Full text of DOI:10.1002/etc.5620211221

Environmental Toxicology and Chemistry, Vol. 21, No. 12, pp. 2675–2684, 2002
2002 SETAC
Printed in the USA
0730-7268/02 $9.00 .00
᭧
ϩ
MOUTHPART DEFORMITIES AND COMMUNITY COMPOSITION OF
CHIRONOMIDAE (DIPTERA) LARVAE DOWNSTREAM OF METAL MINES IN
NEW BRUNSWICK, CANADA
E
RIN O. SWANSBURG,*† WAYNE L. FAIRCHILD,‡ BRIAN J. FRYER,§ and JAN J.H. CIBOROWSKI
†Department of Biological Sciences, University of Windsor, Windsor, Ontario N9B 3P4, Canada
‡Fisheries and Oceans Canada, Gulf Fisheries Centre, P.O. Box 5030, Moncton, New Brunswick E1C 9B6
§Department of Earth Sciences and Great Lakes Institute for Environmental Research, University of Windsor,
Windsor, Ontario N9B 3P4, Canada
࿣
࿣
Department of Biological Sciences and Great Lakes Institute for Environmental Research, University of Windsor,
Windsor, Ontario N9B 3P4, Canada
(
Received 17 October 2001; Accepted 2 May 2002)
Abstract—The effect of metal enrichment on chironomid communities was examined in streams receiving mine drainage from
metal mining operations in New Brunswick, Canada. At five sites receiving mine drainage, metal concentrations were significantly
(p Ͻ 0.05) elevated in water (Zn), periphyton (Cd, Co, Cu, and Zn), and chironomid tissue (Cu, Cd, and Zn) relative to five paired
reference locations. Metal concentrations in chironomid larvae were significantly correlated with concentrations in both water and
periphyton. Chironomid communities were severely affected at sites receiving mine drainage as demonstrated by reduced genera
richness and altered community composition. Sites receiving mine drainage exhibited an increased abundance of metal-tolerant
Orthocladiinae and a reduced abundance of metal-sensitive Tanytarsini relative to reference sites. The incidence of mentum de-
formities was significantly elevated at sites receiving mine drainage (1.43
doubling of that observed at reference sites (0.79 0.22%). Trace metal concentrations at mine-associated streams in New Brunswick
significantly affected the benthic community and have the potential to alter the structure and function of these aquatic ecosystems.
Ϯ 0.24%), with the mean percentage approaching a
Ϯ
Keywords—Chironomid larvae
Metal mining
Mouthpart deformities
Community composition
INTRODUCTION
concentrations of metals may elicit stress in an individual or-
ganism, which may be undetectable at the community assess-
ment level. At the level of the individual, cellular, physiolog-
ical, or morphological changes that may ultimately influence
growth or reproductive success may be better indicators of
impairment [5].
Benthic invertebrates are frequently used to assess the ef-
fects of contaminants in aquatic ecosystems. The limited mo-
bility of invertebrates, their documented sensitivity to a variety
of contaminants, and their close association with contaminant-
accumulating substrates (sediment and periphyton) make them
excellent biomonitoring organisms that are capable of reflect-
ing the location and severity of contamination [1].
Midge (Diptera: Chironomidae) larvae typically comprise
a significant portion of the benthic invertebrate biomass and
much of the diversity in aquatic systems, and are thus com-
monly used in biomonitoring. Chironomidae genera exhibit a
range of sensitivities to contaminants, which permits the de-
tection of subtle effects in metal-enriched environments [2,3].
Consequently, even in the most severely contaminated aquatic
environments, certain chironomid assemblages persist and can
continue to be studied. In addition, chironomids are an im-
portant diet item for predatory insects and benthivorous fish.
As a result, chironomids represent a significant transfer mech-
anism for the movement of contaminants to higher trophic
levels [4].
Hamilton and Saether [6] first proposed the examination of
chironomid deformity levels as an indication of environmental
degradation. A deformity is defined as any morphological fea-
ture that departs from normal configuration [7]. Mouthpart
deformities (mentum, ligula, mandibles, and maxillary palps)
have been observed in many genera inhabiting a variety of
environments. The reported incidence of mentum deformities
ranges from 0 to 83% [8,9] in metal-enriched environments
and 0 to 48% [9–11] in non–metal-enriched environments. As
a result of the range of deformity frequencies, significantly
elevated incidences in metal-enriched environments frequently
are not observed [12]. Small sample sizes and a range of
operational definitions of deformities contribute to the lack of
significant differences observed [12–14]. Frequently, the se-
verity of metal contamination and environmental degradation
are assessed by examining the incidence of deformities in few-
er than 10 larvae per site [8,15].
The objectives of this study were to compare the richness,
community composition, and incidence of mouthpart (men-
tum) deformities in chironomids collected from streams re-
ceiving mine drainage with paired reference streams. Sites
receiving mine drainage are expected to have higher trace
metal concentrations in water, periphyton, and chironomid tis-
sue. Expected effects on the biota were lower chironomid gen-
Exposure to high concentrations of metals can elicit chang-
es in the benthic invertebrate community, such as reduced
abundance, lower diversity, and increased dominance of ben-
thic communities by metal-tolerant taxa [2]. Exposure to lower
* To whom correspondence may be addressed
(swansburge@dfo-mpo.gc.ca). The current address of E. Swansburg
is Faculte´ d’inge´nierie, Universite´ de Moncton, Moncton, New Bruns-
wick E1A 3E9, Canada.
2675

2676
Environ. Toxicol. Chem. 21, 2002
E.O. Swansburg et al.
Table 1. Location of sampling sites (reference and receiving mine
drainage) in New Brunswick, Canada, from which chemical and
biological samples were collected in June and August 1999
Periphyton collection and analysis
Periphyton samples were collected during August 1999.
Palm- to hand-sized cobbles were removed from the steam bed
and periphyton was scrubbed off with nylon brushes. Periph-
yton was rinsed off the cobble surface with stream water into
acid-washed plastic pails. The suspension was allowed to settle
overnight at 4ЊC. Stream water was decanted off the surface
of the settling periphyton, and all visible macroinvertebrates
were removed.
