Feed Containing Novacq Improves Resilience of Black Tiger Shrimp,Penaeus Monodon,to Gill‐associated Virus‐induced Mortality

The ability of Novacq to improve resilience of black tiger shrimp, Penaeus monodon, to infection and mortality induced by gill‐associated virus (GAV) was investigated. Over a 26‐d period, shrimp were fed pellets with or without 10% Novacq. Following this, four replicate tanks, each containing 10 shrimp that had been fed either diet, were maintained as‐is, injected with saline or injected with GAV inoculum (i.e., 40 shrimp for each of the six groups). For shrimp (n = 20) in two of each group of four tanks, survival was monitored daily over 14 d and a pleopod was sampled from each shrimp on Days 0 and 14. For the other two tanks, a pleopod was sampled from each shrimp on Days 0, 3, 7, 10, and 14 to track changes in GAV loads over time. Survival was significantly higher (P < 0.05) from Day 7 onward among the group fed Novacq. GAV infection loads appeared to vary more between individuals in the Novacq diet cohort, but overall were not reduced significantly at any time points post‐challenge compared to shrimp tested from the Control diet cohort.     

High‐quality spontaneous spawning in greater amberjack (Seriola dumerili,Risso 1810) and its comparison with GnRHa implants or injections

Production of sufficient high‐quality eggs of greater amberjack (Seriola dumerili) still constitutes the main bottleneck for commercial production of this species. The main objective of this study was to compare the quality of spontaneous spawn of greater amberjack with those obtained by either GnRHa injection or GnRHa implant protocols. Captive amberjack broodstock were distributed in three circular tanks of 40 m³. Broodstock from Tank 1 were not hormonally induced and spawned spontaneously, whereas those of Tank 2 were intramuscularly injected with GnRHa (20 µg/kg body weight) and those of Tank 3 were given EV‐500 µg GnRHa implants. The number of eggs per spawn obtained in the broodstock without hormonal treatment was larger than in those obtained with injections or implants. Egg quality was best in broodstock with spontaneous spawn, followed by GnRHa‐injected fish and then GnRHa implants. Besides, size of larvae from control and injected broodstock was similar between them and significantly higher (p < 0.01) than those from GnRHa implant spawn. Overall, this study showed that it is possible to obtain very high‐quality spontaneous spawn in greater amberjack, providing the adequate conditions. Furthermore, GnRHa weekly injections lead to similar egg viability and hatching rates than spontaneous spawn and higher fertilization rates than GnRHa hormonal implants, which is better than in previous studies.     

Growth, Survival and Reproductive Performance of Domesticated Australian Stocks of the Giant Tiger Prawn, Penaeus monodon, Reared in Tanks and Raceways

The growth, survival and reproductive performance of domesticated Australian stocks of the giant tiger shrimp Penaeus monodon were evaluated in trials conducted in 1997 and 2003. The 1997 trials assessed the performance of first generation progeny of wild broodstock from the northeast coast of Australia and fourth generation progeny of pond‐reared broodstock, which also originated from northeast coast wild stocks. In these trials, growth and survival of the shrimp were assessed when reared for 17 mo in tanks. Reproductive performance of the shrimp was assessed at 14.5 mo and 17 mo. The 2003 trials assessed the performance of first generation progeny of wild broodstock from the Gulf of Carpentaria (north coast of Australia). In these trials, growth and survival of shrimp were assessed when reared for 14 mo in tanks and raceways. Reproductive performance of the shrimp was assessed at 11 mo, 12 mo, and 15 mo. Growth and reproductive performance of the stocks varied between trials, families, ages and rearing systems. The most pronounced differences in growth and reproductive performance were between the 1997 and 2003 trials. At 11 mo of age, the average wet weight of the shrimp in the 2003 trials (females 117.1 ± 5.8 g; males 87.9 ± 7.6 g) was 200% greater than the average wet weight of shrimp in the 1997 trials (females 55.2 ± 6.8 g; males 41.2 ± 3.4 g). The reproductive performance of the shrimp was also higher in the 2003 trials in terms of the percentage of spawnings hatching (52.0% in 1997; 77.1% in 2003) and mean hatch rate (21.5% in 1997; 31.6% in 2003). Differences in the growth and reproductive performance of the tank‐reared stocks between years were indicative of significant improvements in the rearing environment, diet and husbandry techniques. Variation in the reproductive performance between families was consistent across rearing environments and at different ages and suggests the potential to improve reproductive performance through genetic selection. Notably, this study identified hatch rates of nauplii from the spawned eggs as a key area for future improvement of domesticated stocks reared in tanks and raceways. Future efforts to improve the growth and reproductive performance of domesticated P. monodon could benefit from integrating incremental improvements to husbandry with genetic selection.     

