| Abstract |
Members of the genus Seriola have been notable species for aquaculture production worldwide, with the greater amberjack being one of the most prominent species, due to its cosmopolitan distribution and market acceptability, high growth rates and large size. However, the prerequisite to sustainable aquaculture production -i.e. controlling reproduction in captivity- has been lacking for greater amberjack. The objective of the present doctoral thesis was to describe the reproductive function of wild and captive reared fish, examine the reproductive maturation potential of fish reared in land-based tanks and sea cages and develop broodstock management methods and spawning induction protocols using gonadotropin releasing hormone agonists (GnRHa). The effectiveness of GnRHa injections versus GnRHa implants of sustained release was examined, and the best method was then optimized by examining (a) optimal dose and (b) time of application. The extent of spermiation period and sperm quality parameters of breeders maintained in different locations and facilities was examined using computer assisted sperm analysis (CASA) to examine any variations over time and after spermiation enhancement trials. Finally, the spawning induction method was also evaluated for the first time on first generation (F1) breeders in Spain to ensure uninterrupted production of fingerlings for genetic breeding programs. A comparative study of wild and captive greater amberjacks was carried out to compare the endocrinological status of the pituitary-gonad axis related with reproductive developmental stage. A total of 33 (14 males and 19 females) wild and 24 (12 males and 12 females) captive-reared greater amberjack breeders were sampled at three different phases of the reproductive cycle. The sampled wild females were separated in two categories according to the histological analysis of their ovaries, in vitellogenic in early May and spawning capable in late May-late June. Between these two categories a 3 to 4-fold increase of the gonadosomatic index (GSI) was recorded. At the same time a corresponding increase was recorded for pituitary follicle stimulating hormone (FSH) content and plasma testosterone (T), while almost a 10-fold increase was observed in pituitary luteinizing hormone (LH) content. An increasing trend of plasma 17β-estradiol (E2) and 17,20β-dihydroxy-4- pregnen-3-one (17,20β-P) was also recorded between the two reproductive stages. On the other hand, sampled captive females were separated into four categories of reproductive development, since extended follicular atresia (AT) was observed from late May, with the majority of the fish that had already ceased their reproductive development showing signs of regressed or regenerating ovaries, possibly due to repeated handling. Pituitary content of FSH and LH, fshβ and lhβ expression and plasma levels of FSH and LH remained unchanged, in contrast with the plasma E2 and T that were reduced towards the regenerating phase, and 17,20βP which showed the opposite pattern. No significant hormonal differences, except in 17,20β-P, were recorded between captive and wild vitellogenic females, in contrast to spawning capable females where pituitary LH content, plasma FSH and T were found to be different. Overall, the captive females lagged behind compared to the wilds on reproductive development and this was probably related with the repetitive handling in sea cages. Wild male fish were found to be in corresponding reproductive stages with females, as expected. Pituitary LH content, plasma T, 17,20β-P and GSI of wild males exhibited a 3 to 4-fold increase, while an increasing trend of pituitary FSH content, lhβ expression levels and plasma 11-ketotestosterone (11-KT) was also observed between the developing and spawning capable reproductive stages, while an opposite trend was observed in plasma LH. On the other hand, sampled captive males fell into three categories of reproductive development, showing significant differences from their wild counterparts even from the developing phase in four out of the ten measured parameters. During spawning capable phase, the situation was even worst, since in almost all the measured parameters reduced levels between wild and captive males were recorded, being statistically significant or showing a strong downward trend. Therefore, the captive males apart from the handling stress that could be related with the worst reproductive development compared to the wilds after the 2nd sampling onwards, showed differences from the first sampling, a more pronounced effect of the captivity compared to the females. To develop broodstock management and spawning induction methods for greater amberjack, it was examined the possibility of rearing fish either in tanks or sea cages, and evaluated their spawning performance. The advantages of sea cage rearing and spawning include optimal environmental conditions and welfare, and low cost of fish maintenance. The advantages of tank rearing and spawning include biosecurity, ease of management and handling operations, and control of egg collection. Females reared in tanks supplied with well seawater throughout the year exhibited a significant reproductive dysfunction, such as limited gametogenesis, lower oocyte diameter at the peak of the reproductive season, low female eligibility for spawning induction and almost 0% fertilization success. On the contrary, females in three different broodstocks reared in sea cages for three consecutive years underwent vitellogenesis completely, and almost all females were eligible for spawning induction. Males reared under both conditions exhibited reduced sperm production compared to the wild, but unlike the situation in the females, tank rearing did not seem to have any negative effects on sperm quality parameters compared to rearing in sea cages. Spawning performance (eggs collected and fertilization) was best when fish were reared in sea cages during the year and then moved to land-based tanks for spawning after GnRHa treatment, resulting in the production of large numbers of eggs of adequate quality for commercial larval rearing of the species. The present study examined also two methods for the induction of oocyte maturation/ovulation and spawning, which are based on the induction of endogenous LH release from the pituitary, through the use of