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Some Reproductive Biological Aspects of Gray-Eel Catfish, Plotosus canius Hamilton, spawner in Pattani Bay, Thailand Journal: Songklanakarin Journal of Science and Technology Manuscript ID SJST-0-0.R Manuscript Type: Original Article Date Submitted by the Author: -Sep-0 Complete List of Authors: Amornsakun, Thumronk; price of Songkla University, Pau, Tan; Universiti Malaysia Terengganu, School of Fisheries and Aquaculture Sciences Keyword: Gray-eel catfish, Plotosus canius, Fecundity, Gonadosomatic index

Page of 0 0 0 0 0 0 Some Reproductive Biological Aspects of Gray-Eel Catfish, Plotosus canius Hamilton, Spawner in Pattani Bay, Thailand Thumronk Amornsakun * Boonsong Krisornpornsan Pairat Jirasatian Thodsaphol Pholrat Tan Min Pau and Anuar bin Hassan Fisheries Technology Program, Department of Technology and Industries Faculty of Science and Technology, Prince of Songkla University, Pattani Campus, Pattani, Thailand 000 * Corresponding author E mail: thumronk.a@psu.ac.th thumronk.a@gmail.com Agriculture Technology Program, Department of Technology and Industries Faculty of Science and Technology, Prince of Songkla University, Pattani Campus, Pattani, Thailand 000 Extension office Prince of Songkla University, Pattani Campus Pattani, Thailand 000 Pattani Coastal Aquaculture Station Pattani, Thailand 000 School of Fisheries and Aquaculture Science Universiti Malaysia Terengganu Mengabang Telipot, Kuala Terengganu, Malaysia 00

Page of 0 0 0 0 0 0 Abstract Reproductive biology of gray-eel catfish, Plotosus canius was studied at Pattani Bay by determining its year round changes of gonadosomatic index (GSI). Results shown that the GSI was maximum in October (. %), and minimum in April (0. %). The sampled size of sexually matured female fishes ranged from 0. to 0. cm in total length, and between. to,. g in body weight. The average total length of the female fish is. cm, with an average body weight of. g. The average fecundity was,. egg/fish. The average GSI was 0. % (n=). The fish total length (TL) and body weight (BW) of the matured fish were linearly related, and could be represented by the linear regression as: BW =. TL.. The relationship between body weight (BW) and fecundity (Fe) could also be represented by the linear regression as Fe =. +.BW. The eggs were adhesive demersal egg type, and rounded. The distribution of egg s diameter could be categorized into five groups i.e. group (.%), group (.%), group (.0%), group (. %), and group (. %), with values of.± 0. mm,.±0. mm,.±0. mm,.±0. mm, and.±0. mm respectively. The spawning ground is made of silty clay soil, with sediment s ph of.±0. and water salinity of.0 ± 0. ppt. We also found that the sperms activity was most active in the 0. % sodium chloride s concentration level. Keywords: Reproductive biology; Pattani Bay; GSI; Gray-eel catfish, Plotosus canius

Page of 0 0 0 0 0 0. Introduction The gray-eel catfish, Plotosus canius, is a commercially important species for brackish water fisheries. This species is one of the main species currently being cultured, and is in great demand by the aquaculture industry. Unfortunately, the commercial scale propagation of this fish in hatchery is yet to be standardized. Specifically, there is insufficient information on the propagation procedures of the fish such as type of breeding, stimulating the gonad, larval rearing, and the spawner preparation (Tarnchalanukit et al., ). Recently, several studies focusing on this species have been published, dealing with the maturity characteristic of the male and female fish (Siritanawong, 00), fecundity and egg (Khan et al., 00; Trueman, 00), ovarian development (Laurenson et al., ), density of spermatozoa (Runsirikul et al., 00), and study on the gonadosomatic index (GSI) for the female and male in Songkhla Lake, Thailand (Runsirikul et al., 00). The aim of this study were to investigate the fecundity, the egg diameter, gonadosomatic index (GSI) of mature female fish, sperm activity, and physiochemical sediment parameters of the spawning ground. These parameters will provide useful baseline information for optimization of fry production, and ultimately for the culture management of this fish in the future.

