Environmental Monitoring and Assessment Program Surface Waters Western Pilot Study Field Operations Manual for Wadable Streams Section 11 Benthic Macroinvertebrates


EPA/XXX/X-XX/XXX
April 2001

ENVIRONMENTAL MONITORING AND ASSESSMENT PROGRAM-

SURFACE WATERS:

WESTERN PILOT STUDY
FIELD OPERATIONS MANUAL FOR
WADEABLE STREAMS

Edited by

David V. Peck1, James M. Lazorchak2, and Donald J. Klemm2

1	U.S. Environmental Protection Agency

Regional Ecology Branch
Western Ecology Division
National Health and Environmental Effects Research Laboratory
Corvallis, OR 97333

2	U.S. Environmental Protection Agency

Ecosystems Research Branch
Ecological Exposure Research Division
National Exposure Research Laboratory
Cincinnati, OH 45268

NATIONAL HEALTH AND ENVIRONMENTAL EFFECTS RESEARCH LABORATORY
OFFICE OF RESEARCH AND DEVELOPMENT
U.S. ENVIRONMENTAL PROTECTION AGENCY
RESEARCH TRIANGLE PARK, NC 27711

NATIONAL EXPOSURE RESEARCH LABORATORY
OFFICE OF RESEARCH AND DEVELOPMENT
U.S. ENVIRONMENTAL PROTECTION AGENCY
RESEARCH TRIANGLE PARK, NC 27711

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SECTION 11
BENTHIC MACROINVERTEBRATES

Donald J. Klemm1, James M. Lazorchak1, and Philip A. Lewis1 2

Benthic invertebrates inhabit the sediment or live on the bottom substrates of streams.
Benthic macroinvertebrate assemblages in streams reflect overall biological integrity of the
benthic community. Monitoring these assemblages is useful in assessing the status of the
water body and detecting trend in ecological condition. Benthic communities respond to a
wide array of stressors in different ways so that it is often possible to determine the type of
stress that has affected a macroinvertebrate community (e.g., Klemm et al., 1990). Be-
cause many macroinvertebrates have relatively long life cycles of a year or more and are
relatively immobile, macroinvertebrate community structure is a function of present or past
conditions.

The EMAP-SW benthic macroinvertebrate protocol is intended to evaluate the biologi-
cal integrity of wadeable streams in the United States for the purpose of detecting stresses
on community structure and assessing the relative severity of these stresses. It is based on
the "Rapid Bioassessment Protocol III - Benthic Macroinvertebrates" published by the U.S.
Environmental Protection Agency (Plafkin et al., 1989; Barbour et al., 1999) and adopted for
use by many states. Modifications to the previously published protocol for EMAP-Surface
Waters (Klemm et al.. 1998) for the EMAP-SW Western Pilot Study are summarized in
Table 11-1. The two man kick net procedure of the Rapid Bioassessment Protocol (RBP) is
replaced in the EMAP-SW protocol with a D-frame kick net modified for use by one person
(Figure 11-1). Note this net is modified from that used in previous EMAP and R-EMAP
projects (Klemm et al., 1998), in terms of frame type, mesh size, and dimensions. The
modified protocol still requires only one person and is the preferred macroinvertebrate
collecting method for streams with flowing water (a second person is often used for water
safety and to keep time and record information on the field forms).

U.S. EPA, National Exposure Research Laboratory, Ecological Exposure Research Division, 26 W. Martin Luther King Dr.,
Cincinnati, OH 45268.

2 Current address: 1037 Wylie Road, RR #2, Seaman, OH 45679.

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TABLE 11-1. SUMMARY OF BENTHIC MACROINVERTEBRATE PROTOCOL CHANGES

FOR THE EMAP-SW WESTERN PILOT STUDY
Modifications from Klemm et al. (1998)

14. Two types of samples are collected, a "targeted riffle" sample and a "reach-wide" sample,
replacing the "riffle/run" and "pool/glide" samples. The targeted riffle sample is focused on
riffle areas only (i.e., if no riffle areas are present, the sample is not collected). The reach-wide
sample is collected from transects spaced throughout the reach, as was described in the
previous published protocol.

