19 Indian J. Fish., 62(2): 19-28, 2015 Captive breeding and developmental biology of Sahyadria denisonii (Day 1865) (Cyprinidae), an endangered ish of the Western Ghats, India T. V. ANNA MERCY, S. SAJAN AND V. MALIKA Kerala University of Fisheries and Ocean Studies (KUFOS), Panangad, Kochi - 682 506, Kerala, India e-mail: annamercy2012@gmail.com ABSTRACT Over the past few decades wild population of Sahyadria denisonii (Day 1865), an endemic ornamental barb of the Western Ghats of India has been overexploited for aquarium trade and is presently listed under endangered (EN) category in the IUCN Red List. The present study communicates the irst ever success of captive breeding and early developmental studies of S. denisonii. Life history phases of S. denisonii were classiied into embryonic, larval, juvenile, subadult and adult stages. Spawning season was from November to March in wild and fecundity varied depending on the size and age of breeding pairs. Eggs were obtained through induced breeding using ovaprim hormone at 0.4 ml per kg body weight. Fertilised eggs were adhesive, demersal and attached to any substratum having a diameter of 1184-1312 µm. Hatching took place 36 h after fertilisation at a water temperature of 27.5±0.5ºC. At hatching, mean larval length was 3.5±0.2 mm with high amount of yolk and the yolk sac remained up to 3-4 days. Organogenesis of larvae was completed 15-20 days after hatching. In this paper full developmental sequence from egg to adult stages of S. denisonii in controlled condition is described. Keywords: Captive breeding, Embryonic development, Miss Kerala, Sahyadria denisonii, Western Ghats Introduction The Western Ghats of India is one of the biodiversity hotspots of the world (Myers, 2000) and its range of hills running along India’s west coast (08º 19’08’’ - 21º 16’24’’N to 72º 56’24’’- 78º 19’40’’E) is one of the richest regions in terms of its biological diversity. The Western Ghats extends 1490 km from north to south with a minimum width of 48 km and maximum of 210 km, covering a total area of 136,800 km2 (Molur et al., 2011). The area is drained by 38 east lowing and 27 west lowing major rivers with running water and lacustrine habitats (Abell et al., 2008). Ragahvan and Dahanukar (2013), listed 320 species of freshwater ishes belonging to 11 orders, 35 families and 112 genera including certain secondary freshwater species, which can also live in brackishwater and marine habitats. The Kerala State on the south - western corner of the Indian peninsula is crisscrossed by 44 rivers arising from the Western Ghats (41 west lowing and 3 east lowing), having an immensely rich ichthyofauna of well over 300 species, of which about 50% have ornamental and recreational value (Mercy, 2009). With the high demand and pricing of many ornamental ish, they are being harvested at greater volumes and sold at higher rates, threatening the viability and sustainability of the resources (Chao, 2001; Vagelli and Erdmann, 2002; Cato and Brown, 2003; Lunn and Moreau, 2004). In India, most wild caught aquarium ish are from the Eastern Himalayas and Western Ghats which are ecological hotspots, known for their amazing freshwater biodiversity and endemism (Allen et al., 2010; Molur et al., 2011). Raghavan et al. (2013) reported that around two dozen ishes are regularly being exported from Western Ghats of Kerala, while Liya and Ramachandran (2013) listed Tetraodon travancoricus, Dario dario, Sahyadria denisonii, Botia striata and Carinotetraodon imitator as the major species of ornamental ishes traded from India for the years 2005-2010. Of India’s total live ornamental ish exports to the tune of US $ 1.54 million during 2007-08, S. denisonii accounted for almost 60-65%. Raghavan et al. (2013) recorded that 310,791 numbers of S. denisonii were exported from India during 2005-2012 and the main markets were Singapore (48.63%), Hong Kong (30.52%) and Malaysia (18.4%), with negligible quantities being exported to Germany, United Kingdom and Japan. Among the native ornamental ishes of Western Ghats region, no species has received global fame and hobbyist attention as that of the Redline torpedo ish, S. denisonii. The distribution of S. denisonii is restricted to the southern regions (Kerala and south Karnataka) of the Western Ghats hotspot of India and their population appears to be extremely fragmented in fourteen rivers namely Chalakudy, Periyar, Achencovil, Pampa, Valapattanam, Chaliyar, Kallar, Chandragiri, Bharathapuzha, Bhavani, Karingode, Kuppam, Anjarkandipusha, Kuttiyadi 20 T. V. Anna Mercy et al. (Biju et al., 2000; Shaji and Easa, 2001; Shaji et al., 2000; Raghavan et al., 2010; Mercy et al., 2010 b; 