SEXUAL DISRUPTION IN A SECOND SPECIES OF WILD

SEXUAL DISRUPTION IN A SECOND SPECIES OF WILD

Environmental Toxicology and Chemistry, Vol. 20, No. 12, pp. 2841–2847, 2001
q 2001 SETAC
Printed in the USA
0730-7268/01 $9.00 1 .00
SEXUAL DISRUPTION IN A SECOND SPECIES OF WILD CYPRINID FISH (THE
GUDGEON, GOBIO GOBIO) IN UNITED KINGDOM FRESHWATERS
RONNY
VAN
AERLE,*†‡ MONIQUE NOLAN,§ SUSAN JOBLING,‡ LISETTE B. CHRISTIANSEN,‡ JOHN P. SUMPTER,‡
and CHARLES R. TYLER†
†Environmental and Molecular Fish Biology, School of Biological Sciences, Hatherly Laboratories, University of Exeter, Prince of Wales
Road, Exeter EX4 4PS, United Kingdom
‡Department of Biological Sciences, Brunel University, Uxbridge, Middlesex, UB8 3PH, United Kingdom
§Environment Agency, National Fisheries Laboratory, Bromholme Lane, Brampton, Cambs, PE18 8NE, United Kingdom
( Received 28 November 2000; Accepted 10 May 2001)
Abstract—To establish whether the intersex condition seen in the roach (Rutilus rutilus) in United Kingdom (UK) rivers was
species specific or a more general phenomenon in fish, evidence for sexual disruption was sought in a second cyprinid species, the
gudgeon (Gobio gobio). Gudgeon were collected from the Rivers Aire and Lea (at locations that receive high-volume discharges
of sewage treatment works [STW] effluent and that contain intersex roach) and from two still waters, and their gonads were examined
histologically for evidence of intersexuality (the simultaneous presence of oocytes and testicular tissue). Intersex gonads were found
at all sites, with the highest incidences occurring at one of the still waters (Lakeside Fisheries: 15%) and at sites on the River Aire
(Thwaite Weir, Silsden Bridge, and Knostrop: 14, 13, and 12%, respectively). In the River Lea and Longton Park Lake, the incidence
of intersexuality in gudgeon was 6%. In most cases, intersex gonads were characterized by a few primary oocytes/gonad section
in an otherwise normal testis. However, at some sites on the River Aire (Thwaite Weir and Knostrop), the intersex condition was
more severe. At Thwaite Weir, for example, more than half of the gonad in 40% of the intersex fish was comprised of ovarian
tissue. Elevated concentrations of plasma vitellogenin both in male and in intersex fish indicated that fish had been exposed to
estrogen(s). Some of the gudgeon were found at sites several kilometers downstream of any point discharge of STW effluent;
therefore, the results likely are representative of this species in wild populations found in typical UK river ecosystems. Together
with the findings in the roach, these data on the gudgeon confirm that sexual disruption in fish in UK rivers is not species specific.
Keywords—Sexual disruption
Cyprinid fish
Sewage effluent
Endocrine
Gudgeon
rence of intersexuality (i.e., the simultaneous presence of both
testicular and ovarian tissues in the gonad) in wild populations
of riverine roach (Rutilus rutilus) [21] and exposure to STW
effluents. Evidence that the intersex roach were feminized
males (rather than masculinized females) was derived from the
fact that intersex roach had elevated concentrations of plasma
VTG and that a reciprocal relationship was found between the
number of male fish (with normal-appearing testes) and intersex males caught at any one site. Furthermore, exposure of
juvenile roach to STW effluent during the period of gonadal
differentiation induces formation of an ovarian cavity (i.e.,
female duct) in male fish [22]. To our knowledge, however,
the causative agent(s) of intersexuality in wild roach have yet
to be established [21].
