The impact of sewage sludge exposure on the reproduction of the sand goby, Pomatoschistus minutus

PII:SO269-7491(96)00019-X

Environmental Pollution, Vol. 93, No. 1, pp. 17-25, 1996 Copyright 0 1996 Elsevier Science Ltd Printed in Great Britain. All rights reserved 0269-7491/96 $15.00+0.00

THE IMPACT OF SEWAGE SLUDGE EXPOSURE ON THE REPRODUCTION OF THE SAND GOBY, POMATOSCHISTUS MINUTUS Colin P. Waring,+ Ronald M. Stagg, Karen Fretwell,* H. Anne McLay & Mark J. CostelloS Scottish Ofice Agriculture and Fkheries Department, Marine Laboratory, P.O. Box 101, Victoria Road, Aberdeen AB9 8DB, UK (Received 24 November 1995; accepted 1 February

Abstract Adult sandgobies were exposed to 0.1% sewage sludge for 19 weeks prior to the end of spawning. Exposure to sewage sludge had a significant efiect on male mortality rates but no significant eflects on the gonadosomatic index of males or females. There were no major effects of sludge exposure on testes androgen content or on testes release of androgens after in vitro gonadotrophin stimulation. Fecundity and the number of larvae produced were not sign$cantly aflected by the sludge exposure. There was a tendency for eggs and larvae from sludge-exposed females to have a higher mortality rate. At a population level, however, there was a major reduction in the number of eggs and larvae produced in the sludge-exposed population which reflected a failure of some females to spawn. Of the larvae produced, &I--70% from sludge-exposed parents were lighter and had a larger yolk-sac volume compared to larvae from non-exposed parents which may have indicated impaired yolk utilisation. Copyright 0 1996 Elsevier Science Ltd

1996)

Most studies to date have concentrated on iteroparous species where individuals can spawn over multiple seasons and/or spawn in groups with eggs and sperm released into the water en masse. It is, therefore, difficult to measure the impact of a persistent contaminant on the reproductive success of an individual fish spawning naturally, as opposed to being artificially stripped of gametes, and the effect on an individual’s lifetime reproductive effort. Semelparous species which exhibit paired matings are much more conducive to the measurement of these parameters. One such species is the sand goby, Pomatoschistus minutus, which is abundant in inshore areas of the European continental shelf. Sand gobies participate in paired spawning: males are territorial and guard a nest-site and actively court females which are led into the nests where the eggs are laid on the underside of the nest roof (Hesthagen, 1977; Magnhagen & Kvarnemo, 1989; Lindstriim & Wennstriim, 1994; Kvarnemo, 1995). The short life-cycle of this species means that the probability that a life-time’s reproductive effort occurs in one season is high, since very few individuals survive beyond 22 months (Healey, 1971). To achieve fitness, the reproductive strategy of a species must maximise both the reproductive potential of adults and the survival and viability of their offspring to reach maturity. In the study reported here, this was assessed in sand gobies exposed to sewage-sludge by measuring (i) the reproductive condition of adults, (ii) the quantity of eggs and larvae produced from known pairings, and (iii) estimates of larval viability by measuring larval lengths, weights and yolk-sac volumes. Sewage sludge contains many metal and organic contaminants which may be present at marine disposal sites in significant concentrations, depending on whether the site is accumulative or dispersive (reviewed by Costello & Read, 1994). The use of sewage sludge gives an opportunity to study the effects of a complex mixture of environmentally relevant cantaminants acting simultaneously. The sludge concentrations used in the experiments were based on field measurements at sewage sludge dumping sites (reviewed by Costello & Read, 1994) and have been used in previous studies on the effects of sludge exposure on dab (Limanda linuznda) growth (Houlihan et al., 1994) and immunocompetence (Secombes et al., 1991, 1992).

Keywords: Reproduction, sewage sludge, larvae, Pomatoschistus minutus, fecundity.

