Biochemical and behavioural effects of hydraulic dredging on the target species Chamelea gallina

Fisheries Research 64 (2003) 71–78

Short communication

Biochemical and behavioural effects of hydraulic dredging on the target species Chamelea gallina L. Da Ros a,∗ , N. Nesto a , C. Nasci a , V. Moschino b , D. Pampanin b , M.G. Marin b a

b

Istituto di Scienze Marine, CNR, Castello 1364, I-30122 Venice, Italy Dipartimento di Biologia, Università di Padova, Via Colombo 3, I-30121 Padua, Italy

Received 6 December 2002; received in revised form 22 May 2003; accepted 9 July 2003

Abstract This study is focused on evaluating biological effects on the target species Chamelea gallina as a result of repeated exploitation of clam beds by hydraulic dredging. A biomarker approach was adopted and two physiological indices were applied: adenylate energy charge (AEC) (biochemical level) and reburrowing behaviour (organism level). The aim was to investigate the biological response to various types of fishing, i.e., commercial systems using water at high pressures (HP), and experimental systems at lower pressures (LP) in the field, and to evaluate the response of clams to mechanical stress in the laboratory. Field results showed that HP-treated clams exhibited significantly lower levels of AEC compared with LP treatments. A similar trend was shown by reburrowing behaviour; HP-treated clams reburrowed less. Laboratory results were less clear: a very low level of AEC was measured in both control and treated clams. These poor conditions in foot muscle did not indicate worsening at organism level, as no dead or dying clams were recorded throughout the experiment. However, repeated mechanical stress reduced the percentage of reburrowing clams, which suggested that harvesting may affect reburrowing behaviour. © 2003 Elsevier B.V. All rights reserved. Keywords: Biomarkers; Adenylate energy charge; Reburrowing rate; Hydraulic dredging; Chamelea gallina; Adriatic Sea

1. Introduction Fishing for the striped venus clam Chamelea gallina along the western coast of the Adriatic was changed in the late 1960s by the introduction and rapid acceptance of powerful new gear, the hydraulic dredge. The most evident short/middle-term effect was an immediate increase in landings, which peaked to 100,000 metric tons per year in the early 1980s (Froglia, 1989). ∗ Corresponding author. Tel.: +39-041-2404711; fax: +39-041-5204126. E-mail address: [email protected] (L. Da Ros).

Since their harvests have dramatically decreased, overfishing of the resource was finally recognised as at least one of the factors responsible for the observed decline in natural clam populations. Simultaneous failure in recruitment and widespread mortality in several clam beds along the western coast of the Adriatic have been suggested as concurrent factors (Froglia, 1989, 2000). As a consequence, there was an urgent need to adopt regulations for eco-sustainable management of this important resource, monitoring abundances of the clam population, evaluating its dynamics, and investigating the causes of repeated outbreaks of mass mortalities (Froglia, 2000).

0165-7836/$ – see front matter © 2003 Elsevier B.V. All rights reserved. doi:10.1016/S0165-7836(03)00201-7

72

L. Da Ros et al. / Fisheries Research 64 (2003) 71–78

Studies have demonstrated the deleterious effects of fishing gear on the sea bottom, affecting both abiotic and biotic components (Currie and Parry, 1996; Kaiser et al., 1996; Hall and Harding, 1997; Giovanardi et al., 1998; Pranovi et al., 1998; Hall-Spencer et al., 1999; Veale et al., 2000). However, the use of biomarkers to describe the state of bivalve populations and evaluate the effects of hydraulic dredging as a whole on the biological response of the target species and/or the by-catch, has been a less popular approach (Moore, 1988; Viarengo et al., 1998). More recently, the EU Fair project CT98-4465 offered the opportunity to focus on the effects of repeated disturbance caused by commercial hydraulic dredging on the target species: the hypothesis was that dredging had a detrimental effect on undersized animals, subjecting the exploited clam populations to stress. If commercial hydraulic dredging had significantly negative effects, it might be considered at least a partial cause of increased mortality and a factor contributing towards abnormal susceptibility to normal fluctuations in environmental parameters. In this study, the effects of the mechanical stress undergone by clams when dredged were evaluated in terms of acute variations in two well-known indices of acute stress, adenylate energy charge (AEC) and reburrowing rate (Ivanovici, 1980; Gäde and Meinardus, 1981; Phelps et al., 1985; Phelps, 1989; Moal et al., 1989). As a number of studies have found significant relationships between AEC levels and specific behaviour in bivalves, namely shell closure in mussels (Kramer et al., 1989) and recessing ability in scallops (Maguire et al., 1999a–c, 2002a,b), we decided to examine the AEC level in the foot, which is also the main organ involved in reburrowing of C. gallina. Initially, a preliminary field investigation of the stress caused by different fishing methods was carried out. This was followed by an experimental study, undertaken in the laboratory, for better understanding of the biological response of the animals. The ultimate aim was to evaluate the negative effects of repeated mechanical stress imposed on undersized animals, because the smallest specimens are more likely to be taken up by the hydraulic dredge and then thrown back into the sea more than once before reaching commercial size, due to the high fishing pressure on the clam population.

