Open Literature Review Summary

Chemical Name: Atrazine

CAS No: 1912-24-9

ECOTOX Record Number and Citation:

Hamilton, P.B., G.S. Jackson, N.K. Kaushik, K.R. Solomon, and G.L. Stephenson. 1988.

The impact oftwo applications of atrazine on the plankton communities of in situ

enclosures. Aquat. Toxico!. 13: 123-140 (MRID 47543617).

Purpose of Review:

Determine potential effects of atrazine on an aquatic community. Endpoints from this

study contribute to the weight-of-evidence regarding effects of atrazine on aquatic plant

communities and associated determination of a Level ofConcern (LaC).

Date of Review:

November 24, 2009

Purpose of Study:

This study examines the potential effects oftwo atrazine applications to aquatic plants

and animals (including species interactions and community stability) in experimental lake

enclosures (limnocorrals).

Summary of Study Findings:

Methods: The basic experimental design ofthis study is also reported in Hamilton, et al.

(1987) (1983 study), Hamilton, et al. (1988), and Herman, et al. (1986).

Lake enclosures were constructed in Lake St. George, Ontario, Canada, a 10-ha

mesotrophic dimictic lake surrounding by agricultural land. (Background levels of

atrazine in the lake are not described.) The experiment took place during the ice-free

time between May 1983 and April 1984. The six nylon-reinforced polyvinyl chloride

(PVC) enclosures (5 m x 5 m x 4.5 m deep) had been previously installed (two sources

are referenced for descriptions; it is not mentioned whether the enclosures reach the lake

bottom). Atrazine was applied (product grade, manufacturer, and number oftreatment

enclosures used vs. controls not mentioned, though the latter can be inferred as 3 controls

and 3 treatments from some ofthe article's tables) in two pulses of 0.1 mg/L (100 !!g/L),

on June 1, 1983 and 35 days later on July 6, 1983, by spraying the water surface with a

III1II1III

2087860
hand-pumped backpack applicator (two sources are cited for details, one examining the

mixing process in such applications in enclosures). The study ended 323 days after the

first treatment. To maintain nutrient levels, HP03 and NaN03 were added weekly

(Hamilton, et ai., 1987, is cited for details). One smallmouth bass was added to each

enclosure prior to the first atrazine application to control the planktivorous fish

population (this addition is not mentioned in the 1986 or 1989 papers by Hamilton, et

ai.). The study was terminated 323 days after the first atrazine application.

Baseline samples were collected weekly beginning 3 weeks before the first atrazine

treatment. Hamilton, et ai. (1987) is cited for water quality and atrazine sampling and

analysis method descriptions. Phytoplankton were collected using a 25 mm (outside

diameter) x 3.5 m (long) integrated plankton sampler, with 8 samples collected from each

enclosure and combined and thoroughly mixed before subsampling. I l5 mL subsamples

were taken and preserved with 1 mL of Lugol 's solution, then counted in the laboratory

using Utermohl's inverted microscope technique. A minimum of 300 cells were counted

from each subsample, and cell dimensions were measured for 1-9 individuals of each

species. The cell counts and mean dimensions by species were used to estimate

population numbers and total fresh weight biomass of each species (a 1976 MSc thesis

from the University of Waterloo is cited for details). Zooplankton samples were

collected (for description ofthe procedure, a 1986 paper with two ofthe same coauthors

as the present paper is cited). There were 21 sampling dates in total.

Statistical analyses ofthe data for each sampling day were performed using Student's t

test, with significance detected for p-values < 0.05. When biological data were not

meeting the assumptions of normality and homogeneity of variance well, they were

transformed by 10glO(X+1) (tests for normality and homogeneity of variance are not

described). Differences in the community composition between enclosures were

compared using the Morista coefficient of association (MCA) (a source is cited, but the

MCA is not defined in the article).

Results: The results presented in the article are not redundant with any results described

in the related articles, Hamilton, et al. (1987) (later study), Hamilton, et al. (1989), and

Herman, et ai. (1986).

Atrazine concentration measurement data are not provided.

The phytoplankton community changed seasonally. In the spring (May), diatoms

(Cyclotella stelligera, Fragiiaria crotonensis, Synedra spp.) dominated, followed by a

summer (mid-June through September) shift to green algae (Tetraedron minimum,

Oocystis iacustris, Eiakatothrix geiatinosa), and two additional taxa (Lagerheimia

quadriseta, Sphaerocystis schroeteri) became prominent in the fall. Table 1 lists the

dominant taxa identified in the experiment.