River
Latitude
Longitude
Reference sites
Northwest Miramichi River
McCormack Brook
South Branch Pabineau River
Nepisiquit River
47
47
47
47
47
Њ
Њ
Њ
Њ
Њ
11.184
17.574
26.474
23.502
23.545
Ј
Ј
Ј
Ј
Ј
N
N
N
N
N
65
66
65
66
66
Њ
Њ
Њ
Њ
Њ
53.671
06.512
49.784
11.104
10.959
Ј
Ј
Ј
Ј
Ј
W
W
W
W
W
44 Mile Brook
Samples were centrifuged at 6,000 rpm for 15 min and
remaining water was decanted. Periphyton was freeze-dried
with a vacuum freeze drier for 24 h. Total concentrations of
16 trace elements were analyzed by inductively coupled plas-
ma equipped with an optical emission spectrophotometer at
the University of Windsor, Great Lakes Institute for Environ-
mental Research Analytical Laboratory [16–18].
Sites receiving mine drainage
Little South Branch
Tomogonops River
Mosquito Brook
Austin Brook
Nepisiquit River
47
47
47
47
47
Њ
Њ
Њ
Њ
Њ
17.538
19.012
23.823
23.652
24.064
Ј
Ј
Ј
Ј
Ј
N
N
N
N
N
66
66
65
66
66
Њ
Њ
Њ
Њ
Њ
03.123
06.470
49.165
07.799
07.885
Ј
Ј
Ј
Ј
Ј
W
W
W
W
W
40 Mile Brook
Chironomid collection
era richness, altered community composition, and an increased
incidence of deformed larvae than observed at reference sites.
Benthic samples were collected during June and August
1999, with a 0.1-m2 modified Hess sampler (253-
␮m mesh
MATERIALS AND METHODS
size) and a D-shaped kick net (1-mm mesh size; Wildlife Sup-
ply, Saginaw, MI, USA). Chironomids were handpicked from
shallow pans in the field to ensure the collection of at least
200 individuals at each site. Initially, 200 individuals per site
was deemed adequate, considering that at least 125 individuals
of one genus is recommended to detect a doubling over back-
ground levels of deformities of 3% as statistically significant
Study sites
Rich ore deposits in northeastern New Brunswick have sus-
tained a century-old metal mining industry, which has con-
tributed to elevated levels of metals in many cobble streams
and rivers of this area. Chemical and biological samples were
collected downstream of five metal mining facilities (Heath
Steele Mine, Stratmat Mine, Brunswick 6 Mine, Caribou Mine,
and Wedge Mine; Table 1). At each mine, with the exception
of Caribou Mine, a site suspected of receiving untreated mine
drainage (i.e., percolation of surface water through excavated
waste rock, but not mine tailings process water) was selected
as a replicate of metal enrichment. At Caribou Mine, a suitable
metal enrichment site could not be found immediately down-
stream of mine drainage and therefore a site downstream of
effluent release was selected. A paired reference, or non–metal-
enriched, site was chosen for each site receiving mine drainage,
paired according to similarity in substrate characteristics, can-
opy cover, stream size, and water velocity.
at
ysis where preserved in chilled Carnoy’s solution (anhydrous
ethanol:glacial acetic acid, 3:1, v/v) and stored at 4 C. Re-
p Ͻ 0.05 [13]. Larvae for identification and deformity anal-
Њ
mainders of benthic samples were preserved in Kahle’s so-
lution (water:95% ethanol:100% formalin:glacial acetic acid,
30:15:6:1, v/v/v) and later sorted in the laboratory.
Benthic samples were partitioned into size fractions by rins-
ing through a series of brass sieves (4-mm to 250-␮m mesh).
Under a dissecting microscope, a portion of each size fraction
was sorted. Approximately 300 additional chironomids per site
were sorted from these composite samples, to augment the
number of chironomids examined for deformity analysis to
approximately 500 individuals per site.
Water bodies examined in this study consisted of cold-
water, forested, headwater streams and medium-sized streams
with open canopies. Mean summer water temperatures in these
Chironomid metal analysis
streams seldom exceed 25ЊC. Generally, stream morphology
For metal analysis, chironomid larvae were picked from
trays in the field and placed on dry ice. Approximately 60
chironomid larvae were collected at each site and combined
as a composite sample for metal analysis. Because of the small
available sample size (2–10 mg dry wt), chironomid samples
were analyzed by inductively coupled plasma equipped with
a mass spectrophotometer at the University of Windsor, Great
Lakes Institute for Environmental Research Analytical Lab-
consisted of riffles and runs, with few pools observed. Sub-
strate at all sampling locations consisted of loose cobbles, with
occasional bedrock outcroppings and limited fine sediment.
Cobbles and other hard substrates were coated with a light
film of periphyton. Streams were fast-flowing and well oxy-
genated, with circumneutral pH and low conductivity (
Ͻ0.10
mS/cm).
oratory [16–18]. Samples were maintained at
analysis. Chironomids were dried on a hot plate at 110
Ϫ
80
Њ
C before
C for
Water sampling and analysis
Њ
Water samples were collected in June 1999. With the use
of 500-ml acid-washed polyethylene bottles, samples were col-
lected at a depth equidistant from the surface of the water and
the surface of the substrate. Water samples were acidified to
a 2% HNO3 (v/v) solution within 48 h of collection and stored
60 min in screw-top perfluoroalkoxy vials. Weight and weight
loss (i.e., % moisture) were recorded. Two milliliters of high-
purity 50% HNO3 was pipetted into each vial. The vials were
sealed and placed on a hot plate at 85ЊC for at least 2 h. The
solution was allowed to cool and then 2 ml of H2O2 was added.
Solutions stood for 60 min and then were resealed and placed
at 4ЊC before analysis. Total concentrations of 25 trace ele-
ments were analyzed by inductively coupled plasma equipped
with an optical emission spectrophotometer at the University
of Windsor, Great Lakes Institute for Environmental Research
Analytical Laboratory (Windsor, ON, Canada) [16,17].
on a hot plate at 85
the caps were removed, and then the solutions were evaporated
to dryness on the hot plate at 85 C. The samples were finally
taken to 12-g final solution mass with 1% HNO3 containing
ЊC for 2 h. Solutions were allowed to cool,
Њ

Metal mining and chironomid mouthpart deformities
Environ. Toxicol. Chem. 21, 2002
2677
the inductively coupled plasma–mass spectrometry internal
standards Be, In, and Tl, at concentrations of 5, 0.5, and 1
ngm/g, respectively.
position (log 2 [relative abundances of genera]). Genera were
included in the cluster analysis if their mean relative abundance
averaged across all 10 sites was
sent in at least 4 of the 10 sites.
