Egyptian Journal of Aquatic Research (2013) 39, 125–131

National Institute of Oceanography and Fisheries Egyptian Journal of Aquatic Research

http://ees.elsevier.com/ejar www.sciencedirect.com

FULL LENGTH ARTICLE Spawning frequency, larval development and growth of Muricid gastropod ramosus (Linnaeus, 1758) in the Laboratory at Hurghada, Northern Red Sea, Egypt

Mostafa A.M. Mahmoud *, Tarek A.A. Mohammed, Mohamed H. Yassien

National Institute of Oceanography and Fisheries, Invertebrates Aquaculture Lab., Egypt

Available online 30 July 2013

KEYWORDS Abstract The spawning frequencies; egg capsules clusters, fecundity, growth and development of ; larval stages of Chicoreus ramosus were studied in the laboratory. It lays clusters of egg capsules Spawning; (405 egg capsules) on four frequency times. The capsule length ranged from 19.68 to 20.28 mm. Egg capsule; The number of eggs per capsule varied from 177.8 (±3.58) to 214.3 (±1.61) egg/capsules, where Larval growth; the egg size ranged from 287 · 280.5 to 355.5 · 322 lm. The estimated fecundity of C. ramosus Red Sea was relatively high and reached about 78,748 eggs per throughout the spawning times. After 70 days of hatching from the egg capsules the produced larvae reached about 10.71 mm (±2.16) in length and 5.39 mm (±0.578) in width. The average daily growth rate in length was 0.103 and 0.046 mm/day in width. During the first 2 days, the growth rate was about 0.106 mm/day inside the egg capsules (they feed and nurse on the capsule fluid and on the other small eggs), then it increased to 0.137 mm/day by the 70 days of emerging from the egg capsule because they feed on other juveniles and small gastropods which were placed in the tanks. ª 2013 Production and hosting by Elsevier B.V. on behalf of National Institute of Oceanography and Fisheries.

Introduction the Red Sea whose meat is used as food by the native people. Few investigations have been done on this species in the Red The ramose Chicoreus ramosus (Linnaeus, 1758), is the Sea (e.g. Gohar and Eisawy, 1967) and in the tropical areas largest and most important member of the family in (e.g. Nugranad, 1992; Ruangchoy and Tantichodok, 1992; Xavier-Ramesh et al., 1992; Middelfart, 1996). C. ramosus is * Corresponding author. Address: National Institute of Oceanogra- found widely spread in the Indo-West Pacific, and occurs from phy and Fisheries, Red Sea Branch, Hurghada, Red Sea, Egypt. Tel.: east to South Africa, including Mozambique, Tanzania, Mad- +20 1094576510. agascar, the Red Sea, the Gulf of Oman, Aldabra, Chagos and E-mail address: [email protected] (M.A.M. Mahmoud). Mauritius. It also occurs in eastern Polynesia, southern Japan, Peer review under responsibility of National Institute of Oceanography New Caledonia and Queensland in Australia (Poutiers, 1998). and Fisheries. Sharabati (1985) showed that, C. ramosus was found in inter- tidal rocks, sand and silty areas on shallow reefs in Red Sea. Its meat forms a good substitute to other more expensive protein Production and hosting by Elsevier rich food sources (Xavier-Ramesh et al., 1992).

1687-4285 ª 2013 Production and hosting by Elsevier B.V. on behalf of National Institute of Oceanography and Fisheries. http://dx.doi.org/10.1016/j.ejar.2013.07.002 126 M.A.M. Mahmoud et al.