2
Figure 1 shows the total number of algal taxa found in the samples between Day -20 and

Day 114. The control enclosures always had more algal taxa than the atrazine treatment

enclosures, but the differences were only significant (p<0.05) on Day 27,98, and 114. In

terms oftotal cells numbers, the treatments were often lower than controls, but the

differences were never significant (Figure 2). Fresh weight biomass, however, dropped

47-82% in the treatment enclosures after the second atrazine pulse, with significant

(p<O.05) differences on day 77 (Figure 3).

The green algae were reduced after atrazine treatment (see Figure 2, Figure 3). Total

green algae numbers were significantly (p<0.05) reduced 12 and 26 days after the second

atrazine application. Fresh weight biomass was significantly (p<0.05) reduced on Day

27,49,63,77, and 114, with Chlorophyta biomass at levels >93% reduced compared

with controls between Day 49-77, but recovery was noted the following spring (Day

323). Species-specific significant (p<0.05) declines were noted in: E. gelatinosa (Days

13,27,63,77,98,114), S. schroeteri (Days 27,77,114), T minimum (Days 27, 49, 63,

114), and 0. lacustris (Day 27). The typically summer growing species L. quadriseta

was delayed in its growth in the treatment enclosures, establishing prominent populations

about 47 days later than in controls. No differences between control and atrazine

exposed populations of Scenedesmus acutiformis were noted. See Figure 4.

Dinoflagellate (Pyrrhophyta) numbers were reduced after atrazine exposure, with

significant (p<0.05) differences on Day 1,49,63, and 77 (Figure 2). Gymnodium spp.

were especially affected, with significant reductions in numbers on Day 13,27,49, and

77 (Figure 5). Dinoflagellate biomass was significantly (p<0.05) lower than controls on

Day 49, 77, 98, and 114 (Figure 3). Generally, the dinoflagellates had the lowest

numbers and biomass of any phytoplankton group, and there was high variability in their

data.

The Chrysophyta community, excluding diatoms, consisted mostly ofsmall

unidentifiable monads. Although their biomass and numbers tended to be lower in the

treatment enclosures, no significant differences were found (Figure 2 and Figure 3).

Diatom (BaciIlariophyceae) cell densities were lower in the treatment enclosures from

Day 27-114, but again, no differences were significant (Figure 2). Reductions in diatom

biomass, noted only after the second atrazine treatment, were significant from Day 49-77,

but recovered by Day 114. C. stelligera, F. crotonensis, and Synedra spp. were dominant

Bacillariophyceae species; see Figure 4.

Cryptophyta, in contrast, briefly increased in the atrazine enclosures: numbers increased

on Day 13 (Figure 2), and biomass on Day 13 and 22, although no differences were

observed after that time. Rhodomonas minuta appeared responsible for the increase

(Figure 5), with some contribution from Cyptomonas erosa (including C. erosa var.

3
reflexa). The next spring, Day 323, the Cryptophyta levels were lower in the treatment

enclosures.

In zooplankton, the seasonal pattern ofrotifer growth was similar in control and treatment

enclosures (Figure 6). Kellicottia longispina was dominant in the spring, giving way to

Conochilus and Polyartha in the summer, which both remained persistent for the rest of

the study, though in the fall Cohochilus spp. showed some decline and Kellicottia

bostoniensis became prominent. Mean number ofRotifera taxa was generally somewhat

lower in the treatment enclosures, but the difference was only significant (p<0.05) on

Day 63 (Figure 1). Ofthe microzooplankton, no differences were found in the speciesspecific numbers ofKeratella cochlearis, K. longspina, K. bostoniensis, or Chromogaster

spp., Polyartha spp. had higher numbers in the treatment enclosures on Day 62, and

Conochilus spp. increased until Day 98 in the controls but only until Day 62 in the

treatments, then declined significantly (p<0.05) in the treatments. See Figure 6.