Page of 0 0 0 0 0 0. Materials and methods Reproductive biology of the P. canius was studied for a one-year cycle, starting from October st, 00 to September 0 th, 0 at Pattani Bay, Thailand. Field experiments were carried out using Complete Randomized Design method, and data analyzed using SPSS/PC + program. The Plotosus canius sexual dimorphism of male fish and female fish are shown in Figure and Figure, respectively. Ten samples of female fish were collected per month for the reproductive study. Eleven sampling stations around at Pattani bay were used in order to determine physio-chemical parameters of sediment, and the water salinity at the spawning ground. All sampling were done in triplicate. The sexual maturity of P. canius was studied by determining its fecundity, egg s diameter, and GSI. Fecundity estimation was made by using a gravimetric method (Tarnchalanukit et al., ). Egg s diameter was measured using ocular microscope (Amornsakun et al., 0). GSI was calculated by using the formula (weight of ovary / weight of body) x 00 (Tarnchalanukit et al., ). The sperm motility of the male fish were also studied, by investigating the saline solution s effect on sperm motility. Three samples of matured male fish obtained from Pattani bay were injected with suprefact acetate (Buserline) 0 µg/kg mixed with motilium (Domperidone) 0 mg/kg (Amornsakun, ). The fish abdomen was dissected after hours of injection to take the testis. Later, the sperm were mixed with sodium chloride solution at different concentration levels such as 0, 0., 0., 0. and % for 0 seconds (Westin and Nissling, ). The movement of sperm was observed and then recorded. The data were then subjected to the analysis of variance for movement percentage of sperm in each sodium chloride solution, and followed by study on differences in each sodium chloride solution using the Duncan s new multiple range test (Walpole and Myers, ).

Page of 0 0 0 0 0 0. Results Gonadosomatic index (GSI) is a strong indicator of a fish maturity that strongly relate to the fish reproductive potential. The result of this work indicate that the GSI of Plotosus canius was at the maximum level in October (. %), and was at the minimum level in April (0. %) as shown in Table and Figure. The sampled size of sexually matured female fishes used in this work ranged from 0. to 0. cm in total length, and between. to,. g in body weight. The average total length of the female fish is. cm, with an average body weight of. g. The average fecundity was,. egg/fish. The average GSI was 0. % (n=) as in Table and Figure. The total length (TL) and body weight (BW) of matured fish were linearly related, and could be represented by the linear regression as: BW =. TL -., R = 0., n=..figure. Meanwhile the relationship between body weight (BW) and fecundity (Fe) could also be represented by the linear regression as: Fe =. +.BW, R = 0., n=...figure. The egg s type were adhesive demersal egg, and rounded. The distribution of the fish egg s diameter could be categorized into five groups i.e. group (.%), group (.%), group (.0%), group (. %), and group (. %), with values of.± 0. mm,.±0. mm,.±0. mm,.±0. mm, and.±0. mm respectively (mean ±SD, n=0) (Table ). By observing the egg s diameter of group, and, lead us to conclude that the fish is ready for spawning

Page of 0 0 0 0 0 0 The physio-chemical parameters of the sediment at the spawning ground (in August through October) such as ph, Organic matter, Phosphorus, Potassium, Calcium, Magnesium, Sand, Silt, and Clay were found as follows:.±0.,.±.0 g kg -,.±. mg kg -,,0.±. mg kg -,,.±,. mg kg -,.±. mg kg -,.0±. %,.0±. % and.0±. %, respectively. Water salinity was.0 ± 0. ppt (mean ±SD, n=). The results of physio-chemical sediment, and water salinity were not significantly different (P>0.0) among the eleven stations (Table and Figure ). The testis of P. canius (Figure ) were removed in order to study the sperm motility. The average percentage of sperm motility in each treatment were 0 %,. %,.0 %, 0. % and. %, respectively. The movement of sperms detected at NaCl concentration level of 0. % was significantly higher as compared to other NaCl concentration levels (P<0.0) (Table ).

