Long-term cryopreservation of spermatophore of the giant freshwater prawn, Macrobrachium rosenbergii (de Man)

Long‐term cryopreservation of the giant freshwater prawn, Macrobrachium rosenbergii, spermatophores using glycerol (Gly) and ethylene glycol (EG) as cryoprotective agents (CPAs) was studied. The tolerance of sperm to cryopreservation was evaluated on the basis of sperm survival and fertilizing ability. The survival of the sperm was determined by trypan blue staining, while the fertilizing ability was assessed from artificial insemination of the cryopreserved spermatophores. The rates of embryo survival on day 5 after spawning and of spermatophores capable of producing embryos survived to hatching were determined. Storage of spermatophores at ?20°C without CPA for a short period of up of 1–5 days decreased the sperm survival significantly and did not preserve fertilizing ability. Preservation at ?20°C in the presence of 10% or 20% Gly or of 10% or 20% EG offered a simple and efficient short‐term storage up to 10 days. For a long‐term storage, cryopreservation in the presence of 20% EG at ?196°C was more efficient than at ?20°C. High sperm survival rates and high fertilizing ability were recorded from those cryopreserved at ?196°C for up to 150 days. High sperm survival rates with moderate levels of fertilizing ability were obtained from those cryopreserved at ?20°C for not more than 30 days. The results indicate that preservation at ?196°C with 20% EG is a suitable procedure for long‐term storage of the giant freshwater prawn spermatophores.     

Low-cost diet for monoculture of giant freshwater prawn (Macrobrachium rosenbergii de Man) in Bangladesh

An experiment was conducted for 3 months in 12 experimental ponds, each of 30 m², with a view to develop a low‐cost diet for monoculture of Macrobrachium rosenbergii in ponds. Three experimental diets (30% protein) were formulated using fish meal, meat and bone meal, mustard oilcake, sesame meal and rice bran in different combinations partially replacing fish meal by meat and bone meal and sesame meal and assigned to treatments T₁, T₂ and T₃ respectively. A commercial golda feed (Starter‐III) was assigned to T₄ (reference diet). Each treatment had three replicates. Juveniles of M. rosenbergii (2.90±0.21 g) were stocked at the rate of 40 000 ha⁻¹. Prawns were fed three times daily at the rate of 10% and 5% of their body weight at the beginning and for the last 2 months respectively. The ponds were provided with aeration during the night using air pumps. The ranges of water quality parameters recorded in different ponds were: temperature 28.9–32.5°C, dissolved oxygen 5.1–8.1 mg L⁻¹ and pH 6.4–7.7. The results showed that the weight gain of prawns fed diet 1 was significantly higher (P<0.05) than those fed diets 2 and 3, but was not significantly different from those fed diet 4 (reference diet). The feed conversion ratio (FCR) values of diets ranged between 2.21 and 2.96 with diets 1 and 4 showing significantly lower (P<0.05) FCR values. The survivals (%) ranged between 68% and 78% with prawns fed diets 1 and 4 showing significantly higher survival. The production of prawn ranged between 921 and 1428 kg ha⁻¹ and diet 1 resulted in a significantly high (P<0.05) production. A simple economic analysis showed that diet 1 generated the maximum net profit of Tk 159 178 ha⁻¹. The results of the study showed that a diet containing 20% fish meal, 10% meat and bone meal, 15% mustard oilcake, 15% sesame meal, 35% rice bran, 4% molasses and 1% vitamin–mineral premixes may be recommended to the farmers for monoculture of M. rosenbergii in ponds.     

Optimization of stocking density of freshwater prawn Macrobrachium rosenbergii (de Man) in carp polyculture in Bangladesh

A study was conducted to optimize stocking density of freshwater prawn, Macrobrachium rosenbergii, in carp polyculture for 3 months in 10 experimental ponds of 80 m². Five stocking densities of prawn, 2500, 5000, 7500, 10 000 and 12 500 ha^?1, were assigned to treatments T₁, T₂, T₃, T₄ and T₅ respectively. The densities of catla, Catla catla, rohu, Labeo rohita and silver carp, Hypophthalmicthys molitrix, were 2500, 5000, and 2500 ha^?1, respectively, in each treatment. Each treatment had two replicate ponds. The mean initial weights of prawn, catla, rohu and silver carp were 1.1±0.02, 8.28±0.1, 25.2±1.1 and 36.32±1.2 g respectively. A pelleted diet containing 30% protein was prepared using fish meal, meat and bone meal, mustard oilcake, rice bran, wheat bran and molasses, and was fed twice daily at a rate of 5% of fish biomass. Water quality parameters were measured fortnightly and the ranges of temperature, pH and dissolved oxygen were 27.5–1.3°C, 6.9–8.6 and 4.5–8.6 mg L^?1 respectively. Feed conversion ratios ranged from 2.05 to 2.20 among the treatments. Per cent survival (%) of prawns ranged from 72% to 78%, while it varied from 80% to 93%, 90% to 95% and 90% to 92% for catla, rohu and silver carp respectively. The results showed that there were no significant differences among the weight gains of prawn and carp in different treatments. However, the overall total production of prawn and fish together was significantly (P<0.05) higher in T₃ and T₄ compared with other treatments. The total production for 3 months ranged between 2618 and 2916 kg ha^?1. The production of prawn was significantly higher (361.3 kg ha^?1) in T₅ with a highest stocking density of 12 500 prawn ha^?1. Although there was no significant difference (P>0.05) between the total production of prawn and fish together in T₃ and T₄, the highest net profit (Tk. 69 006 ha^?1) was obtained in T₄. Therefore, from the result of the study it may be concluded that a stocking ratio of 4:1:2:1 of prawn:catla:rohu:silver carp at a total density of 20 000 ha^?1 may be recommended for prawn–carp polyculture in ponds.     