The crustaceans, like the rotifers, had similar seasonal patterns in the control and atrazine

treated enclosures. Bosmina longirostris and Diacyclops bicuspidatus were dominant in

the spring, while Ceriodaphnia lacustris and Daphnia galeata mendotae became

dominant filter feeders in late June through the fall, and Diaptomus oregonensis was the

dominant cyclopoid in June through late fall. Little effect of atrazine treatment on the

number of macrozooplankton (Crustaceae) taxa, although the reduction from treatment

on Day 77 was significant (p<0.05) (Figure 1). Cladocera populations were consistently

46.6% lower in the atrazine exposed enclosures after Day 13, but no differences were

significant. The reduced cladoceran numbers were due to significant (p<0.05) declines in

B. longirostris (Figure 6), which began showing reductions after Day 13 and had

significant reductions on Day 48, 62, and 76. Atrazine did not have a noticeable effect on

the total number of copepods, but D. oregonensis was consistently less abundant in

treatment enclosures after Day 48. Immature crustacea (copepodites and nauplii) were

not initially affected by atrazine, but at Day 114 there were significantly fewer immature

zooplankters in the treatment enclosures, and a similar difference was found the

following spring (Day 323).

Morista coefficients of association (MCA) are presented in Table 2 for the phytoplankton

communities. The control corrals did not have very similar communities throughout the

experiment, although they became somewhat more similar from Day 77-114. Controls

C2 and C3 had the most dissimilarity, while C 1 was initially similar to C2 (until Day 27)

and then became more similar to C3 (Day 27-63). The atrazine treated enclosures had

more similar community assemblages to each other, with A2 and A3 consistently similar

until Day 114, and Al showing some divergencc after Day 63. Differences between

controls and treatments manifested after treatment, with large differences after the second

atrazine treatment especially (MCA from 0.21-0.36 after Day 27), but differences after

the first treatment and before the second treatment were less dramatic than within-control

group differences.

4
MCA for the microzooplankton (Rotifera) are presented in Table 3. These communities

remained similar among the control enclosures through Day 49, with enclosure Cl

diverging after that. The atrazine-treated enclosures remained similar throughout the

experiment. There were no clear differences between the atrazine-treated and control

communities, since there was no consistent trend, and lower mean MCAs occurred only

on Day 63, 98, and 125 (of 0.64, 0.52, and 0.57, respectively) with at least one enclosure

being highly dissimilar on each ofthese days.

Macrozooplankton communities were variable in how similar they were (Table 4). The

controls were similar only until Day I, while the atrazine-treated enclosures were similar

until Day 77, after which A1 diverged from the other treatments. The control and

atrazine-treated enclosures differed from one another consistently, though not

dramatically, after Day 63 .

1 r" r •I 1 I ~r-r--"r--Ti--"T--

20 40 60 80 100 120

DAV$ AFTER TREAT..... ENT

Figure 1. Mean number of phytoplankton, rotifer, and crustacean species enumerated in sample

counts. Vertical bars represent one standard deviation for each sample date. Solid line=Controls,

Dashed line=Atrazine treated enclosures. Single arrow represents the first atrazine application,

double arrow the second atrazine application. (study figure 1)

5
TorAL

80

60

4.0

20

'~ .1 OlRYSC'?HYTA

.~-, n: ~. "B !"f r~ • t 1 t

f:

1- ,

.~ t. 4t t

l/I 80 Q;:

GO·

:I 40

20-

§

ao -\

60":

4.o~

2°1

80

6.0

4,0·

20 -

:JActI...L'AP.iO"HYCEAE. II

-20 -6 13 27 "'9 63 77 98 1I4323

----1.1 I J I j-+--A-"'+---S-'~ADAyS AFTER TREATMEN T H9B31

Figure 2. Mean numbers of total phytoplankton, Chlorophyta (3-8 taxa), Chrysophyta (2-10 taxa),

Bacillariophyceae (0-11 taxa), Cryptophyta (2-8 taxa), and Pyrrophyta (0-3 taxa). Solid bars=lake

stations (not described in the article text), Blank bars=controls, Slashed bars=atrazine enclosures.

Numbers are log transformed with error bars representing 2 standard deviations. Single arrow

represents the first atrazine application, double arrow the second atrazine application. (study figure

2)

6
30 .