Productivity benefits of selectively breeding Black Tiger shrimp (Penaeus monodon) in Australia

Advances in the domestication and selective breeding of Australian Black Tiger shrimp, Penaeus monodon, opens the opportunity for world producers to reconsider the benefits of farming this species. Just over a decade ago this species was the world's most farmed shrimp species, however, difficulty in its domestication, in part, led to the widespread establishment of Penaeus vannamei (Pacific White shrimp) as the most farmed shrimp species in the world. This study empirically evaluates the productivity benefits of commercially domesticating P. monodon against production from wild broodstock of the same species. The evaluation compared the relative production from commercial ponds stocked with the progeny of wild P. monodon broodstock and ponds stocked with the progeny of domesticated stocks. The production data were from 164 ponds of domesticated stocks and 30 ponds of wild stocks, collected over 4 years (2009–2013) from two separate farm sites of the same Australian shrimp farming company. The wild stocks were sourced from the east coast of Australia. The results suggested that the productivity of the selectively bred stocks was 39% greater compared with production from wild stocks given equivalent amounts of feed and other inputs. Furthermore, productivity was additionally enhanced depending on the choice of feeds and whether stocking took place in September rather than later in the year (i.e. in early spring rather than late spring/early summer in the Southern hemisphere). This suggests that there is significant potential to further enhance the productivity of P. monodon farms via integrating advances in domestication, feeds and management practices.     

Pigment deficiency correction in shrimp broodstock using Spirulina as a carotenoid source

In this study, an effort was made to cure the pigment deficiency syndrome (PDS) exhibited by pond‐reared Fenneropenaeus indicus (H.Milne Edwards) broodstock, using the blue‐green algae Spirulina as a carotenoid source in diet. The PDS symptoms included bleaching of ovary and reduced early larval performance. Biochemical analysis indicated reduced quantity of total carotenoids in egg and nauplii from the females with PDS. When PDS females were fed 30 g kg^?1Spirulina‐supplemented diet immediately after the appearance of symptoms, the egg and nauplii carotenoid values improved gradually and attained the pre‐PDS values after a minimum of 4 week period. Spirulina supplementation also improved significantly (P < 0.05) the spawn quality (hatch per spawn, nauplii per spawn, nauplii viability) and larval quality indices (P < 0.05). The egg and nauplii protein values exhibited a trend similar to carotenoids, confirming the influence of latter on protein stability previously reported by researchers. The study confirms the bioavailability of carotenoids from Spirulina to shrimp broodstock and recommends its regular inclusion in diet to preclude carotenoid deficiency‐related problems in shrimp hatcheries.     

Comparative costs of postlarval production from wild or domesticated Kuruma shrimp, Penaeus japonicus (Bate), broodstock

The present study compares the relative costs of stocking ponds with postlarvae from wild or domesticated Kuruma shrimp, Penaeus japonicus (Bate), broodstock. Wild broodstock were obtained from a commercial trawler. Domesticated broodstock were reared to harvest size (wet weight = 25 g) on a commercial farm and then transferred to controlled environment tanks where they were grown to a final wet weight of 50 g. The costs of stocking ponds with postlarvae were calculated from the observed reproductive output, the costs of purchasing wild broodstock and the costs of domesticated broodstock production in the controlled environment facility. Domesticated P. japonicus spawned comparable numbers of eggs to wild broodstock of similar size, but hatching success was significantly lower. A total of 12 domesticated P. japonicus broodstock would be needed to produce the postlarvae to stock a 1-ha pond, compared to only six wild broodstock. However, the much higher relative costs of wild broodstock means that the cost of using their postlarvae would be Aus$851 per pond compared to Aus$390 for domesticated broodstock. The present authors conclude that the use of domesticated P. japonicus broodstock could be a cost-effective alternative to wild broodstock in Australia and in other countries where P. japonicus is farmed.     