2013 b). Raghavan et al. (2010) and Silas et al. (2011) observed that over-exploitation of freshwater ishes from the wild for aquarium trade is the main reason for severe depletion of their population. The existing export business of freshwater ornamental ishes from India is not sustainable since the present trade is completely dependent on wild collection (Dahanukar et al., 2004; Raghavan et al., 2013). Over the last few decades, wild population of S. denisonii has declined due to various reasons (Dahanukar et al., 2004; Mercy et al., 2010a; Raghavan et al., 2013) and IUCN’s Freshwater Biodiversity Assessment of the Western Ghats has categorised this species as Endangered (Ali et al., 2010). In a bid to ensure long term availability of S. denisonii, the government of Kerala is currently in the process of inalising various measures aimed at checking the uncontrolled, unorganised, unscientiic nature of wild collection and export of this species (Raghavan et al., 2010). Captive breeding is a conservation strategy that is widely used for the recovery and reintroduction of endangered species (Kelley et al., 2006). Efforts were taken to develop captive breeding technology for indigenous ish species from Western Ghats of India (Ogale, 2002; Padmakumar et al., 2004; Swain et al., 2008; Mercy, 2009). It is envisaged that if S. denisonii can be produced in captivity at a commercial level which will deinitely boost the share of the species in the trade to a large extent and will naturally lead to the conservation of its germplasm. In this context, captive breeding technology for S. denisonii was successfully developed for the irst time and the early embryonic as well as larval development stages were documented. Materials and methods Development of broodstock under simulated natural condition A water lowing habitat was constructed near the Iruvanchipuzha, tributary of River Chaliyar, in such a way that water from the same river is pumped to overhead storage tank (10x10x2 m). Water was allowed to low continuously through the artiicial habitat (10x3x2 m) by gravitational force. All the materials used in the artiicial habitat, such as sand, bottom sediments, rocks and plants were brought from the same river from where the ishes were collected. A diagrammatic representation of the habitat is given in Fig. 1. Fifty juveniles of S. denisonii in the size range 6-8 cm were collected from the Iruvanjipuzha Tributary of River Chaliyar in Calicut District of southern Kerala during September 2008. The collected ishes were transported in oxygen illed polythene bags and stocked in running water habitat. They were fed with protein rich artiicial food, bloodworms and earthworms ad libitum. Captive breeding Twenty ive males and 15 females of S. denisonii collected from the wild were segregated and reared in glass tanks (3x2x2 ft.). As there is no distinct sexual dimorphism in this species; the only dimorphic character observed was that female has a slightly wider body than the male when sexually mature. In a fully ripe female (size 10 cm and above), eggs are seen extruded under slight pressure and male ish (size 9 cm and above) is identiied by the low of milt on applying gentle pressure on the abdominal region towards the genital openings (Mercy et al., 2013b). Maturity of the ripe eggs was determined by observing the position of migratory nucleus of the eggs under a microscope (Rottmann et al., 1991). Induced breeding experiments were conducted during December 2009 to March 2010, coinciding with the natural spawning of this species in nature. From the life history parameters, it was evident that the ish bred during November-April months (Mercy et al., 2010a, 2013a, Solomon et al., 2011). Since the broodishes are very sensitive and handling stress can lead to their mortality, the ish were anaesthetised before handling using clove oil (30 mg l-1) (Sajan et al., 2012). The anaesthetised male (10.4±1.6 cm, 15.5±2.7 g) and female (12.1±1.5 cm, 21.5±3.4 g) broodishes were injected intramuscularly between the anterior region of dorsal in and lateral line using a 1ml syringe with single dose of ovaprim (SGnRH+ Domperidone- Syndel laboratory, Canada) at a rate of 0.4 ml kg-1 body weight by late evening (around 18.00 hrs). Breeding set comprising male and female in 2:1 ratio were injected and were kept together in glass tanks (150 l) provided with continuous aeration. It was observed that induced broodishes did not release eggs or milt by their own. After 10-12 h of latency period, ish were inspected for readiness for ovulation. Both males and females were stripped, and the eggs were dry fertilised using milt collected (Harvey and Carolsfeld, 