V,

l/',;

<t

:; ~,0-":

c)

m 20-~

30 '

20

ID

CR'rPTOPHYTA

PYRRQPffYTA

-20 -6 Il 27 491 lH 77

--t.'--"_.•-t---J---------l---J__1\----1-

()n'l'S lonER 1 Rl:Ar"'tNI (''183J

Figure 3. Mean fresh weight biomass for total phytoplankton, Chlorophyta, Chrysophyta,

Bacillariophyceae, Cryptophyta, and Pyrrophyta are presented for outside (solid), control (blank),

and atrazine (slash) enclosures. Biomass data are log transformed with error bars representing 2

standard deviations. Single arrow represents the first atrazine application, double arrow the second

atrazine application. (study figure 3)

7
CONTROL ATRAIINI::

f~M~~10~H~~~ GtLAlr~I~~

__ ,"" ~~:~~~;~~ ~T r.fg':::~'OFi.· l: R 1

.

1;>0

? r,

6 II

i . ] I ,i I . I

5 fl.. -......'.". '. .1·.... .' '.' '.. / .r... - -·t '.--.'1 I I' , /'" "r-' .', """"/T-;.!ii"'" I" ,: ~L t.');.['"> / ~{,"~'". i".'

"

I I~' " t· I '~1' l . i

o ,._..J---l-....L_. .,. L

-;>D (1 ~~O 60 SO

'-I 6 _

-;',"

Figure 4. Mean numbers for the common phytoplankton (Elakatothrix gelatinosa, Sphaerocystis

schroeteri, and Tetraedron minimum, top; Oocystis lacustris, Lagerheimia quadriseta, and

Scenedesmus acutiformis, middle; and eyclotella stelligera, Fragillaria crotonensis, and Synedra spp.,

bottom). Vertical bars represent one standard deviation for each sample date. Control populations

are on the left and atrazine-exposed populations are on the right. Single arrow represents the first

atrazine application, double arrow the second atrazine application. (study figure 4)

8
:~,t>-<~~+/i '5

2

CONTROL ATRAZINE

== 1iAd;~8;;;·t1fr.B'~

:>

:~, :

i~[~~~' IJ ':l'll ·--:3~O-----L.60 -~1:?o 0 '-L:2:-C-,O'-------30~O-·--"-~O-~':_~-~

Z

:=::- t~:UB;''l,W::J"l5h ...

-20 0 30 GO SO t:?l) -Zll " :SO ftO 90 120

.. *DAYS AFTEH TR~ATME~T

Figure 5. Mean numbers for the common phytoplankton (Cryptomonas erosa and Rhodomonas

minuta, top, and Peridinium sp. and Gymnodinium spp., bottom). Vertical bars represent one

standard deviation for each sample date. Control populations are on the left and atrazine-exposed

populations are on the right. Single arrow represents the first atrazine application, double arrow the

second atrazine application. (study figure 5)

9
Figure 6. Mean numbers for the common zooplankton (Keratella cochlearis, Kellicottia longispina,

and Polyartha spp., top; Kellicottia bostoniensis, Chromogaster spp., and Conochilus spp., top-middle;

Daphnia galeata mendotae, Bosmina longirostris, and Ceriodaphnia lacu.stris, bottom-middle; and

Diacyclops b. thomasi, Diaptomus oregonensis, and Tropocyclops p. mexicanus, bottom). Vertical bars

represent one standard deviation for each sample date. Control populations are on the left and

atrazine-exposed populations are on the right. Single arrow represents the first atrazine application,

double arrow the second atrazine application. (study figure 6)

10
Table 1. The common plankton taxa encountered throughout the experiment. a=very common,

"statistically affected" after atrazine exposure, b="A weak statistical effect", and c=no apparent

effect. (no significance results mentioned in original caption) (study table I)

Phytoplankton

Green algae (Chlorophyta)

Elakatothrix gelatinosa Wille a

Lagerheimia quadriseta (Lemmermann) G.M. Smith b

Ooeystis laeustris Chodat a

Seenedesmus aeuttformis Schroeder b

Sphaeroeystis sehroeteri Chodat a

Yellow flagellates (Chrysophyta)

Bitriehia ehodatii (Reverdin) Chodat

Chromulina minuta Doflein

Dinobryon divergens Imhof

Dinobryon soeiale Ehrenberg

Mallomonas sp.

Monosiga sp.

Pseudokephyrion spp.