Muricids are commercially important gastropods and are Materials and methods exploited by fishermen for shell craft industries. Few species belonging to the Chicoreus are listed as endangered Four mature specimens of C. ramosus (Fig. 1a) were collected under Wild life Act 1972 (Jagadis et al., 2013). The study by snorkeling at 2–3 m depth from the inshore area at Hurgh- of the spawn, egg capsules and embryos of gastropods is ada City, Red Sea during January 2010. The collected important not only for studies of reproductive biology in were placed in a 2 m3 capacity rectangular fiberglass tank in general, but also for its implications for dispersal and bioge- the wet laboratory at NIOF, Red Sea Branch. Continuous flow ography and (Pastorino et al., 2007). Muricidae of well aerated sea water was used. embryos are maintained within protective capsules and fed Water temperature (C), salinity (&) and pH were mea- on nutritive sources during development into planktotrophic sured daily inside the tanks using multiparameter survivor 4, veligers, veliconcha or metamorphosed juveniles (Romero where the water temperature was controlled in the tanks using et al., 2004). heaters and the water temperature ranged between 19 and Encapsulation is a common strategy observed among mar- 24 C. The above mentioned parameters are listed in Table 1. ine caenogastropods. Although the capsule confers protection Three aspects of the spawns were observed; number and for embryos, its rigidity and strength pose a significant chal- size of egg capsules, number and size of eggs and time of lenge as the larvae hatch (Gallardo et al., 2013). Also encapsu- egg development. Egg capsule numbers were counted in each lation of early ontogenetic stages is facilitated by the secretion spawning time by eyes from the tank walls, and their dimen- of capsule laminae around embryos within the capsule gland, sions such as length and width were measured with a vernier and by sculpturing, hardening and attachment of capsules by caliper of 0.01 mm accuracy. The number of eggs per capsule the ventral pedal gland (D’Asaro, 1988). was measured, by breaking the capsule, with an ocular C. ramosus is one of the gastropods undergoing laboratory micrometer under an optical microscope based upon 8–10 evaluations as candidates for aquaculture (Nasution and capsules in each spawning time. Developmental stages from Roberts, 2004). Bussarawit and Ruangchoy (1991) stated egg to hatching were observed daily with a stereoscopic that, the hatching of C. ramosus occurs within 25–28 days microscope. after spawning and it spawns from June to December in For the study of fecundity in C. ramosus, the number of egg Thailand. capsules in a single cluster was counted and for counting the Nugranad (1992) studied the experimental rearing of larval total number of eggs, 40 egg capsules, mostly from the middle stages of C. ramosus in Thailand from September to April. He portion of each egg clusters were used. A small slit was made showed that eggs are laid within vase shaped capsules clustered near the apical plate, and with a pipette the eggs with albumin together ranging from 205 to 712 capsules per cluster, the num- fluid were sucked out and counted using a binocular micro- ber of eggs per capsule varied from 116 to 353 eggs for spawn- scope according to Xavier-Ramesh et al. (1992). ing in capacity, and from 217 to 734 eggs for the wild spawn Three capsules were dissected daily in the first week of and the growth rate of the larvae reached to 1295 lmin spawning in each spawning time to obtain the egg measure- 15 days and to 1375 lm after 20 days. ments and their development and metamorphosis. Then four The present work aims to determine the spawning fre- capsules were dissected weekly to follow the larval quencies; egg capsule clusters, fecundity, growth and develop- development up to 70 days of hatching from the egg capsules. ment of larval stages of C. ramosus under laboratory During incubation, the larval development was observed daily. conditions. The juveniles were placed in small tanks to allow the daily

a b

7 cm 7 cm

c d

1mm e

Figure 1 Showing the dorsal and ventral view of Chicoreus ramosus (a); the animal during releasing of egg capsules in the 1st frequency (b); the shape of egg capsules (c), unfertilized eggs (d) and fertilized eggs before hatching (e). Spawning frequency, larval development and growth of Muricid gastropod Chicoreus ramosus (Linnaeus, 1758) 127

The total number of the egg capsules of C. ramosus reached Table 1 The controlled environmental conditions (tempera- to 405 capsules throughout the four batches of spawned cap- ture, salinity and pH) in the laboratory during the period of sules (118, 126, 97 and 64 capsules, respectively). Each capsule study. is vase-shaped with convex and narrow concave sides (Fig. 1c). Months Temperature (C) Salinity (&)pHIt has cream white color with a middle aperture on the apical Nov. 22.62 39.75 8.05 part and contains a gelatinous material to keep and protect the Dec. 21.95 40.65 8.03 egg masses inside. Jan. 19.86 41.10 7.98 Feb. 19.56 42.05 7.98 Egg number, size and fecundity Mar. 21.21 40.86 7.98 Apr. 22.84 40.54 8.03 May 23.75 40.81 8.01 The average number of eggs per capsule ranged between Jun. 24.08 40.93 8.02 177.8 ± 3.58 and 214.3 ± 1.61 eggs/capsule (n = 10 capsules). Jul. 4.22 40.95 8.01 However, the ratio of fertilized eggs ranged between 37.59% and 43.87% of the average recorded eggs per capsule in addi- tion to the observation of few unfertilized nurse eggs (Table 3). The fertilized egg sizes ranged from 287 · 280.5 to 355.5 · observations and measurements during their growth. Eggs and 322 lm (based on 8–10 eggs in each spawning time). It was ob- early larval stages were photographed with a camera linked to served that, only 12–27 eggs are developed to the embryonic a binocular stereoscopic microscope and their dimensions were stage in each capsule (Fig. 1d and e), while the remaining eggs measured in micron using micrometer while the other late were used by the developed embryos as food. The estimated to- stages dimensions were measured in millimeters using a vernier tal fecundity of C. ramosus reached about 78,748 eggs through- caliper of 0.01 mm accuracy, standard deviations (SD) are gi- out the four spawning times. This estimated fecundity varied ven in parentheses. among the different spawning times. The average fecundity ranged between 20,787 and 24,469 eggs (Table 2). Results Larval development and growth Spawning frequencies and egg capsules C. ramosus was developed entirely within an egg capsule up to Only one female lays egg capsule clusters four times during the juvenile stage in about 40 days. The earlier larvae reached period of March 21 to May 29 on the tank walls in the labora- about 380–440 lm on the 3rd day after hatching. The prevelig- tory (Fig. 1a and b). They secreted gelatinous materials inside er stage appeared on 8th day and reached 610–670 lmin and outside the capsules for the protection of egg masses. The length (Fig. 2a), then its growth rate increased and developed egg capsules were spawned in clusters. These lasted for a per- into veliger larvae on the 12th day where they reached iod of time ranging from 5 to 9 days per each cluster 820–955 lm in length (Fig. 2b and c). The veliger larvae were frequency. The average length of the egg capsules ranged be- a translucent and have no calcified shell. A well-developed veli- tween 19.68 and 20.28 mm (Table 2) throughout the recorded ger larva was found to have a developed foot (1.15–1.22 mm) frequencies. on the 29th day of hatching (Fig. 2d and e). With further devel-