Information regarding intersexuality in wild fish in UK rivers is limited to studies on the roach, and nothing is known
regarding the effects (if any) of exposure to STW effluents on
sexual development in other fish species. A wide variety of
patterns for sexual differentiation and development in fish have
been found (reviewed in [23,24]); hence, the effects of endocrine-active chemicals (EACs) on different species may differ. One study has also shown that fish species can differ in
their sensitivity to steroidal estrogens (as assessed by VTG
induction) [9]. To establish whether the intersex condition in
fish in UK rivers was species specific or a more general phenomenon, evidence for sexual disruption was investigated in
a second cyprinid fish, the gudgeon (Gobio gobio). The gudgeon was chosen for study because it has a different ecological
niche and reproductive strategy compared with the roach. Gud-
INTRODUCTION
Studies on United Kingdom (UK) sewage treatment works
(STW) effluents have established that most are strongly estrogenic to fish, inducing the synthesis of the estrogen-dependent
yolk precursor vitellogenin (VTG) [1]. More recently, the phenomenon of estrogenic STW effluents has been shown to occur
in other European countries, including Norway [2], Sweden
[3], Denmark [4], Germany [5], and France [6], as well as in
the United States [7], albeit the estrogenic potency appears to
vary greatly. A wide variety of natural and synthetic compounds are discharged into the aquatic environment that are
capable of mimicking endogenous estrogens, including natural
and synthetic steroidal estrogens [3,8,9], phyto (i.e., plant)and myco (i.e., fungal)-estrogens [10], and a wide variety of
man-made industrial compounds, such as plasticizers [11], surfactants [12,13], bisphenol-A [14,15], and various pesticides
and herbicides [16–18]. Some of the chemicals responsible for
the estrogenic effects observed in caged fish exposed to STW
effluent in the UK and Sweden have been identified, and they
include the synthetic estrogen, ethinylestradiol, the natural steroidal estrogens estradiol-17b and estrone, and some alkylphenolic chemicals [3,8].
The estrogenic activity of STW effluents in UK rivers has
been shown to persist for considerable distances downstream
of the effluent discharges [19,20], and in these rivers, a relationship has been established between the widespread occur* To whom correspondence may be addressed
([email protected]).
2841
2842
Environ. Toxicol. Chem. 20, 2001
R. van Aerle et al.
Fig. 1. Locations of the sites from which gudgeon were collected. Population equivalents (PE) are a measure of the strength of the influent; a
PE of one is the amount of organic biodegradable load that has a biochemical oxygen demand of 60 g of oxygen per day. STW 5 sewage
treatment works.
geon live on the bottom of the river, close to the river sediments, and spawn at least four times within one breeding season, whereas the roach live midwater and spawn only once a
year [25,26]. Gudgeon generally spawn during May and June
at water temperatures of between 15 and 188C, but occasionally, they also spawn during late summer [25,26].
MATERIALS AND METHODS
Selected sites/rivers
The two rivers sampled for gudgeon were the River Aire
in Yorkshire (northeast England) and the River Lea (southeast
England) (Fig. 1). The River Aire receives both domestic and
industrial discharges, with a major contribution from the woolscouring industry, whereas the major discharges into the River
Lea are predominantly domestic in origin [20,27]. On the River
Aire, gudgeon were collected from a site upstream of any
known STW, at Silsden Bridge, and at three sites of increasing
distance downstream of a large STW effluent discharge at
Marley (population equivalent [PE], 112,000)—namely,
Crossflats, Thwaite Weir, and Knostrop, respectively. Thwaite
Weir was also downstream (13 km) of a very large STW at
Esholt (PE, 674,717), and the site at Knostrop (Swillington
Bridge) was downstream of an additional STW at Knostrop
(PE, 847,000). On this stretch of the River Aire, between 5
and 30% of the flow of the river is made up by STW effluents.
The locations of the river sampling sites were the same as
those for previous studies on intersexuality in wild roach [21].
A major consideration in river site selection was that physical
barriers existed between the sites, thus severely limiting the
movement of fish between the designated sites [21]. On the
River Lea, gudgeon were obtained from one site only, Harpenden, which is downstream of East Hyde STW (PE,
142,000). The mean percentage flow of the River Lea made
up of STW effluent discharge at this sampling site is between
50 and 80%, and it may rise to 100% during the summer
months. We were not able to find any UK rivers that were
inhabited by gudgeon but did not receive STW effluents; therefore, two lakes (Longton Park Lake, near Stoke-on-Trent, and
Lakeside Fisheries in The Midlands) were selected as control
sites, because they did not receive any STW effluent (Fig. 1).
Longton Park Lake was the only site that received no sewage
effluent whatsoever. Lakeside Fisheries received some sewage
effluent directly from cesspool overspills.