Page of 0 0 0 0 0 0 Discussion Gray-eel catfish, Plotosus canius is a commercially important species for brackish water fisheries. The egg s type were adhesive demersal egg and rounded. The female fish size at sexual maturity was. cm in average total length, with. g in average body weight, and the average fecundity was,. egg/fish. For handling purpose, this fish can be considered as a medium size fish. The relatively low fecundity indicates that the natural behavior of the spawner is to take care of the newly hatched larvae by constructing a nest for spawning (Tarnchalanukit et al., ). Comparatively, the fecundity of the P. canius was lesser than the other species such as Siamese gourami, Trichogaster pectoralis which was reported to have, egg/fish, and sexually matured size was at.0 cm in total length and.0 g in average body weight (Amornsakun et al., 00). In addition Amornsakun et al. (00) reported the size of sexually matured female Climbing perch, Anabas testudineus, was.0 cm in total length and.0 g in body weight, and the fecundity was,0. egg/fish. Fecundity varies with different species depending on age, length-weight, environmental, and some other parameters. (Ghafari and Jamili, 00; Shivasanthini et al., 00; Zalina et al., 0). We found that the GSI of matured gray-eel catfish was. %, which is comparatively similar to the other fishes. Normally the GSI value of catfish was to % (Tarnchalanukit et al., ). Several studies have reported that the GSI of Siamese gourami was 0.%, while climbing perch was 0.% (Amornsakun et al., 00; Amornsakun et al., 00).

Page of 0 0 0 0 0 0 The spawning season of P. canius in Pattani Bay and Songkhla Lake, Thailand are at the same time period, i.e. in October (Runsirikul et al., 00). Diameter of fish egg for spawning time were in the range of.-. mm. It size is bigger than various other fish such as red-tail catfish, Mystus wyckioides (. mm), Siamese gourami, Trichogaster pectoralis (0. mm), climbing perch, Anabas testudineus (0. mm), and snake head fish, Channa striatus (0. mm) (Amornsakun, ; Amornsakun et al., 00; Amornsakun et al., 00; Amornsakun et al., 0). Loya and Fishelson () have suggested that, to be suitable for brackish water fish rearing, the main soil texture environment at spawning ground should be classified as silty clay with the sediment s ph at neutral (Pattani Bay ph is.±0.), and water salinity not more than 0 ppt (Pattani Bay salinity is.0 ±0. ppt). In order to survive and propagate successfully, the fish s sperm need to be able to tolerate some salt concentration in the water. In this work, it was shown that the movement of sperms in the 0. % sodium chloride (NaCl) concentration level was higher than the any other sodium chloride (NaCl) concentration levels, which means that the most suitable concentration of sodium chloride solution for sperm dilution is 0. %. For a good fertilization rate, the spermatozoa should be able to enter the micropyle of ovary within seconds. Kudo et al. () reported that European catfish sperm entered to micropyle of egg within 0-0 seconds after fertilization, and minutes later, the activity of spermatozoa slowed down significantly. In conclusion, for a sexually matured gray-eel catfish, the average total length and body weight were. cm and. g, respectively, while the average fecundity of the fish was,. egg/fish. When the fish is ready to spawn, the egg s diameter will ranged from. to. mm. The GSI of gray-eel catfish will varied as a function of time. Normally, as the GSI reached.% (in October), the fish is

Page of 0 0 0 0 0 0 ready to spawn. The result also shown that geographical location is not an important factor, since P. canius at Pattani Bay and P. canius at Songkhla Lake (Runsirikul et al., 00) spawned at the same time, which is in October. Concerning the effect of concentration of NaCl solution on the movement of sperm, it was shown that concentration of NaCl solution at 0.% has higher sperm s movement count than the other concentrations. Especially important for spawning is the environment at the spawning ground which should have neutral sediment s ph (.±0.), and water salinity of.0±0. ppt. which is considered similar to the other brackish water fish s requirement.

Page 0 of 0 0 0 0 0 0 Acknowledgements I am grateful to the National Research Council of Thailand (SAT 00S) for financial support of the field work. I also thank Miss Veeranut Kuntiyapiruk and Miss Siriporn Sopolpongsathon for assistance in research. I would also like to thank Assoc. Prof. Dr. Buhri Bin Arifin for his editorial assistance in preparing this manuscript. References Amornsakun, T.. Some aspects in early life stages of larval red-tail catfish, Mystus wyckioides. Songklanakarin Journal of Science and Technology. (), 0-0. Amornsakun, T., Sriwatana, W. and Promkaew, P. 00. Some aspects in early life stage of Siamese gourami, Trichogaster pectoralis (Regan) larvae. Songklanakarin Journal of Science and Technology. (), -. Amornsakun, T., Sriwatana, W. and Promkaew, P. 00. Some aspects in early life stage of climbing perch, Anabas testudineus larvae. Songklanakarin Journal of Science and Technology. (), 0-. Amornsakun, T., Sriwatana, W. and Promkaew, P. 0. Some aspects in early life Stage of snake head fish, Channa striatus larvae. Songklanakarin Journal Science and Technology. (), -. Ghafari, S.M. and Jamili, S. 00. Certain aspects of the reproductive biology of berzem, Barbus pectoralis in Karoon River. Journal of Fisheries and Aquatic Sciences., -. Khan, M.S.A., Alam, M.J., Rheman, S., Mondol, S. and Rahman, M.M. 00. Study on the fecundity and GSI of brackish water catfish Plotosus canius (Hamilton-Buchanan). Journal of Biological Sciences. (), -.