INTRODUCTION There is increasing concern regarding environmental contaminants impacting on the reproductive success of fish populations. In a series of studies it has been shown that exposure of various teleost species to lake waters containing bleached kraft mill effluent has led to reduced gonad size, decreased plasma gonadal steroid concentrations, altered age to maturity, decreased egg size and decreased larval growth rates (McMaster et al., 1992; Munkittrick et al., 1992a,b). Similarly, in a marine flatfish, the sole Parophrys vetulus, inhabiting contaminated estuaries there has been a significant impact on a whole suite of reproduction-related parameters (Johnson et al., 1988; Casillas et al., 1991; Collier et al., 1992). *Deceased, and to whose memory this work is dedicated. TPresent address and address for correspondence: Ministry of Agriculture, Fisheries and Food, Fisheries Laboratory, Pakefield Road, Lowestoft, Suffolk NR33 OHT, UK. tPresent address: Environmental Science Unit, Trinity College, Dublin 2, Republic of Ireland. 17

18

C. P. Waring et al.

MATERIALS AND METHODS Fish collection and maintenance During November and December 1989 adult sand gobies (weighing between 2 and 6 g) were collected by beam trawl from Firemore Bay, Loch Ewe, Ross and Cromarty, Scotland. The fish were maintained in the laboratory in four 1100 litre tanks (50-60 fish per tank) containing 1 litre of clean natural sediment and supplied with seawater (salinity: 3335 parts per thousand) at a flow-rate of 5 litres min-‘. Fish were fed to satiation once a day with mysids and finely chopped squid. Fish were exposed to a natural photoperiod and water temperature fluctuations (from 6.9”C in December to 14.1”C in June). Water pH was in the range 7.9-8.1 and dissolved oxygen (DO) was >95% saturation (both parameters measured twice per week). Water total ammonia was measured occasionally and was in the range 0.14.2 mg litre-‘. Sewage sludge The sewage sludge used in these experiments was from a single batch from the Shieldhall sewage works, Glasgow and was stored at 4 f 1°C. The sludge is derived from both industrial and domestic sources and contains a variety of heavy metal and organic contaminants as shown in Table 1. It is not known if cold storage altered sludge composition. The organics were analysed using gas chromatography with mass spectrometry or gas chromatography with electron capture detection and the metals using standard atomic absorption analysis techniques. Full details of the methodologies are reported by Roddie et al. (1990). Fish were exposed to sewage sludge from the end of January 1990 onwards. The sludge was added to two of the tanks once on each week day (Monday-Friday) to give an initial concentration of 0.1% wet sludge (v/v). With a 90% flushing time of 19 h, the nominal daily average concentration was 0.032%. After sewage sludge addition, the water DO concentrations did not fluctuate significantly from 93-95% saturation and water total ammonia concentrations peaked at 0.5-0.6 mg litre-’ 1 h after addition and declined exponentially to 0.14.2 mg litre-’ within 4 h. To control for disturbance, the non-sludge exposed population had an equivalent volume of seawater added to their two tanks. Mortalities were recorded daily throughout the course of the experiment. Gonads were dissected and weighed and the gonadosomatic index (GSI) was determined (gonad weight/body weight x 100%). Effect of sludge exposure on male reproductive PbysiologY From March to June 1990, male gobies were sampled to evaluate any effect of sewage sludge exposure on reproductive physiology. The small size ( < 6 g) and limited numbers of sexually mature males precluded measurement of plasma gonadal steroid concentrations. Therefore testes androgen concentrations and testes responsiveness to in vitro gonadotrophin (GTH) stimu-

lation were evaluated as indices of male reproductive physiology. Not enough females were available to allow for a comparable study. For testes androgen concentrations, males were periodically sampled from the tanks and anaesthetised in MS 222 (0.1 g litre-I), killed, and the testes removed, weighed and snap frozen on dry-ice and stored at -20°C. Testes were homogenised in 1 ml ethyl acetate and centrifuged at 5000 g for 5 min and the supematant was removed. This procedure was repeated twice and the pooled ethyl acetate extracts were dried under a stream of nitrogen. The ethyl acetate was replaced with 1 ml of 0.1 M phosphosaline buffer (pH 7.0) and the testosterone and 1 1-ketotestosterone concentrations were measured using radioimmunoassay (RIA) as described by Simpson and Wright (1977) and Zhao and Wright (1985). In March (after 8-9 weeks of exposure to sewage sludge), testes were removed from eight control and six sludge-exposed males to evaluate the responsiveness of testes to GTH stimulation. Testes were removed on ice, minced to approximately 1 mm3 pieces, weighed and Table 1. Concentrations of measured contaminants sewage sludge from Shieldhall

