Hatchery Cultivation of the Common Cockle (Cerastoderma Edule L.): from Conditioning to Grow-Out

Aquaculture Research 2015, 46, 302–312 doi:10.1111/are.12178

Hatchery cultivation of the common cockle (Cerastoderma edule L.): from conditioning to grow-out

Anna Elisabeth Pronker1, Frank Peene1, Silke Donner1, Sander Wijnhoven2, Pieter Geijsen1, Peter Bossier3 & Nancy Marie Nevejan3 1Roem van Yerseke B.V., Yerseke, The Netherlands 2Monitor Taskforce, Royal Netherlands Institute for Sea Research (NIOZ–Yerseke), Yerseke, The Netherlands 3Laboratory for Aquaculture & ARC, Ghent University, Gent, Belgium

Correspondence: A Pronker, Roem van Yerseke B.V., Yerseke, The Netherlands. E-mail: [email protected]

Mauritania in West Africa up to the western Abstract Barents Sea (Tebble 1966). It inhabits intertidal This study describes for the ﬁrst time the cultiva- and subtidal areas with densities up to 60 000 tion of Cerastoderma edule on a commercial scale. juveniles m2 (Jensen 1992). It can grow to a

A protocol to grow F2 generation cockles was maximum size of 5 cm (Tebble 1966). The com- developed, which led to fine-tuning experiments mon cockle is a culinary appreciated shellfish spe- for broodstock conditioning and spat growth. cies and used to be an important export product Broodstock animals were conditioned with diets for the Netherlands, particularly for the fisheries of Isochrysis galbana (T-Iso) or Tetraselmis suecica, industries situated around Yerseke (South-West whereas a third group was not fed. The best diet, Netherlands) (Stichting ODUS 2001; Provincie T. suecica, induced 12 females out of 100 animals Zeeland 2006). The Dutch cockles were mainly to spawn a total of 3 380 000 eggs. The non-fed fished in the Dutch Wadden Sea in the North. group did not spawn. Between 1998 and 2000, an average of Cockle spat (4.9 1.0 mm) grew best when 14 000 tonnes of cockle meat was fished annually given a mixed diet of C. muelleri, T-Iso and Sceleto- (LEI 2001). Since 2005 there is a ban on commer- nema costatum, or a mixture of P. tricornutum and S. cial hydraulic fishing of wild stocks in the Wadden costatum at a concentration of 240 cells ll1 day1, Sea, after it was proclaimed as a national reserve. resulting in a tripling of their wet weight after Beukema and Dekker (2006) showed that the total 14 days. The impact of density, burrowing substrate cockle biomass in the Dutch Wadden Sea was not and food availability on cockle spat growth (41 days sufficient to sustain both wintering bird popula- old, 5.6 1.2 mm) was studied for 11 weeks. Best tions and the commercial fisheries. results were obtained by culturing spat at ad libitum Other important cockle regions are the Eastern food conditions at 500 ind m2, resulting in an Scheldt and the Western Scheldt in the South- average growth rate of 168 lm day1, an average Western part of the Netherlands, although the final size of 19.0 1.9 mm and a total final bio- cockle production in these waters is much lower mass of 1040 g m2. than in the Dutch Wadden Sea, by a factor 10 and 20, respectively. The total cockle biomass Keywords: hatchery, nursery, grow-out, Ceras- available in these waters is evaluated yearly toderma edule, microalgae diets before dredging is allowed, to make sure enough cockleswillbeleftasfoodforwaterbirds. During the last 10 years, hydraulic dredging Introduction for cockles only took place in the Eastern Scheldt Cerastoderma edule (Linnaeus 1758) is a common in 2001 and 2006 and in the Western bivalve in European waters and can be found from Scheldt from 2002 to 2005 (CBS & PBL 2011;