Fish sampling
The gudgeon were electrofished from rivers during September 1995 and August 1998 and Seine-netted from the control lakes between September 1997 and March 1998 (Table 1).
To obtain sufficient numbers of fish, it was sometimes necessary to sample a site on two or even three occasions. The
fish were anesthetized in 2-phenoxyethanol (1:2,000, v/v), and
blood samples were collected from the caudal sinus into 1-ml
heparinized syringes. The blood was centrifuged and the resulting plasma withdrawn and frozen on dry ice for transportation to the laboratory. Plasma was maintained frozen at
2808C before VTG analysis. Total length, total weight, and
gonadal weight were determined for each of these fish, and
scales were removed for age determination. The gonadosomatic index (GSI) was calculated as gonadal weight/(total body
wt 2 gonadal wt) and expressed as a percentage. The gonads
were fixed in Bouin’s fixative for 6 h and transferred to 70%
methylated spirit in preparation for histological processing.
Gonadal histology
Gonads from each fish were divided into three equal pieces,
and transverse portions (thickness, 3–5 mm) were cut from the
center of each piece to provide a total of six portions per fish.
The portions were then dehydrated and embedded in paraffin
wax, and sections were cut at 5 mm, collected on slides, and
stained with Harris’s hematoxylin and eosin (Merck Eurolab,
Poole, Dorset, UK). The slides were mounted with DPX mountant (Merck Eurolab, Poole, Dorset, UK) and analyzed by light
microscopy. The degree of intersexuality was classified according to the mean number of oocytes per section (from six
transverse sections, one from each of the anterior, mid, and
posterior pieces of each of the two gonads). The physical appearance of the gonadal duct was not used in quantifying sexual disruption, because in male cyprinid fish, the gonadal duct
varies in shape and form during sexual maturity [28]. Thus,
because the fish were obtained at different times during the
year, it was very difficult to directly compare duct development
between fish from the different sampling sites (see Results).
The prevalence of intersexuality in this paper is presented as
a percentage of the population sampled at any given site. The
rationale for this is that we do not know if the intersex condition at the different sites occurred as a consequence of feminization of male fish, masculinization of female fish, or some
combination of both.
Environ. Toxicol. Chem. 20, 2001
Sexual disruption in wild cyprinid fish
2843
Table 1. Details on the river and still water sites from which gudgeon were sampled, showing the influent strength as population equivalents
(PE) of the nearest upstream sewage treatment works (STW) discharging into the river, sampling dates, and the fish caught
PE of STW
immediately Dilution
upstream
factor
River/still water
site/location
River Aire, Silsden Bridge
—
—
Gonad- Total Age in years
no. of
(mean
al
fish
[range])
Sampling date
sexa
Sept. 1995
Aug. 1998
River Aire, Crossflats
112,000
17.6
Sept. 1995
River Aire, Thwaite Weir
674,717
7.1
Sept. 1995
May 1998
Aug. 1998
River Aire, Swillington Bridge
847,000
3.7
Sept. 1995
Aug. 1998
River Lea, East Hyde
130,393
1.8
Sept. 1995
Oct. 1998
Lakeside Fisheries
—
—
Feb. 1998
Mar. 1998
Longton Park Lake
—
—
Nov. 1997
Fork length
(cm)
Weight (g)
14.0 6 1.1
14.2
21.6
10.9 6 0.9
3.2
16.0 6 3.4
F
I
M
F
I
M
5
1
1
3
1
4
2.8 (2–3)
2.0 (2)
3.0 (3)
NMc
NM
NM
11.5 6 0.3b
11.5
13.5
9.7 6 0.3
7.0
11.1 6 0.8
F
M
F
I
M
F
I
M
F
I
M
31
1
6
5
10
—
2
21
13
8
40
2.9 (2–4)
3.0 (3)
2.8 (2–3)
3.0 (3)
2.9 (2–3)
—
4.0 (4)
3.8 (3–5)
NM
NM
NM