Ͼ1.0% and if they were pre-
Chironomid identification and deformities
Differences in genera richness (number of chironomid gen-
era) and mean incidence of deformities between sites receiving
mine drainage and their paired reference site were assessed
Heads of individual chironomid larvae were severed and
mounted ventral side up beside the body on microscope slides
with one-tailed paired-comparison t tests (p Ͻ 0.05). Richness
in CMC-9AF௡ aqueous mounting medium (Masters, Bensen-
and deformity data (percent) were logarithmically transformed
(log 10) before analysis.
ville, IL, USA). Chironomid larvae were identified to the level
of genus [19]. Ten percent of the identified larvae were ran-
domly selected and reexamined to verify correct designation.
Mouthparts of each larva were examined for deformities
of the mentum (one or more missing or additional teeth) during
taxonomic identification. Deformed larvae were later reex-
amined to ensure correct designation. Individuals displaying
broken, chipped, or worn teeth were classified as damaged not
deformed. Gaps in the mentum were classified as deformities
if their surface was smooth, rather than jagged, which indicates
breakage. Other structures were not examined for deformities.
Data were expressed as percentage of individuals deformed,
with standard error determined according to the binomial dis-
tribution.
RESULTS
Trace element analysis of water, periphyton, and
chironomid larvae
Metal concentrations in water, periphyton, and chironomid
larvae (Tables 2 to 4) generally were greater at sites receiving
mine drainage than at their paired reference site (Fig. 1). At
sites receiving mine drainage, concentrations of some metals
in water (Mn and Zn), periphyton (Cd, Co, Mn, Pb, and Zn),
and chironomid larvae (Cd, Pb and Zn) were at least five times
greater than concentrations at reference sites. Cadmium, Co,
Cr, Cu, Ni, Pb, and V were undetected in water but were at
detectable levels in periphyton and chironomid larvae (Fig. 1).
Three principal component (PC) factors were extracted
from metal concentrations in water, accounting for 84.8% of
the variation in the original data (Table 5). Concentrations of
Ca, Mg, Na, and Sr were strongly associated with values of
PC-I, which did not significantly differ between sites receiving
Statistical analysis
Principal component analyses (Statistica௡, StatSoft, Tulsa,
OK, USA) were performed to identify trends in metal con-
centration (water, periphyton, and chironomid tissue), and rel-
ative abundance of dominant genera data according to sites
receiving mine drainage and reference sites. Data for several
elements were excluded from all statistical analyses because
their concentrations were undetectable in most samples. When
concentrations of other elements in individual samples were
below detectable limits, the detection value was used in sta-
tistical analyses. Mean relative abundance (percentage) of gen-
era was examined across all 10 sites, and the nine most abun-
dant taxa were selected. All data were logarithmically trans-
formed before analysis (log 10 for metal concentration data
and log 2 for abundance data).
mine drainage and reference sites (p Ͻ 0.97; Table 5). Iron,
Mn, and Zn concentrations were associated with values of PC-
II, and the mean value of scores for this factor was significantly
higher at sites receiving mine drainage than at reference sites
(p Ͻ 0.05; Table 5). Concentrations of Fe, Mn, and Zn were
most different between mine and reference locations at Heath
Steele and Caribou. Barium and K concentrations were strong-
ly associated with values of the PC-III (Table 5) and the score
for PC-III was significantly higher at sites receiving mine
drainage than at reference sites (p Ͻ 0.05).
Principal component analysis was performed from a cor-
relation matrix. Factor loadings were varimax rotated and met-
al concentrations or relative abundances of individual genera
were said to be associated with a specific principal component
Two principal component factors described metal concen-
trations in periphyton, accounting for 84.1% of the variation
in the original data (Table 5). Concentrations of Cd, Co, Cu,
Mn, Pb, and Zn were strongly associated with values of PC-
I, and the mean value of scores for this factor was significantly
higher at sites receiving mine drainage than at paired reference
when the loading of that genus on a component was
Ͼ0.600.
Factor scores for each site generated from principal component
analysis were then analyzed with a one-tailed paired compar-
sites (p Ͻ 0.05; Table 5). Trace metal concentrations of these
ison
5) and sites receiving mine drainage (
adjusted values).
Correlations among metal concentrations in water, periph-
yton, and chironomid tissue at sites receiving mine drainage
and reference sites were determined with Pearson’s correlation
coefficients (Statistica). Metal concentrations that were below
detection limits were replaced by detection values. Data were
logarithmically transformed (log 10) before analysis.
The composition of chironomid communities at sites was
estimated from both handpicked and composite chironomid
samples. The total number of chironomids sorted from com-
posite samples in the laboratory was added to the number of
chironomids that had been picked from the site in the field.
Absolute estimates of abundance were not determined because
of differences in the sampling effort expended among sites.
Cluster analysis (Euclidean distances grouped by Ward’s meth-
od) was used to group sites on the basis of community com-
t
-test to test for differences between reference sites (
n
ϭ
elements were most different between mine and reference lo-
cations at Heath Steele and Caribou. Concentrations of Cr, Ni,
and V were strongly associated with values of PC-II, which
did not differ significantly between sites receiving mine drain-
n
ϭ
5) (with Bonferroni-
p
age and reference sites (p Ͻ 0.60; Table 5).
Two principal component factors were extracted from metal
concentrations in chironomid tissue, accounting for 76.0% of
the variation in the original data (Table 5). Concentrations of
Fe, Mn, and Pb were strongly associated with PC-I, scores of
which did not differ significantly between sites receiving mine
drainage and reference sites (p Ͼ 0.07; Table 5). Concentra-
tions of Cu, Cd, and Zn were positively associated with PC-
II, but Ba concentration was negatively associated with this
factor (Table 5). The scores for PC-II were significantly dif-
ferent at sites receiving mine drainage and reference sites
(
p
Ͻ
0.05).