Wijnhoven, Escaravage, Herman, Smaal & Hum- (F1-generation) were conditioned with a mixture mel 2011. of different algae species: D. tertiolecta, P. tricornu- Due to the poor availability of dredgeable cockle tum, T. suecica, S. costatum, C. muelleri and T-Iso at populations in the Netherlands, the interest of the a ratio of 1:1:3:3:3:5 (based on cell-count), ad libi- Dutch shellfish industry in developing hatchery tum. An equal distribution of males and females production and land-based grow-out techniques was assumed, based on the findings of Boyden for the common cockle has increased considerably. (1971) and own field surveys. When the animals At the moment, there is no commercial hatchery had well-developed gonads, they were carefully that can provide cockle seed for grow-out, either washed in filtered seawater and placed on a 1-cm on land or in the tidal zone. Moreover, information mesh screen at the bottom of a 40 L spawning on hatchery techniques for this species is very tank with running seawater (2 L min1 at 8°C). scarce. Broodstock conditioning experiments for After 30 min at 8°C, spawning was initiated by cockle production are not described to our knowl- instantly raising the water temperature from 8°C edge, nor the use of native Dutch algae species for to 19°C at which temperature the animals were spat culture of bivalves in general and C. edule in kept during the entire 24-h spawning period. Fer- particular. Large scale, land-based cultivation of tilization took place in the spawning tank, immedi- C. edule from the Eastern Scheldt seems technically ately after the release of the gametes. Fertilized feasible, but still many questions have to be eggs were retained on a 60 lm sieve after passing answered before this culture becomes economically through a 150 lm sieve to remove debris. Unfer- viable. tilized and fertilized egg samples were taken for measurements at two occasions (n = 30). Eggs were transferred to 130 L conical larvae tanks Materials and methods with 19°C seawater and light aeration. No algae For both culturing algae and cockles, 5 lm filtered were added. Twenty-four hour after fertilization, and UV-treated seawater was used from the East- D-larvae were retained on a 60 lm sieve, counted, ern Scheldt. The water had a salinity of approxi- measured and restocked at a density of 8 mately 28 g L1. larvae mL1. Continuous micro-algae cultures (Sea-Cap The larvae were reared on a mixture of T-Iso, systemâ)ofDunaliella tertiolecta (D. tertiolecta, P. lutherii and C. muelleri (1:1:1 ratio, based on CCMP364), Tetraselmis suecica (T. suecica, CCAP66/4), cell count) at a concentration of 80 cells lL1. Sceletonema costatum (S. costatum, CCY9929), Phaeo- Water flow was kept constant at 0.7 L min1 and dactylum tricornutum (P. tricornutum, CCY0818), the flow-through tanks were cleaned three times a Chaetoceros muelleri (C. muelleri, CCMP1316), Chaetoc- week with freshwater and a mixture of diluted eros calcitrans (C. calcitrans, CCAP 1010/11), Pavlova acetic and hydrochloric acid. lutherii (P. lutherii) and Isochrysis galbana affinis Tahiti Larvae were measured after every spawning (T-Iso, CCAP 927/14) were grown in 220 L plastic event, by collecting approximately 100 larvae at bags, supplemented with filtered (5 lm) and pasteur- varying intervals till day post fertilization (DPF) 9. ized seawater from the Eastern Scheldt, enriched with The size and growth rate of the larvae, was modified Walne medium (Walne 1970) and CO2. The described by fitting a linear trend line through the pH of the cultures was kept between 7.5 and 8.5 by averages of the three batches, according to Dela- regulating the supply of CO2. Algae were grown unay, Marty, Moal and Samain (1992). Nine days under 24-h light conditions, with constant aeration, after fertilization, most of the larvae were settled at a temperature of 21 1°C. Algae in the exponen- or in the process of settling. They were transferred tial growth phase were harvested daily and cell to 5-cm deep downwellers with a diameter of concentrations were counted using a Burker€ counting 40 cm at densities of approximately 10 individu- cell. als cm2. Filtered seawater of 19°C, was added at a flowrate of 200 mL min1. From this stage up to a size of 1 cm, the young cockles received ad Spawning and growth of F generation cockles 2 libitum a mixture of different algae species consist- This protocol was followed at three different ing of T-iso, P. lutherii, C. muelleri, C. calcitrans, occasions (May, July and August 2007). A group S. costatum and T. suecica. The growth of spat was of 100, one-year old hatchery-reared cockles followed from DPF 9 till DPF 51 and was modelled

© 2013 John Wiley & Sons Ltd, Aquaculture Research, 46, 302–312 303 Cultivation of the common cockle A E Pronker et al. Aquaculture Research, 2015, 46, 302–312 by fitting a 4th order polynomial through the data instantly from 12°Cto19°C and was kept at for each batch. Average spat sizes were calculated 19°C during the entire spawning period. Females according to this model. An extra fourth batch, starting to spawn were immediately removed reared according to the same procedure, provided from the spawning tank and placed individually extra measurements for day 43 and 57, to help to in 300 mL beakers with filtered seawater at identify the transition period from spat to juvenile 19°C. The number of eggs released by each cockle. Actual growth rate was determined by tak- female was counted by suspending them in 1 L ing the derivative of the average growth curve. To of filtered seawater and counting a 50 lL estimate the time needed to produce commercial subsample. sized cockles (20 mm), growth was measured until the cockles were 134 days old. Shell lengths (ante- Effect of algae diets on spat growth rior-posterior axis) were measured to the nearest 1.0 mm. Seven different micro-algae diets were tested for their nutritional value for cockle spat (Fig. 4). The mixture of T-Iso, C. muellerii and S. costatum was considered a Effect of conditioning diets on fecundity balanced diet and served as positive control (PosC) at Broodstock animals with an average size of a concentration of 240 cells lL1 day1 and nega- 3.0 cm, were collected at the end of February tive control at 80 cells lL1 day1. The other treat- 2010 at ‘Speelmansplaat’, an intertidal area in the ments received 240 cells lL1 day1, except for Eastern Scheldt. To check the gonadal maturation treatments receiving D. tertiolecta, which was given at the start of the experiment, a subsample of 100 at a ratio of 1 to 5 (D. tertiolecta = 5 9 T-Iso). All cockles was induced to spawn. As no animals algae species, except for T-Iso and C. muelleri, are spawned, the remaining animals were randomly native to the Netherlands, making them suitable for divided into three experimental groups of 150 ani- use in open, land based grow-out systems without mals each. They were placed in trays of 25 L, sup- environmental risks. ported by a 2-cm mesh screen, 4 cm off the For each treatment, three small upwellers bottom. Filtered seawater of 12°C was provided at (18 cm high, 10 cm diameter) were placed in a a constant rate of 0.25 L min1. Aeration was 20 L bucket, which in its turn stood in a 40 L provided by air stones and the tanks were rinsed tank. Each upweller contained spat with an aver- weekly with freshwater. age size of 4.9 1.0 mm and a total live weight One group was starved (negative control), of 3.0 g with approximately 150 individuals per whereas other two groups received either T-Iso (Iso silo. From the upweller, the water flowed into the treatment) or T. suecica (Tetra treatment) for 40 L tank. Water-airlifts were positioned in the 6 weeks. The cockles were fed daily 4.5% of dry 40 L tanks to recirculate the water back into the meat weight in dry weight of algae. The Iso treat- bucket and to give extra aeration. Fresh seawater ment received 5.87 9 1010 cells day1, whereas the was added to the buckets at a constant rate of Tetra treatment only 5.87 9 109 cells day1,based 8.3 L h1. The total renewal of water in one sys- on an algae dry weight of 20 and 200 pg cell1 tem was 4.8 times per day1. Micro-algae were respectively (Utting & Spencer 1991). This resulted added continuously from a 30 L feeding bucket in a concentration of, respectively, 127 and over a period of 22 h using a peristaltic pump at a 12.7 cells lL1 in the conditioning tanks. The daily flowrate of 21 mL min1. Temperature was kept amount of algae (diluted with 5 lm filtered seawater constant at 19°C. Feeding buckets were cleaned to obtain a total volume of 90 L) was given to the daily and upwelling systems weekly, with fresh cockles at a constant rate of 300 mL min1,overa water and a mixture of diluted acetic and hydro- period of 22 h, by using a peristaltic pump. chloric acid. After 6 weeks of conditioning, 100 individuals Significant differences in spat weight between from each group were carefully cleaned with treatments were tested for at t = 7 and filtered seawater and placed in spawning tanks. t = 14 days using a student’s t-test with a signifi- The base of the spawning tank was covered cance level of p < 0.05. Weekly growth rates were with a black plastic sheet to easily see the calculated using the formula: %Gr7 ¼ 100 ð ð Þ ð Þ Þ released gametes. Spawning was initiated by pro- ln Wt tþ7 ln Wt t (Curatolo, Ryan & Mercer viding a thermal shock: temperature was raised 1993).