12.3
13.5
14.3
14.4
14.5
—
13.0
12.7
11.6
11.1
12.0
6 0.2
6
6
6
6
6
F
I
M
F
I
M
7
2
7
14
4
15
3.0 (2–4)
2.5 (2–3)
3.0 (1–4)
3.1 (1–4)
3.8 (3–4)
3.5 (2–5)
12.9
13.0
14.0
10.6
13.3
12.0
F
I
M
F
I
M
19
3
19
22
2
18
2.7 (1–4)
3.0 (2–4)
2.4 (1–3)
3.8 (3–5)
4.5 (4–5)
3.9 (3–5)
F
I
M
F
I
M
12
7
4
20
3
20
F
I
M
14
3
33
6 0.7
0.0
0.2
0.7
0.8
0.3
16.5
21.5
24.8
27.0
25.4
—
23.8
22.8
19.2
18.1
21.3
6
6
6
6
6
0.3
1.0
2.9
3.9
1.5
6
6
6
6
6
6
0.4
0.5
0.6
0.5
2.3
0.3
20.3
20.2
27.5
16.6
19.6
21.7
6
6
6
6
6
6
1.2
2.1
3.1
1.9
1.8
1.6
15.3
16.2
15.4
11.6
12.3
11.7
6
6
6
6
6
6
0.4
0.2
0.3
0.3
0.3
0.2
29.9
36.8
30.6
17.0
21.2
18.1
6
6
6
6
6
6
1.7
4.7
1.6
1.2
2.4
0.9
3.0 (2–4)
2.8 (2–4)
3.0 (3)
3.8 (3–5)
3.0 (3)
3.4 (3–5)
11.0
10.8
10.1
11.4
10.3
11.0
6
6
6
6
6
6
0.3
0.4
0.3
0.2
0.4
0.2
14.8
13.5
11.0
17.4
12.9
14.7
6
6
6
6
6
6
1.8
1.4
1.0
1.3
1.3
0.9
1.6 (1–2)
2.0 (2)
1.6 (1–2)
10.5 6 0.2
10.5 6 0.3
10.3 6 0.1
6 0.4
6 0.4
6 0.2
6 1.7
6 2.4
6 1.0
11.1 6 0.4
12.0 6 1.4
11.1 6 0.3
I 5 intersex, F 5 female, M 5 male.
Expressed as mean 6 standard error of the mean.
c NM 5 not measured.
a
b
Measurement of plasma VTG
Concentrations of VTG in plasma samples were determined
using an enzyme-linked immunosorbent assay for carp (Cyprinus carpio) VTG that has been validated for measuring
gudgeon VTG [29,30].
Statistical analyses
Differences in plasma VTG concentrations were analyzed
using analysis of variance (ANOVA) with SigmaStaty (SPSS
UK, Woking, Surrey, UK). Data were not normally distributed,
and a Kruskal-Wallis one-way ANOVA on ranks was performed. In cases of significant differences between the groups,
post-hoc tests according to Dunn’s method were used to identify
groups that differed significantly from the control group (i.e.,
Longton Park Lake), which received no sewage discharges.
RESULTS
Meristic measurements
In total, 400 gudgeon were collected from the different
sampling sites. The number of fish sampled from the different
sites and their meristic measurements are shown in Table 1.
The mean (6 standard error of the mean) length of the fish
ranged between 10.4 6 0.1 cm (Longton Park Lake) and 13.5
6 0.2 cm (River Lea). The mean weight of the fish ranged
between 11.2 6 0.2 g (Longton Park Lake) and 24.1 6 1.0 g
(River Lea). The age range of the fish caught was between
greater than one and five years old, and the most common year
class was greater than three. Fish of all ages were maturing,
indicating that gudgeon were fully mature at the sites studied
in the second (i.e., greater than one) year of their life.
Gonadal development
All gonads dissected from the fish appeared macroscopically to be either male (i.e., testis) or female (i.e., ovary).
Histological examination of the gonads, however, revealed that
a proportion of the male fish were, in fact, intersex, as defined
by the simultaneous presence of both testicular tissue and ovarian tissue, specifically oocytes. Histological sections of a normal
testis, a normal ovary, and intersex gonads with either primary
and/or secondary oocytes are shown in Figure 2.
2844
Environ. Toxicol. Chem. 20, 2001
R. van Aerle et al.
Fig. 2. Histological sections of (a) a normal male testis (3214), (b) a normal female ovary in vitellogenesis (354), (c) an intersex gonad
containing primary oocytes only (category A, 3214)), and (d) a grossly intersex gonad containing both primary and secondary (vitellogenic)
oocytes (category C, 354)). CA 5 cortical alveolus–stage oocyte, PO 5 primary oocyte, Sc 5 spermatocytes, Sg 5 spermatogonia, SO 5
secondary (vitellogenic) oocyte, Sz 5 spermatozoa, St 5 spermatids, Tt 5 testicular tissue, * 5 degenerative tissue.