15. The number of kick samples in the targeted riffle sample is 8. The number of kick samples in
the reach-wide sample is increased from 9 (transects B through J) to 11 (Transects A through
K).

16. Each sample type is prepared as a single composite sample. For the reach-wide sample, all
kick samples are combined into a single composite sample, replacing the "RIFFLE" composite
and the "POOL" composite samples.

17. The sampling device is changed from a rectangular kick net to a D-Frame design. Mesh size
is decreased from 595 ^m to 500 //m. Net width is decreased from 18 in to 12 in (50 cm to 30
cm).

18. The area of each kick sample is reduced from 0.5 m2 to 0.09 m2 (1 ft2).

19. The time for each kick sample is increased from 20 seconds to 30 seconds.

Modifications from Western Pilot Study Year 2000 Activities:

1. Clarified procedure for collecting at sampling points choked with vegetation.

2. Field form has been modified to record the microhabitat type (pool, glide, riffle, rapid) for each
reachwide kicknet sample.

The "biomorphs" (refer to Figure 2-1) collect kick net samples for benthic macro-
invertebrates at sampling points located on each cross-section transect (termed the "reach-
wide" sample) and from riffle habitats located within the sampling reach (termed the "tar-
geted riffle" sample). Kick net samples are collected at the same time as periphyton sam-
ples (Section 8). Samples collected as part of the "reach-wide" sample are combined into a
single composite for the stream reach, while those collected for the "targeted riffle" sample
are combined into a separate composite.

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1.5 m long, 2-piece detachable handle 	>

Figure 11-1. Modified D-frame kick net. (Not drawn to scale.)

11.1 SAMPLE COLLECTION
11.1.1 Reach-Wide Sample

The index sample design for collecting the reach-wide sample for benthic
macroinvertebrates is shown in Figure 11-2. This design was used in the EMAP and
R-EMAP stream studies in the mid-Atlantic region (refer to Section 1 for project descrip-
tions).

A kick net sample is collected from each of the eleven cross-section transects
(Transects "A" through "K") at an assigned sampling point (Left, Center, or Right). These

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TRANSECT SAMPLES (1 per transect)

Sampling point of esch transect (1/4,1/2, 3/4) selected systematically after random start

Modified kick net (500 |jm mesh)

1 ft2 quadrat sampled for 30 sec

Combine all kick net samples collected from
riffles and runs and from pools

COMPOSITE REACHWIDE
SAMPLE



SIEVING

500 |jm mesh
Remove as much debris and fine
sediment as possible

COMPOSITE INDEX SAMPLE

500-mL or 1-L aliquots
Fill no more than 50% full with sample
Preseive with 95% ethanol to final
concentration of 70%

Figure 11-2. Index sampling design for benthic macroinvertebrate reachwide sample.

176

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Rev. 2, April 2001 Page 5 of 20

points may have been assigned when the sampling reach was laid out (Figure 11-2; refer
also to Section 4; Table 4-3). If not, the sampling point at Transect "A" is assigned at
random using a die or other suitable means (e.g., digital watch). Once the first sampling
point is determined, points at successive transects are assigned in order (Left, Center,
Right). These are the same sampling points as those used for periphyton samples (Section
8). At transects assigned a "Center" sampling point where the stream width is between one
and two net widths wide, pick either the "Left" or "Right" sampling point instead. If the
stream is only one net wide at a transect, place the net across the entire stream width and
consider the sampling point to be "Center". If a sampling point is located in water that is too
deep or otherwise unsafe to wade, select an alternate sampling point on the transect at
random.

The procedure for collecting a kick net sample at each transect is described in Table
11-2. At each sampling point, determine if the habitat is a "riffle/run" or a "pool/glide". Any
area where there is not sufficient current to extend the net is operationally defined as a
pool/glide habitat. Record the dominant substrate type (fine/sand, gravel, coarse substrate
(coarse gravel or larger) or other (e.g., bedrock, hardpan, wood, aquatic vegetation, etc.)
and the habitat type (pool, glide, riffle, or rapid) for each kick net sample collected on the
Sample Collection Form as shown in Figure 11-3. As you proceed upstream from transect
to transect, combine all kick net samples into a bucket or similar container labeled "REACH-
WIDE", regardless of whether they were collected using the "riffle/run" or "pool/glide" proce-
dure.