300.0 Effect of burrowing substrate and density on growth of juveniles 250.0 A batch of 1500 juvenile cockles, (41 days old, 200.0 average size 5.6 1.2 mm) originating from the same spawning event was randomly divided into 150.0 ﬁve groups. One group was placed in an upweller on a mesh-screen on top of the sediment to pre- Size (um) 100.0 vent them from burrowing into the sediment at a y = 16.75x + 79.34 density of 1000 ind m2, equivalent to a total bio- 50.0 mass of 70 g m2. A second group on the other R² = 0.93 0.0 hand, was allowed to bury into the sediment and 0246810 was stocked at the same density. This set-up was DPF repeated both indoors and outdoors. A last group batch 1 batch 2 batch 3 was placed inside on a mesh-screen on top of the sediment at a density of 500 ind m2, equivalent Figure 1 Average growth curve of cockle larvae (C. edule) from day 1 till metamorphosis (DPF 9) to a total biomass of 35 g m 2. The cockles (n = 3). indoors received ad libitum, a mixture of the algae species D. tertiolecta, P. tricornutum, T. suecica, to mortality and culling. After metamorphosis mor- S. costatum, C. muelleri and T-Iso at a ratio of tality dropped to practically zero. approximately 1:1:3:3:3:5 (based on cell-count). Young spat was transferred into downwellers at The cockles outside received the wastewater from densities of 10 ind cm 2 with an extended food the indoor hatchery rearing systems. No extra food choice. From day 9 till day 57, the spat grew from was added. At each sampling occasion, at least an average of 226 to 6700 lm, the equivalent of 100 individuals were measured. Shell lengths 4 mm month 1 (Fig. 2a). A maximum growth (dorsal-ventral axis) were measured to the nearest rate of almost 230 lm day 1 was reached at DPF 1.0 mm. The experiment lasted 11 weeks. 40 (Fig. 2b). Following up the growth till DPF 134, the equation of the growth curve calculates that 9 months are needed to obtain commercial Results sized cockles of 20 mm (Fig. 3).

Growth F2 generation Effect of conditioning diets on spawning success At the three spawning sessions, males usually started to spawn ﬁrst, soon followed by the females. Feeding broodstock cockles during their condition- One hundred cockles produced on average, a total ing period, increased spawning success (Fig. 4). of 1.37 0.60 million eggs, of which 63.5 5.4% Moreover, starved cockles yielded no eggs. Best were recovered as D-larvae after two days. Released results were obtained by providing the cockles a diet eggs had an average size of 70.0 5.1 lm while of T. suecica: 12 females spawned a total of including the gelatinous membrane they had a 3 380 000 eggs. This is 3.5 times more eggs than diameter of 139.0 12.8 lm. After fertilization, that were obtained from females fed T-Iso. The average egg size without and with membrane number of spawning females in the latter treatment increased to 73.7 4.8 lm and 144.0 14.5 lm was similar (13), but 77% of the females spawned respectively. From DPF 1 (24 h after fertilization) less than 10 000 eggs whereas this was true for until DPF 9, the cockle larvae grew from only 25% of the females receiving T. suecica.In 109.0 4.5 lm (D-larvae) to 226.6 22.1 lm addition, more males spawned in the Tetra (pediveliger larvae), which equals an average treatment than the Iso treatment (20 versus 9). growth rate of 16.7 lm day1 (Fig. 1). At day 9, almost all of the pediveliger larvae had lost their Effect of algae diets on spat growth velum and were settled or in the process of settling, also indicated by the development of gills, a foot and Cockle spat of the treatments DS, DPS and PosC an eyespot. From DPF 1 till after metamorphosis, 2 grew from an initial weight of 3.0 g to an average of the batches lost 20% and 62% of the larvae, due of 5.8 0.2 g, 5.6 0.4 g and 5.4 0.5 g

(a) (b)

Figure 2 (a) Average cockle spat growth expressed in length (mm) from DPF 9 to DPF 41 or 57 (n = 4) (b) Aver- age cockle spat growth rate (GR) in mm day1.