Organics (ng gg’ dry wt) cr-Hexachlorocyclohexane /I-Hexachlorocyclohexane y-Hexachlorocyclohexane Hexachlorobenzene Aldrin Dieldrin Endrin op DDE

31 288 1079 133 145 519 28 18

PP DDE

64

PP DDD

592

op DDD op DDT

PP DDT

Heptachlor cis-Heptachlor epoxide a-Endosulfan D-Endosulfan PCB C28 PCB C52 PCB Cl01 PCB Cl18 PCB Cl38 PCB Cl53 PCB Cl80 cis-Permethrin trans-Permethrin Cypermethrin Fenvalerate Deltamethrin Metals @g g-’ dry wt) Cadmium Chromium Copper Lead Nickel Zinc Mercury Solids (% wet wt)

53 ~6 4

810 < 2.5 18 158 56 156 34 37 676 68 5 2343 2422 <20 < 10 < 10

3 442 335 572 41 744 1.5 2.45

in the

Sewage sludge and reproduction of the sand goby

split into three equal portions and placed in sterile plastic petri dishes containing 5 ml of incubation medium [comprising (g litre-‘): 7.5 NaCl, 0.23 KCl, 0.07 MgS0+7H20, 0.2 MgC12.6Hz0, 0.5 CaC12, 1.O glucose, and 0.02 M HEPES and 200 IU ml-’ penicillin and streptomycin, pH 8.01. Partially purified Pacific salmon GTH @GA-GTH; Syndel Laboratories, Vancouver, Canada) were added at 0, 1.0, or 2.5 pg ml-’ and the testes were incubated for 24 h at 10 f 1°C. A 1 ml aliquot was then removed and extracted with ethyl acetate and the testosterone and 1I-ketotestosterone concentrations measured using the RIAs described above. Spawning studies At the end of March 1990, fish of both sexes were removed from the stock tanks and transferred to a glass aquaria system. Individual males (weight range: 2-6 g) were randomly allocated to their own 12.5 litre aquarium containing 200 ml clean sediment with a water flow-rate of 250 ml min-I. Each male in a tank was supplied with a nest of either a broken clay pot or a large bivalve shell. Two females (3-6 g) were assigned for each male and kept together in separate aquaria. All aquaria were aerated and the water quality parameters both before and after sludge addition were similar to those of the 1100 litre stock tanks described above. Females were individually presented to a male on a 47 day rota system and nests were checked once daily for eggs. On discovery of eggs, the female was removed and its length and weight recorded. The male was then provided with a new nest and its second female. Both male and female gobies which had previously been exposed to sludge in the holding tanks continued to be treated with sludge in the glass aquaria. Sewage sludge-exposed males were presented only with sludge-exposed females. Both sexes were fed daily throughout the spawning period. Eggs and larvae Batches of eggs, still adhering to the nest, were transferred to 2 litre glass beakers containing 1.5 litres filtered (1 pm mesh) seawater at 10 f 1°C under continuous illumination and aeration. The water was renewed daily with filtered seawater. Water DO was checked twice per week and was always greater than 93% saturation. When hatching commenced, the aeration was reduced to prevent damage to the larvae and a 50% renewal of seawater was carried out daily. Under these conditions, water DO was maintained at > 85% saturation. When all hatching was completed, the number of eggs laid in each batch and the number of dead eggs remaining were counted under a dissecting microscope. The number of larvae hatched in each 24 h period was recorded (termed a ‘hatch-day’ group) for each batch of eggs produced. The larvae from each hatch-day were then gently transferred to separate 1 litre plastic beakers containing 750 ml of filtered seawater. Twenty larvae from each hatch-day group from each batch of eggs were randomly sampled and sacrificed for measurements of larval lengths, weights and yolk-sac volumes.