Generally, metal concentrations were highest in periphyton,
followed by chironomid larvae, and lowest in water (Fig. 1).

2678
Environ. Toxicol. Chem. 21, 2002
E.O. Swansburg et al.
Table 2. Total concentrations of trace elements (
␮
g/L) in water at reference sites (Ref.) and sites receiving mine drainage (Mine) at five metal
mining facilities (Heath Steele, Stratmat, Brunswick 6, Wedge, and Caribou mines) in New Brunswick, Canada
Heath Steele Stratmat Brunswick 6 Wedge Caribou
Metal
Ref.
Mine
Ref.
Mine
ND
466.8
ND
ND
6.8
Ref.
Mine
Ref.
Mine
Ref.
Mine
Ag
Al
As
B
Ba
Be
Bi
NDa
ND
ND
ND
2.53
ND
ND
ND
233.4
ND
ND
2.8
ND
ND
ND
ND
3.29
ND
ND
ND
ND
ND
ND
5.58
ND
ND
ND
154.3
ND
ND
5.83
ND
ND
ND
ND
3.23
ND
ND
ND
ND
ND
ND
3.76
ND
ND
ND
ND
ND
ND
5.13
ND
ND
ND
ND
ND
ND
3.59
ND
ND
ND
ND
ND
ND
ND
ND
Ca
Cd
Co
Cr
Cu
Fe
K
Mg
Mn
Mo
Na
Ni
Pb
Sn
Sr
4,173
ND
ND
ND
ND
105
352.2
734.4
2,428
ND
ND
ND
ND
337.5
544.9
809.4
87.72
ND
1,466
ND
ND
2,239
ND
ND
ND
ND
79.86
468.5
808.2
1,802
ND
ND
ND
ND
150.7
627.8
780.4
38.01
ND
1,442
ND
ND
ND
11.75
ND
ND
6,026
ND
ND
ND
ND
137.2
421.1
1,230
57.81
ND
4,615
ND
ND
4,857
ND
ND
ND
ND
180.7
410.4
1,170
241.2
ND
1,527
ND
ND
3,763
ND
ND
ND
ND
63.57
365.7
871.6
4,118
ND
ND
ND
ND
78.66
446.7
922
5,203
ND
ND
ND
ND
38.9
406.1
877.2
13,281
ND
ND
ND
ND
62.18
556.9
1,577
55.88
ND
2,287
ND
ND
ND
34.95
ND
ND
151.3
6.32
8.69
3.48
9.86
2.44
ND
1,550
ND
ND
ND
ND
1,363
ND
ND
ND
10.54
ND
ND
ND
1,646
ND
ND
ND
ND
1,620
ND
ND
ND
18.8
ND
ND
ND
1,268
ND
ND
ND
ND
12.9
ND
ND
108.7
ND
31.05
ND
ND
18.57
ND
19.79
ND
ND
237.3
17.24
17.35
20.99
Ti
V
Zn
ND
ND
2.56
ND
ND
ND
ND
ND
3.03
5.2
75.25
14.72
a ND
ϭ
not detected.
In periphyton, Zn concentrations were 104 to 106 times higher
than in water and 10 to 100 times higher than in chironomid
larvae. Compared to metal concentrations in chironomid lar-
vae, Cu and Pb concentrations were 10 to 100 times higher in
periphyton, whereas Cd concentrations were only 1 to 10 times
higher in periphyton. Zinc concentrations in chironomid larvae
were significantly correlated with both metal concentrations
in water (r ϭ 0.84; p Ͻ 0.05) and periphyton (r ϭ 0.79; p Ͻ
0.05). Cadmium, Cu, and Pb concentrations in chironomid
larvae were significantly correlated with metal concentrations
these associations were not as strong as the associations for
Zn ( 0.68, 0.36, and 0.52, respectively).
r
ϭ
Chironomid community composition
Approximately 4.1
104 chironomid larvae were collected
ϫ
at the 10 study sites. Identification of a subset of approximately
5,000 slide-mounted larvae yielded 48 genera belonging to
five tribes or subfamilies (Table 6). The collected larvae were
comosed of 49% Chironominae, 39% Orthocladiinae, 11%
Tanypodinae,
Ͻ1% Diamesinae, and Ͻ1% Pseudochironom-
in periphyton. Although statistically significant (p Ͻ 0.05),
inae. The genera Cricotopus and Orthocladius are difficult to
Table 3. Total concentrations of trace elements (
␮g/g dry wt) in periphyton at reference sites (Ref.) and sites receiving mine drainage (Mine) at
five metal mining facilities (Health Steele, Stratmat, Brunswick 6, Wedge, and Caribou mines) in New Brunswick, Canada
Heath Steele
Stratmat
Brunswick 6
Wedge
Caribou
Metal
Ref.
Mine
Ref.
Mine
Ref.
Mine
Ref.
Mine
Ref.
Mine
Al
As
Ca
Cd
Co
Cr
Cu
Fe
K
Mg
Mn
Na
Ni
Pb
V
13,911
18.57
8,526
2.11
29.4
25.5
15.4
20,523
5,543
4,485
6,481
2,561
18.85
ND
34.21
38,488
851.1
5.97
5,380
4,664
12,409
62.51
3,908
8.36
158.8
18.52
17.54
16,517
1,848
1,694
26,826
268
43,029
12,296
14,994
6.82
2,924
4.02
14,473
22,825
96.18
9,550
48.04
162.6
39.87
1,323
19,186
4,265
4,287
151.9
3,556
16.57
340.8
30.12
23.88
4,467
7.47
9.67
ND
21.48
5,838
6,076
3,155
3,291
134.8
3.91
109.8
6.98
1,328
83.58
7,241
49.77
585.5
61.92
475.6
89,918
3,350
4,607
35.74
5,406
3.4
11.25
23.2
ND
12,951
3,274
3,503
1,320
894
15.61
ND
47.49
138.9
11.07
9,355
1.77
1.35
2.75
15.72
15.36
692.8
11.04
25.11
18.33
NDa
ND
1,304
54,648
8,718
5,967
13,247
690
1,766
5,978
1,330
2,589
159.9
1.44
ND
ND
248.3
40,746
18,300
1,571
1,742
959
5,859
6,343
1,968
158,682
33,419
354.7
116.6
1,387
41.3
18,213
472.6
574.7
345.6
25.17
636.4
60.65
54.41
33.32
15.3
7.9
558.2
22.02
585.9
13.22
13.69
39.46
308.7
74.43
40.34
35.95
Zn
202.4
5,377
2,048
25,740
a ND
ϭ
not detected.