20 y = 2E-07x4 – 7E-05x3 + 0.0075x2 – 0.0839x 18 R² = 0.99315 16

8 Size (mm)

0 Figure 3 Cockle growth at hatch- 0 25 50 75 100 125 150 ery conditions till 134 days past DPF fertilization (DPF).

12 were even more pronounced. Spat fed on DS, DPS T-Iso diet Tetra diet and PosC weighed 7.4 0.6 g, 7.2 0.5 g and 10 8.5 1.4 g, respectively (equivalent to a biomass 2 8 of 943–1076 g m ), which was highly signiﬁ- cantly different from those fed on DP, P and NegC 6 (4.0 0.1 g, 4.3 0.3 g and 4.8 1.0 g, respectively). After 14 days spat fed with PS was 4 signiﬁcantly heavier (6.8 1.3 g) than those fed

spawning females (n) 2 on DP or NegC. The weekly growth rate in the ﬁrst week varied 0 <100 100 - 500 500 - 800 from 26.3% (DP) to 66.7% (DS), but declined con- no. eggs (x1000) siderably during the second week for all diets (3.0% (DP)–45.2% (PosC)) (Fig. 6). However, the Figure 4 Spawning result of C. edule female brood- decline was less for diets PosC and PS, where the 9 stock after 6 weeks conditioning with T-Iso (5.87 spat’s growth rate dropped with only 22% and 1010 cells day1 9 100 ind1)orT. suecica (5.87 9 25% respectively. 109 cells day-1 9 100 ind1).

Effect of density and burrowing substrate on respectively (Fig. 5). This is signiﬁcantly heavier cockle growth than the cockles grown on P and DP, that reached an average weight of 3.9 0.4 g and 3.9 Best results were obtained when cockle spat was 0.1 g respectively. At day 14, the differences in cultured inside at ad libitum food conditions and at weight between the diet groups observed at day 7, a density of 500 ind m2 (Fig. 7a). It resulted in

B l 1 10 drastically to an average of 59 m day . At high t = 0 B B B densities, there was no difference in growth rate 8 t = 7 t = 14 and total biomass at day 100 between cockles that abc c c bc A 6 ab were able to bury into the sediment or not. A a aA Cockles cultured outside at densities of 500 4 2 Weight (g) ind m which were able to bury into the sedi- 2 ment had an average growth rate of 65 lm day1 0 between DPF 41–103 and reached an average size PS P DP DS DPS PosC NegC of 9.7 1.4 mm and a total biomass of 2 Figure 5 Effect of different microalgae diets on cockle 229 g m at DPF 103. At similar densities and spat growth (n = 3). culture conditions, cockle spat that was not able to bury into the sediment reached an average size of 7.1 1.4 mm, an average growth rate of 70.0 25 lm day 1 and a total biomass of 105 g m 2 60.0 (Fig. 7b). 50.0 40.0 30.0 Discussion GR (%) 20.0 Broodstock animals in this study were conditioned 10.0 ° ° 0.0 at 12 C and initiated to spawn at 19 C. It is t = 0 t = 7 t = 14 known that spawning can occur at temperatures PS DP DPS NegC around 13°C (Boyden 1971) although another P DS PosC study reported 15.44°C as minimum spawning Figure 6 Weekly growth rate (%) of cockle spat, sub- temperature (Rueda, Smaal & Scholten 2005). No mitted to seven different diets during 2 weeks (n = 3, premature spawning was detected during this only positive st.dev indicated in graph). study, but it would probably be safer to condition broodstock animals at temperatures well below an average growth rate of 168 lm day1 between 12°C to avoid premature shedding of gametes. DPF 41 and 118, an average size of 19.0 There are several ways to obtain gametes from 1.9 mm at DPF 118 and a total biomass of cockles. Boyden (1971) used strip-spawning to 1040 g m2. Cockles cultured under similar food obtain between 7000 and 20 000 ova per individ- conditions, but at 1000 ind m2 had an average ual cockle, but noted that gametes obtained from growth rate of 270 lm day1 between DPF 40 naturally spawning individuals had a better qual- and 60 and a total biomass of 400 g m2 at DPF ity although the initiation of spawning by using 60, keeping up with the low density treatment. temperature stimulus or by the addition of After DPF 60, however, growth rates decreased gametes was not reliable. In this study, only a

(a)Nursery inside (b) Nursery outside 20000 20000 in sand 16000 16000 on sand

12000 on sand low 12000 density size (um)

on sand Size (um) 8000 high density 8000 in sand high density 4000 4000 40 60 80 100 120 140 40 60 80 100 120 DPF DPF

Figure 7 (a) Nursery inside cockle growth at low densities (500 ind m2) and high densities (1000 ind m2) with ad libitum food availability, on and in sand. (b) Outside nursery cockle growth (500 ind m2) at pond conditions, on and in sand.