If it is impossible to sample at the sampling point with the modified kick net following
either procedure, spend about 30 seconds hand picking a sample from about 0.09 m2 (1 ft2)
of substrate at the sampling point. For vegetation-choked sampling points, sweep the net
through the vegetation for 30 seconds. Place the contents of this hand-picked sample into
the "REACH-WIDE' sampling container.

11.1.2 Targeted Riffle Sample

Figure 11-4 illustrates the sampling design for the targeted riffle sample. Table 11-3
presents the procedure for selecting individual sampling points within the available riffle
macrohabitat units located within the sampling reach. Note that if the total available area of
riffle habitat is less than 8 ft2 (i.e., such that 8 non-overlapping kick net samples cannot be
collected), do not collect a targeted riffle sample. There may be stream reaches where
more than one 1 ft2 kick net sample is collected from a single riffle unit. The objective for

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Rev. 2, April 2001 Page 6 of 20

TABLE 11-2. PROCEDURE TO COLLECT KICK NET SAMPLES FOR THE REACH-WIDE

1. At each cross-section transect, beginning with Transect "A", locate the assigned sampling point
(Left, Center, or Right as you face downstream) as 25%, 50%, and 75% of the wetted width,
respectively. If you cannot collect a sample at the designated point because of deep water or
unsafe conditions, relocate the point on the transect nearby.

2. Attach the 4-ft handle to the kick net. Make sure that the handle is on tight or the net may
become twisted in a strong current, causing the loss of part of the sample.

3. Determine if there is sufficient current in the area at the sampling point to fully extend the net. If
so, classify the habitat as "riffle/run" and proceed to Step 4. If not, use the sampling procedure
described for "pool/glide" habitats (Step 9).

NOTE: If the net cannot be used, spend 30 seconds hand picking a sample from about
0.09 m2 (1 ft2) of substrate at the sampling point. For vegetation-choked sampling
points, sweep the net through the vegetation within a 0.09 m (1 ft2) quadrat for 30 sec-
onds. Place the contents of this hand-picked sample into the "REACH-WIDE" sampling
container. Go to Step 15.

Riffle/Run Habitats:

4. With the net opening facing upstream, position the net quickly and securely on the stream
bottom to eliminate gaps under the frame. Avoid large rocks that prevent the sampler from
seating properly on the stream bottom.

NOTE: If there is too little water to collect the sample with the kick net, randomly pick up
10 rocks from the riffle and pick and wash the organisms off them into a bucket labeled
"REACH-WIDE" which is half-full of water.

5. Holding the net in position on the substrate, visually define a rectangular quadrat that is one
net width wide and one net width long upstream of the net opening. The area within this
quadrat is -0.09 m2 (1 ft2). Alternatively, place a wire frame of the correct dimensions in front
of the net to help delineate the quadrat to be sampled.

6. Hold the net in place with your knees. Check the quadrat for heavy organisms, such as mus-
sels and snails. Remove these organisms from the substrate by hand and place them into the
net. Pick up any loose rocks or other larger substrate particles in the quadrat. Use your hands
or a small scrub brush to dislodge organisms so that they are washed into the net. Scrub all
rocks that are golf ball-sized or larger and which are over halfway into the quadrat. Large rocks
that are less than halfway into the sampling area are pushed aside. After scrubbing, place the
substrate particles outside of the quadrat.

7. Keep holding the sampler securely in position. Start at the upstream end of the quadrat,
vigorously kick the remaining finer substrate within the quadrat for 30 seconds (use a stop-
watch).

8. Pull the net up out of the water. Immerse the net in the stream several times to remove fine
sediments and to concentrate organisms at the end of the net. Avoid having any water or
material enter the mouth of the net during this operation.

9. Go to Step 14.

(continued)

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Rev. 2, April 2001 Page 7 of 20

Pool/Glide habitats:

10. Visually define a rectangular quadrat that is one net width wide and one net width long at the
sampling point. The area within this quadrat is -0.09 m2 (1 ft2). Alternatively, lay a wire frame
of the correct dimensions in front of the net at the sampling point to help delineate the quadrat.