1993). Fertilised eggs were transferred to a glass tank (10 l) and aerated gently. Ten trials were made successfully during the season. Water quality parameters were monitored at weekly intervals (Table 1). Embryonic and larval developments were documented and photographed with binocular stereo microscope (LABOMED) with digital camera (Canon Power Shot-A570). The hatchery produced juveniles (F1 generation) were reared in the hatchery at Thiruvambady and hundred numbers of juveniles were transported live to the hatchery at the College of Fisheries, Ernakulam during June 2010. They were acclimatised and reared in outdoor rectangular cement tanks (10 t capacity) and were fed with the same food as before. Water quality parameters were maintained similar to the wild (Table 1). During January 2012, three 21 Kerala River & Lakes N Arabian Sea Inlet Outlet Fig. 1. Schematic representation of the modiication of habitat for captive breeding of Sahyadria denisonii Table 1. Water quality parameters of the wild habitat and broodstock rearing tank Parameters Range Water temperature (oC) 26 - 28 pH 7.0 -7.5 Dissolved oxygen (mg l-1) 5.0- 6.8 Total alkalinity (mg l-1) 20 - 25 Hardness (mg l-1) 20 - 25 Ammonia (mg l-1) <0.01 Nitrate (mg l-1) <0.01 pairs of S. denisonii of F1 generation were successfully bred following the same protocol as above. Instead of clove oil, MS-222 (150 mg l -1 ) was used as anaesthetic agent (Mercy et al., 2013a) Results and discussion The results of this study gave a better understanding about the breeding protocol of S. denisonii. In hatchery, broodstock management is one of the major aspects for successful induced breeding of any ish species. Proper care of broodstock is very important for assuring the production of eggs, fry and ingerlings (Siddik et al., 2013). In the present study, S. denisonii was successfully bred under captivity by artiicial fertilisation, using ovaprim as inducing agent. Ovaprim was used as an inducing agent for Labeo dussumieri (Kurup, 1994); Channa striatus (Haniffa et al., 2000); Systomus 22 Captive breeding and developmental biology of Sahyadria denisonii sarana (Chakraborty et al., 2003); Mystus montanus (Arockiaraj et al., 2003), Schizothorax richardsonii (Agarwal et al., 2007); Garra surendranathanii (Thamby, 2009) and Dwakinsia ilamentosus (Mercy, 2009). Ovaprim has been accepted as the most superior inducing agent for carps with high spawning success, percentage of fertilisation and hatching rate (Nandeesha et al., 1990). Fertilised eggs (Fig. 2.2) of S. denisonii were heavily yolked, transparent, spherical and yellow in colour. The eggs were sticky and seen stuck to the glass surface similar to Puntius species. Egg incubation and hatching is better performed in glass tanks under slight continuous aeration. According to Balon (1975), S. denisonii is a lithophil (morphotype), open substratum spawner and falls under ethological class of non-guarders as per the eco-morphological classiication. According to Mercy et al. (2013b) absolute fecundity of S. denisonii ranged from 293 to 967ova per female, while Solomon et al. (2011) recorded it as 376 to 1098. The embryonic period started just after fertilisation and ended when the embryo acquired all the organ systems as in other ishes (Table 2; Fig. 2). In the present breeding trial, latency period was observed as 12.78±0.83 h, fertilisation rate as 86.11±5.23% and hatching rate as 85.89±2.98%. The higher rates of artiicial fertilisation during the present investigation may be related to the dry method of fertilisation where the viability of the spermatozoa remains high. However, the percentage of fertilisation is also related to the maturity and weight of ish (Haque and Ahmed, 1991; Agarwal et al., 2007). In the present study, breeding was performed at an ambient temperature of 26.0-28.0°C. This range of temperature is suitable for breeding of most indigenous small ishes (Islam and Chowdhury, 1976; Akhteruzzaman et al., 1992; Siddik et al., 2013). Embryonic phases Formation of embryo : The fertilised eggs (Fig. 2.2) were spherical, orange brown in colour and adhesive with a size of 1184-1312 µm dia. Fertilised eggs were sticky due to the sticky secondary membrane from the follicular envelope. Egg development started immediately after fertilisation and it activated the cytoplasmic movements. Yolk free cytoplasm begins to stream towards the animal pole gradually segregating the blastodisc from the vegetal cytoplasm. Within 20 min, the streaming movement of the protoplasm towards the lower pole is completed and a blastodisc is formed (Fig. 2.4). The irst cleavage commenced 20 min after fertilisation when the blastodisc was divided into