the Great Lakes
science, New York, N.Y. Region leeches (Annelida: Hirudinea). Mich. Acad. 24:37.
HOLME, N.A. & A.D. MCINTYRE. 1971. Methods for the Study of Marine LOVELL, L. & R.G. VELARDE. 1991. Regional Standardization of Taxon-
Benthos, IBP Handbook No. 16. Blackwell Scientific Publications, omy. In Proc. Symposium, Biological Criteria: Research and Reg-
Oxford, England. ulation, EPA 440/5-91/005. Off. Water, U.S. Environmental Pro-
LEWIS, P.A. 1972. References for the Identification of Freshwater Mac- tection Agency, Washington, D.C.
roinvertebrates, EPA-R4-F2-006. U.S. Environmental Protection CUFFNEY, T.F., M.E. GURTZ & M.R. MEADOR. 1993. Guidelines for the
Agency, Cincinnati, Ohio. Processing and Quality Assurance of Benthic Invertebrate Samples

14
BENTHIC MACROINVERTEBRATES (10500)/Data Evaluation, Presentation, and Conclusions

Collected as Part of the National Water-Quality Assessment Pro- EPLER, J.H. 2001. Identification Manual for the Larval Chironomidae
gram, Open-File Rep. 93-407. U.S. Geological Surv., Raleigh, N.C. (Diptera) of North and South Carolina. St. Johns River Water
BARBOUR, M.T. & J. GERRITSEN. 1996. Subsampling of benthic samples: A Management District, Palatka, Fla. Special Publication SJ2001-
defense of the fixed count method. J. N. Amer. Benthol. Soc. 15:386. SP13.
BOWMAN, M.F. & R.C. BAILEY. 1997. Does taxonomic resolution affect MERRITT, R.W., K.W. CUMMINS & M.B. BERG. 2008. An Introduction to
the multivariate description of the structure of freshwater benthic the Aquatic Insects of North America, 3rd ed. Kendall/Hunt Pub-
macroinvertebrate communities? Can. J. Fish. Aquat. Sci. 54:1802. lishing Co., Dubuque, Iowa.
KATHMAN, R.D. & R.O. BRIKHURST. 1999. Guide to the Freshwater MERRITT, R.W., K.W. CUMMINS & M.B. BERG. 2009. An Introduction to
Oligochaetes of North America. Aquatic Resources Center, Thomp- the Freshwater Insects of North America. Kendall/Hunt Publishing
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DOBERSTEIN, C.P., J.R. KARR & L.L. CONQUEST. 2000. The effect of THORP, J.H. & A.P. COVICH, eds. 2010. Ecology and Classification of
fixed-count subsampling on macroinvertebrae biomonitoring in North American Freshwater Invertebrates. Academic Press, Inc.,
small streams. Freshwater Biol. 44:355. New York, N.Y.

10500 D. Data Evaluation, Presentation, and Conclusions

There are two basic approaches to evaluating pollution’s ef- 2. Quantitative Data Evaluation
fects on aquatic life. One approach is to make a qualitative
inventory of benthic fauna “above (upstream) and below (down- Statistical data-evaluation methods and mathematical descrip-
stream)” or “before and after” the suspected or known polluted tions of community structure are valuable tools in data analysis.
areas, thereby determining species’ presence or absence. Then, Analyzing biological data commonly begins with the calculation
via an understanding of various species’ responses to certain of descriptive statistics (mean, standard deviation, standard error,
pollutants and habitat degradation, determine the significance of and confidence intervals). Analysis proceeds by applying robust
damage or change. The other approach is to make a quantitative statistical methods of comparison (Chi-square, Student’s t, re-
analysis of the numbers of individuals, species, and structure gression, correlation, analyses of variance, or nonparametric
(abundance and composition) of the aquatic community affected equivalents).1,2
by pollution, and then compare that data with reference infor- Mathematical expressions (e.g., numerical indices of commu-
mation. The two approaches are integrated in most pollution nity structure) are useful in characterizing and describing aquatic
surveys because each provides valuable interpretative informa- communities. These expressions usually are based on the struc-
tion. tural and functional stability of the system.2
Diversity indices, although limited, condense considerable
biological data into single numerical values.2–9 Unfortunately,
1. Qualitative Data Evaluation useful information may be lost by condensing biological data.3,10
Select methods for analyzing multivariate benthic community
No two aquatic organisms react identically to a pollutant data using two important criteria: the methods should test spe-
cific effect-related hypotheses suggested by the data quality
because of complex relationships between genetic factors and
objectives and study design, and the methods should objectively
environmental conditions. However, certain taxa are relatively
identify relationships among variables. Use methods that make
sensitive to certain types of pollution (e.g., siltation and turbid-
a priori assumptions about relationships among variables only
ity, organic enrichment, acidity, heavy metals and other indus-
secondarily for presentations, not for primary analysis.
trial toxic wastes, oil production, agricultural products, radioac-
More powerful multivariate statistical analyses generally are
tive wastes, and thermal effects). For example, operculate snails, less subject to criticism and may be more appropriate for some
immature stages of certain mayflies, stoneflies, caddisflies, riffle bioassessment studies.1,10 Recommended data analyses ap-
beetles, hellgrammites, many marine amphipods, mysids, bi- proaches are regression of species (or taxa) richness on abun-
valve larvae, and echinoderms are sensitive to many pollutants. dance, analysis of variance (ANOVA) followed by linear orthog-
Pollution-tolerant macroinvertebrates (e.g., certain sludge onal contrasts,11 various other multivariate approaches (e.g.,
worms, midge larvae, leeches, pulmonate snails, and some cluster techniques and ordination, analyzing principal compo-
polychaetes) usually multiply under organically enriched condi- nents, ANOVA, discriminate analyses), and macroinvertebrate
tions. Facultative organisms (those that tolerate moderate pollu- community metrics2,12 for assessing biomonitoring data and
tion) include most snails, sowbugs, scuds, and blackfly larvae. water quality.
Tolerant organisms may be found in either clean or polluted When statistically evaluating data collected in pollution sur-
situations, so their presence is not definitive. However, a popu- veys, it always is beneficial to identify the sources of variability
lation of tolerant organisms combined with an absence of intol- commonly found. Variability in macroinvertebrate data comes
erant ones is a good indication that pollution is present. The same from sampling methods and organism distribution. Perhaps the
species may well react differently or be present in different major source is sampling error. Organisms generally are clus-
numbers in different geographical areas and throughout the year. tered in relation to habitat distribution; therefore, random sam-