11. Inspect the stream bottom within the quadrat for any heavy organisms, such as mussels and
snails. Remove these organisms by hand and place them into the net or into a bucket labeled
"REACH-WIDE". Pick up any loose rocks or other larger substrate particles within the quadrat
and hold them in front of the net. Use your hands (or a scrub brush) to rub any clinging organ-
isms off of rocks or other pieces of larger substrate (especially those covered with algae or
other debris) into the net. After scrubbing, place the larger substrate particles outside of the
quadrat.

12. Vigorously kick the remaining finer substrate within the quadrat with your feet while dragging
the net repeatedly through the disturbed area just above the bottom. Keep moving the net all
the time so that the organisms trapped in the net will not escape. Continue kicking the sub-
strate and moving the net for 30 seconds. NOTE: If there is too little water to use the kick net,
stir up the substrate with your gloved hands and use a sieve with 500 ^m mesh size to collect
the organisms from the water in the same way the net is used in larger pools.

13. After 30 seconds, remove the net from the water with a quick upstream motion to wash the
organisms to the bottom of the net.

All samples:

14. Invert the net into a plastic bucket marked "REACH-WIDE" and transfer the sample. Inspect
the net for any residual organisms clinging to the net and deposit them into the "REACH-
WIDE" bucket. Use watchmakers' forceps if necessary to remove organisms from the net.
Carefully inspect any large objects (such as rocks, sticks, and leaves) in the bucket and wash
any organisms found off of the objects and into the bucket before discarding the object. Re-
move as much detritus as possible without losing any organisms.

15. Place an "X" in the appropriate substrate type box for the transect on the Sample Collection
Form.

Fine/sand: not gritty (silt/clay/muck < 0.06 mm diam.) to gritty, up to ladybug sized (2 mm
diam.)

Gravel: fine to coarse gravel (ladybug to tennis ball sized; 2 mm to 64 mm diam.)

Coarse: Cobble to boulder (tennis ball to car sized; 64 mm to 4000 mm)

Other: bedrock (larger than car sized; > 4000 mm), hardpan (firm, consolidated fine
substrate), wood of any size, aquatic vegetation, etc.). Note type of "other" substrate in
comments on field form.

16. Thoroughly rinse the net before proceeding to the next sampling location. Proceed upstream
to the next transect (including Transect K, the upstream end of the sampling reach) and repeat
Steps 1 through 9. Combine all kick net samples from riffle/run and pool/glide habitats into the
"REACH-WIDE" bucket.

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	Rev. 2, April 2001 Page 8 of 20	

SAMPLE COLLECTION FORM - STREAMS

1 by (Initial):

SITE ID:

..W.XX£l±r.	fflf	

DATE:

.7./.Q././.2 ,0,0,1

WATER CHEMISTRY

Sample ID



Transect

.JL

Comments

REACH-WIDE BENTHOS SAMPLE

Sample ID

No,of Jars

Comment

r« rtAJ*t(cr	&TH6H, =. Sim Al l kJmlry

TRANSECT

B

1

5UBSTRATF. CHAN

POC)!
Gliue
RtlfR-
Nauitl

O F
G5 ^
0<=

no

~	p
O'S

~	•»

X

~ =
~ «=
Q »

~	B|

~	«

E'
D°

~ =
~ o

D°

n bi

~'»

8 &
~ c
O o

~ »
Ho

n

O RA

KSa
Do
~ o

~	p

~	«
P "i

O RA

Of
Do
S3 c
Do

~ »
D«

S3 «
~ f«

®G

~ <:
~ »

Og

58
~ »*

TARGEt'ED RIFFLE BENTHOS SAMPLE

Q»

~	c

~	o

~ «"

n bi
~»«

X'

~	o

~	c

DO

82»
Do

~	m

~	«*

D o
D<=

Do

Do

~	W

~	><*

~ f

Qe
Dc

po

D «
D »

~ I*

Sample ID

JL3JL-£j2JL

Mo. of Jars

,	,	L

Comment

NEAREST
TRANSECT

•35 Fine/Sand

J Grave!