Long‐lasting Effect Against White Spot Syndrome Virus in Shrimp Broodstock,Litopenaeus vannamei,by LvRab7 Silencing

The white spot syndrome virus (WSSV) remains the most devastating viral pathogen of shrimp culture worldwide. Gene silencing by RNA interference (RNAi) using double stranded RNA (dsRNA) has been considered a powerful tool for conferring protection against WSSV when viral genes are silenced, as documented in several shrimp species. However, this effect is not long lasting. Our results provide the first evidence that long‐term silencing of the LvRab7 endogen produced antiviral effect against WSSV, which endured at least 21 d after dsRNA treatment (dat). Until now, the most efficient way to implement RNAi with dsRNA into the shrimp is by injection. Consequently, its application to broodstock in hatcheries is possible, minimizing the risk of vertical transmission of the virus. We show that the expression of Rab7 in hemocytes is lowest at 2 dat and finally recovers to basal status. In contrast, in gills and pleopods, gene expression silencing continued for at least 21 d. We challenged Litopenaeus vannamei broodstock with WSSV at 7, 14, or 21 dat reaching mortality rates of 0, 40, and 27%, respectively. In conclusion, the LvRab7 gene silencing is progressive and effective against WSSV. However, further studies are necessary to elucidate the functions of Rab7 in shrimp cells before applying this methodology at a commercial level.     

The impact of dietary supplementation with astaxanthin on egg quality and growth of long snout seahorse (Hippocampus guttulatus) juveniles

This study investigated the effect of dietary astaxanthin supplementation on egg quality and juvenile growth of long snout seahorse (Hippocampus guttulatus). Captive breed seahorse broodstock were fed four diets composed of frozen shrimp [Atlantic ditch shrimp, Palaemonetes varians) used as a vector to deliver artificial diets with increasing levels of astaxanthin (0, 75, 100 and 125 mg kg^?1 dry weight)]. The results indicated that the astaxanthin uptake into eggs from the enriched shrimp diets was highly efficient. Females fed unsupplemented astaxanthin diet produced similar‐sized eggs with lower concentration of astaxanthin than females fed diets with astaxanthin. The lower concentration of astaxanthin in the eggs was correlated with the production of smaller juveniles in comparison with the juveniles hatched from parents fed supplemented astaxanthin diets. Juvenile growth and survival was limited by their size on release from the male's pouch as at the end of 28‐day postparturition juveniles produced with the diet with no astaxanthin were still significantly smaller (P < 0.05) than those produced from parents fed astaxanthin‐supplemented diets. These results demonstrate a significant benefit of dietary astaxanthin supplementation in long snout seahorse diets in terms of improved egg quality and juvenile growth and survival.     

Cloning,characterization, expression analysis and RNAi of Retinoblastoma‐like gene from black tiger shrimp (Penaeus monodon)

Retinoblastoma (Rb) is a multifunctional regulator involved in several key cellular processes, such as cell cycle control, cell differentiation, tumorigenesis and senescence. In this study, an Rb‐like gene, PmRBL, was cloned from black tiger shrimp (Penaeus monodon). The full‐length cDNA sequence of PmRBL is 4,069 bp with an open reading frame of 3,243 bp, which encodes 1,080 amino acids. Quantitative real‐time PCR (qRT‐PCR) analysis indicated that PmRBL was highly expressed in the gills, hepatopancreas and ovaries of P. monodon. The highest PmRBL expression levels were observed in stage III of the ovarian development in P. monodon. RNA interference experiments were conducted to examine the expression profiles of PmRBL, PmCDK2 and PmCyclin E. The knockdown of PmRBL in the ovary and hepatopancreas by dsRNA‐RBL was successful. After dsRNA‐RBL was injected into the shrimp, the relative expression levels of PmCDK2 and PmCyclin E were upregulated at 12–72 hr in the ovaries and hepatopancreas. The localization and level of PmRBL expression in the ovary and hepatopancreas were investigated through in situ hybridization, which revealed consistent results with those of qRT‐PCR. Therefore, PmRBL, PmCDK2 and PmCyclin E may be involved in vitellogenin synthesis and ovarian maturation in P. monodon.     