Intersex gudgeon were found at all the sites studied (Fig.
3a). The incidence of intersexuality was highest at Lakeside
Fisheries (15%), followed by Thwaite Weir, Silsden Bridge,
and Knostrop, at which the incidences of intersexuality were
14, 13, and 12%, respectively. No intersex fish were found at
Crossflats, although 97% of all the fish sampled (n 5 32) at
this site were female. In both the River Lea and Longton Park
Lake, the incidence of intersexuality was 6%.
Intersexuality in each fish was divided into the following
categories according to its severity. Category A contained fish
with a maximum of five primary oocytes per section. Category
B contained fish with more than five primary oocytes per section. Category C consisted of fish with both primary and secondary oocytes comprising a high proportion (.50%) of the
gonad. The classification for the degree of intersexuality used
with the gudgeon in this study differed from that established
for the roach [21], because unlike in the roach, in which a
gradient was observed in the severity of intersexuality, only
two basic patterns of intersexuality were observed in the gudgeon. In the first pattern (i.e., categories A and B), the gonad
appeared as a normal testis, with the exception of primary
oocytes being located at the periphery of the gonad and/or in
clusters within the gonad. In the second (i.e., category C), the
intersex gonad contained a high proportion of female tissue
($50%) interspersed within the testicular tissue. These intersex gonads generally were oddly shaped and contained both
primary and secondary oocytes. However, oocytes more mature than those in early to mid-vitellogenesis were not observed. Often, the gonads that were grossly intersex contained
eosinophilic degenerative tissue (Fig. 2).
At all sites except Crossflats, category A intersex fish were
observed, varying in number from 2 (of 15) fish at Silsden
Bridge to 10 (of 36) fish at Lakeside Fisheries. At Thwaite
Weir and at Knostrop, category B intersex fish occurred (1 of
Fig. 3. (a) Percentage of male, female, and intersex gudgeon captured
at the different sampling sites. Numbers above the bars give the numbers of fish in each group. (b) Numbers of fish in the different intersex
categories.
Sexual disruption in wild cyprinid fish
Fig. 4. Gonadosomatic index (GSI) in female, male, and intersex
gudgeon collected from the different sampling sites. Numbers are
given as the mean 6 standard error of the mean (error bars). Statistical
analyses of the GSI data were complicated by the fact that fish were
collected from some of the field sites at different times of the year.
15 and 3 of 6 fish, respectively). The greatest severity of intersexuality was found in gudgeon collected from Thwaite
Weir, for which 6 of 15 intersex fish appeared to be almost
completely sex-reversed (category C) (Fig. 3b).
The GSI in female fish from the different sites ranged between 2.4 6 0.2 (Thwaite Weir) and 10.6 6 0.6 (Lakeside
Fisheries) (Fig. 4). No significant differences were observed
in the GSI in male fish collected from the different sites, which
ranged between 0.5 (Crossflats) and 2.2 6 0.2 (Lakeside Fisheries). The GSI in intersex fish was either similar to that in
males or intermediate between that in the males and that in
the females, ranging between 1.3 6 0.3 (Harpenden) and 2.0
6 1.3 (Silsden Bridge).
Plasma VTG
Plasma concentrations of VTG in the gudgeon sampled are
shown in Figure 5. The plasma concentration of VTG in female
fish was higher than that in males and intersex fish (p , 0.05
throughout) and ranged between 4.2 6 3.5 mg/ml (Silsden
Bridge) and 1.0 6 0.2 mg/ml (Lakeside Fisheries). In males,
plasma VTG concentrations were very low (,20 ng/ml) only
at Longton Park. Plasma VTG concentrations in males from
the river sites ranged between 0.2 mg/ml (Crossflats) and 21.2
6 12.5 mg/ml (Knostrop) and were significantly higher in
males from Thwaite Weir and Knostrop than in males from
Longton Park Lake (p , 0.05). At Lakeside Fisheries, male
gudgeon also had higher plasma VTG concentrations (5.1 6
3.2 mg/ml) than those sampled from Longton Park. Plasma
concentrations of VTG in intersex fish were generally similar
to those in males at the different sites and ranged between
10.0 6 0.0 ng/ml (Longton Park) and 22.0 6 8.1 mg/ml
(Thwaite Weir).