two blastomeres (Fig. 2.5) followed by second cleavage at the 40th min after fertilisation (Fig. 2.6). The sixteen cell stage (Fig. 2.8) was noticed within 80 min and 32 cell stage at 100-110 min post-fertilisation. Cell division continued rapidly after the eight blastomere stage (Fig. 2.7). Morula (Fig. 2.9) and blastula (Fig. 2.10) stage occurred at 180th and 210th min respectively. Cell division took place more or less synchronously during the early stages of the blastula period. Blastoderm Table 2. Embryonic developmental stages of Sahyadria denisonii Developmental stages Duration Figure Features Eggs freely ooze out Fertilized eggs 00:00 h. 00:00 h Sticky and orange in colour Spherical, orange coloured, demersal and adhesive Blastodisc stage Two cell stage Four cell stage Eight cell stage Sixteen cell stage Morula stage Blastula stage Dome stage Late blastula stage Blastopore stage Yolk plug stage 00:10 h 00:20 h 00:40 h 00:60 h 01:20 h 03:00 h 03:30 h 06:00 h 08:00 h 08:30 h 09:00 h 2.01 2.02 2.03 2.04 2.05 2.06 2.07 2.08 2.09 2.10 2.11 2.12 2.13 2.14 Head - tail bud stage Tail free stage Eye vesicle stage Twitching stage Just before hatching Hatching 17:00 h 18:00 h 19:00 h 30:00 h 34:00 h 36:00 h Just hatched larvae Hatchling 3.54 ± 0.21 mm TL 36:00 h 36:00 h. 2.16 2.17 2.18 2.19 2.20 5.21 2.22 2.23 2.24 2.25 Cytoplasm streams toward animal pole First cleavage Second cleavage Third cleavage Fourth cleavage Blastodermal cells formed after cleavage, arranged in group in the animal pole Crown like blastodern spread over yolk at animal pole Bblastoderm expanded to the equator of the yolk sphere; pre-gastrula stage Blastoderm extended towards the vegetal pole Large yolk plug protrudes from the blastopore. Yolk invasion completed by gradual spreading over the germ layer. Rudimentary head and tail appeared and became differentiated. Head and tail rudiment visible Yolk at the tail end elevated to inside to free tail from yolk mass. Eye vesicle observed Vigorous movements before hatching. Continuous beating of the caudal region especially around middle region of the body The egg shell breaks up and the tail emerges out irst, followed by the head; 3.54 ± 0.21 mm in total length Larvae hatch out from egg capsule Newly hatched larvae non-pigmented and had an average total length of 3.54 ± 0.21 mm and yolk sac of 1 mm dia. 23 T. V. Anna Mercy et al. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 Fig. 2. Embryonic and larval development stages of Sahyadria denisonii 38 39 Captive breeding and developmental biology of Sahyadria denisonii was observed at 5 h post-fertilisation, (hpf) followed by dome stage (Fig. 2.11) at 1 h. At dome stage, the blastoderm formed a dome-like shape due to bulging of the yolk towards the animal pole and the epiboly continued during the gastrulation period. In the gastrula period, extensive cell movements were observed, including involution, convergence and extension, producing the three primary germ layers and the embryonic axis. Gastrulation began with cell involution at around 50% epiboly. At 75% epiboly stage (Fig. 2.12), the embryonic shield became less distinctive, as compared to shield stage and its cells got repacked to elongate the shield along the animal pole axis. At 90% epiboly stage, the yolk plug was clearly seen in the vegetal pole (Fig. 2.13 and 2.14) and was noticed at 9 hpf. The segmentation period was observed at 15 h which was characterised by the sequential formation of the somites, and this period lasted up to hatching. During this period, the embryo elongated along the animal pole axis, the tail bud became longer and rudiments of the primary organs became visible (Fig. 2.15). Differentiation of embryo: The head and tail bud of the embryo (Fig. 2.16) became clearly distinguishable after 17 h. In this stage, the optic primordium had a prominent horizontal crease and tail bud became more prominent. At 19 hpf, the eye vesicle was observed, the embryo increased in size and eighteen myotomes were clearly visible (Fig. 2.17). Tail bud started to separate from the yolk sac at 22 hpf (Fig. 2.18). The yolk extension got clearly delimited from the yolk ball as the tail straightened out. Twitching movements of the embryo started by 23 hpf. At 26 h, tail end was free from the rest of the yolk sac (Fig. 2.19) and at 30 h, embryo started to twist or rotate completely inside the egg membrane. After 34 h, movements of the embryo within the egg membrane were very frequent. Close to hatching, the twitching and lashing of the embryo inside the egg capsule became rapid (Fig. 2.20). The egg shell broke at 36 h due to rapid shaking movements of the body and the head emerged out