Effects of different diets and seawater systems on egg production and quality of the broodstock Babylonia areolata L. under hatchery conditions

The broodstock of spotted Babylon snail, Babylonia areolata, were conditioned in indoor tanks under two seawater systems (static system and flow‐through system) and five types of foods (fish, shrimp, squid, green mussel and formulated diet). High survival and good feeding was found from broodstock conditioned in all treatments of seawater systems and diets. All broodstock showed active behaviour with no signs of stress as exhibited by active movement and feeding, protrusion of siphonal tube and constant egg laying throughout the experiment. Two‐way analysis of variance showed that there were no significant differences (P>0.05) in the total yield of egg capsules, size of egg capsules, size of newly hatched larvae and growth of larvae among broodstock B. areolata conditioned in the seawater system and diet treatments, but there were significant differences (P>0.05) in the frequency of egg capsule laying, number of fertilized eggs/embryos per capsule, total yield of fertilized eggs/embryos and final survival of larvae among broodstock B. areolata conditioned in the diet treatments. We present initial evidence that the seawater system had a stronger influence on the egg production and quality of broodstock B. areolata than conditioning diets.     

White spot syndrome virus (WSSV) prevalence associated with disease resistance among wild populations of black tiger shrimp,Penaeus monodon (Fabricius)

White spot disease caused by white spot syndrome virus (WSSV) is the major issue of huge economic destruction globally in the shrimp aquaculture industry. In the present investigation, WSSV prevalence associated with disease resistance was studied among wild black tiger shrimp, Penaeus monodon (Fabricius) from four distant geographic locations along the East coast of India during 2009–2010. Results suggested that the WSSV prevalence in wild P. monodon was the highest (56.2%) in Chennai, Tamil Nadu followed by Digha, West Bengal (10.9%), Visakhapatnam, Andhra Pradesh (0.6%) and Chilika, Orissa (0%). Quantitative data suggested that the mean copy number of WSSV among these four places was 1.4 × 10⁶, 4.6 × 10⁴, 1.6 × 10² and 2.3 × 10² copies μg^?1 shrimp genomic DNA respectively. The disease resistant prevalence using the 71 bp microsatellite DNA marker was the highest among Chilika, Orissa (63.6%) and Visakhapatnam, Andhra Pradesh (63.5%). Higher WSSV prevalence in Chennai, Tamil Nadu and Digha, West Bengal corresponded to lower disease resistant prevalence (24% and 40.2%). Conclusively, probably collection of broodstock of P. monodon from places like Chilika and Visakhapatnam would be a much safer approach for the development of specific pathogen‐resistant shrimp aquaculture.     

Breeding and larval rearing of the rabbitfish,Siganus guttatus (Bloch)

Females of Siganus guttatus reared to sexual maturity in canvas tanks were induced to spawn by using human chorionic gonadotropin (HCG, Ayerst) at 500 IU/fish or about 2 IU/g body weight. The amount of HCG used depended on the initial mean egg diameter; the smaller the diameter, the more HCG was used. Fish with oocytes characterized by germinal vesicle migration (mean egg diameter ≥ 0.47 mm) spawned without HCG injection. Fertilization and hatching rates for both treated and untreated fish were more than 90%. The larvae were reared to metamorphosis using rotifers from day 2–17, rotifers + newly hatched Artemia nauplii from day 18–20 and rotifers + newly hatched Artemia nauplii + artificial feed from day 21–35. In addition, Isochrysis galbana was introduced to the rearing tanks from day 1–10 and Chlorella sp. and/or Tetraselmis sp. from day 1–35. Survival rates of larvae tended to be lower as the broodstock became older.     