GnRHa either in the form of sustained-release delivery systems (implants) or injections (acute release). The stock (n = 28) consisted of wild fish captured in 2011 and each GnRHa administration method was conducted in two replicates with 6 and 8 fish per tank, at a 1:1 sex ratio. Fish were given a GnRHa injection once a week (three administrations), or a GnRHa implant every 2 weeks (two administrations). Mean daily relative fecundity was significantly higher in the implanted fish compared to the injected fish. Total relative fecundity was also significantly higher in the implanted fish compared to the injected ones, but there were no differences in the quality of eggs in terms of fertilization, 24-h embryo survival, hatching and 5-d larval survival. The number of females with fully vitellogenic oocytes eligible for induction of spawning decreased from the initial to the final sampling, from 7 to 6 females for the GnRHa implant treatment and from 7 to 3 females for the GnRHa injection treatment. The use of GnRHa implants apparently promoted the proper endocrine changes leading to multiple cycles of oocyte maturation, ovulation and spawning, and thus producing larger numbers of eggs. In addition, the use of GnRHa implants may be more appropriate in greater amberjack than multiple injections because (a) it also stimulates vitellogenesis and (b) involves less handling of the fish. After the successful control of oocyte maturation, ovulation and spawning using hormonal therapies in the form of controlled-release delivery systems of GnRHa, the objective was to optimize the protocol and (a) determine the most efficient GnRHa implant dose and (b) examine the extent of the spawning induction period in greater amberjack reared in sea cages throughout the year, based on the resulting spawning kinetics, egg production and quality. Wild-caught greater amberjack were maintained in two commercial facilities, and were transferred to onshore tanks for spawning after treatment with GnRHa implants (a) of different doses (25 and 75 μg kg-1 body weight), or (b) at different times during the expected spawning season (June-July). Both doses were equally effective, and two consecutive implantations in 2 weeks resulted in mean (±SD) total relative fecundity of 185,221± 91,311 to 199,492 ± 16,973 eggs kg-1 in 11-18 spawns, without any significant differences in egg quality. Another noteworthy observation from the present study was that “spent” females or with smaller remaining vitellogenic (Vg) oocytes after spawning induction in the onshore tanks, recovered to their initial reproductive stage –in terms of maximum Vg oocyte diameter- after a “resting” period of 14-28 days in sea cages, while their repetitive handling at the time of reproductive stage evaluation didn’t caused any negative effects on their reproductive development. Therefore, it was shown that a protocol where breeders are maintained in sea cages, are taken at different times and are induced to spawn with a GnRHa implant of a recommended dose of 50 μg kg-1 , can be used successfully during a period of at least 2 months when seawater temperature is 19-24°C. In many fish species, the F1 fish face reproductive dysfunctions in captive conditions, such as lower reproductive success, production of unfertilized eggs and lack of spawning, compared to the wild-caught breeding animals. To examine this possibility in greater amberjack, hatchery-produced fish were examined during the expected spawning season and were hormonally induced to spawn. The results showed that hatchery-produced greater amberjack undergo normal gametogenesis and can be induced to undergo maturation, ovulation and spawning after multiple administrations of GnRHa implants, over an extended spawning period lasting from May to September in the Canary Islands, Spain. The use of GnRHa-delivery systems resulted in multiple spawns of fertilized and viable eggs. The developed reproduction control method shows great potential to advance the commercial production of greater amberjack, by enabling the use of hatchery-produced broodstocks for further breeding selection. However, more research is needed to provide solutions for selective breeding in greater amberjack, due to impossible use of the in vitro fertilization protocol in this species. Finally, sperm quality was evaluated using CASA before and after GnRHa administration and the extent of the spermiation period was determined. Males were in spermiation throughout the monitoring period from the 30th of May until the 18th of July, at temperatures between 19-24ºC. However, lower sperm motility duration, density and survival under cold storage were observed from early July onwards. Sperm quality did not recover after the end of spawning induction experiment in tanks and the return of the fish for 14-28 days to the sea cage in mid-July, which could be related to the high temperatures of this period. An improvement trend was observed in the quality of the milt collected on day 7 after a single GnRHa administration, but a significant decrease was observed on day 21 in sperm density, survival under cold storage and straight line velocity (VSL). On the contrary, a double GnRHa administration spaced 14 days apart maintained the same sperm quality for a longer period of 29 days. Further experimentation needs to be done in greater amberjack to ensure adequate sperm production for high fertilization success. Considering all the above, the captive greater amberjack is sensitive to handling at the time before the onset of spawning period regarding normal reproductive development. In males, there is a clear effect of confinement on reproductive development. In contrast, when the fish are left unhandled before the spawning period the repetitive handling do not cause any negative effect on their reproductive development. Interestingly, the females recover shortly to their initial reproductive stage after spawning induction after a resting period in sea cages, opposite to the males. A protocol where fish are maintained in sea cages throughout the year, and transferred to onshore tanks after spawning induction with GnRHa implants at a dose of 50 μg GnRHa kg-1 body weight (BW) seems to be best in the Mediterranean. Both females and males, at temperatures ranging from 19-24°C in June-July, can be used to ensure adequate egg production.
|
Search