Metal mining and chironomid mouthpart deformities
Environ. Toxicol. Chem. 21, 2002
2679
Table 4. Total concentrations of trace elements ( g/g dry wt) in chironomid larvae from reference sites (Ref.) and sites receiving mine drainage
␮
(Mine) at five metal mining facilities (Heath Steele, Stratmat, Brunswick 6, Wedge, and Caribou mines) in New Brunswick, Canada. Concentrations
of Ca, K, Mg, and Na are reported as percentage of the dry mass
Heath Steele
Ref. Mine
1.7 3.3
Stratmat
Brunswick 6
Wedge
Caribou
Mine
Metal
Ref.
Mine
Ref.
Mine
Ref.
Mine
Ref.
As
Ba
Bi
Ca (%)
Cd
Co
Cr
Cu
Fe
K (%)
Mg (%)
Mn
Na (%)
Ni
Pb
Rb
Sr
V
0.9
17.9
ND
0.2
3.6
1.4
1.7
13
3.6
34.1
0.1
0.2
20.3
3.8
3.2
23.8
ND
0.1
8.3
18.2
0.2
2.0
17.4
ND
0.2
0.2
2.5
5.8
13
1.6
41.3
ND
0.1
1.2
2.6
3.5
10.6
ND
0.1
0.6
0.5
0.9
10
2.0
6.3
ND
0.2
37.2
2.7
1.4
115
591
0.8
0.1
401
0.4
2.3
1.6
1.7
3.5
0.5
98.8
NDa
0.1
0.5
0.6
0.3
6
295
0.5
0.1
5.0
0.2
0.1
4.7
3.2
0.5
62
759
0.4
0.1
133
0.2
1.3
13.8
1.1
0.9
0.6
Ͻ
0.2
3.6
5.2
13.3
1.4
10.6
3.6
1.0
31
2.4
29
1,530
45
6,417
153
1,288
0.6
0.1
160
1,229
2,072
2,591
597
1.1
0.1
0.6
0.1
541
0.4
1.4
6.0
2.8
6.9
2.4
1.1
0.2
0.6
0.1
0.9
0.2
0.5
0.2
155
0.6
6.0
3.5
11.3
7.2
6.8
81
Ͻ
135
1,590
1,504
2,707
138
0.3
1.1
ND
1.2
1.3
0.2
1.0
1.4
81.8
3.7
8.9
2.0
0.4
5.5
0.8
5.1
0.6
2.9
23.9
8.8
4.3
2.4
0.6
1.3
2.4
7.1
4.1
0.7
10.2
3.6
131.4
4.3
3.7
5.3
0.9
3.3
Zn
45
436
141
1,691
308
929
197
113
1,813
a ND
ϭ not detected.
distinguish as larvae, and thus were classed jointly (i.e., Cri-
cotopus Orthocladius). Cricotopus Orthocladius sp. was the
most abundant taxon, representing 23% of chironomid larvae
examined.
site receiving mine drainage for all five pairs of sites examined.
The largest difference in genera richness between mine drain-
age and reference sites occurred at Heath Steele (13 genera).
The smallest difference was observed at Stratmat (one genus).
/
/
Chironomid richness ranged from 12 to 27 genera per site.
Genera richness was greater at the reference site than at the
Sites receiving mine drainage had a mean (
[SE]) richness of 16 2 genera, which was significantly lower
than the mean at corresponding reference sites (23 1;
0.025; 5, paired comparison test).
Ϯ standard error
Ϯ
Ϯ
p Ͻ
n
ϭ
t
Community composition differed between sites receiving
mine drainage and reference sites at both the tribe or subfamily
level and the genera level. Larval Tanypodinae, Chironomini,
and Tanytarsini were relatively more abundant at reference
sites, whereas larval Orthocladiinae were relatively more abun-
dant at sites receiving mine drainage (Fig. 2).
The nine most relatively abundant chironomid genera ac-
counted for 60 to 93% of all chironomids at the study sites.
Relative abundances of Larsia
sectra Stempellinella, and Microtendipes were highest at ref-
erence sites, whereas abundances of Polypedilum Cricotopus
Orthocladius Eukiefferiella, and Thienemanniella were great-
, Thienemannimyia, Microp-
,
,
/
,
est at sites receiving mine drainage. Principal component anal-
ysis extracted three factors, accounting for 77.6% of the
variation in the original data (Table 7). Relative abundances
of Larsia, Thienemannimyia, Eukiefferiella, and Polypedilum
were most strongly associated with the values of PC-I, which
accounted for 33% of the variation. Relative abundances of
the predatory tanypodines Larsia and Thienemannimyia were
positively associated with each other but negatively associated
with the relative abundances of Eukiefferiella and Polypedil-
um (Table 7).
The relative abundance of Micropsectra
, Stempellinella,
Microtendipes, and Cricotopus Orthocladius was most
/
Fig. 1. Mean concentrations (
in (top) water ( g/L), (middle) periphyton (
chironomid larvae ( g/g dry wt) at reference sites and sites receiving
mine drainage ( 5) associated with metal mining facilities in New
Brunswick, Canada. Metals are indicated on the axis and were
ordered according to concentrations in water at reference sites. Scales
are logarithmic and scales and units of measurement differ among
panels.
Ϯ
1 standard error) of trace elements
strongly associated with PC-II. Relative abundances of the
Tanytarsini taxa (Micropsectra and Stempellinella) were pos-
itively associated with PC-II, whereas the relative abundances
of orthoclads (Cricotopus/Orthocladius) were negatively as-
sociated with values of PC-II. Relative abundance of Thiene-
manniella was positively associated with values of PC-III.
␮
␮
g/g dry wt), and (bottom)
␮
n
ϭ
x

2680
Environ. Toxicol. Chem. 21, 2002
E.O. Swansburg et al.