Salpingoeeafrequentissima (Zacharias) Lemmermann

Diatoms (Bacillariophyceae)

Cyclotella stelligeri (Cleve et Grunow) Van Heurck b

Fragilaria erotonensis Kitton b

Synedra sp. b

Cryptoflagellates (Cryptophyta)

Cryptomonas caudata Schiller c

Cryptomonas erosa Ehrenberg c

Cryptomonas erosa var. rejlexa Marsson c

Cryptomonas ovata Ehrenberg

Rhodomonas minuta Skuja c

Dinoflagellates (Pyrrophyta)

Gymnodinium spp. a

Peridinium cf. inconspieuum Lemmermann

Zooplankton

Rotifers (Rotifera)

Chromogaster spp.

Conochilus spp. c

Kellicottia bostoniensis (Rousselet) c

Kellicottia longispina Kellicott c

Keratella cochlearis (Muller) c

Cladocerans

Bosmina longirostris (O.F. Muller) a

Ceriodaphnia laeustris (Birge) c

Copepods

Daphnia galeata mendotae Birge c

Diacyclops bicuspidatus thomasi S.A. Forbes c

Diaptomus oregonensis Libbjeborg b

Tropocyclops prasinus mexicanus Kiefer c

11
Table 2. Comparisons of Morista coefficients of association for the phytoplankton community within

the control (Ct, C2, C3) and atrazine (At, A2, A3) enclosures, and between control and atrazine

enclosures (mean, minimum, and maximum shown). (study table II)

Control Atrazine Control vs. Atrazine Cn=9i

Days CI-C2 CI-C3 C2-C3 AI-A2 AI-A3 A2-A3 Mean Min. Max.

-20 0.99 0.99 0.97 0.77 0.99 0.79 0.90 0.69 1.00

-6 0.64 0.57 0.85 0.97 0.94 0.93 0.85 0.6H 0.96

I 0.50 0.83 0.37 0.62 0.65 0.94 0.63 0.22 0.86

13 0.78 0.32 0.08 0.67 0.58 0.70 0.58 0.17 0.87

27 0.58 0.38 0.52 0.66 0.62 0.33 0.54 0.35 0.81

49 0.01 0.26 0.01 0.46 0.80 0.57 0.21 0.02 0.72

63 0.13 0.71 0.28 0.81 0.85 0.93 0.36 0.18 0.67

77 0.14 0.90 0.15 0.39 0.42 0.96 0.18 0.09 0.25

98 0.36 0.56 0.40 0.48 0.11 0.64 0.24 0.03 0.72

114 0.04 0.08 0.80 0.52 0.50 0.17 0.21 0.01 0.47

Table 3. Comparisons of Morista coefficients of association for the rotifera community within the

control (Ct, C2, C3) and atrazine (At, A2, A3) enclosures, and between control and atrazine

enclosures (mean, minimum, and maximum shown). (study table III)

Control Atrazine Control vs. Atrazine Cn=9i

Days Ci-C2 Ci-C3 C2-C3 Ai-A2 Ai-A3 A2-A3 Mean Min. Max.

-20 0.98 1.00 0.96 0.99 0.94 0.90 0.95 0.85 0.99

-6 1.00 0.84 0.80 1.00 0.74 0.76 0.89 0.67 0.99

1 0.92 0.69 0.91 0.99 0.96 0.93 0.91 0.73 0.99

13 0.97 0.67 0.81 0.96 0.86 0.69 0.77 0.37 0.99

27 0.98 0.89 0.96 1.00 1.00 1.00 0.95 0.87 1.00

41 0.65 0.49 0.75 0.99 0.61 0.68 0.71 0.42 0.99

49 0.98 0.79 0.89 0.92 0.70 0.75 0.81 0.62 0.99

63 0.54 0.28 0.51 0.41 0.36 0.97 0.64 0.19 0.99

77 0.64 0.55 0.79 0.91 0.94 0.99 0.80 0.53 0.97

98 0.41 0.96 0.62 1.00 0.87 0.87 0.52 0.23 0.92

114 0.32 0.70 0.91 1.00 0.94 0.95 0.81 0.63 0.91

125 0.75 0.88 0.81 0.95 0.95 0.95 0.57 0.18 0.94

139 0.63 0.34 0.74 0.91 0.84 0.96 0.68 0.45 0.90

12
Table 4. Comparisons of Morista coefficients of association for the crustacean community within the

control (CI, C2, C3) and atrazine (AI, A2, A3) enclosures, and between control and atrazine

enclosures (mean, minimum, and maximum shown). (study table IV)

Days

Control Atrazine Control vs. Atrazine (n=9)

Ci-C2 Ci-C3 C2-C3 Ai-A2 Ai-A3 A2-A3 Mean Min. Max.