© 2013 John Wiley & Sons Ltd, Aquaculture Research, 46, 302–312 307 Cultivation of the common cockle A E Pronker et al. Aquaculture Research, 2015, 46, 302–312 moderate temperature rise from 12°Cto19°C was Lucas 2000), overall, T. suecica has only low used, in order to induce the release of only ripe amounts of dietary lipids and similar amounts of gametes. Cockles are very suitable for a ‘mass protein in comparison with T-Iso (Volkman, Jeffrey, spawning’ technique. The eggs are surrounded by Nichols, Rogers & Garland 1989; Brown 1991). a protective jelly layer, which consists of a fibrous However, T. suecica contains twice the amount of network imbedded in gelatinous material. This carbohydrates (12.6% DW versus 6% DW in T-Iso) layer prevents polyspermy (Hagstrom€ 1959) (Volkman et al. 1989; Brown 1991) which are the whereas the increase in size caused by the layer preferred catabolized substrate to fuel cellular promotes egg-sperm encounters at low sperm den- metabolism in bivalves during gametogenesis sities (Farley & Levitan 2001). The advantage of (Bayne et al. 1975; Newell & Bayne 1980; Mathieu this practice is that eggs are immediately fertilized & Lubet 1993). Comparison between the average after release and reduced vitality of the gametes is egg production during the production of the F2 gen- avoided. In addition, the jelly layer also protects eration (1.37 million eggs) and the egg production the eggs until they have reached D-larvae stage during the conditioning experiment (1.3–3 million from shear stress, which can lead to deformations eggs for Iso and Tetra treatment respectively), (Thomas, Edwards, Bolton, Sewell & Zande 1999). strengthens our observation that T. suecica is indeed In this study, no deformed D-larvae were observed. a good diet for cockle broodstock. Broodstock conditioning increased drastically Fertilization rates of 99% were reported for C. la- the number of gametes produced. A total of 22% marcki by Yankson and Moyse (1983). In this (T-Iso) and 32% (Tetra) of the animals spawned, study, no fertilization rate was recorded because whereas none from the starvation treatment. This clumping of the eggs made visual observation can be explained by the fact that the egg quantity difficult. The hatching rate was quite low (63.5%), (and quality) of C. edule is determined during compared with the 98% hatching rate of C. la- gametogenesis, which can be as short as 4 weeks marcki reared in the laboratory (Yankson & Moyse (Boyden 1971; Honkoop & van der Meer 1998). 1983) although the average egg size (70.0 Instead of the gonads redeveloping to a certain 5.1 lm) in this study was very comparable to pre- degree immediately after spawning and maintain- vious work (Labour 1938; Creek 1960; Yankson ing in this stage over winter, as happens in many & Moyse 1983; Honkoop & van der Meer 1998). other lamellibranches, gametogenesis in C. edule in This low hatching rate might be caused by clump- temperate climates immediately precedes spawning ing of the eggs, due to the gelatinous egg mem- (Boyden 1971). This characteristic allows hatch- brane and the high concentration of eggs in the ery managers to control the egg quality by creat- spawning tank and sieve which possibly led to ing optimal conditions for gametogenesis. oxygen depletion or bacterial infection (Yankson & The number of eggs produced by bivalves is influ- Moyse 1983) or might be an effect of insufficient enced by the quantity and quality of the condition- egg reserves as described by Krauter, Castagna ing diet (Bayne, Gabbott & Widdows 1975; and Van Dessel (1982) for Mercenaria mercenaria Heasman, O’Conner & Frazer 1996; Soudant, and Argopecten irradians. However, the big size of Marty, Moal, Robert, Quere, Le Coz & Samain the D-larvae in comparison with other studies does 1996; Utting & Millican 1997; Nevejan, Courtens, not support these hypotheses. The D-larvae at day Hauva, Gajardo & Sorgeloos 2003; Liu, Alabi & 1 (24 h after fertilization) measured 109.0 Pearce 2008). The nutritional value of algae for 3.6 lm, 36% larger than the D-larvae obtained by bivalves is thought to be mainly determined by the Labour (1938) and Creek (1960). amount of essential fatty acids although some sta- The larval growth fitted a linear growth pattern ted that their importance might be overrated (Fig. 1) as observed in other bivalve species (Pech- (Thompson, Guo & Harrison 1993). After brood- enik, Eyster, Widdows & Bayne 1990; Delaunay stock conditioning, females of the Tetra treatment et al. 1992; Nevejan, Saez, Gajardo & Sorgeloos spawned a maximum number of 790 000 eggs, 2003). The daily growth rate of 16.7 lm day1 whereas the ones that received the Iso treatement was very similar to the growth rate of almost only released 393 000 eggs. Although chemical 16 lm day1 of C. edule measured by Kingston composition and thus food value of algae can vary (1974) at 20°C. greatly depending on culturing conditions (Ibarrola, Larval survival from D-larvae till settlement was Navarro, Iglesias & Urrutia 1999; Leonardos & highly variable and ranged between 38% and