Table 5. Identification of metals from concentrations in water,
periphyton, and chironomid larvae loading on principal components
(PC) and the percentage of total variance explained by each component
at mine drainage vs 0.39% at reference). Mean percentage
(
Ϯ
SE;
at sites receiving mine drainage. This was significantly higher
0.05; one-tailed paired-comparison test) than the mean
n ϭ 5) of total deformed individuals was 1.43 Ϯ 0.24%
(
p
Ͻ
t
Principal
Variance
component
Parameter
Factor loadings
explained (%)
incidence observed in larvae at reference sites and (0.79
0.22%).
Ϯ
Metal concentrations in water
PC-I
Ca
Mg
Na
Sr
Fe
Mn
Zn
Ba
K
0.94
0.90
0.76
0.98
0.95
0.87
0.76
0.82
0.69
38.0
DISCUSSION
Trace element analysis of water, periphyton, and
chironomid larvae
PC-II
27.0
19.0
Sites downstream of waste rock piles, at metal mine sites
in New Brunswick, received significant metal input, even up-
stream of effluent release locations. Metals accumulated at the
base of the food chain and in organisms feeding on these basal
resources. Significantly elevated concentrations of Zn were
observed in water, periphyton, and chironomid larvae. Despite
concentrations being undetectable in water samples, Cd, Cu,
and Pb were significantly elevated in both periphyton and chi-
ronomid larvae, indicating that these metals are indeed enter-
ing receiving waters from waste rock piles on mine property.
Metal concentrations in periphyton were 104 to 106 times
higher than observed in water, suggesting that periphyton ef-
fectively bioaccumulates trace metals [20]. Considering the
importance of periphyton in lotic food webs [21], the presence
of elevated metal concentrations in periphyton may have im-
portant implications. Periphyton is a principal food source for
many aquatic organisms, which consume predominately algae
(scrapers and grazers) and detritus associated with algae (col-
lectors and gatherers). Consequently, invertebrates associated
with periphyton likely ingest and retain substantial concentra-
tions of metals. Metals accumulated by primary consumers
may be transferred to higher trophic levels, in both aquatic
and terrestrial ecosystems [2].
Metals such as Cd, Cu, and Zn are readily accumulated
from periphyton by benthic invertebrates [22,23]. Kiffney and
Clements [23] argued that invertebrates in the Arkansas River
(USA) accumulated Cd, Cu, and Zn primarily through diet, as
indicated by the correlation between bioaccumulated metals
and levels in periphyton. However, in this study, strong cor-
relations between Zn concentrations in chironomid larvae and
concentrations in both water and periphyton were found, mak-
ing it difficult to discern the source of Zn uptake by larvae.
Significant correlations between Cd, Cu, and Pb concentrations
in periphyton and chironomid larvae suggest that periphyton
is a source of uptake for these elements.
PC-III
Metal concentrations in periphyton
PC-I
Cd
Co
Cu
Pb
Mn
Zn
Cr
Ni
V
0.89
0.75
0.79
0.81
0.73
0.93
0.93
0.80
0.91
49.0
PC-II
25.0
Metal concentrations in chironomid larvae
PC-I
Fe
0.90
0.89
0.86
0.71
0.85
0.75
0.84
38.9
37.1
Pb
Mn
Ba
Cd
Cu
Zn
PC-II
Ϫ
No significant difference was observed in mean factor
scores of PC-I or PC-III between sites receiving mine drainage
and reference sites (one-tailed paired-comparison
0.05). Relative abundances of the taxa associated with these
factors (Larsia Thiennemannimyia Eukiefferiella Polype-
dilum, and Thienemanniella) were only substantially different
between two of the five pairs of sites, Brunswick and Wedge,
respectively. Relative abundance of taxa associated with PC-
II differed significantly between sites receiving mine drainage
t tests; p Ͼ
,
,
,
and reference sites (one-tailed paired-comparison
t test; p Ͻ
0.03), with the greatest differences between mine and reference
sites apparent at Heath Steele, Brunswick, and Caribou.
The 18 chironomid genera included in the cluster analysis
accounted for
Ͼ92% of chironomids identified at all 10 sites.
Cluster analysis generated two relatively distinct groups of
sites (Fig. 3). Sites receiving mine drainage at Heath Steele,
Brunswick, and Caribou mines had similar chironomid com-
munities, characterized by higher relative abundances of Cri-
cotopus/Orthocladius and Psectrocladius. Chironomid com-
munities at all reference sites, and two sites receiving mine
drainage (Wedge and Stratmat mines) were characterized by
Chironomid community composition
According to Armitage and Blackburn [24], specific iden-
tification of chironomid larvae is as effective at distinguishing
degrees of metal contamination as examining whole macro-
invertebrate assemblages. However, few studies have focused
on the effects of metals on chironomid communities beyond
the subfamily level [3,9–11,25,26], obscuring the potential for
chironomid larvae to reflect metal exposure through loss of
richness and altered community composition.
higher relative abundance of Microtendipes, Paratendipes,
Rheotanytarsus, and Stempellinella
.
Chironomid deformities
The menta of 5,042 chironomid larvae were examined for
deformities ( 500 per site). A total of 56 deformed individuals
In this study, sites with elevated metal concentrations sup-
ported significantly fewer chironomid genera than their paired
reference site, confirming the susceptibility of chironomid di-
versity to reflect metal enrichment. Chironomid genera rich-
ness at sites receiving mine drainage was comparable to genera
richness reported at other metal-enriched streams [9,26–28],
whereras genera richness at reference sites was two to nine
ϳ
belonging to 15 different genera was observed in samples
(Table 6). The differences in the incidence of mentum defor-
mities between sites receiving mine drainage and references
sites were largest at Heath Steele mine (1.69% at mine drainage
vs 0.20% at reference) and smallest at Caribou mine (0.58%

Metal mining and chironomid mouthpart deformities
Environ. Toxicol. Chem. 21, 2002
2681
Table 6. The number of chironomid genera collected at reference sites (Ref.) and sites receiving mine drainage (Mine) at five metal mining
facilities (Heath Steele, Stratmat, Brunswick 6, Wedge, and Caribou mines) in New Brunswick, Canada. Values in parentheses represent the
number of deformed larvae
Health Steele
Stratmat
Brunswick 6
Ref. Mine
Wedge
Caribou
Taxa
Ref.