2. Materials and methods 2.1. Field study In February 2000, samples of C. gallina were dredged from two natural clam beds (Jesolo, latitude 45◦ 30.6 N, longitude 12◦ 41.3 E; Lido, latitude 45◦ 23.9 N, longitude 12◦ 22.6 E) along the western coast of the North Adriatic, just off the Lagoon of Venice (Fig. 1). The two sites were chosen on the basis of their different grain size sediments (Brambati et al., 1988). Commercial-sized clams (26 ± 1 mm long) were collected using two fishing methods: (1) dredging at high water pressure (inlet pressure value 2.5 bar) and mechanically sieved for sorting (high pressure (HP) samples), to verify the biological responses of clams to the fishing system normally used by commercial vessels; (2) dredging at low water pressure (inlet pressure value 1 bar), without sorting (lower pressure (LP) samples), to obtain a less manipulated sample. 2.2. Laboratory study In September 2000, a shaking experiment was set up to simulate the repeated disturbance caused to juvenile clams by fishing. Undersized specimens (13–19 mm long) obtained by dredging were acclimatised in the laboratory for 4 days in tanks with sea water at 17 ± 1 ◦ C and 35‰ and fed with Isochrysis galbana. Mechanical stress conditions due to fishing were simulated using a vortex mixer: clams were arranged in a box (volume 3600 cm3 ), subjected to daily mechanical shaking (1–3 days) for 10 min at the maximum speed (40 Hz); the vortex was balanced equally each time by processing the same number of clams (100). Stress was applied once a day over 3 days. On day 1 a control and a stressed sub-sample were analysed, the remaining clams were returned to seawater, and on day 2 were re-stressed, sub-sampled and analysed. The same procedure was repeated on day 3. 2.3. Adenylate energy charge The striated muscle of 20 individuals (10 in the laboratory study) was immediately removed and frozen in liquid nitrogen (on board for the field study, in the laboratory for the shaking experiment). The samples

L. Da Ros et al. / Fisheries Research 64 (2003) 71–78

73

Fig. 1. Sampling sites.

were then analysed according to the method of Moal et al. (1989). The AEC was calculated using the following equation (Atkinson, 1968): AEC = (ATP + 0.5ADP)/ (ATP + ADP + AMP). Samples were compared using the non-parametric Kruskal–Wallis test (STATISTICA 5.5 package).

were compared using the Gehan and Wilcoxon test (Gehan, 1965). The percentage of clams reburrowing over a fixed time period was also determined, and samples were compared using the G-test.

2.4. Reburrowing rate

3.1. Field study

Within 1 h, clams from the field study were transferred to the laboratory in refrigerated boxes (at ±4 ◦ C and 100% humidity). Twenty individuals were placed in aquaria (42 cm × 24 cm × 26 cm) containing fine particles obtained by sieving natural beach sediment. The aquaria were kept in a thermostatic chamber simulating original water temperatures. A web cam was used for continuous observation of reburrowing behaviour. The camera was set to record one frame every 30 min for a period of 10 h. Subsequently, the RT50 (time required for reburrowing by 50% of the clams) was calculated according to the method of Kaplan and Meier (1958), and reburrowing curves

Similar results were obtained for the AEC response in samples from both stations, Lido and Jesolo (Table 1). Significantly higher mean values were always shown by the LP samples compared with the HP ones (respectively, 0.83 vs. 0.73, P < 0.05, at Jesolo; 0.79 vs. 0.74, P < 0.001, at Lido). Reburrowing rates are shown in Table 2 and Fig. 2. The trends, expressed both as percentage of reburrowing clams and reburrowing curves, are similar in the two stations. At Lido, the LP clams reburrowed more quickly than the HP ones, although the difference was not significant (90 vs. 65%, respectively). At Jesolo, the LP sample had a significantly higher reburrowing

3. Results

74

L. Da Ros et al. / Fisheries Research 64 (2003) 71–78

Table 1 Field study. AEC in C. gallina collected at Lido and Jesolo in February 2000 using different fishing systemsa Stations

LP

HP

Statistical comparison (Kruskal–Wallis test)

Lido Jesolo

0.835 ± 0.009 0.789 ± 0.013

0.735 ± 0.019 0.0738 ± 0.019

P < 0.001 P < 0.05

a

n = 20. Values are mean ± S.E. LP: low pressure; HP: high pressure and mechanical sorter.