irst, followed by the tail (Fig. 2.21 to 2.24). Hatching of eggs continued up to 40 h, probably the temperature and oxygenation of the water played an important role in incubation as reported by Laurila et al. (1987) and Dwivedi and Zaidi (1983). Larval phase Larval phase can be further subdivided into yolk sac stage, pre-lexion stage, lexion stage and post-lexion stage (Fig. 2.25 to 2.39) according to the classiication by Pena and Dumas (2009). Yolk sac larva/eleuthero embryo: The newly hatched larvae (Fig. 2.25) were non-pigmented with an average total length of 3.5±0.2 mm and yolk sac of 1mm dia. 24 Hatchlings had non- pigmented eyes and were devoid of distinct mouth and ins. Head seemed to be in contact with yolk sac and optic vesicles. The yolk sac is large and was found attached to the body along its ventral side. A narrow in fold was found to be differentiated as a thin membrane, surrounding the caudal region and extended up to the yolk sac. The crawling movements of the hatchlings were detected immediately after hatching. Two day old larva: On the second day, the larvae appeared to be more active; and inhabited the bottom of the tank as a group with average total length of about 4.1±0.2 mm (Fig. 2.28). The head portion seemed to be further developed. On closer observation, the heart was seen pulsating rhythmically. Pigmentation started in the eyes (Fig. 2.26). Normally in ish, the shape of the yolk sac undergoes signiicant changes during organogenesis (Winnicki et al., 2001). In case of S. denisonii during embryogenesis, the yolk sac got divided into two parts, proximal portion with spherical shape and caudal part with cylindrical shape. Yolk sac became slightly reduced in size, with 3 mm in length towards the ventral side of the body (Fig. 2.27). Ventral and caudal in folds were separated by the formation of anus. Formation of pectoral in folds was also seen. Three day old larva: Larvae on the third day, reached an average length of 4.5±0.2 mm. Yolk sac was seen further reduced. Mouth and anus became distinguishable. Pectoral ins with in rays started to move vigorously (Fig. 2.29). Head became more prominent and free movements of the eye balls were also noticed Melanophores appeared around the head and snout (Fig. 2.30). Larvae exhibited vigorous movements upwards to the column water and inally sank to the bottom. All the active larvae got accumulated together into a lump-like ball showing strong thigmotactic behaviour. Pre-lexion Stage : The pre-lexion stage began on day 4 when the larva started its irst external feeding. Average length (TL) at this stage was 5.1±0.2 mm. Pre-lexion stage extended from a period of about 4 to 8 days; till it reached an average TL of 6.8±0.4 mm. Yolk sac got reduced and became tube like with 2.2 mm length and 0.5 mm width. At the onset of the pre-lexion stage the eye became fully pigmented and the mouth and anus opened. Alimentary canal was found to contain a yellowish luid. Air bladder was noticed and occurrence of pigmentation on the air bladder started (Fig. 2.31 to 2.32). Opercular movements was also noticed. Caudal in became modiied with rudimentary in rays. Pigments were found more concentrated on the anterior region of the body and also in caudal in fold. Vigorous movements of the mouth was noticed as if the larva was in search of food. Eye balls exhibited rotating movements and larvae showed grouping behaviour when 25 T. V. Anna Mercy et al. all of them accumulated into a bundle towards the latter stage. Melanophores also appeared in the middle and in the notochord region. One week old larva was very active and started free swimming. Egg yolk was completely absorbed at this time. Average body length of the larvae increased to 7 mm and at this stage larvae fed entirely on exogenous food. Flexion stage: The lexion stage started from day 08 (average TL 07.4±0.3 mm) and extended to day 17 (average TL 10.4±1.5 mm). The complement of the notochord lexion was characterised by the anterioposterior orientation of the caudal rays. A key event was the development of lexion in the ventral side of the spinal cord in the notochord associated with the tail in. On day 9 or 10, the single chambered air bladder (Fig. 2.32) was transformed into double chambered (Fig. 2.33). Furthermore, together with the disappearance of the larval in fold and development of the ins a change in swimming style was also observed (Fig. 2.34). Similar observations were also reported in other teleost larvae (Van Snik et al., 1997; Gisbert, 1999; Gisbert et al., 2002). Post-lexion stage: The post-lexion stage was observed from day 17 (average TL 10.4±1.5 mm) and extended to day 26 (average TL 16.7±2.1 mm). After