19

The larvae used for weighing were individually washed in glass-distilled water for 30 s, blotted dry and placed on a Perkin-Elmer Ad-2Z autobalance, and measurements were taken at 30, 60 and 90 s intervals and the initial weight at 0 s estimated using regression analysis. Larvae for length and for yolk-sac volume measurements were anaesthetised in 10 mg litre-’ MS222 and placed on a microscope graticule. Total lengths were measured and the yolk-sac volumes estimated by multiplying the length along the greatest dimension of the yolk-sac by the height (or measurement perpendicular to the long axis) and converting the area obtained in mm2 to nl using the formulae of Ryland et al. (1975). Statistical analysis Adult mortality rates were analysed using probit analysis and linear regressions. The GSI data and testes androgen contents were analysed using a 2-way analysis of variance with time and treatment as factors, and the in vitro androgen secretion data were analysed using a 2-way analysis of variance with treatment and GTH dose as factors. When ANOVAR indicated significance, Student-Neuman-Keuls tests were used as a multiple range test. Egg and larval production from batches of eggs were analysed using students t-tests. Larval lengths, weights and yolk-sac volumes were analysed using t-tests to compare control and sludge-exposed values for each hatch-day group. In all cases, significance was accepted when p < 0.05.

RESULTS Effect of sewage sludge on adult gobies Cumulative mortality rates in adult female sand gobies were around 30% at the end of the experimental period (Fig. 1) and there was no effect of sewage treatment. Control males had a 40% mortality rate at the end of the experiment whereas males exposed to sewage sludge had a higher final mortality rate of 60% (Fig. 1). For both males and females there was a significant increase in the GSI as the reproductive season progressed (Fig. 2) and for both sexes sewage sludge exposure had no significant impact on their GSI. Sludge-exposed males had a significantly higher GSI in April compared to controls but not in other months (Fig. 2). Testes testosterone and 1 1-ketotestosterone contents are shown in Table 2. For both androgens, exposure to sewage sludge had no effect on the concentrations measured in the testes. However, whilst there was no difference in the basal in vitro release of testosterone or 1l-ketotestosterone from testes, testes from sludge-exposed males released a significantly lower amount of testosterone at the lowest GTH concentration compared to controls (Table 3). However, at the highest GTH concentration this difference was not apparent, nor was there any difference in the 1 1-ketotestosterone released from testes under GTH stimulation between treatments (Table 3).

20

C. P. Waring et al.

Reproductive success

The number of eggs laid, how many were fertile and the larvae produced by individual females are shown in Table 4. For virtually all of these parameters there was a trend for a reduction in the numbers and survival of eggs and larvae from the sludge-exposed population, but none of these individual differences were statistically significant. Sixteen control goby females were available for breeding and laid a total of 12 batches of eggs over an 8 week period out of 11 weeks of experimentation. Only one female produced a second batch of eggs. Twenty sludge-exposed females were available for spawning and they produced six batches of eggs in a 2 week period out of the 11 weeks available. None of these females produced a second batch of eggs. The sewage sludgeexposed females that did not spawn contained eggs and had a normal GSI value. When the numbers of eggs and larvae produced are expressed per unit (g) female available to spawn in the population (i.e. including those that did and did not spawn) then a dramatic reduction in both the numbers of eggs and larvae produced from the sludge-exposed population was apparent (Fig. 3).

20

FEMALE 15

g +

7

10

z

5

0

n

rrlov

DeC Jan Feb Mar Apr lu

Jun

Month

Larvae

5

Not all eggs hatched at the same time and larvae appeared from a batch of eggs over a 7 day period (mean value being 6.8 days for both the control and sludge-exposed populations). However, 6&80% of larvae hatched within the first 2 days and there was no significant difference in the larval hatching profiles from the control and sludge-exposed populations (Fig. 4). Larval lengths did not significantly differ between hatchday groups from control and sludge-exposed populations except in the day 5 group when larvae from sludgeexposed parents were significantly shorter (Fig. 5(a)).

4

MALE

3 z ;j ci 2

1

0

I;

~

Nov Dee Jan Feb Mar I 4pr May Month

Fig. 2. Female and male goby GSI data from the period November-June. Open bars represent controls and filled bars represent the sludge-exposed group. Data are means ( f SE) of S-10 observations. *p < 0.05 compared to controls.

Table 2. Monthly variations in testis testosterone and llketotestosterone contents in control aud sludge-exposed male sand gob&. AU values are expressed as pg steroid mg-’ testis and represent tbe means f SE of 5-15 observations

Spawning Period

5

10

15

20

Testosterone

1 1-Ketotestosterone

Weeks of Experiment

Fig. 1. Cumulative mortality rates for male and female sand gobies throughout the course of the experiment. Squares represent sludge-exposed males, circles represent control males, normal triangles represent sludge-exposed females and upside-down triangles represent control females. The period of spawning activity is shown.