% reburrowed clams

Reburrowing curves - Lido 100 80 60 40

n.s.

20 0 1

2

3

4

5

6 hours

7

LP RT50 = 2

8

9

10

11

10

11

HP RT50 = 2

Reburrowing curves - Jesolo % reburrowed clams

100 80 60 40 *

20 0 1

2

3

4

5

6 hours

LP RT50 = 4

7

8

9

HP RT50 = n.d.

Fig. 2. Field study. Reburrowing curves in C. gallina collected at Lido and Jesolo in February 2000 using different fishing systems (n = 20). Statistical comparison (Gehan and Wilcoxon test): ∗, P < 0.05; n.s., not significant. LP: low pressure; HP: high pressure and mechanical sorter.

L. Da Ros et al. / Fisheries Research 64 (2003) 71–78 Table 2 Field study. Percentage of reburrowed clams over a 10 h period in C. gallina collected at Lido and Jesolo in February 2000 using different fishing systemsa Stations

LP

HP

Statistical comparison (G-test)

Lido Jesolo

90 70

65 45

P > 0.05 P > 0.05

n = 20. LP: low pressure; HP: high pressure and mechanical sorter.

75

tween controls and treated samples at any time, although the general trend was towards a decrease in the burrowing rate of treated clams. This was particularly evident after the 48 h treatment. Considering the reburrowing curves (Fig. 3), no significant differences were observed between control and treated samples at any time.

a

Table 3 Laboratory study. AEC in C. gallina subjected to mechanical shakinga Days

C

T

Statistical comparison (Kruskal–Wallis test)

1 2 3

0.250 ± 0.044 0.425 ± 0.057 0.454 ± 0.060

0.272 ± 0.037 0.173 ± 0.039 0.419 ± 0.049

P > 0.05 P < 0.01 P > 0.05

a

n = 10. Values are means ± S.E. C: control; T: treated.

rate (P < 0.05) than the HP sample (70 vs. 45%, respectively). Both indices show relatively higher values in samples from Lido than Jesolo. 3.2. Laboratory study In both controls and treated specimens, the AEC values were low and within a very narrow range (0.17–0.45) (Table 3). There was no significant difference between treated and control samples, either at time 0 or after 48 h. Only after 24 h did controls show a significant higher value (P < 0.01) compared with the treated specimens (0.42 and 0.17, respectively). Table 4 shows the percentage of clams reburrowing over a 10 h period. Values were very similar, and ranged from 70 to 90% of successfully burrowed clams. No significant differences were apparent beTable 4 Laboratory study. Percentage of reburrowed clams over a 10 h period in C. gallina subjected to mechanical shakinga Days

C

T

Statistical comparison (G-test)

1 2 3

85 80 90

80 75 70

P > 0.05 P > 0.05 P > 0.05

a

n = 20. C: control; T: treated.

4. Discussion The AEC ratio, as an acute response to environmental modifications (from minutes for micro-organisms to hours for molluscs), has been used extensively to evaluate the effects on specific organisms of various kinds of environmental and human disturbance, such as anoxia, variations of temperature and salinity, and exploitation of coastal and shelf sea (Wiebe and Bancroft, 1975; Wijsman, 1976; Ivanovici, 1980; Gäde and Meinardus, 1981). Referring specifically to the possible disturbing action of fishing, only recently have Maguire et al. (2002a) found decreasing AEC levels in the adductor muscle of Pecten maximus captured by spring-loaded dredges, in comparison with diver-harvested samples, and after being treated in an experimental device simulating the fishing gear operating system. Skjoldal (1981) also determined AEC levels in tropical zooplankton from the Great Barrier Reef and found that Eucalanus subcrassus had significantly lower AEC values when collected during a period of 30 min than during 3 min towing, indicating capture stress. The AEC levels recorded in our field study for the foot of C. gallina ranged from 0.73 to 0.83, and were similar to those obtained by Maguire et al. (1999a) for the adductor muscle of P. maximus (0.9). Also, the results found by various authors in several species of marine molluscs, analysed in their whole soft body, were in the same range. Moal et al. (1989) found AEC levels of 0.73 in Crassostrea gigas harvested off Brest (Brittany, France). Similarly, Gäde and Meinardus (1981) measured AEC levels in the range of 0.88–0.9 for Cardium edule, and Ivanovici (1980) showed AEC levels of 0.8–0.9 in the snail Pyrazus ebeninus from Jervis Bay (New South Wales). In our field results, AEC levels showed highly significant differences between HP and LP samples in the two study areas, together with low inter-individual variability. Observations demonstrated both the