Mine
Ref.
Mine
2
Ref.
69
Mine
Ref.
Mine
Ablabesmyia
Larsia
Paramerina
Thienemannimyia
Macropelopia
Nilotanypus
Procladius
Diamesa
Pagastia
Potthastia
26
116
2
2
20
35
1
1
1
8
3
13
5
2
69
19
1
12 (1)
1
14 (2)
32 (1)
1
8
6
1
1
59 (2)
1
16
165 (1)
73 (2)
1
2
2 (1)
6
3
1
5
24
19
1
12
19
Chironomus
2
6
Cryptochironomus
Demicryptochironomus
Glyptotendipes
Microtendipes
Nilothauma
1
1
3
1
1
1
1
1
32
3
111
11
31 (1)
5
2
3
Parachironomus
Paralauterborniella
Paratendipes
Phaenopsectra
Polypedilum
Saetheria
1
24
2
12
38
151 (2)
17
2
44 (1)
8 (1)
52
5
2
1
1
6
131
17
133 (2)
208 (4)
63 (3)
307 (3)
21
1
1
Tribelos
Xenochironomus
Cladotanytarsus
Heterotanytarsus
Micropsectra
Paratanytarsus
Rheotanytarsus
Stempellina
Stempellinella
Tanytarsus
Pseudochironomus
Brillia
1
1
1
6
61 (1)
58 (1)
36
98 (1)
1
32
3
56
7
6
8
1
17 (1)
82 (1)
129
1
88
1
5
4 (2)
2
2
15
50
28
7
2
52
17
2
2
13
2
35 (1)
22 (2)
3
4
1
10
1
20
33
Corynoneura
3
65
12
2
2
22
1
7
22
2
260 (4)
90
1
42
2
9
5
9
Cricotopus
/
Orthocladius
375 (5)
118 (4)
16
29
225
3 (1)
Eukiefferiella
Heterotrissocladius
Krenosmittia
Lopescladius
2
1
1
1
2
1
1
7
5
Parakiefferiella
Parametriocnemus
Psectrocladius
Pseudosmittia
Rheocricotopus
Symposiocladius
Synorthocladius
Thienemanniella
Sum
2
2
14
27
10
9 (1)
8 (1)
1
1
21
12
1
1
78
1
2
3
7
1
46
519
19
13
7
507
21
3
489
25
1
534
12
37 (2)
502
20
2
501
27
6
466
20
21
515
23
495
13
1.21
514
18
0.58
Generic richness
Proportion deformed
0.20
1.69
1.00
1.73
1.40
1.81
0.97
0.39
times higher than previously reported for mine-effect studies
[9,28]. Undoubtedly, the sampling effort employed in this
algal taxa (e.g., blue-green algae). Consequences of taxa loss
are influenced by the role and abundance of metal sensitive
taxa in the community. Loss of taxa representing a unique
niche or whose presence influences the abundance of other
species will adversely affect ecosystem processes [31].
At sites receiving mine drainage, community composition
supported a greater relative abundance of genera reported as
metal-tolerant, and a decreased abundance of metal-sensitive
taxa. Increased relative abundances of orthoclad larvae (e.g.,
study (i.e.,
richness.
n ϳ 500 per site) contributed to a greater genera
Low genera richness at sites receiving mine drainage may
have resulted from the loss of sensitive taxa due to both direct
and indirect effects of metal toxicity. Zinc concentrations in
water at sites receiving mine drainage of this study were similar
to concentrations causing a 50% reduction in survival of larval
Tanytarsus in laboratory tests [29]. Gower et al. [30] suggested
that reduced richness in metal enriched environments resulted
from fewer available food sources because of the replacement
of palatable species of algae (e.g., diatoms) by metal-tolerant
Cricotopus, Orthocladius, and Eukiefferiella spp.) have fre-
quently been observed downstream of metal mining and elec-
troplating industries [2,3,26–28,30,32]. At sites receiving mine
drainage in this study, the relative abundance of Cricotopus
/

2682
Environ. Toxicol. Chem. 21, 2002
E.O. Swansburg et al.
Fig. 2. Mean (
ronomid tribes or subfamilies at reference sites (open) and sites re-
ceiving mine drainage (solid) ( 5) associated with metal mining
facilities in New Brunswick, Canada.
Ϯ 1 standard error) percent relative abundance of chi-
Fig. 3. Similarity of reference sites (
drainage (
࿞
R) and sites receiving mine
n
ϭ
࿞
M) associated with metal mining facilities (Heath Steele
[HEA], Stratmat [STR], Brunswick 6 [BRU], Wedge [WED], and
Caribou [CAR] mines) in New Brunswick, Canada, according to the
percent relative abundance of chironomid genera as determined by
cluster analysis. Distances between clusters were measured by Eu-
clidean distances and clusters were grouped according to Ward’s method.
Orthocladius was significantly higher, whereas other orthoclad
taxa (Eukiefferiella and Psectrocladius) showed trends toward
increased abundance relative to reference sites. Surber [32]
suggested that perhaps the ability of orthoclad taxa to tolerate
metals was only indirectly responsible for their dominance at
metal-contaminated sites. Rather, the dietary preference of or-
thoclade taxa for metal-tolerant blue-green algae, which dom-
inate the periphyton of these sites, could explain their domi-
nance.
Reduced abundances of larval Tanytarsini (e.g., Microp-
sectra and Tanytarsus spp.) are commonly observed in metal-
enriched environments, resulting in their designation as metal
sensitive [26,30]. However, Kiffney and Clements [33] ob-
served increased abundance of larval Tanytarsini in conditions
of low (i.e., 0.001 mg Cd/L, 0.010 mg Cu/L, and 0.108 mg
Zn/L) metal contamination, suggesting that these larvae may
exhibit moderate metal tolerance. Clements et al. [28] also
found that the abundances of Micropsectra and Stempellinella
spp. were only reduced at highly metal-contaminated sites. At
sites receiving mine drainage of the present study, the relative
abundances of Micropsectra and Stempellinella spp. were re-
duced by a factor of 2 and 10, respectively, consistent with
the designation of others of Tanytarsini as sensitive to metal
enrichment.
drainage with lower metal concentrations (Wedge and Stratmat
mines) had community assemblages resembling those at ref-
erence sites. Two of the three sites with reduced richness and
dominance of metal-tolerant taxa were operating at the time
of sampling, whereas mining at the third site had been ter-
minated for a significant period of time. Presumably at this
site, significant drainage from waste rock is still affecting water
quality 15 years after mine closure.