two weeks, the larvae completely metamorphosed; and the average body length increased to 11.2±1.5 mm. Changes during this stage included the development of dorsal in having 6 - 7 in rays. A vertical black band was formed over the body and an inferior mouth was clearly observed. Fin elements became evident in the dorsal, caudal, anal and pelvic ins during the post-lexion stage. Body pigmentation increased in the middle part of the body. Larval feeding and weaning : The newly hatched larvae (3.5±0.2 mm) fed on its yolk content up to 3 to 4 days after hatching. Infusoria (pure culture of Paramecium) was given as irst exogenous feed. The larvae were fed with freshly harvested paramecium at every 3 h interval. About 40-50% water exchange (30 l glass tanks) was done every day after exogenous feeding. After 10 days, the larvae were stocked in glass tanks (50 l) with an approximate stocking density of 10 larvae l-1 and were fed with live micro-worms (Panagrellus spp.) cultured using bread or oats medium. No substratum was provided during irst two weeks of larval rearing and thereafter larval tanks were provided with 2 or 3 small pieces of clean rock as substratum for the larvae. Hatchlings were gradually weaned to artiicial feeds (Artemia lakes), 12-14 days after hatching. Uneaten food was siphoned out from the bottom of each tank every morning before irst feeding and dead ish were counted and preserved to analyse percentage of mortality. Dissolved ammonia plays a vital role during larval rearing of S. denisonii and daily water quality assessment is necessary. In nature, post-larval stages of S. denisonii exhibit a bottom dwelling habit and they rely on bottom deposited diets such as detritus, algae and diatom that grows on pebbles and stones. Similar indings were also noticed by Costa and Fernando (1967). Feeding experiments suggested that artiicial feed can be used for rearing post-larvae as well as juvenile stages of S. denisonii in aquaria (Mercy and Sajan, 2014) and it will be beneicial for the development of better larval rearing techniques for S. denisonii under hatchery condition. Juvenile phase One month old juveniles (average TL 29.8±2.3 mm) of S. denisonii showed vertical body banding pattern. Juveniles of S. denisonii possessed four vertical black cross band at nuchal, sub dorsal, supra anal and caudal positions (Fig. 2.35 to 2.36). Later, after disappearance of vertical cross bands, a lateral black bar started to appear from posterior margin of the eye region and extended towards the caudal peduncle. A transverse black and yellow band was observed in the tip of each lobe of the caudal in. The anterior base of dorsal in exhibited a black and red blotch (Fig. 2.36). Sub-adult and adult phase The juveniles (Fig. 2.37) reached an average TL of 35.8±5.3 mm size after rearing for three to four months in iber tanks (200x100x50 cm). A red streak started to appear from the tip of the snout, parallel to the black stripe already formed posteriorly. It extended up to the middle of body below the dorsal in. This sub-adult stage (Fig. 2.38) externally resembled adults in presence of these stripes. The four vertical cross bands disappeared and their body became greenish brown on the dorsal side and silvery in the ventral side (Fig. 2.38). Juveniles and sub-adults kept under ibre tank condition and in artiicial habitat exhibited size variation during growth. This may be due to the availability of natural food and environmental conditions as has been reported by Hecht and Pienaar (1993) and Wankowski and Thorpe (1979) in Salmo salar. The ish became adult (Fig. 2.39) on reaching a size of TL 7.8±2.2 cm. The results clearly demonstrated the possibility of using both clove oil and MS-222 as anaesthetics and synthetic ish breeding hormone ovaprim for effective induced breeding and seed production of S. denisonii under captive condition. This captive breeding technology has also been transferred to the ornamental ish breeders from different states by the Marine Product Export Development Authority, Kochi (Anon., 2014). The present work generated information on the early life history and developmental stages and also on commencement of irst feeding time for larval rearing. The indings of the present Captive breeding and developmental biology of Sahyadria denisonii 26 study can be used in induced breeding of S. denisonii in hatcheries and in conservation of this critically endangered valuable species. Cato, J. C. and Brown, C. L. 2003. Marine ornamental species: Collection, culture, and conservation. Iowa State University Press, Ames, Iowa, 310 pp. Acknowledgements Chakraborty, B. 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