3

"? Coarse
£

c other: Note in
Comments

OF/S

ait;

DC

DO

~ F/s

BG

DC
OO

6

O F/S

~	G
BC

~	O

DBS
BG

~ C

DO

~ F/S
OG
BC
DO

D F/S
OG
BC
DO

~	F/S
B G

~	C
DO

~	F/S
KG

~	C
DO

SUBSTRATE SIZE CLASSES
F/S - iadybug or smaller (<-,2 mm)

G - Iadybug to tennis ball (2 to 64
mm)

C - tennis ball to car sized (64 to
4000 mm)

O - bedrock, hardpan, wood, etc

Additional Benthos Comments





COMPOSITE PERIPHYTON SAMPLE

Sample SD

> g.O.O, f, ?,£>,

Composite Volume (ml)

,	,IA&

Assemblage ID
(50-mL lube, preserved)

Number of transects sampled (0-11):

/ ,/

Chlorophyll
(GF/F filter)

Biomass

{GF/F Filter)

Sample Voi (mU

_JHCL

Flag

Sampro Vo!. (ml)

IX

Flag

Sample Vol 'ml.!

X£

Flag

Flag codes: K - Sample not collected: U - Suspect sample; Ft. F2. etc. ~ misc. flag assigned by field crew. Explain ail Hags in comment sections.

31443

M 03/26/2001 2001 Sample Collection	^

Figure 11-3. Sample Collection Form (page 1), showing information for the reach-wide and
targeted riffle benthic macroinvertebrate samples.

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	Rev. 2, April 2001 Page 9 of 20	

COMPOSITE INDEX SAMPLE

500-mL or 1-L aliquots

Fill no more than 50% full of sample

Preserve with 95% ethanol to final
concentration of 70%

Figure 11-4. Index sampling design for benthic macroinvertebrate targeted riffle sample.

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Rev. 2, April 2001 Page 10 of 20

TABLE 11-3. LOCATING SAMPLING POINTS FOR KICK NET SAMPLES:
TARGETED RIFFLE SAMPLE

1. Before sampling, survey the stream reach to estimate visually the total number (and area) of
riffle macrohabitat "units" contained in the defined stream reach. To be considered as a unit,
the area of the riffle must be greater than 1 ft2.

A. Do not sample poorly represented habitats. If the reach contains less than 8 ft2 of riffle
macrohabitat, then do not collect a targeted riffle sample.

B. If the reach contains more than one distinct riffle macrohabitat units but less than eight,
allocate the eight sampling points among the units so as to spread the effort throughout
the reach as much as possible. You may need to collect more than one kick sample from
a given riffle unit.

C. If the number of riffle macrohabitat units is greater than eight, skip one or more habitat
units at random as you work upstream, again attempting to spread the sampling points
throughout the reach.

2. Begin sampling at the most downstream riffle unit, and sample units as they are encountered
to minimize instream disturbance.

3. At each unit, exclude "margin" habitats by constraining the potential sampling area. Margin
habitats are edges, along the channel margins or upstream or downstream edges of the riffle
macrohabitat unit. Define a core area for each riffle unit as the central portion, visually estimat-
ing a "buffer" strip circumscribing the identified unit. In some cases, the macrohabitat unit may
be so small that it will not be feasible to define a core area and avoid an edge.

4. Visually lay out the core area of the unit sampled into 9 equal quadrats (i.e., a 3 * 3 grid). For
each macrohabitat type, select a quadrat for sampling at random from the following list of
locations (right and left are determined as you look downstream):

Lower right quadrat
Lower center quadrat
Lower left quadrat
Right center quadrat
Center quadrat
Left center quadrat
Upper right quadrat.

Upper center quadrat.

Upper left quadrat

5. Collect the kick sample in the center of the selected quadrat as described in Table 11-4.

6. If a second sample is required from a single macrohabitat unit, select additional quadrats at
random from the list in Step 4.

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TABLE 11-4. COLLECTING A KICK NET SAMPLE FROM WADEABLE STREAMS FOR THE

TARGETED RIFFLE COMPOSITE SAMPLE

1. Beginning at the most downstream riffle unit within the sampling reach, locate the sampling
point within the macrohabitat unit as described in Table 11-3.