DISCUSSION
This study demonstrates the presence of the intersex condition in a second species of cyprinid fish, the gudgeon, in
UK freshwaters. The low incidence of intersexuality in wild
gudgeon (6%) seen at Longton Park Lake (a control site receiving no known effluent discharges) was perhaps not surprising given that intersexuality has similarly been reported at
Environ. Toxicol. Chem. 20, 2001
2845
Fig. 5. Plasma vitellogenin concentrations in female, male, and intersex gudgeon collected from the different sampling sites. Numbers
are given as mean 6 standard error of the mean (error bars). Statistical
analyses of plasma vitellogenin were complicated (especially for the
females) by the fact that fish were collected from some of the field
sites at a different time of the year than fish were collected at the
others. *Significant differences (p , 0.05) from the male fish at Longton Park.
low levels in other cyprinid fish, including the roach [21,31]
and bream (Abramis brama) [32]. Whether the low level of
intersexuality in wild gudgeon (as in roach) in UK waters is
a natural phenomenon or has resulted from exposure to chemicals (e.g., in the case of Longton Park Lake, as a possible
consequence of exposure to diffuse sources such as pesticide/
herbicides from agricultural runoff) is not known. A previous
study documenting the reproductive biology of a captive stock
of gudgeon reported an incidence of intersexuality of only 1%
[33]. The reason for the higher incidence of intersex fish (15%)
at Lakeside Fisheries compared with Longton Park is unclear,
but at Lakeside Fisheries, overspills from private cesspools
and industrial discharges are suspected to occur.
The major difference in gudgeon from the various sites on
the River Aire was in the degree rather than the incidence of
intersexuality. At most sites, only intersex fish in category A
(less than five primary oocytes per section) occurred. At Knostrop and Thwaite Weir, however, intersex gudgeon with a higher level of gonadal disruption (category B and/or C) were also
found. The most severely affected fish were those collected
from Thwaite Weir, in which the gonads (i.e., testes) in 6 of
15 intersex fish were comprised of more than 50% (and often
more than 75%) ovarian tissue. The distribution pattern of
intersexuality in gudgeon living in the River Aire was similar
to that established for the roach (in which the greatest severity
in the intersex condition occurred at Thwaite Weir [21]). In
roach sampled on the River Aire, oocytes in intersex fish were
observed that were in the mid to late stages of vitellogenesis.
In contrast, the most mature oocytes observed in intersex gudgeon were in early vitellogenesis. This difference between
species is probably a function of the time of year at which the
gudgeon were collected rather than of an inability by oocytes
in intersex gudgeon to reach full maturity. Vitellogenesis in
the gudgeon starts around May, depending on the water temperature [26,33], and most fish in this study were sampled
earlier during the reproductive season than this. The low incidence of intersex gudgeon from Harpenden on the River Lea
mimics the situation seen in the roach [21]. The percentage
2846
Environ. Toxicol. Chem. 20, 2001
flow of the river comprised of STW effluent at the sampling
site on the River Lea was higher (a mean annual figure of
between 50 and 80%) compared with the sites on the River
Aire (between 5 and 30%), yet the incidence (and severity) of
the intersex condition was higher in gudgeon in the River Aire.
The reasons for this are not known. However, the influent
entering Harpenden STW is predominantly domestic in origin,
whereas the influents into the STW on the River Aire contain
considerable inputs from industry. A possible link between a
high severity of the intersex condition and exposure to effluent
from STW that receive a high level of industrial influent is in
common with that seen for roach living in the Rivers Aire and
Nene [21]. Furthermore, studies on a marine flatfish, the flounder (Platichthys flesus), in UK estuaries have shown a higher
incidence of intersexuality in fish caught in estuaries receiving
appreciable industrial discharges [34]. A recent study that exposed juvenile roach to a treated STW effluent with primarily
a domestic influent during the time of sexual differentiation
(i.e., for a period of five months) did not induce female sex
cells (i.e., oocytes) in the testes of male fish [22], reinforcing
the suggestion that intersexuality (as defined by the presence
of both oocytes and male sex cells within the gonad) may
result from exposure to chemicals other than (or in addition
to) steroidal estrogens.