Page of 0 0 0 0 0 0 Kudo, S., Linhart, O. and Billard, R.. Ultrastructural studies of sperm penetration in the egg of the European catfish, Silurus glanis. Aquaculture Living Resource. -. Laurenson, L.J.B., Neira, F.J. and Potter, I.C.. Reproductive biology and larval morphology of the marine plotosid Cnidoglanis macrocephalus (Teleostei) in a seasonally closed Australian estuary. Hydrobiologia., -. Loya, Y. and Fishelson, L.. Ecology of fish breeding in brackish water ponds near the Dead Sea (Israel). Journal Fish Biology., -. Runsirikul, J., Laongsiriwong, L. and Donyadon, Y. 00. Study on reproductive biology of canine catfish-eel, Plotosus canius. In Seminar coastal aquaculture 00, Nevada Grand Hotel, Ubolratchatani province, 0- June 00. p 0. (In Thai) Shivasanthini, K., Charles, G.A. and Shutharsan, S. 00. Fecundity studies of Gerres abbreviates (Bleeker, 0) from the Jaffna lagoon, Sri Lanka. Journal of Fisheries and Aquatic Sciences., -. Siritanawong, B. 00. Study on water quality and environment at spawning ground of canine catfish-eel, Plotosus canius. Faculty of Agriculture Technology, Rajapat Petchaburi Institute. 0 p. (in Thai) Tarnchalanukit, W., Chuapoehuk, W., Suraniranat, P. and Na Nakorn, U.. Pla Duk Dan Culture. Thailand: Faculty of Fisheries, Kasetsart University. pp. (in Thai) Trueman, W.T. 00. Methods for the Hatchery production of the freshwater jewfish of eel tailed catfish, Tandanus. Native Fish Australia Technical Report. p. 0

Page of 0 0 0 0 0 0 Walpole, R.E. and Myers, R.H.. Probability and Statistics for Engineers and Scientists. nd Edition. Macmillan Publishing Co., Inc., New York, U.S.A. 0 p. Westin, L. and Nissling, A.. Effect of salinity on spermatozoa mortality, percentage of fertilization eggs and egg development of Baltic cod (Gadus morhua), and implications for cod stock fluctuations in the Baltic. Marine Biology. 0, -. Zalina, I., Saad, C.R., Christianus, A. and Harmin, S.A. 0. Induced breeding and embryonic development of climbing perch (Anabas testudineus, Bloch). Journal of Fisheries and Aquatic Science. (), -0.

Page of 0 0 0 0 0 0 Figure. Sexual dimorphism of male fish a) Head of male fish b) Urogenital papillae of male fish b a

Page of 0 0 0 0 0 0 Figure. Sexual dimorphism of female fish a) Head of female fish b) Urogenital papillae of female fish a b

Page of 0 0 0 0 0 0 GSI (%) 0 Month Figure. Gonadosomatic index (GSI, %) of the gray eel -catfish in year round

Page of 0 0 0 0 0 0 Figure. Ovary of gray eel-catfish

Page of 0 0 0 0 0 0 Body weight (g) 00 00 000 00 00 00 00 0 Figure. Body weight-total length relationship y =.x -. R² = 0. 0 0 0 0 0 0 0 0 Total length (cm)

Page of 0 0 0 0 0 0 Number of egg (egg) 00 000 00 000 00 000 y =.x +. R² = 0. 00 0 0 00 00 00 00 000 00 00 Body weight (g) Figure. Fecundity-body weight relationship