Effect of different diets on larval production, quality and fatty acid profile of the marine ornamental shrimp Lysmata amboinensis (de Man, 1888), using wild larvae as a standard

The present work evaluates the influence of broodstock diets [Marine Cuisine^®– MC, MC supplemented with highly unsaturated fatty acid- (HUFA) enriched Artemia biomass – MC + AB, and MC supplemented with squid – MC + S] on larval production, newly hatched and early zoeal stage survival and fatty acid profile of newly hatched larvae of Lysmata amboinensis . These parameters are compared with those from larvae hatched from embryos spawned in the wild. The number (±SE) of larvae produced with MC and MC + S (1077 ± 219 and 1103 ± 184, respectively) was similar to that in broodstock carrying embryos spawned in the wild (1224 ± 111), while those fed MC + AB displayed significantly lower values (1044 ± 161). Larvae produced with MC + AB displayed lower survival for all starvation periods, while larvae spawned in the wild displayed the highest survival. No larvae resisted 144 h of starvation and none moulted to zoea II. The fatty acid comparison revealed that larvae from embryos spawned in the wild displayed the highest levels of DHA, as well as higher DHA/EPA and n -3/ n -6 ratios. These results suggest that broodstock diets commonly used to promote ornamental shrimp's maturation (based on mixed frozen components) are far from being optimal.     

Analysis of viral load between different tissues and rate of progression of white spot syndrome virus (WSSV) in Penaeus monodon

At present the most common and most devastating disease of shrimp is caused by the white spot syndrome virus (WSSV), which has spread throughout the world mainly by different species of crustaceans carrying the virus. After experimental injection of Penaeus monodon with a known copy number of WSSV in the abdominal muscle, the rate of viral progression in different tissues at 12, 24, 36 and 48 hpi (hours post infection) was assessed using quantitative real‐time PCR. At 12 hpi the viral load was highest in haemocytes followed by pleopod, muscle and gills whereas at 48 hpi, the gills, the main target of WSSV, showed the highest viral load followed by pleopod, muscle and haemocytes. Viral copy number in the haemocytes was the lowest beyond 12 hpi indicating a remarkable reduction in the rate of viral replication in haemocytes compared with other tissues. The viral load in haemocytes, though increased again beyond 36 hpi, never surpassed the load in the other tissues. The real‐time PCR assay with its high sensitivity and wide dynamic range make it ideal for detecting low‐level WSSV infections that can occur in apparently healthy P. monodon.     

Mourilyan virus pathogenicity in kuruma shrimp (Penaeus japonicus)

The bunyavirus Mourilyan virus (MoV) occurs commonly in Black tiger (Penaeus monodon) and kuruma shrimp (Penaeus japonicus) farmed in eastern Australia. There is circumstantial evidence of MoV causing mortalities among P. japonicus moved from farm ponds to tanks for rearing as broodstock. To directly assess its pathogenic potential, independent cohorts of pond- (n = 24) or tank-reared juvenile (n = 21) P. japonicus were challenged intramuscularly with a cephalothorax tissue homogenate of P. monodon containing high loads of MoV (1.48 ± 0.28 × 10⁸ MoV RNA copies/µg total RNA). In each trial, mortalities accumulated gradually among the saline-injected controls. Mortality onset occurred 12–14 days earlier in the pond-reared shrimp, possibly due to them possessing low-level pre-existing MoV infections. Despite the time to onset of mortality differing, Kaplan–Meier survival analyses confirmed mortality rates to be significantly higher in both the pond- (p = .017) and tank-reared shrimp (p = .031) challenged with MoV. RT-qPCR data on shrimp sampled progressively over each trial showed high loads of MoV to establish following challenge and discounted GAV and other endemic viruses from contributing to mortality. Together, the data show that acute MoV infection can adversely compromise the survival of juvenile P. japonicus.     