2. Position the kick net quickly and securely on the stream bottom so as to eliminate gaps be-
tween the frame and the stream bottom. If necessary, rotate the net so the narrower side is
against the bottom.

3. Hold the sampler firmly in position on the substrate. Define a quadrat immediately upstream
from the mouth of the net having a width equal to the width of the net frame (total area = 0.09
m2).

4 Hold the net in place with your knees and pick up any loose rocks or other larger substrate
particles in the quadrat. Use your hands or a small scrub brush to dislodge organisms so that
they are washed into the net. Scrub all rocks that are golf ball-sized or larger and which are
over halfway into the quadrat. Large rocks that are less than halfway into the sampling area
are pushed aside. After scrubbing, place the substrate particles outside of the quadrat.

S. Keep holding the sampler securely in position. Start at the upstream end of the quadrat,

vigorously kick the remaining finer substrate within quadrat for 30 seconds (use a stopwatch).

7. Pull the net up out of the water. Immerse the net in the stream several times to remove fine
sediments and to concentrate organisms at the end of the net. Avoid having any water or
material enter the mouth of the net during this operation.

8. Invert the net into a plastic bucket marked "TARGETED RIFFLE" and transfer the sample.
Inspect the net for any residual organisms clinging to the net and deposit them into the
"TARGETED RIFFLE" bucket. Use watchmakers' forceps if necessary to remove organisms
from the net.

9. Record the nearest transect location in the box for the sample on the Sample Collection Form.
Also note the dominant substrate type by checking the appropriate box on the Sample Collec-
tion Form.

Fine/sand: not gritty (silt/clay/muck < 0.06 mm diam.) to gritty, up to ladybug sized (2 mm
diam.)

Gravel: fine to coarse gravel (ladybug to tennis ball sized; 92 mm to 64 mm diam.)
Coarse: Cobble to boulder (tennis ball to car sized; 64 mm to 4000 mm)

Other: bedrock (larger than car sized; > 4000 mm), hardpan (firm, consolidated fine
substrate), wood of any size, aquatic vegetation, etc.). Note type of "other" substrate in
comments on field form.

(continued)

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	Rev. 2, April 2001 Page 12 of 20	

TABLE 11-4. (Continued)

10.	Thoroughly rinse the net before proceeding to the next sampling location (either the next riffle
unit or a different quadrat location within the same riffle unit).

11.	Repeat steps 1-10 at subsequent riffle sampling points until 8 kick samples have been col-
lected and placed into the "TARGETED RIFFLE" bucket.	

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Rev. 2, April 2001 Page 13 of 20

selecting sampling points within the available riffle macrohabitat units is to allocate points
throughout the sampling reach as much as possible.

Procedures for collecting a point sample using the kick net from riffle macrohabitat
units are presented in Table 11-4. At each sampling point, a quadrat having a total area of
0.09 m2 (1 ft2) is sampled. Because the reach-wide and targeted riffle samples are
collected in the order they are encountered during a single pass through the reach, it is very
important to rinse the kick net thoroughly between samples to avoid carryover and possible
cross-contamination of the targeted riffle sample and the reach-wide sample.

11.2 SAMPLE PROCESSING

After collecting kick net samples for both the reach-wide and targeted riffle samples,
prepare two composite index samples from the contents of the "REACH-WIDE" and
"TARGETED RIFFLE" buckets as described in Table 11-5. Record tracking information for
each composite sample on the Sample Collection Form as shown in Figure 11-3. A set of
completed sample labels, including the label that is used if more than one jar is required for
a single composite sample, is shown in Figure 11-5. Note that each composite sample has
a different sample number (barcode). The ID number is also recorded on a waterproof label
that is placed inside the jar (Figure 11-5, lower right). If more than one jar is used for a
composite sample, a special label (Figure 11-5, lower left) is used to record the ID number
assigned to the sample. DO NOT use two different barcode numbers on two jars containing
one single sample. Blank labels for use inside of sample jars are presented in Figure 11-6.
These can be copied onto waterproof paper.