March April May June

Ctrl

Sludge

Ctrl

Sludge

3.8+ 1.9 7.1zkl.5 3.8zkO.3 4.9 f 0.7

4.4* 1.0 4.9kO.5 3.3kO.3 3.3 f 1.2

l.OhO.4 1.2zkO.6 1.6kO.8 1.3zto.4

1.3zto.3 1.3zkO.2 1.0+0.2 1.4zko.4

21

Sewage sludge and reproduction of the sand goby

Larvae from sludge-exposed parents were significantly lighter than corresponding controls in the day 1 hatchday group (Fig. 5(b)) and a similar trend (p=O.O9) was apparent for the day 2 hatch-day groups. Similarly, larvae from sludge-exposed parents had a significantly greater yolk-sac volume than controls for the hatch-day 1 group (Fig. 5(c)) and a similar trend (p =0.08) was apparent in the hatch-day 2 data.

DISCUSSION A wide variety of environmental contaminants are known to affect fecundity and larval production in fish (Carlson, 1971; Speranza et al., 1977; Struhsaker, 1977; Bengtsson, 1980). However, in the present study, sewage sludge-exposure of adults for 4-5 months did not significantly affect the fecundity or larval production of those individual females that did spawn. However, at a population level there was a marked reduction in egg and larval production in the sludge-exposed population. This reduced population fitness was due to an inhibition of spawning in some of the females (around 50%). The reason for the apparent failure of many sludge-exposed females to spawn is not clear. Spawning and nonTable 3. The effect of in vitro GTH stimulation (O-2.5 pg ml-‘) on androgen secretion in testes from control (n = 8) and sludgeexposed males (n = 6). AU vahw are in pg steroid released mg-’ testis and represent the means iSE. *p < 0.05 compared to control 0 pg GTH Testosterone 11-Ketotestosterone

Ctrl Sludge Ctrl Sludge

7.0+ 4.8 f 2.5 f 3.0 f

1.7 2.5 0.6 1.2

1.0 pg GTH

2.5 pg GTH

56.8zt 13.1 16.3 & 3.7* 7.ozt 1.9 4.8 f 0.9

70.1+ 15.1 77.0 zt 28.0 14.7*4.2 13.4zt4.2

spawning females had similar GSI values indicating that all the sludge-exposed females had undergone vitellogenesis. Whether oocytes in the non-spawning females had completed vitellogenesis and undergone final maturation is not known. However, data from other studies of the long-term effects of environmental contaminants on female ovarian development have shown that exposure to aromatic and chlorinated organic contaminants can negatively affect both immature and vitellogenic oocytes (Nagler et al., 1986; Casillas et al., 1991: Johnson et al., 1994) causing an inhibition of final

400 2 E IL” +oo = 9 u B ; 200 $

W P

100

“2

0 0

.I

2

’4

3

5

6

7

6

9

Weeks of Spawning

Table 4. Indices of reproductive success from batches of eggs produced by control (n = 12) and sludge-exposed (n = 6) females. Data represent means f SE Control

Sludge

EGGS Number of eggs (a) per batch (b) per g female (wet weight)

14381’~140 757 f 77

1340*258 655 f 78

% Eggs fertile

82.7 f 8.1

79.5 f 6.6

Egg incubation

(“days)

LARVAE Number of larvae (a) per batch (b) as % of total eggs laid (c) as % of fertile eggs (d) per g female (wet weight) Hatch duration (days) % Larvae died at hatch

202&7

969 f 64.6% 70.5 f 5201t

202 11.3 10.2 115

1921Ic4

662+ 155 50.6*9.5 64.0* 10.6 342 f 86

6.8 zto.7

6.8 f 1.o

12.5 f 3.9

29.9* 13.4

rl I

2

3

4

5

6

weeks of Spawning

Fig. 3. Total numbers of eggs and larvae produced (expressed as per g female available to spawn) by the control (open bars) and sludge-exposed (filled bars) populations over the 8 weeks of spawning activity.