80%. Nine days after fertilization larvae settled at 24-methylenecholesterol (Gatenby, Orcutt, Kreeger, an average size of 238 22 lm. This is much fas- Parker, Jones & Neves 2003). Wikfors, Patterson, ter than observed by Labour (1938) and Creek Ghosh, Lewin, Smith and Alix (1996) noted that Tet- (1960) who reported that it takes 2–5 weeks in the raselmis strains yielded the most rapid oyster growth natural environment for pediveliger larvae to meta- in post-set oysters Crassostrea virginica because of the morphose into spat at an average size of 270 lm. higher content in sterols 24-methylcholesterol and/ Kingston (1974) who reared C. edule in the labora- or 24 methylenecholesterol. tory at different temperatures observed that the fast- Downwelling systems are the preferred grow-out est growing larvae metamorphosed after 20– systems for young cockles until they have reached a 24 days at 20°C at a size of 317–351 lm, after size of at least 5 mm to prevent them from drifting being exclusively fed on I. galbana. He also showed away by using a thin mucus thread (Yankson 1986; that cockle larvae were only able to complete their De Montaudouin 1997). After this period, they can metamorphosis into spat under conditions that per- be placed in upwelling systems or sandy ponds for mitted a growth rate in excess of 4 lm day1. further grow-out. According to Creek (1960), juve- This is the first study to assess the suitability of nile cockles reach a size of 600–700 lm after about various microalgae diets for C. edule spat growth. 3 weeks, and need another 3 months to reach a size All well-performing diets contained S. costatum, of 1.5 mm. In this study, however, spat reached an one of the most commonly used algae in aquacul- average size of 327 lm after 3 weeks and reached ture and easy to grow in outdoor algae ponds. It is 11.7 1.8 mm after 3 months. also one of the most common algae species found When algae were sufficiently available, growth in the Scheldt Estuary (Rijstenbil 1987). of juveniles slowed down considerably when cock- Taking into account the decline in growth rate les reached an average size of 1.0 cm and a total during the second week, the diet PS would seem biomass of 400 g m2 at densities of 1000 the preferred choice when indigenous algae are ind m2. At densities of 500 ind m2, however, used. The presumption that the PosC diet, which growth rate remained high up to a total biomass included T-Iso and C. muelleri would be the best diet of 1040 g m2 with an average size of 1.9 cm. proved to be true, although not significantly differ- This difference of final biomass is an indication of ent from the PS diet. Continuation of the experi- density dependent competition. The result is con- ment over a longer time period would have been sistent with Mazon-Su astegui, Ruız-Ruız, Parres- useful to see whether the differences in perfor- Haro and Saucedo (2008), who reported that mance remain on the long-term. At an algae con- hatchery growth of Crassostrea corteziensis spat centration of 8 cells ll1 (NegC), the cockle spat was inversely related to stocking density. Kamer- were underfed. The Iso-equivalent of 50 cells ll1 mans, van der Veer, Karczmarski and Doeglas (PosC, PS) seemed appropriate, leading to fast (1992) on the other hand, found no indication of growth and no pseudo-faeces production. density effects on growth under natural conditions A diet consisting of exclusively S. costatum defi- at densities up to 1984 ind m2 (size range nitely needs to be tested as an alternative diet for 14.5–23.6 mm) in an area of a few square metres. C. edule. P. tricornutum, another native, common However, the growth rate measured was much and easy-to-culture alga rich in protein (Brown, lower (1.14 mm month1) than recorded in this Jeffrey & Garland 1989) turned out to have a very study, explaining the difference with our findings low food value for C. edule. According to Albento- the average growth rates in the indoors nursery sa, Perez-Camacho, Labarta and Fernandez-Reiriz was 5.0 mm month1 for the size class of (1996) this is caused by its tough cell wall, mak- 0–10 mm and 3.6 mm month1 for size class ing it hard to digest. Another explanation of the 10–20 mm (at 500 ind m2). This is very compa- good performance of diets containing S. costatum rable to growth rates of a continuous immersed in comparison with P. tricornutum could be that wild population of C. edule in the Wadden Sea, the former contains approximately 38% measured by Guevara and Niell (1989) being 24-methylenecholesterol (Robert, Chretiennot- 5.5 mm month1 and 3.5 mm month1 respec- Dinet, Kaas, Martin-Jezequel, Moal, Le Coz, Nic- tively. Under food limiting conditions (outdoor cul- olas, Bernard, Connan, Le Dean, Le Gourrierec, ture), cockles that were able to bury into the Leroy & Quere 2004) whereas P. tricornutum sediment grew slightly faster as they could benefit consists of 80-99% brassicasterol and almost no from benthic algae growing on the sediment

© 2013 John Wiley & Sons Ltd, Aquaculture Research, 46, 302–312 309 Cultivation of the common cockle A E Pronker et al. Aquaculture Research, 2015, 46, 302–312 surface. Benthic algae can constitute up to 88% of Acknowledgements the juveniles’ diet (Sauriau & Kang 2000). On a We are grateful to the staff membersof Roem van practical level this means that during the grow- Yerseke B.V. for their support. NIOZ-Yerseke is the out phase, benthic algae are a valuable extra food former Centre for Estuarine and Marine Ecology of source for young cockles, without extra time and the NIOO-KNAW; this is Monitor Taskforce Publi- effort needed to grow them. cation Series 2012-02. The study was funded by Under natural conditions cockles achieve their Roem van Yerseke B.V. and financially supported maximum growth in the first year after settlement by the Province of Zeeland, which we gratefully to ensure maximum size before excessive predation acknowledge. severely limits their numbers (Seed & Brown 1978). Depending on the moment of recruitment and standing crop biomass they can grow to a size References between 4 and 18 mm in their first year (Seed & Albentosa M. Perez-Camacho A. Labarta U. & Fernandez- Brown 1978; Kristensen 1957; Hancock 1973). Reiriz M.J. (1996) Evaluation of live micro algal diets After their first year, growth slows down, espe- for the seed culture of Ruditapes decussatus using physi- cially after their third winter, by that time they ological and biochemical parameters. Aquaculture 148, would have reached a size of more than 26 mm 11–23. (Jensen 1992). Therefore, for commercial purposes Bayne B.L. Gabbott P.A. & Widdows J. (1975) Some it is important to achieve maximum growth dur- effects of stress in the adult on the eggs and larvae of ing the first growing season. By introducing hatch- Mytilus edulis L. Journal of the Marine Biological Associa- ery reared cockle seed in grow-out ponds in early tion of the United Kingdom 55, 675–689. spring, advantage can be taken from this first-year Beukema J.J. & Dekker R. (2006) Annual cockle growth spurt, making it possible to produce Cerastoderma edule production in the Wadden Sea usually fails to sustain both wintering birds and a commercial sized cockles (20 mm) in one year. commercial fishery. Marine Ecology Progress Series 309, 189–204. Conclusion Boyden C.R. (1971) A comparative study of the reproductive cycles of the cockles Cerastoderma edule and This study proves for the first time that cockles C. glaucum. Journal of the Marine Biological Association can be produced successfully in commercial num- of the United Kingdom 51, 605–622. bers and in reliable succession of batches. Besides Brown M.R. (1991) The amino acid and sugar composi- from using broodstock from the wild, it was dem- tion of 16 species of microalgae used in mariculture. onstrated that also hatchery produced cockles (F1) Aquaculture 145,79–99. can act as broodstock animals, thereby closing the Brown M.R. Jeffrey S.W. & Garland C.D. (1989) Nutri- production cycle and opening a window of oppor- tional aspects of microalgae used in mariculture: a lit- erature review. CSIRO Marine Laboratories Report tunities for future broodstock selection. It was pos- 205, 44 pp. sible to obtain several millions of eggs and larvae CBS & PBL (2011). Kokkels in Waddenzee en Zeeuwse at each spawning event, though this part of the Delta, 1990-2010 (versie 04). Website available from production cycle still needs further improvement. www.compendiumvoordeleefomgeving.nl (accessed on Two algal diets were tested for broodstock condi- 17-06-2011), Centraal Bureau voor de Statistiek (CBS, tioning, concluding that T. suecica was better than Den Haag), Planbureau voor de Leefomgeving (PBL, I. galbana (T-Iso) and indicating the possible impor- Den Haag/Bilthoven), Wageningen UR (Wageningen). tance of carbohydrates during gametogenesis. Creek G.A. (1960) The development of the lamellibrach A mixed diet of P. tricornutum and S. costatum, Cardium edule L. Proceedings of the Zoological Society of – both naturally present in the Eastern Scheldt, ful- London 135, 243 260. filled the dietary requirements of spat. This infor- Curatolo A. Ryan M.J. & Mercer J.P. (1993) An evaluation of the performance of Manila clam spat (Tapes mation suggests that it is possible to grow philippinarum) fed on different rations of spray-dried hatchery reared cockle spat till commercial size by algae (Tetraselmis suecica). Aquaculture 112, 179–186. using enriched water from the Eastern Scheldt. De Montaudouin X. (1997) Potential of bivalves’ second- To conclude, above hatchery model indicates that ary settlement differs with species: a comparison 100 broodstock animals allows an estimated produc- between cockle (Cerastoderma edule) and clam tion of 1 million cockles in only 9 months, making (Ruditapes philippinarum) juvenile resuspension. Marine C. edule a very promising new aquaculture species. Biololgy 128, 639–648.