A Microsatellite DNA Marker Developed for Identifying Disease‐resistant Population of Giant Black Tiger Shrimp,Penaeus monodon

White spot disease caused by white spot syndrome virus (WSSV) poses major problems that result in huge economic losses each year in shrimp aquaculture throughout the world. In the present study, microsatellite‐based DNA fingerprints have been compared between naturally occurring WSSV disease‐resistant and susceptible populations of giant black tiger shrimp, Penaeus monodon, to find DNA markers. For the first time, we report here a microsatellite locus, which, after amplification by polymerase chain reaction, provides a highly statistically significant DNA fingerprint of 71 bp, only in disease susceptible populations but not in disease‐resistant shrimp populations, whereas a 317 bp band is common in both. The absence of the former DNA marker will be very useful to identify disease‐resistant broodstock of P. monodon for marker‐assisted selection in breeding programs to generate disease‐free shrimps (P. monodon) in the aquaculture industry.     

Acrosome reaction in the sperm of the black tiger shrimp Penaeus monodon (Decapoda, Penaeidae)

Owing to the problem of male infertility in the domesticated shrimp Penaeus monodon, this study was conducted to reveal the morphological events of an acrosome reaction (AR) of sperm of this highly valuable species. The AR observed in an in vitro incubation of sperm with egg water (EW) and that during actual spawning was compared. Under transmission and scanning electron microscopy, sperm taken from the female thelycum was composed of a posterior main body, a central cap and an anterior single spike. Upon contact with EW, the sperm underwent two phases of AR: acrosomal exocytosis and spherical mass formation. The former was composed of a degeneration of the spike, swelling of the cap region and rupture of the acrosomal pouch. The latter began with polymerization of materials within the subacrosomal region and ended with re‐configuration of the subacrosomal region into an electron‐dense spherical mass. The AR of the sperm observed during spawning revealed similar morphological events, with degeneration of the spike upon contact with the vitelline envelope and formation of the spherical mass while penetrating into jelly material produced by protruding cortical rods. The results suggest the presence of AR inducers derived from the vitelline envelope and cortical rods of the egg. This study forms the basis for an evaluation of infertility regarding to AR in the domesticated P. monodon male.     

Examination of broodstock diet effects on egg production and egg quality in flame angelfish (Centropyge loriculus)

This study was conducted to determine the effects of broodstock diet on flame angelfish (Centropyge loriculus) reproduction and generate baseline spawning performance (mean fecundity, egg fertilization, egg viability) and egg biochemistry data for this species. Sixteen flame angelfish broodstock pairs were maintained on one of three formulated diets or a raw diet (consisting of squid, shrimp, fish eggs and algae) to determine the effect of diet composition on the fecundity, egg fertilization rates and egg viability over a 7‐month period. Although all the diets supported similar mean spawn frequency and fecundity over the duration of the experiment, the mean fertilization rates and egg viability were significantly greater among pairs receiving the raw diet. Egg fatty acid composition was generally reflective of the maternal diet and similar to the values reported for other tropical species. In particular, dietary factors such as n‐3 highly unsaturated fatty acid and ARA level (which were elevated in the raw diet and eggs from fish receiving that treatment) are correlated with egg quality and are therefore worthy of future investigation. Because of the ease of conditioning, adaptability to varied formulated diets and continuous, sustained reproductive output, flame angelfish are an excellent model for examining the factors affecting marine fish egg and larval quality.     

Highly efficient double‐stranded RNA transfection of penaeid shrimp using cationic liposomes

Yellow head virus (YHV) is a viral pathogen of shrimp that has caused high economic loss in the Thai shrimp farming industry, and the RNA interference mechanism has potential as a virus control strategy, as previous studies have shown that injection of virus specific double‐stranded RNA (dsRNA) can inhibit viral replication in shrimp. However, to date, complete inhibition viral replication has not been achieved. This study sought to determine whether cationic liposomes mixed with an antiviral dsRNA (YHV‐Pro‐dsRNA) were able to enhance protection against YHV. The results showed that injection of a YHV‐Pro‐dsRNA/cationic liposome complex markedly increased protection against YHV as compared with naked YHV‐Pro‐dsRNA, suggesting that cationic liposomes enhance the uptake of protective dsRNA into shrimp cells.