Table 2 Average egg capsule sizes and numbers of spawning days. Spawning times Egg capsules Number Number of spawning days Fecundity Length (mm) Width (mm) 1st time 19.95 ± 0.28 5.51 ± 0.15 118 9 22,113 2nd time 19.96 ± 0.27 5.50 ± 0.14 126 6 24,469 3rd time 19.68 ± 0.28 5.51 ± 0.15 97 6 20,787 4th time 20.28 ± 0.36 5.51 ± 0.15 64 5 11,379 Total – – 405 26 78,748

Table 3 Average numbers and sizes of eggs inside the egg capsule. Spawning times Average number of eggs Fertilized egg Average number of eggs/capsule (n = 10) Max Min Fertilized eggs obtained Length (mm) Width (mm) No. % 1st time 187.4 ± 2.39 197 176 77 41.09 287.0 ± 9.90 280.5 ± 0.71 2nd time 194.2 ± 2.29 203 179 73 37.59 306.0 ± 5.66 282.5 ± 2.12 3rd time 214.3 ± 1.61 219 205 87 40.60 294.0 ± 1.63 281.0 ± 1.15 4th time 177.8 ± 3.58 197 158 78 43.87 355.5 ± 22.52 322.0 ± 1.63 128 M.A.M. Mahmoud et al.

a b c

160 µm 240 µm 240 µm d e

240 µm

Figure 2 Showing the larval developmental stages of Chicoreus ramosus inside the egg capsules before shell formation; pre-veliger larvae (a), larvae after formation of cilia and eyespots in lateral view (b) and ventral view (c); larvae with fully developed foot in dorsal (d) and ventral view (e).

Table 4 Shell length, growth rate in length per day, shell width and growth rate of width of C. ramosus juvenile from 1 day before emerging (b.e.) from capsule to 70 days. Time Shell length (mm) Growth rate in Shell width (mm) Growth rate length (mm/day) in width (mm/day) 1 day (b.e.) 1.54 ± 0.031 – 1.03 ± 0.009 – 2 day 1.86 ± 0.091 0.106 1.14 ± 0.019 0.035 15 day 3.35 ± 0.467 0.115 2.25 ± 0.229 0.085 25 day 4.45 ± 0.273 0.110 2.69 ± 0.174 0.044 70 day 10.71 ± 2.160 0.137 5.39 ± 0.578 0.046

opment these larvae formed transparent shells that measured growth rate increased gradually and measured 0.155 mm/day about 1.54 mm (±0.031) in length and 1.03 mm (±0.009) in in length and 0.085 mm/day in width by the 15 days of releas- width, on the 39th day (Table 4, Figs. 3a and 4). ing from capsule and measured 0.110 mm/days in length and After 40 days from egg fertilization, only 11–14 embryos 0.044 mm/day in width by the day 25. Finally, by 70 days of per capsule were released and emerged from the egg capsules emerging from the egg capsule, the growth rate reached as juveniles. The juvenile shells, on the 2nd day after emerging 0.103 mm/days in length and 0.046 mm/day in width (Table from the egg capsules, appeared to be a translucent. The aver- 4). They feed on other juveniles and small gastropods which age length was 1.86 mm (±0.09) and the average width was were placed in the tanks. 1.14 mm (±0.02) (Fig. 3b). On the 15th day the juvenile grew and measured an average length of 3.35 mm (±0.47) and aver- age width 2.25 mm (±0.25) (Fig. 3c). On the 25th day after Discussion hatching from capsules the length reached to 4.45 mm (±0.27) and the width reached to 2.69 mm (±0.17) With over 1600 extant described species, the Muricidae is one (Fig. 3d). After 70 days, the recorded length increased to be of the most species-rich and morphologically diverse families 10.71 mm (±2.3) and the width was 5.39 mm (±0.65) (Table of molluscs. As predators of molluscs, polychaetes, anthozo- 4 and Fig. 3e). ans barnacles and other invertebrates, they form an important During the first 2 days, the growth rate was relatively low component of many benthic communities (Barco et al., 2010; 0.106 mm/day in length and 0.035 mm/day in width due to Merle et al., 2011). Genus Chicoreus comprises many different the limited available food within the capsule. They feed and species, however C. ramosus is one of the most widely distrib- nurse on the capsule fluid and on the other small eggs. The uted species (Poutiers, 1998). According to Bouchet (1989), the Spawning frequency, larval development and growth of Muricid gastropod Chicoreus ramosus (Linnaeus, 1758) 129