Chironomid deformities
This study documents one of the lowest deformity fre-
quencies at both reference and contaminant-receiving sites in
the published literature. We were able to document significant
elevations in incidence of deformities despite the low fre-
quency because we examined more individuals than is typi-
cally done in deformity analysis. The background incidence
of deformities observed at our reference sites (0.79%) falls in
the low end of the range (0–5%) of mentum deformities fre-
quently reported at reference locations [10,34]. Natural inci-
dences of mentum deformities previously have been observed
Changes in chironomid communities were consistent with
metal concentrations in water and periphyton. Sites with higher
metal concentrations (Heath Steele, Brunswick 6, and Caribou
mines) had the lowest taxa richness and highest relative abun-
dances of metal-tolerant taxa. Conversely, sites receiving mine
to be
with a frequency of
taminated [37].
Ͻ1% based on subfossil records [35,36], and thus sites
Ͼ1% are considered by some to be con-
The incidence of deformities at sites receiving mine drain-
age also falls within the range routinely observed by others at
reference sites. Low levels of deformities could be due to
several factors. First, different chironomid genera are reputed
to exhibit differential sensitivity to deformity expression
[12,38,39]. Therefore, our estimate of the incidence of defor-
mities could be conservative, considering that this metric was
expressed as a percentage pooled across all genera. Similarly,
tolerance to metal exposure also could contribute to lower
incidence of deformities at sites receiving mine drainage. Met-
al tolerance has been observed in midge larvae [40–42] and
has resulted in altered responses to biomonitoring indices [43]
and lower deformity frequencies [15]. The comparison of the
incidence of deformities observed in this study to that observed
in others may also be inappropriate, considering the dissimi-
larity in habitats. Most deformity analysis studies have focused
on chironomid communities of soft-sediment lotic and lentic
Table 7. Identification of dominant chironomid genera (based on
relative abundance) loading on principal components (PC) and the
percentage of total variance explained by each component
Principal
Factor
Variance
component
Parameter
loading explained (%)
PC-I
Thienemannimyia
Polypedilum
Larsia
Eukiefferiella
Cricotopus/Orthocladius
Micropsectra
Stempellinella
Microtendipes
Thienemanniella
0.96
0.77
0.72
0.68
0.94
0.87
0.66
0.52
0.82
33.0
31.0
13.0
Ϫ
Ϫ
Ϫ
PC-II
PC-III

Metal mining and chironomid mouthpart deformities
Environ. Toxicol. Chem. 21, 2002
2683
the Natural Sciences and Engineering Research Council of Canada
(NSERC; J.J.H. Ciborowski and B.J. Fryer), Fisheries and Oceans
Canada through the Maritimes Region and the Toxic Chemicals Pro-
gram (W.L. Fairchild), and by an NSERC postgraduate scholarship
(E.O. Swansburg). We thank G. Lindsay and R. Parker (Environment
Canada, Atlantic Region) for assistance in selecting metal mining
facilities for this study. Access to sites on mine premises was gen-
erously granted by Noranda and Breakwater Resources.
environments. This is one of the few to report chironomid
deformity levels in cobble streams and rivers. Lastly, differ-
ences in the classification of deformities are apparent in the
literature [12,14], and this contributes to observed variation
in the incidence of deformities.
Regardless of the low incidence of deformities measured,
a consistent increase occurred at sites receiving mine drainage,
ranging from 1.25 to more than 8 times above background.
Incidences averaged across all sites receiving mine drainage
approached a doubling of that observed across all reference
sites, implying that elevated metal concentrations were indeed
associated with higher levels of deformities than are normally
encountered in these populations. Although the manifestation
of deformities has been used as an indicator of contaminant
exposure, deformities also may indicate a lower fitness of the
deformed individuals [13]. Higher body burdens of contami-
nants, slower growth and development, and lower emergence
rates have been observed in deformed individuals than in nor-
mal individuals [11,33,34,44].
A significant elevation in mentum deformities was detected
when using a conservative definition of deformity and a large
sample size. The necessary sample size is a function of the
baseline incidence of deformities and the effect size. Consid-
ering the low baseline incidence of mentum deformities, 500
individuals, not the 200 previously assumed as sufficient, were
necessary to provided adequate power to evaluate deformity
frequencies.
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Conclusions
The presence of metal mining facilities was significantly
associated with elevated metal concentrations in water, pe-
riphyton, and chironomid larvae in receiving streams. Mine-
affected sites have significantly less diverse chironomid com-
munities, with larval Tanytarsini most sensitive and larval Or-
thocladiinae least sensitive to metal enrichment. The incidence
of deformities at sites receiving mine drainage was double that
observed at reference sites.
Considering the importance of chironomids in lotic food
webs, significant alterations in community composition and
metal bioaccumulation could affect other trophic levels, par-
ticularly in the transfer of contaminants through food. Trace
metal concentrations at mine-associated streams in New Bruns-
wick affect the benthic community and thus have the potential
to alter the structure and function of these aquatic ecosystems.
This is the first study examining the incidence of defor-
mities in chironomids of metal-enriched, cobble-bottomed
streams. A significant elevation in mentum deformities was
detected when using a conservative definition of deformity
and a large sample size. Because of the low incidence of back-
ground deformity incidences observed in this study, the rec-
ommendation is made that subsequent studies employ larger
sample sizes than currently employed.
Finally, the biological impact of metal enrichment on these
streams could not have been assessed with water quality in-
formation alone. And although the approach applied in this
study effectively evaluated the impact of metal enrichment, it
was quite labor intensive. As a result, preliminary work has
been done on a laboratory bioassay incorporating field-col-
lected metal-enriched periphtyon as a food source for tradi-
tional bioassay organisms (e.g. chironomids, gammarids, and
others).
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