Delaunay F. Marty Y. Moal J. & Samain J.F. (1992) Marine Biological Association of the United Kingdom 23, Growth and lipid class composition of Pecten maximus 119–144. (L.) larvae grown under hatchery conditions. Journal of LEI (2001) De Nederlandse Schelpdiersector. Projectcode Experimental Biology and Ecology 163, 209–219. 65149. url:http://www.kokkels.nl/data/pdf/70698.pdf. Farley G.S. & Levitan D.R. (2001) The role of jelly coats Leonardos N. & Lucas I.A.N. (2000) The nutritional in sperm-egg encounters, fertilization success and selec- value of algae grown under different culture conditions tion on egg size in broadcast spawners. The American for Mytilus edulis L. larvae. Aquaculture 182, 301–315. Naturalist 157 (6), 626–636. Liu W. Alabi A.O. & Pearce C.M. (2008) Broodstock con- Gatenby C.M. Orcutt D.M. Kreeger D.A. Parker B.C. Jones ditioning in the basket cockle, Clinocardium nuttallii. V.A. & Neves R.J. (2003) Biochemical composition of Journal of Shellfish Research 27 (2), 399–404. three algal species proposed as food for captive fresh- Mathieu M. & Lubet P. (1993) Storage tissue metabolism water mussels. Journal of Applied Phycology 15,1–11. and reproduction in marine bivalves – a brief review. Guevara J.M. & Niell F.X. (1989) Growth rates in Invertebrate Reproduction and Development 23, 123–129. a continuously immersed population of Cerastoderma Mazon-Su astegui J.M. Ruız-Ruız K.M. Parres-Haro A. & edule L. Scientia Marina 53, 483–489. Saucedo E. (2008) Combined effects of diet and stock- Hagstrom€ B.E. (1959) Further experiments on jelly free ing density on growth and biochemical composition of sea urchin eggs. Experimental Cell Research 17, spat of the Cortez oyster Crassostrea corteziensis at the 256–261. hatchery. Aquaculture 284,98–105. Hancock D.A. (1973) The relationship between stock Nevejan N. Courtens V. Hauva M. Gajardo G. & Sorge- and recruitment in exploited invertebrates. Rapport loos P. (2003) Effect of lipid emulsions on production et Proces-verbaux des Reunions, Conseil international pour and fatty acid composition of eggs of the scallop Argo- L’Exploration de la Mer 164, 113–131. pecten purpuratus. Marine Biology 143, 327–338. Heasman M.P. O’Conner W.A. & Frazer A.W. (1996) Nevejan N. Saez I. Gajardo G. & Sorgeloos P. (2003) Temperature and nutrition as factors in conditioning Energy versus essential fatty acids: what do scallops brood stock of the commercial scallop Pecten fumatus larvae (Argopecten purpuratus) need most? Comparative Reeve. Aquaculture 143,75–90. Biochemistry and Physiology Part B: Biochemistry and Honkoop P.J.C. & van der Meer J. (1998) Experimentally Molecular Biology 134, 599–613. induced effects of water temperature and immersion time Newell R.I.E. & Bayne B.L. (1980) Seasonal changes in on reproductive output of bivalves in the Wadden Sea. the physiology, reproductive condition and carbohy- Journal of Experimental Biology and Ecology 220, 227–246. drate content of the cockle Cardium (=Cerastoderma) Ibarrola I. Navarro E. Iglesias J.I.P. & Urrutia M.B. (1999) edule (Bivalvia: Cardiidae). Marine Biology 56,11–19. Time-course of digestive-enzyme acclimation in the Pechenik J.A. Eyster L.S. Widdows J. & Bayne B.L. cockle Cerastoderma edule. Marine Biology 135,47–56. (1990) The influence of food concentration and tem- Jensen K.T. (1992) Dynamics and growth of the cockle, perature on growth and morphological differentiation Cerastoderma edule, on an intertidal mud-flat in the Dan- of blue mussel Mytilus edulis (L.) larvae. Marine Biology ish Wadden Sea: effects of submersion time and density. 136,47–64. Netherlands Journal of Sea Research 28, 335–345. Provincie Zeeland (2006) Aquacultuur in Zeeland: Kamermans P. van der Veer H.W. Karczmarski L. & De blauwe revolutie. Province of Zeeland, Zierikzee, 76 Doeglas G.W. (1992) Competition in deposit- and pp. suspension-feeding bivalves: experiments in controlled Rijstenbil J.W. (1987) Phytoplankton composition of outdoor environments. Journal of Experimental Biology stagnant and tidal ecosystems in relation to salinity, and Ecology 162, 113–135. nutrients, light and turbulence. Netherlands Journal of Kingston P. (1974) Some observations on the effects of Sea Research 21, 113–123. temperature and salinity upon the growth of Cardium Robert R. Chretiennot-Dinet M.J. Kaas R. Martin-Jezequel edule and Cardium glaucum larvae in the laboratory. V. Moal J. Le Coz J.R. Nicolas J.L. Bernard E. Connan Journal of the Marine Biological Association of the United J.P. Le Dean L. Le Gourrierec G. Leroy B. & Quere C. Kingdom 54, 309–317. (2004) Amelioration des productions phytoplancto- Krauter J.N. Castagna M. & Van Dessel R. (1982) Egg niques en ecloserie de mollusques: caracterisation des size and larval survival of Mercenaria mercenaria (L.) microalgues fourrage. Ifremer, DRV/RST/RA/LPI/ and Argopecten irradians (Lamarck). Journal of Experi- 2004-05, 150 p. mental Marine Biology and Ecology 56,3–8. Rueda J.L. Smaal A.C. & Scholten H. (2005) A growth Kristensen I. (1957) Differences in density and growth in model of the cockle (Cerastoderma edule L.) tested in the a cockle population in the Dutch Wadden Sea. Archives Oosterschelde estuary (The Netherlands). Journal of Sea Neerlandaises de Zoologie 12, 351–453. Research 54, 276–298. Labour M. (1938) Notes on the breeding of some lamelli- Sauriau P.G. & Kang C.K. (2000) Stable isotope evidence branchs from Plymouth and their larvae. Journal of the of benthic microalgae-based growth and secondary