76

L. Da Ros et al. / Fisheries Research 64 (2003) 71–78

Reburrowing curves - 1st day

% reburrowed clams

100 80 60 40

n.s.

20 0 1

2

3

4

5

6 hours

C1 RT50 = 4

% reburrowed clams

100

7

8

9

10

11

10

11

10

11

T1 RT50 = 3.5

Reburrowing curves - 2nd day

80 60 40 n.s. 20 0 1

2

3

4

5

6 hours

C2 RT50 = 4.5

% reburrowed clams

100

7

8

9

T2 RT50 = 5

Reburrowing curves - 3rd day

80 60 40

n.s.

20 0 1

2

3

4

5

C3 RT50 = 2

6 hours

7

8

9

T3 RT50 = 2

Fig. 3. Laboratory study. Reburrowing curves in C. gallina subjected to mechanical shaking (n = 20). Statistical comparison (Gehan and Wilcoxon test): n.s., not significant. C: control; T: treated.

effectiveness of the AEC level in the foot as an indicator of mechanical stress due to the fishing practice used (water jet pressure and sorting) and its sensitivity, confirming that it is a rapid, precise method of evaluating the effects of environmental perturbation on these organisms. Unfortunately, laboratory simulation of repeated fishing did not provide similarly clear results in the undersized clams. Also in

undersized scallops significant cumulative effects on the AEC level were observed when dredging was repeated after 24 h, but not after 48 h (Maguire et al., 2002b). The very low AEC values in both controls and treated specimens on the first day of the experiment may indicate poor conditions in foot muscle, but did not reveal worsening in organismic level, as no dead or dying clams were recorded throughout

L. Da Ros et al. / Fisheries Research 64 (2003) 71–78

the experiment. This condition may reflect an inadequate acclimatisation period—also suggested by the increased value recorded in the controls on the second and third days of the experiment, ultimately masking the effects of mechanical stress in the treated clams. Moreover, the lack of a clear trend in the treated clams (decreasing values observed on the second day and subsequent recovery on the third day) may indicate that the AEC level in muscle is not a good index of the whole physiological status of undersized clams when kept in experimental laboratory conditions. The ecological relevance of burrowing behaviour in clams becomes apparent when we consider their need to avoid predation (Doerding, 1982; Pearson et al., 1981). On this basis, a rapid bioassay has been developed and successfully used in toxicity sediment tests (Phelps et al., 1985; Phelps, 1989). It gives an indication of the indirect mortality resulting from increased exposure to predators (Phelps, 1989) and can provide a short-term response (minutes to hours) to the introduction of chemical or environmental perturbation. The reburrowing rates found in our field study for C. gallina (RT50 = 2–4 h) were lower than those calculated for Mya arenaria, which is considered a suitable organism for rapid burrowing bioassays in estuarine sediments, having an average burrowing speed in controls of ET50 (elapsed time for 50% population) of 0.45 h (Phelps, 1989). Protothaca staminea, used in a toxicity test with Cu-enriched sediment, also showed higher burrowing rates in controls (ET50 = 0.2–1 h) (Phelps et al., 1985); similarly, Ensis siliqua and E. ensis from North Wales had with a maximum ET50 of 0.25 h (Henderson and Richardson, 1994). These considerations were helpful in highlighting how long C. gallina can be left on the seabed after any kind of sediment perturbation, and show that it is a very responsive organism for evaluating mechanical stress. In general, samples from Lido seemed to be less stressed than those from Jesolo. The different grain size distribution in sediments (sandier at Lido, muddier at Jesolo) may have influenced the speed and proper operation of the hydraulic dredge, reflecting a different impact on the seabed (more severe on muddy bottoms). The greater percentage of mud in sediments at Jesolo than at Lido (14.70 vs. 1.79%) (Bianchi, pers. commun.) may contribute to poor environmental conditions for C. gallina, which prefers to live in sandy seabeds near the coastline where medium, fine and very fine sands