-20 0.96 0.87 0.80 0.68 0.99 0.65 0.86 0.71 0.99

-6 0.86 0.78 0.99 0.94 0.99 0.89 0.91 0.70 0.99

1 0.89 0.87 0.91 0.99 0.95 0.93 0.94 0.88 1.00

13 0.31 1.00 0.31 0.98 0.90 0.85 0.78 0.34 1.00

27 0.67 0.95 0.47 0.93 0.73 0.61 0.75 0.43 0.97

41 0.90 0.52 0.69 0.87 0.90 0.99 0.80 0.48 0.98

49 0.96 0.62 0.78 0.88 0.96 0.83 0.70 0.16 0.96

63 0.92 0.33 0.56 0.70 0.53 0.68 0.68 0.22 0.96

77 0.40 0.64 0.86 0.34 0.36 0.42 0.47 0.08 0.90

98 0.72 0.81 0.61 0.13 0.10 0.92 0.46 0.07 0.88

114 0.94 0.29 0.48 0.16 0.07 0.99 0.55 0.06 0.94

125 0.90 0.62 0.51 0.04 0.03 0.01 0.39 0.02 0.84

139 0.58 0.90 0.71 0.31 0.08 0.94 0.49 0.20 0.88

13
Data Summary Table:

Effect Magnitude of Nominal Day of Time Did Comments

Effect Atrazine Study Effect Weighted Recovery

relative to Concentration Occurred Average Occur - if

controls (flg/L) Atrazine so. when

Concentration (~ee

from Day 0 to comment)?

When Effect

Occurred

Change in ,

phytoplankton Yes (the community >93% following From text, composition (signiticant 100-200 I-lg/L Day 49 130 flg/L spring, p.128. (decrease in at p<0.05)

green algae Day 323)

biomass)

Decrease in From

total number of 20% lOOllg/L Day 1 1001lg/L No Fig.l and

phytoplankton text, p.

taxa 126.

Change in

number of No effect 100-200 flg/L NA NA NA From

zooplankton Fig.l

taxa

Change in

rotifer No effect 100-200 Ilg/L NA NA NA From

community Table 3.

composition

Change in

crustacean

zooplankton 45% (not community statistically 100 Ilg/L Day 13 100 Ilg/L No From text,

composition significant) p. 133

(decrease in

Cladocera

numbers)

Decrease in NR From text, immature (significant 100-200 Ilg/L Day 114 169.6Ilg/L No

zooplankton at p<0.05) p. 133

Note: NA=Not Applicable, NR=Not Reported

Bolded values are those data points specified in the "Cosm Data Points and Exposure Profiles" spreadsheet.

Since no atrazine measurements were reported, it was assumed that no degradation

occurred and concentrations were always nominal (100 /-lg/L on Day 0, 200 /-lg/L starting

Day 35). Time weighted average concentrations were then computed as the cumulative

average over all elapsed days; see linked Excel table below. Note that Herman et af.

1986 and Hamilton, et af. 1989 papers provide atrazine concentrations and may be used

to provide more specific TWA's.

14
Day Cone TWA 0 100 100.0 1 100 1000 2 100 1000 3 100 100.0 4 100 1000 5 100 100.0 6 100 100.0 7 100 1000 8 100 1000 9 100 1000