a b d

c e

Figure 3 The shell growth and metamorphosis; inside the capsule (a), 2 days after hatching (b); 15 days after hatching from the capsule (c), 25 days after hatching (d) and 70 days after hatching (e).

12.00 10.71

( (1958) and related to the capsules described by Xavier-Ramesh 10.00 et al. (1992) and Ruangchoy and Tantichodok (1992) off And- mm 8.00 omon Sea Coast of Thailand. They stated that, the capsules of th ( M. virginus and C. ramosus contained a large number of egg 6.00 4.45 grow capsules (25–267 capsules). 3.35 The number of eggs is depending on the availability of food 1.86 1.54 and the environmental influences (Webber, 1977), which may verage A be considered as controlling factors such as water temperature, 0.00 salinity and pH (D’Asaro, 1991; Abdelsalam and Tanida, One day befor After 2 days After 15 days After 25 days After 70 days hatching 2013). During the present study, the obtained number of egg capsules (405 capsules) are within the range (205–1214 cap- Figure 4 Average shell length growth during 70 days of hatching sules) of Ruangchoy and Tantichodok (1992) and Xavier- from the capsules. Ramesh et al. (1992) but is very high when compared with Chicoreus sp. given by Nugranad (1992) (Table 5). non-planktotrophic development in mollusks are observed in The egg mass and fecundity in the present study is relatively two different situations: the first; when the larva hatches as a high (mean of 78,532 eggs/individual) compared to (11,70 swimming non-feeding veliger and metamorphosis will occur 0eggs/individual) Gohar and Eisawy (1967) but lies in the ob- after a few hours to a few days in the plankton (lecithotrophic tained range (68,360–15,8000 eggs/individual) of Ruangchoy development), the second one; when metamorphosis occurs be- and Tantichodok (1992). The number of eggs per capsule is rel- fore emerging from the capsule, often said to be intracapsular atively low (177–214 egg/capsule) in the present study but lies metamorphosis. C. ramosus belongs to the second situation; it within the range obtained by the later authors who stated that, spent a long time (40 days) where larvae metamorphosis was the capsules provide the egg, larvae and embryos with protec- done within the capsules before they emerged as young juve- tion, survival and play a vital role in their nourishing (Kohn, niles. Xavier-Ramesh et al. (1992) reported that, the mortality 1983). of C. ramosus decreased in the months of February and Khalil and Mousa (2013) stated that for the successful March, where the maximum spawning is associated with the propagation of any species, larval rearing is considered a ma- prevailing low temperature in January. On the other hand, jor bottleneck. Nugranad (1992) stated that, larvae growth rate according to Kasinathan et al. (1975), spawning extended from is relatively slow and reached about 4 mm length by the fifth September to December in the gastropods Thais margaraticole month. While the growth rate observed in the present study in both field and laboratory. The spawning of C. ramosus in is relatively higher than this rate and ranged between 4.15 the present study extended from March to late May in the and 4.80 mm lengths by the 25th day after hatching from the laboratory. However, the maximum value was observed in capsules then reached more than 12.75 mm by the third month March and April but the minimum value was in May. after hatching from the capsules. The growth rate in the pres- Moreover, the shapes of the egg capsules were closely sim- ent study is faster than that obtained by Nugranad (1992) dur- ilar and to these of Murex virginus as described by Natarajan ing the first 3 months of hatching (Table 6). The growth rate of 130 M.A.M. Mahmoud et al.

Table 5 Number of egg capsules, eggs per capsules and fecundity in the present study and the previous works on C. ramosus. Number of egg capsules Number of egg capsules Fecundity Species References 12 278–367 4093 C. ramosus Nugranad (1992) 205–1214 132–735 68,360–1,58,000 C. ramosus Ruangchoy and Tantichodok (1992) 67–698 25–267 – C. ramosus Xavier-Ramesh et al. (1992) 405 177–214 78,748 C. ramosus The present study

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