C. P. Waring et al.

22

0

2

4

6

6

10

12

14

Hatch-Day Group

Fig. 4. Hatching curves of larvae from control (circles) and sludge-exposed (squares) parents. Data represent means ( f SE) of 5-10 observations.

maturation and ovulation, so that some females fail to spawn (Collier et al., 1992, 1993). A failure of some females to spawn may also have been due to an effect of sludge exposure on male gobies. Indeed, 88% of males in the control population bred successfully, whereas only 40% of sludge-exposed males bred successfully. Male sand gobies actively court females with a courtship display to attract females to their nest site (Fonds, 1973; Hesthagen, 1977). Sludge exposure may have had an adverse affect on male sexual behaviour such that females were not attracted and, therefore, failed to spawn. It was noted that male gobies had a greater sensitivity to sewage sludge judging from their mortality rates and the rate of mortality appeared to increase during the spawning season. Testosterone and 11-ketotestosterone have been implicated as having a major controlling role in the sexual behaviour of a wide variety of male teleosts with territorial and courtship behaviour (reviewed by Borg, 1995). We found little evidence for impaired androgen physiology in sludge-exposed male sand gobies. Testes testosterone and 11-ketotestosterone contents were similar to controls and there was no indication of an impairment in the ability of testes from sludge-exposed males to secrete androgens in response to maximal GTH stimulation. However, the fact that secretion was reduced at the lowest GTH concentration used may indicate an effect on the affinity of GTH for its receptor. Furthermore, we did not measure conjugated androgens, and glucuronidated 5Breduced androgens may have an important role to play in male-female spawning synchronisation in gobies. In another goby species, the black goby (Gobius jam), the territorial males release etiocholanone glucuronide which functions as a potent attractant for ovulated females (Colombo et al., 1980). It is not inconceivable that sludge exposure impacted on the pheromonal-mediated synchronisation of spawning

between the sexes. Recently, sublethal concentrations of various organic contaminants have been shown to alter the metabolism of glucuronidated androgens in male teleosts (Truscott et al., 1992; Singh et al., 1994) and interfere with the pheromonal synchronisation of spawning between the sexes (Moore & Waring, 1996). Although compounds in sewage effluents can have oestrogenic effects in male teleosts (Jobling & Sumpter, 1993; Purdom et al., 1994; Jobling et al., 1995) it is not known if the oestrogenic compounds interfere with steroidogenesis in males and/or affect their behaviour. Also, it is not known how many of the compounds of concern occur in sewage sludge. Sewage sludge contains a variety of organochlorine pesticide residues which have been reported to be oestrogenic to fish (Sumpter et al., 1996). In the present study, water, sediment, and fish were sampled at the end of the experiment to monitor the dispersion of metals and organics into the water column and accumulation in fish tissues. However, technical problems prevented these measurements. In a parallel experiment, in essentially identical conditions, sewage sludge additions at 0.1% resulted in water concentrations of 0.6 ng litre-i hexachlorobenzene (peaks to 2.2-3.0 ng litre-’ 1 h after addition of sludge), 0.3 ng litree’ dieldrin (peaks to 1.4 ng litre-‘), 0.9 ng litre-’ lindane (peaks to 1.6 ng litre-I), and a relatively stable concentration of 0.4 ng litre-’ a-hexachlorocyclohexane (M. J. Costello, unpublished data). Although these water concentrations are low, it is probable that the bioaccumulation of these lipophilic compounds after 3 months of exposure would be significant (Saito et al., 1992). Indeed, the fact that the fecundity, and numbers of viable eggs and larvae produced were reduced in sludge-exposed spawning females and that larval morphological characteristics were altered from sludge exposed parents suggests this to be the case. Exposure of parent fish to a wide variety of contaminants can have a negative impact on their larvae in terms of larval survival and incidence of deformities (Black et al., 1988; Spies & Rice, 1988; Casillas et al., 1991; Mac & Edsall, 1991; Walker et al., 1994). In the present study, there was no major effect of sludge exposure of parent fish on larval lengths except in the hatch-day 5 group. This affected only 45% of the total number of larvae produced. However, 6&70% of larvae from sludge-exposed parents were significantly lighter and had a larger yolk-sac volume. There is evidence that an enlarged yolk-sac volume at hatch can occur due to metabolic disturbances in the embryo which prevents efficient use of yolk reserves (Lonning, 1977; von Westernhagen, 1988). An inefficient or reduced use of yolk reserves for somatic growth would mean lighter larvae with more yolk reserves at hatch. The ecological significance of this is difficult to determine. Cushing (1974) and Ware (1975) suggest that, since the best survival strategy for larvae is rapid growth, the consequences of being small at hatch may be significant. If being light with a bigger yolk-sac impairs swimming ability then this may make the larvae more vulnerable to predation.