77

normally exceed 90%, with a redox potential (Eh) greater than 300 mV (Barillari et al., 1978). Moreover, looking at the results of the laboratory simulation, the shaking treatment seems to have affected the reburrowing behaviour of the juveniles, as treated samples showed a reduction in the percentage of reburrowing clams compared with controls, especially after 3 days’ consecutive treatment, although differences were not statistically significant. In our field study, both AEC and reburrowing rate revealed greater stress in HP samples, emphasising the impact caused by the fishing method on clams. From reduced well-being in commercial-size clams, we infer that undersized individuals are subjected to similar or greater stress levels, juveniles being in general more vulnerable and more sensitive to environmental variations (His et al., 2000). When returned to the sea after passing through the sieve on board, these juveniles can no longer quickly reburrow and are thus more vulnerable to predation by crabs and gastropods, as suggested by Hall-Spencer et al. (1999). Although our overall results indicate that these stress indices can be used to measure the impact of hydraulic dredging on C. gallina, further research should aim at studying the relationship between AEC level in the foot and AEC contents of the whole organism. Moreover, the influence of endogenous and environmental variables (reproductive period, temperature, salinity, dissolved oxygen, food availability, etc.) on the biological responses of juveniles to mechanical stress should be carefully evaluated in field studies.

Acknowledgements This study was carried out with financial support from the Commission of the European Communities, Agriculture and Fisheries (FAIR) specific RTD programme, CT98-4465, ECODREDGE.

References Atkinson, D.E., 1968. The energy charge of the adenylate pool as a regulatory parameter: interaction with feed-back modifiers. Biochemistry 7, 4030–4034. Barillari, A., Boldrin, A., Mozzi, C., Rabitti S., 1978. Alcune relazioni tra natura dei sedimenti e presenza della vongola

78

L. Da Ros et al. / Fisheries Research 64 (2003) 71–78

Chamelea (Venus) gallina nell’Alto Adriatico presso Venezi., Atti Ist. Veneto Sci., CXXXVI. Brambati, A., Ciabatti, M., Fanzutti, G.P., Marabini, F., Marocco, R., 1988. Sedimentological map of the Northern Adriatic Sea. In: Brambati, Uras (Eds.), National Council of Research “Progetto Finalizzato Oceanografia e Fondi Marini”. Istituto Geografico De Agostini, Novara. Currie, D.R., Parry, G.D., 1996. Effects of scallop dredging on a soft sediment: a large-scale experimental study. Mar. Ecol. Prog. Ser. 134, 131–150. Doerding, P.H., 1982. Reduction of the sea star predation by the burrowing response of the hard clam Mercenaria mercenaria (Mollusca: Bivalvia). Estuaries 5, 310–315. Froglia, C., 1989. Clam fisheries with hydraulic dredges in the Adriatic Sea. In: Caddy, J.F. (Ed.), Marine Invertebrate Fisheries: Their Assessment and Management. Wiley, New York, pp. 507–524. Froglia, C., 2000. Il contributo della ricerca scientifica alla gestione della pesca dei molluschi bivalvi con draghe idrauliche. Biol. Mar. Medit. 7 (4), 71–82. Gäde, G., Meinardus, G., 1981. Anaerobic metabolism of the common cockle Cardium edule. Mar. Biol. 65, 113–116. Gehan, E.A., 1965. A generalized Wilcoxon test for comparing arbitrarily singly-censored samples. Biometrica 52, 203–223. Giovanardi, O., Pranovi, F., Franceschini, G., 1998. “Rapido” trawl fishing in the Northern Adriatic: preliminary observations of the effects on macrobenthic communities. Acta Adriatica 39 (1), 37–52. Hall, S.J., Harding, M.J.C., 1997. Physical disturbance and marine benthic communities: the effects of mechanical harvesting of cockles on non-target benthic infauna. J. Appl. Ecol. 34, 497– 517. Hall-Spencer, J.M., Froglia, C., Atkinson, R.J.A., Moore, P.G., 1999. The impact of Rapido trawling for scallops, Pecten jacobaeus (L.), on the benthos of the Gulf of Venice. ICES J. Mar. Sci. 56, 111–124. Henderson, S.M., Richardson, C.A., 1994. A comparison of the age, growth rate, growth rate and burrowing behaviour of the razor clams, Ensis siliqua and E. ensis. J. Mar. Biol. Assoc. UK 74, 939–954. His, E., Beiras, R., Seaman, M.N.L., 2000. The assessment of marine pollution—bioassays with bivalve embryos and larvae. Adv. Mar. Biol. 37, 1–178. Ivanovici, A.M., 1980. The adenylate energy charge in the estuarine mollusc, Pyrazus ebeninus. Laboratory studies of responses to salinity and temperature. Comp. Biochem. Physiol. A 66, 43– 55. Kaiser, M.J., Hill, A.S., Ramsay, K., Spencer, B.E., Brand, A.R., Veale, L.O., Prudden, K., Rees, E.I.S., Munday, B.W., Ball, B., Hawkins, S.J., 1996. Benthic disturbance by fishing gear in the Irish Sea: a comparison of beam trawling and scallop dredging. Aquat. Conserv. 6, 269–285. Kaplan, E.L., Meier, P., 1958. Nonparametric estimation from incomplete observations. J. Am. Stat. Assoc. 53, 457–481.