Both the proportions of gudgeon that were intersex and the
severity of the condition were not as high as those found in
the roach at the same study sites [21]. This is perhaps surprising given that the gudgeon is a demersal species that lives
on the river/lake bottom and, therefore, likely has greater exposure to EACs that concentrate in the sediment, which many
of the known EACs do [27], whereas roach live at midwater.
Most gudgeon collected, however, were caught in stretches of
water that had stony beds rather than soft sediments. Furthermore, predisposition to endocrine disruption depends on many
factors in addition to their habitat, including diet (e.g., roach
feed on the detritus from STW effluents, but gudgeon do not),
bioaccumulation of chemicals, and the pathway/effectiveness
of chemical metabolism. Recent studies have shown that the
pathways of metabolism for octylphenol, a known estrogen
mimic, differ in roach compared with the rainbow trout (Oncorhynchus mykiss) [35]. Furthermore, fish show different sensitivities to chemicals that are known to be able to disrupt
gonadal development, such as steroidal estrogens [9].
Analyses of the GSIs in gudgeon from the different sampling sites are complicated by the fact that fish were collected
at various times during the year. The GSI data, however, suggest that the gonad size in intersex fish was similar to that in
males, reinforcing the hypothesis that intersex fish result from
the feminization of males. That the fish collected from Crossflats on the River Aire were almost exclusively females (97%
of 32 fish) is intriguing. Some fish species school in monosex
populations for periods of the year, with mixing of the sexes
occurring only during the time of spawning. Not enough is
known regarding the reproductive biology of the gudgeon to
know whether this occurs in this species. However, at no other
sites sampled on the River Aire, or elsewhere, were monosex
populations seen, indicating that the situation at Crossflats was
probably unusual. We cannot rule out the possibility that this
almost exclusively female population of gudgeon was a consequence of the complete sex reversal of some of the males
at this site.
The maximum concentration of VTG measured in maturing
female gudgeon (1 mg/ml) is similar to that seen in other
R. van Aerle et al.
cyprinid fish with similar patterns of ovarian development [36].
In male fish from the control site at Longton Park Lake, plasma
VTG concentrations were very low indeed (,20 ng/ml), confirming that VTG is predominantly a female plasma protein
in this species. Analyses of plasma VTG concentrations in fish
from the different sampling sites, as with analyses of the GSI
data, are complicated by the fact that fish were collected at
different times of the year. Nevertheless, the higher concentrations of plasma VTG in males and intersex fish at all the
riverine sites confirm that the gudgeon had been exposed and
responded to estrogenic substances in those rivers. In the intersex gudgeon, category C fish (the most severely intersex)
had higher plasma titers of VTG compared with fish having
less disrupted gonads (categories A and B) (p , 0.001). An
elevated plasma concentration of VTG in male gudgeon at
Lakeside Fisheries confirmed that the water body at this site
was contaminated with environmental estrogens.
In summary, the observations on intersexuality, taken together with the GSI data and elevated plasma VTG concentrations, provide evidence that populations of wild gudgeon
inhabiting rivers and some still waters in the UK are subject
to endocrine disruption, which, in turn, results in the disruption
of sexual differentiation/development, probably as a result of
exposure to estrogenic and/or other gender-disrupting chemical
contaminants. Furthermore, some of the gudgeon were found
at sites that were several kilometers downstream of any point
discharge of STW effluent; therefore, the results are probably
representative of this species in wild populations found in
typical river ecosystems. However, what is also clear from this
work is that, despite an extensive field study (.400 fish) we
cannot be definitive in our statements regarding the prevalence
and causation of intersexuality in gudgeon in UK rivers. Very
extensive field surveys will be required to more fully appreciate the condition of intersexuality in wild fish populations.
Acknowledgement—R. van Aerle and S. Jobling were funded by
grants from the Department of the Environment, Transport and the
Regions and C.R. Tyler and J.P. Sumpter by the Natural Environmental
Research Council. We would like to thank the UK Environment Agency Fisheries Teams for catching the fish and the Environment Agency
as well as members of the Fish Research Group at Brunel University
for their excellent assistance throughout the field work.
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