Check to be sure that the prenumbered adhesive barcoded label is on the jar and
covered with clear tape, and that the waterproof label is in the jar and filled in properly. Be
sure the inside label and outside label describe the same sample. Replace the cap on each
jar and seal them with plastic electrical tape. Check to make sure the cap is properly
marked with site number, habitat type (reach-wide or targeted riffle). Place the samples in a
cooler or other secure container for transporting and/or shipping the laboratory (see Section
3). The container and absorbent material should both be suitable for transporting ethanol.
Check to see that all equipment is in the vehicle.

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Rev. 2, April 2001 Page 14 of 20

TABLE 11-5. PROCEDURE FOR PREPARING COMPOSITE SAMPLES FOR

^^^^^^^^^^^^^3|NTHICMACROINV|RT|BRATgJ^^^^^^^^^^^^=

1. Pour the entire contents of the "REACH-WIDE" bucket through a sieve with 500 jjm mesh
size). Remove any large objects and wash off any clinging organisms back into the sieve
before discarding.

2. Using a wash bottle filled with stream water, rinse all the organisms from the bucket into the
sieve. This is the composite reach-wide sample for the site.

3. Estimate the total volume of the sample in the sieve and determine how large a jar will be
needed for the sample (500-mL or 1-L). Avoid using more than one jar for each of the
composite samples.

4. Fill in a "REACH-WIDE" (or "TARGETED RIFFLE") sample label with the stream ID and date of
collection. Attach the completed label to the jar and cover it with a strip of clear tape.

5. Wash the contents of the sieve to one side by gently agitating the sieve in the water. Wash the
sample into a jar using as little water from the wash bottle as possible. Use a large-bore funnel
if necessary. If the jar is too full pour off some water through the sieve until the jar is not more
than % full, or use a second jar if a larger one is not available. Carefully examine the sieve for
any remaining organisms and use watchmakers' forceps to place them into the sample jar.

If a second jar is needed, fill in a sample label that does not have a pre-printed ID num-
ber on it. Record the ID number from the pre-printed label prepared in Step 4 in the
"SAMPLE ID" field of the label. Attach the label to the second jar and cover it with a strip
of clear tape.

6. Place a waterproof label with the following information inside each jar:

Stream Number

Type of sampler and mesh size used
Habitat type (riffle or pool)

Name of stream

Date of collection
Collectors initials
Number of transect samples
composited

7. Completely fill the jar with 95% ethanol (no headspace) so that the final concentration of
ethanol is between 75 and 90%. It is very important that sufficient ethanol be used, or the
organisms will not be properly preserved.

NOTE: Prepared composite samples can be transported back to the vehicle before
adding ethanol if necessary.

8. Replace the cap on each jar. Slowly tip the jar to a horizontal position, then gently rotate the
jar to mix the preservative. Do not invert or shake the jar. After mixing, seal each jar with
plastic tape.

9. Repeat Steps 1 through 8 for the "TARGETED RIFFLE" bucket.

10. Store labeled composite samples in a container with absorbent material that is suitable for use
with 95% ethanol until transport or shipment to the laboratory.

186

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EMAP-Western Pilot Study Field Operations Manual forWadeable Streams, Section 11 (Benthic Macroinvertebrates),
Rev. 2, April 2001 Page 15 of 20

REACH-WIDE BENTHOS

uuyypqo . c? 9
ww/v/vr L. ¦ *¦ ¦

7 III 2001
500000

TARGETED RIFFLE BENTHOS
WXXP99-J? _f. JUL
Ol I O! I 2001
800000

BENTHOS

(Jteach Wide) Targeted Riffle
WXXP99 -
-------
EMAP-Western Pilot Study Field Operations Manual forWadeable Streams, Section 11 (Benthic Macroinvertebrates),
Rev. 2, April 2001 Page 16 of 20

BENTHOS IDENTIFICATION

Site Number

Stream

Collection Date

Sampler

Habitat Type

Collector(s)

Number of Transects

BENTHOS IDENTIFICATION

Site Number

Stream

Collection Date

Sampler

Habitat Type

Collector(s)

Number of Transects

BENTHOS IDENTIFICATION

Site Number

Stream

Collection Date

Sampler

Habitat Type

Collector(s)

Number of Transects

BENTHOS IDENTIFICATION

Site Number

Stream

Collection Date

Sampler

Habitat Type

Collector(s)

Number of Transects

BENTHOS IDENTIFICATION

Site Number

Stream

Collection Date

Sampler

Habitat Type

Collector(s)

Number of Transects

BENTHOS IDENTIFICATION

Site Number

Stream

Collection Date

Sampler

Habitat Type

Collector(s)

Number of Transects

Figure 11-6. Blank labels for benthic invertebrate samples.