Kramer, K.J., Jenner, H.A., de Zwaart, D., 1989. The valve movement response of mussels: a tool in biological monitoring. Hydrobiologia 188/189, 433–443. Maguire, J.A., Cashmore, D., Burnell, G.M., 1999a. The effect of transportation on the juvenile scallop Pecten maximus (L.). Aquat. Res. 30, 325–333. Maguire, J.A., Fleury, P.G., Burnell, G.M., 1999b. Some methods for quantifying quality in the scallop Pecten maximus (L.). J. Shell. Res. 18 (1), 59–66. Maguire, J.A., O’Connor, D.A., Burnell, G.M., 1999c. An investigation into behavioural indicators of stress in juvenile scallops. Aquacult. Int. 7, 169–177. Maguire, J.A., Coleman, A., Jenkins, S., Burnell, G.M., 2002a. Effect of dredging on undersized scallops. Fish. Res. 56, 155– 165. Maguire, J.A., Jenkins, S., Burnell, G.M., 2002b. The effects of repeated dredging and speed of tow on undersized scallops. Fish. Res. 58, 367–377. Moal, J., Le Coz, L.R., Samain, J.F., Daniel, J.Y., 1989. Nucleotids in bivalves: extractions and analysis by high-performance liquid chromatography (HPLC). Comp. Biochem. Physiol. B 93 (2), 307–316. Moore, M.N., 1988. Cellular and histopathological effects of a pollutant gradient—summary. Mar. Ecol. Prog. Ser. 46, 109– 110. Pearson, W.H., Woodruff, D.L., Sugarman, P.C., Olla, B.L., 1981. Effects of oiled sediment on predation on the littleneck clam, Protothaca staminea, by the Dungeness crab, Cancer magister. Estuar. Coast. Shelf Sci. 13, 445–454. Phelps, H.L., 1989. Clam burrowing bioassay for estuarine sediment. Bull. Environ. Contam. Toxicol. 43, 838–845. Phelps, H.L., Pearson, W.H., Hardy, J.T., 1985. Clam burrowing behaviour and mortality related to sediment copper. Mar. Pollut. Bull. 16 (8), 309–313. Pranovi, F., Giovanardi, O., Franceschini, G., 1998. Recolonization dynamics in areas disturbed by bottom fishing gears. Hydrobiologia 375–376, 125–135. Skjoldal, H.R., 1981. ATP concentration and adenylate energy charge of tropical zooplankton from waters inside the Great Barrier. Reef. Mar. Biol. 62, 119–123. Veale, L.O., Hill, A.S., Hawkins, S.J., Brand, A.R., 2000. Effects of long-term physical disturbance by commercial scallop fishing on subtidal epifaunal assemblage and habitats. Mar. Biol. 137, 325–337. Viarengo, A., Pons, G., Fabbri, R., Mozzone, S., Evangelisti, V., Capri, F., 1998. Evaluation of the physiological status of Chamelea gallina using biomarkers. Biol. Mar. Medit. 5 (3), 1090–1101. Wiebe, W.J., Bancroft, K., 1975. Use of the adenylate energy charge ratio to measure growth state of natural microbial communities. Proc. Natl. Acad. Sci. USA 72, 2112–2215. Wijsman, T.C.M., 1976. Adenosine phosphate and energy charge in different tissues of Mytilus edulis under aerobic and anaerobic conditions. J. Comp. Physiol. 107, 129–140.