Page of 0 0 0 0 0 0 Figure. Sampling station in Pattani bay

Page 0 of 0 0 0 0 0 0 Figure. Testis of gray eel-catfish

Page of 0 0 0 0 0 0 Table Gonadosomatic index (%) of gray eel catfish in year round at Pattani bay, Thailand Sample Jan Feb Mar April May June July Aug Sep Oct Nov Dec 0. 0. 0. 0. 0. 0... 0.. 0. 0. 0. 0. 0. 0. 0. 0.. 0. 0.. 0. 0. 0. 0. 0. 0. 0. 0. 0. 0.0. 0. 0. 0. 0. 0. 0. 0.. 0.. 0.. 0. 0. 0... 0. 0. 0. 0. 0. 0. 0..0 0. 0. 0. 0. 0. 0. 0.. 0. 0. 0.. 0. 0. 0. 0. 0. 0. 0. 0.. 0. 0.. 0... 0. 0. 0. 0. 0. 0. 0. 0. 0. 0. 0. 0. 0.. 0. 0. 0.. 0.. 0.. 0. 0 0. 0. 0. 0. 0. 0. 0... 0. 0. 0. Mean.0 a. a. a 0. a. a. a. b.0 b. b. c.0 a 0. a SD... 0...... 0...0

Page of 0 0 0 0 0 0 Table Body weight (g) total length (cm) fecundity (egg) and gonadosomatic index (GSI, %) of matured gray-eel catfish Sample Body weight (g) Total length (cm) Fecundity (egg) GSI (%). 0...... 0.. 0.. 00... 0 0.... 0... 0.. 0... 0.. 0 0........ 0 0.. 0. Mean... 0. SD....0 Min. 0.. Max. 0..

Page of 0 0 0 0 Table Diameter of gray-eel catfish egg (mm) (N=0) Sample 0 0 0......0..................0............0..0............0........0....0.0...0..0...0.0.....0.0.0....0.0.0...0.0.0...0..................0....0....0..0..0........0....0...0........0.....0......0.....0..0.............0.0...........................................0................................................. 0............0.......0.0............0.....0..0...0.0.0.0.0.0.0.0...0.0..0..0...0......0..............0..............0...............................0....0.0.0............0..0......0.........................0........0.0.....0.0........0..............0.........0....0.........0.0..0..

Page of 0 0 0 0 Table The average of physio-chemical sediment at the spawning ground in August-October Station ph OM(g/kg) P(mg/kg) K(mg/kg) Ca(mg/kg) Mg(mg/kg) Sand(%) Silt(%) Clay(%) Water salinity (ppt). a. abc 0. d. ab. bcd 0. cde 0.0 a. bc. bcd. ab. a. bcd. abc. cd. a.0 de. b 0. ab 0. b. abc. a. bcd. a. abc 0. ab. cde 0. a. d. a. abc. a. bcd. abc 000. abcd. abc. ab. a. bc. bc. a.0 a. ab. abc. bcd. a 0.0 bcde. b. ab. bc. abc. a. a. ab. a. abc. a. b 0. ab 0. b.00 c. a. cd. c. bc. abcd.0 de. a. c. b. bc. a 0. abcd. c. abc. ab. abc. a. c. bc. bc. a. bcd. c. e. cd. e. a. abc 0. bcd. a 0. b. d. b.0 abcd. abc 0. abcd. a. ab. cd. a. a. d. b 0. d 0. d. e 0. a. a. d. ab Means with the same superscript in the same row are not significantly different (P>0.0) Remark. Lam Nok o'.''n 0o'.''E Ban Datoa o'.''n 0o'.''E Ban Talosamilae o'.''n 0o'.''E Ban Budi o'.''n 0o'.''E Yamu river moouth o'.''n 0o'.'' Daloa School o'.''n 0o0'.''E Ban Tunlong Luloa o'.''n 0o'.''E Parae o'.''n 0o'.'' Freezer company o'.''n 0o'.''E 0 Pattani fishing port o'.00''n 0o'.''E Ban Lam Nok o'.''n 0o'.''E

Page of 0 0 0 0 0 0 Table Percentage of sperm movement in different sodium chloride solution (NaCl) Sample Sodium chloride solution (%) 0 0. 0. 0. 0 0 0 Mean 0 a. b. b 0. c. b SD 0.0.0..0 Means with the same superscript in the same row are not significantly different (P>0.0)