production in the suspension feeder Cerastoderma edule Utting S.D. & Spencer B.E. (1991) The hatchery culture (Mollusca, Bivalvia) in the Marennes-Oleron Bay. of bivalve mollusc larvae and juveniles. Ministry of Hydrobiologia 440, 317–329. Agriculture, Fisheries and Food (MAFF), Directorate of Seed R. & Brown R.A. (1978) Growth as a strategy for Fisheries Research, Lowestoft, Laboratory leaflet num- survival in two marine bivalves, Cerastoderma edule ber 68, 31 pp. and Modiolus modiolus. Journal of Animal Ecology 47, Volkman J.K. Jeffrey S.W. Nichols P.D. Rogers G.I. & 283–292. Garland C.D. (1989) Fatty acid and lipid composition of Soudant P. Marty Y. Moal J. Robert R. Quere C. Le Coz 10 species of microalgae used in mariculture. Journal of J.R. & Samain J.F. (1996) Effect of food fatty acid Experimental Marine Biology and Ecology 128, 219–240. and sterol quality on Pecten maximus gonad composi- Walne P.R. (1970) Studies on the food value of nineteen tion and reproduction process. Aquaculture, 36, genera of algae to juvenile bivalves of the genera Ostrea, 361–378. Crasostrea, Mercenaria and Mytilus. Ministry of Agricul- Stichting ODUS (2001) Uit de schulp. Visie op duurzame ture, Fisheries and Food; Fishery investigations 26,1–62. ontwikkeling van de Nederlandse schelpdiervisserij. Wijnhoven S. Escaravage V. Herman P.M.J. Smaal A.C. Stichting Ontwikkeling Duurzame Schelpdiervisserij & Hummel H. (2011) Short- and mid-long term effects (ODUS), Yerseke, 48 pp. (in Dutch). of cockle-dredging on non-target macrobenthic species: Tebble N. (1966) British Bivalve Seashells. A Handbook for a before-after-control-impact experiment on a tidal Identification, The British Museum, London, p. 211. mudflat in the Oosterschelde (The Netherlands). Marine Thomas F.I.M. Edwards K.A. Bolton T.F. Sewell M.A. & Ecology 32, 117–129. Zande J.M. (1999) Resistance to shear stress: the role Wikfors G.H. Patterson G.W. Ghosh P. Lewin R.A. Smith of echinoderm egg extracellular layers. The Biological B.S. & Alix J.H. (1996) Growth of post-set oysters, Bulletin 197,7–10. Crassostrea virginica, on high-lipid strains of algal Thompson P.A. Guo M. & Harrison P.J. (1993) The influ- flagellates Tetraselmis spp. Aquaculture 143, 411–419. ence of irradiance on the biochemical composition of Yankson K. (1986) Observations on byssus systems in three phytoplankton species and their nutritional value the spat of Cerastoderma glaucum and C. edule. Journal for larvae of the Pacific oyster (Crassostrea gigas). Mar- of the Marine Biological Association of the United ine Biology 117, 259–268. Kingdom 66, 277–292. Utting S.D. & Millican P.F. (1997) Techniques for the Yankson K. & Moyse J. (1983) Artificial fertilization and hatchery conditioning of bivalve broodstocks and the rearing of Cerastoderma lamarcki (Reeve) in the labora- subsequent effect on egg quality and larval viability. tory. Journal of Molluscan Studies 12a, 209–213. Aquaculture 155,45–54.