23

Sewage sludge and reproduction of the sand goby 300

b.

a

250

s

0

1

2

3

4

5

1

0

7

6

3

2

4

5

6

7

Hatch-Day Group

Hatch-Day Group

C.

10

5 0

1

2

3

4

5

6

7

Hatch-Day Group

Fig. 5. Lengths (a), weights (b) and yolk-sac volumes (c) of larvae from control (circles) and sludge-exposed (squares) parents in the different hatch-day groups. Data represent means ( f SE) of 10-50 observations. ‘p < 0.05 compared to control value.

The high metabolic

cost of inefficient swimming may also reduce the metabolic reserves available for somatic growth. On the other hand, a larger yolk-sac may mean that the larvae would have more yolk reserves available to them during the transition to erogenous feeding. In conclusion, sewage sludge exposure of adult sand gobies had few significant effects on an individual female’s fecundity or larval production but, at a population level, there was a significant inhibition of spawning in 50% of the sludge-exposed females. This spawning inhibition significantly impacted on the overall reproductive fitness of the sludge-exposed population. Future experiments should examine the mechanism of this inhibition and the relative impacts on the reproductive status of the sexes. The larvae from the sludge-exposed population were lighter and had a larger

yolk-sac volume which may have serious implications on their survival ability and ultimately on population recruitment. Further research on the impact of sewage sludge on fish reproduction is therefore warranted.

ACKNOWLEDGEMENTS This work was funded by the Department of the Environment-Water Research Centre grant number DOE PECD 7/7/263. We thank Katherine Kelly, Anne Taylor, Melanie Harding, Murdo MacIver, Philip MacLachlan and Dr Norrie Ramsay for their help with fish collection and husbandry and for interesting discussions and comments during the long stormy days in the West Highlands.

24

C. P. Waring et al.

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Borg, B. (1995). Androgens in teleost fishes. Comp. Biochem. Physiol., 109C, 219-245.

Carlson, A. R. (1971). Effects of long-term exposure of carbaryl (Sevin) on survival, growth, and reproduction of the fathead minnow (Pimephales promelas). J. Fish. Res, Bd Can., 29, 583-587.

Casillas, E., Misitano, D., Johnson, L. L., Rhodes, L. D., Collier, T. K., Stein, J. E., McCain, B. B. & Varanasi, U. (1991). Inducibility of spawning and reproductive success of female English sole (Parophrys vetulus) from urban and nonurban areas of Puget sound, Washington. Mar. Environ. Res., 31, 99-122.

Collier, T. K., Stein, J. E., Sanborn, H. R., Horn, T., Myers, M. S. & Varanasi, U. (1992). Field studies of reproductive success and bioindicators of maternal contaminant exposure in English Sole (Parophrys vetulus). Sci. Total Environ., 116, 169-185.

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Colombo, L., Marconato, A., Belvedere, P. C. & Friso, C. (1980). Endocrinology of teleost reproduction: a testicular steroid pheromone in the black goby Gobius jozo L. Boll. Zool., 47, 355-364.

Costello, M. J. & Read, P. (1994). Toxicity of sewage sludge to marine organisms: a review. Mar. Environ. Res., 37, 2346. Cushing, D. H. (1974). In The Early Life History of Fish, ed. J. H. Blaxter, Springer-Verlag, New York, pp. 103-I 11. Fonds, M. (1973). Sand gobies in the Dutch Wadden sea (Pomatoschistus Gobiidae, Pisces). Neth. J. Sea Res., 6, 417478.

Healey, M. C. (1971). Gonad development and fecundity of the sand goby Gobius minutus Pallas. Trans. Am. Fish. Sot., 3, 52&526.

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