188

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EMAP-Western Pilot Study Field Operations Manual forWadeable Streams, Section 11 (Benthic Macroinvertebrates),
Rev. 2, April 2001 Page 17 of 20

EQUIPMENT AND SUPPLIES FOR BENTHIC MACROINVERTEBRATES

OTY

ITFM

Modified kick net ( D-frame with 500 jjm mesh) and 4-ft handle (Wildco #425-C50)

Spare net(s) and/or spare bucket assembly for end of net

Watch with timer or a stopwatch

Buckets, plastic, 8- to 10-qt capacity, labeled "REACH-WIDE" and "TARGETED
RIFFLE"

Sieve with 500 |jm mesh openings

Sieve-bottomed bucket, 500 jjm mesh openings

2 pr.

Watchmakers' forceps

Wash bottle, 1-L capacity labeled "STREAM WATER"

Small spatula, spoon, or scoop to transfer sample

Funnel, with large bore spout

4 to 6
each

Sample jars, HDPE plastic with screw caps. 500-mL and 1-L capacity, suitable for
use with ethanol

2 gal

95% ethanol, in a proper container

2 pr.

Rubber gloves, heavy rubber

Cooler (with suitable absorbent material) for transporting ethanol and samples

Composite Benthic sample labels, with preprinted ID numbers (barcodes)

Composite Benthic sample labels without preprinted ID numbers

Blank labels on waterproof paper for inside of jars

Sample Collection Form for site

Soft (#2) lead pencils

Fine-tip indelible markers

1 pkg.

Clear tape strips

4 rolls

Plastic electrical tape

Knife, pocket, with at least two blades

Scissors

Pocket-sized field notebook (optional)

1 Pkg.

Kim wipes in small self-sealing plastic bag

1 copy

Field operations and methods manual

1 set

Laminated sheets of procedure tables and/or quick reference guides for benthic
macroinvertebrates

Figure 11 -7. Equipment and supply checklist for benthic macroinvertebrates.

189

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EMAP-Western Pilot Study Field Operations Manual forWadeable Streams, Section 11 (Benthic Macroinvertebrates),
Rev. 2, April 2001 Page 18 of 20

Cuffney, T.F, M.E. Gurtz, and M.R. Meador. 1993. Methods for Collecting Benthic Inverte-
brate Samples as Part of the National Water-Quality Assessment Program. U.S.
Geological Survey Open-File Report 93-406, Raleigh, North Carolina.

Klemm, D.J., P.A. Lewis, F. Fulk, and J.M. Lazorchak. 1990. Macroinvertebrate Field and
Laboratory Methods for Evaluating the Biological Integrity of Surface Waters.
EPA/600/4-90/030. U.S. Environmental Protection Agency, Environmental Monitoring
Systems Laboratory, Cincinnati, Ohio.

Klemm, D.J., J.M. Lazorchak, and P.A. Lewis. 1998. Benthic Macroinvertebrates. pp. 147-
182 IN.: J.M. Lazorchak, D.J. Klemm, and D.V. Peck (Eds.). Environmental Monitoring
and Assessment Program-Surface Waters: Field Operations and Methods for Measur-
ing the Ecological Condition of Wadeable Streams. EPA/620/R-94/004F. U.S. Envi-
ronmental Protection Agency, Washington, D.C.

Plafkin, J.L., M.T. Barbour, K.D. Porter, S.K. Gross, and R.M. Hughes. 1989. Rapid

Bioassessment Protocols for Use in Streams and Rivers: Benthic Macroinvertebrates
and Fish. EPA/440/4-89/001. U.S. Environmental Protection Agency, Assessment
and Watershed Protection Division, Washington, D.C.

190

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NOTES

191

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NOTES

192

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