15
BENTHIC MACROINVERTEBRATES (10500)/Data Evaluation, Presentation, and Conclusions

ples often show high variability among replicates. In statistical 10. SMITH, W., V.R. GIBSON, L.S. BROWN-LEGER & J.F. GRASSLE. 1979.
analyses of quantitative data, large numbers of samples often are Diversity as an indicator of pollution. Cautionary results from
required to detect statistically significant differences. Exercise microcosm experiments. In J.P. Grassle, G.P. Patil, W. Smith &
care when using parametric statistical methods because the basic C. Taille, eds. Ecological Diversity in Theory and Practice. Inter-
national Publ. House, Fairland, Md.
assumption of normal distribution is not always true. Data often
11. HOKE, R.A., J.P. GIESY & J.R. ADAMS. 1990. Use of linear orthog-
have to be transformed before being tested. Do not assume that onal contrast in analysis of environmental data. Environ. Toxicol.
a statistically significant difference is ecologically significant. Chem. 9:875.
Also, do not assume that lack of statistical significance indicates 12. PLAFKIN, J.L., M.T. BARBOUR, K.D. PORTER, S.K. GROSS & R.M.
lack of ecological significance.13 HUGHES. 1989. Rapid Bioassessment Protocols for Use in Streams
and Rivers: Benthic Macroinvertebrates and Fish, EPA-440/40-89-
001. Off. Water, Assessment and Watershed Protection Div., U.S.
3. Data Presentation
Environmental Protection Agency, Washington, D.C.
13. PARKHURST, D.F. 2001. Statistical significance tests: Equivalence
Data presentation may take many forms. The basic techniques and reverse tests should reduce misinterpretation. BioScience
include tables, bar graphs (horizontal and vertical), pie diagrams, 51(12):1051.
pictorial charts (ideographs), line graphs, frequency distribution 14. CHAPMAN, P.M. 1996. Presentation and interpretation of sediment
tables and graphs, histograms, frequency polygons, and cumu- quality triad data. Ecotoxicol. 5:1.
lative frequency polygons. These may be superimposed on maps.
Several reports that may be useful in analyzing macroinverte-
6. Bibliography
brate data have been included in the bibliography. Methods for
interpreting benthic invertebrate data with measures of contam-
BECK, W.M. 1955. Suggested method for reporting biotic data. Sewage
ination and toxicity are available.14 Ind. Wastes 27:1193.
INGRAM, W.M. 1960. Effective methods for collecting and recording data
4. Conclusions from water pollution surveys. In C.M. Tarzwell, compiler. Biolog-
ical Problems in Water Pollution, p. 260. U.S. Dept. Health, Edu-
cation & Welfare, Cincinnati, Ohio.
Despite detailed data quality objectives, field methodology, INGRAM, W.M. & A.F. BARTSCH. 1960. Graphic expression of biological
and laboratory analysis and data presentation, it often requires data in water pollution reports. J. Water Pollut. Control Fed.
extensive professional experience, skill, and knowledge of the 32:297.
scientific literature to draw defensible conclusions from a data PIELOU, E.C. 1966. The measurement of diversity in different types of
set. Even in the best circumstances, more than one conclusion biological collections. J. Theor. Biol. 13:131.
can sometimes be drawn from a study. When more than one CAIRNS, J., JR. 1971. A simple method for the biological assessment of
conclusion is possible, it is appropriate to present all options. the effects of waste discharges on aquatic bottom-dwelling organ-
isms. J. Water Pollut. Control Fed. 43:755.
ERMAN, D.C. & W.T. HELM. 1971. Comparison of some species impor-
5. References tance values and ordination techniques used to analyze benthic
invertebrate communities. Oikos 22:240.
1. GREEN, R.H. 1979. Sampling Design and Statistical Methods for ORLOCI, L., C.R. RAO & W.M. STITELER, eds. 1978. Multivariate Meth-
Environmental Biologists. John Wiley & Sons, New York, N.Y. ods in Ecological Work. International Cooperative Publ. House,
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roinvertebrate Field and Laboratory Methods for Evaluating the CONOVER, W.J. 1980. Practical Nonparametric Statistics, 2nd ed. John
Biological Integrity of Surface Waters, EPA-600/4-90-030. Envi- Wiley & Sons, New York, N.Y.
ronmental Monitoring Systems Lab., U.S. Environmental Protection POLLARD, J.E. 1981. Investigator differences associated with a kicking
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