10 100 100.0

11 100 1000

12 100 100.0

13 100 100.0

14 100 1000

15 100 1000

16 100 1000

17 100 1000

18 100 1000

19 100 100.0

20 100 1000

21 100 1000

22 100 1000

23 100 1000

24 100 1000

25 100 1000

26 100 100.0

27 100 100.0

28 100 1000

29 100 1000

30 100 100.0

31 100 100.0

32 100 1000

33 100 100.0

34 100 100.0

35 200 1028

36 200 105.4

37 200 1079

38 200 110.3

39 200 1125

40 200 114.6

41 200 1167

42 200 118.6

43 200 1205

44 200 122.2

45 200 1239

46 200 1255

47 200 127.1

48 200 1286

49 200 130.0

50 200 131.4

51 200 1327

52 200 1340

53 200 1352

54 200 136.4

55 200 1375

56 200 1386

57 200 1397

58 200 140.7

59 200 1417

60 200 142.6

61 200 1435

62 200 1444

63 200 1453

64 200 1462

65 200 1470

66 200 1478

67 200 1485

68 200 1493

69 200 1500

70 200 150.7

71 200 1514

72 200 1521

73 200 1527

74 200 1533

75 200 1539

76 200 154.5

77 200 155.1

78 200 1557

79 200 1563

80 200 1568

81 200 1573

82 200 1578

83 200 1583

84 200 1588

85 200 1593

86 200 1598

87 200 1602

88 200 1607

89 200 1611

90 200 1615

91 200 162.0

92 200 162.4

93 200 162.8

94 200 1632

95 200 1635

96 200 1639

97 200 164,3

98 200 1646

99 200 165.0

100 200 1653

101 200 1657

102 200 1660

103 200 1663

104 200 1667

105 200 167.0

106 200 1673

107 200 1676

108 200 167.9

109 200 168.2 15 110 200 1685

111 200 1688

112 200 1690

113 200 1693
Reviewer Comments:

This study provided a relatively comprehensive examination ofthe phytoplankton and

zooplankton community responses to the two pulses of atrazine in the lake enclosures.

As in the other articles on the same enclosure experiment, although only one atrazine

treatment level is examined, 3 replicates for treatments and controls in a semi-natural

experimental setting provide a somewhat reasonable sample size for statistical analysis

(i.e., more than 2 replicates found in many ofthe 'cosm studies). In addition, it is unclear

whether atrazine was present in the controls as no information was provided on

surrounding land use or background pesticide concentrations in Lake St. George where

the enclosures were located. As in the other studies published by Hamilton, there is some

cause for concern regarding enclosure effects. In this article, bar graphs of biomass and

population numbers for the major groups of plankton also seem to provide open-water

sample analyses, which for the diatoms and dinoflagellates show much stronger

differences between open and enclosed samples than between atrazine-exposed and nonexposed samples. Since, at least in these particular variables, atrazine effects were

outweighed by enclosure effects, other parts ofthe study (which do not compare to openwater samples) may be compromised by the confounding effects of enclosure.

Another issue raised by information in this study that is not provided in the other

Hamilton, et al., studies is higher trophic level effects caused by the presence of

planktivorous fish. The methods state that one smallmouth bass was added to each

enclosure to control these populations; otherwise, there is no mention ofthe fish, and the

other studies fail to mention fish anywhere. It is unclear whether the enclosures were

similar in terms ofthe number ofplanktivorous fish present during pre-treatment and

whether the planktivorous population changed with treatment.

The following presentation deficiencies in the methods and results sections were noted:

no background atrazine concentrations are provided, nor are data of measured atrazine

concentrations in the treatment enclosures provided; zooplankton sampling was not

described; the Morista coefficient of association was not defined; and the species list in

Table 1 is incomplete, missing Tetraedron minimum and Polyartha sp., which are both

listed in the Figures (4 and 6, respectively) and in the body text. In particular, the openwater samples displayed in the bar graph figures, which provide insights into the potential

limitations ofthe experimental methodology, are not described in the methods and so

their data cannot be precisely interpreted.

Description of Use in Document (QUAL, QUAN, INV): Qualitative

Rationale for Use: Results confirm a shift in phytoplankton community structure

following two applications of atrazine (35 days apart) at 100 Ilg/L. Significant declines

in the Chlorophyta community were observed, beginning 13 days after atrazine

16
application. Recovery ofthe phytoplantkon community structure did not occur until 323

days after the first atrazine application.

Limitations of Study: Enclosure effects, evident in diatom and dinoflagellate

populations (biomass, numbers), may have influenced other variables for which no openwater samples were analyzed. Only one treatment level was tested and there is a lack of

data on the potential presence of atrazine in the control enclosures. Higher trophic level

endpoints were not analyzed, although feedbacks with planktivorous fish populations

were possible.

Primary Reviewer: Sara Hendrix, M.S., Staff Scientist, Cambridge Environmental Inc

Secondary Reviewer: Anita Pease, M.S., Senior Biologist, Environmental Fate and

Effects Division, U.S. Environmental Protection Agency.

17