Bacterial microflora associated with farmed freshwater prawn Macrobrachium rosenbergii (de Man) and the aquaculture environment

The initial counts of bacteria associated with farmed giant freshwater prawn (Macrobrachium rosenbergii de Man) as well as with the water and sediment from two farms located at Kottayam district in Kerala (India) were determined. A total of 367 randomly selected isolates were characterized and identified. Prawn samples yielded mean microbiological counts of 4.92 log CFU g^?1 of shell with muscle and 7.78 log CFU g^?1 of intestine at 30°C. Bacterial numbers in the intestine of prawn were much higher than those in the pond water. Motile and non‐motile aerobic Gram‐negative bacteria together with Enterobacteriaceae accounted for 60–70% of the mesophiles isolated from water and prawn. However, the microorganisms isolated in significant numbers from shell with muscle and intestine of prawn were not recovered from the growing waters. Among Gram‐positive bacteria, Micrococcus, Bacillus, coryneforms, and Arthrobacter were found. Faecal coliforms and enterococci were found in significant numbers in M. rosenbergii. The rearing practices such as feeding and pond fertilization could have influenced the microflora in prawn. The commensal microflora of freshwater prawn included opportunistic pathogens such as Aeromonas spp. and Streptococcus spp. Storage of prawn at elevated temperatures can permit their survival and growth leading to quality loss. Care should be exercised during handling and processing to prevent contamination of edible meat.     

罗氏沼虾胚胎附着机制的初步研究

甲壳动物十足目的大多数种类,其受精卵自产出后就附着于亲体腹部,并完成胚胎发育过程直至孵化,这对于胚胎的发育、保护和幼体散布有重要的生物学意义。胚胎的附着机制较为复杂,与刚毛、卵柄、卵索、卵膜和黏液腺有关:卵柄和卵索的形成决定了胚胎发育的顺利进行;卵膜的变化影响着胚胎的附着;黏液腺在胚胎附着过程中形成卵柄和外层卵膜,胚胎是依靠卵柄或卵索附着于腹肢的刚毛上,进而完成胚胎发育过程。本研究采用常规组织学和扫描电镜的方法研究了罗氏沼虾(Macrobrachium rosenbergii)胚胎及其附着结构。结果表明,胚胎附着与母体腹肢的结构、黏液腺分泌的黏液相关,也与胚胎表面结构、次级卵膜的形态和结构密切相关。原肢底节和内肢刚毛密而长,基部有细小的分支,有利于胚胎的附着,外肢刚毛羽状则有利于胚胎的托附和保护。抱卵雌虾腹肢的原肢和内肢上布满腹肢黏液腺分泌的黏液;携卵刚毛中空、壁厚,刚毛外壁由微绒毛状结构组成,扩大了携卵刚毛的表面积,以利胚胎附着。胚胎外表面由初级卵膜和次级卵膜组成,次级卵膜主要由卵巢黏液腺分泌的黏液参与形成。排卵时,受精卵依靠坚韧的次级卵膜相互粘连,同时,部分受精卵在刚毛上滑动、旋转,并借助次级卵膜逐渐形成了卵柄或卵索固着在腹肢刚毛上。从受精卵到胚胎孵化,胚胎之间最主要的连接方式是依靠次级卵膜的连接。胚胎的表面有许多褶皱和黏液,利于胚胎的附着以及胚胎对溶氧和小分子物质等的通透。以上结构与附着相适应,保证了胚胎发育的顺利进行和幼体的正常孵化。本研究旨为进一步认识甲壳动物胚胎附着机制、胚胎发育和人工育苗的研究提供参考。[中国水产科学,2007,14(1):67-73]     

Comparison of larval performance between Thai and Vietnamese freshwater giant prawn, Macrobrachium rosenbergii (de Man): a preliminary study

This study compared the performance of larvae of giant freshwater prawn (Macrobrachium rosenbergii) from Thailand and Vietnam. Thai broods were collected from a farm and Vietnamese broods were collected from a river. Larvae from these two sources were raised in identical environments, and their rates of development and survival as well as their ability to withstand environmental stresses were assessed. The first postlarval stage was observed in both strains on day 22. All larvae (100%) from Thai broods completed metamorphosis by day 36, but only 89% of the Vietnam strain completed metamorphosis by day 45. The stages of development were more uniform in larvae from the Thai stock than the Vietnamese stock. Survival differences were observed within the first week (92% in the Thai group vs. 55% in the Vietnamese group). These differences were most pronounced beginning in week 5 (52% and 16% respectively) and week 7 (33% and 2% respectively). No clear difference was found in brood size and fecundity between the two groups. No difference in survival was observed between the two groups of larvae challenged with salinity, oxygen or formalin. Differential development rates of larvae in the same batch would naturally create conditions favourable for cannibalism and impaired ability to capture food. The earlier and more uniform rate of development, and subsequent higher rate of survival of Thai stock compared with the Vietnamese wild stock is suggested to be due to inadvertent selection in the domesticated stock.     

On the breeding behaviour and reproduction of the freshwater prawn,Macrobrachium rosenbergii (de Man 1879) (Decapoda‐Crustacea) under laboratory conditions

The giant freshwater prawn, Macrobrachium rosenbergii is one of the most common decapod species, and now getting more attention from the aquaculturists world wide due to its high market demand. It is commercially important because of its size as well as its eating flesh qualities. The breeding behaviour, reproduction and hatching of this species were observed for about 8 months during 2009. Juveniles (0.55 ± 0.177 g total weight and 2.7 ± 0.12 cm, total length) were reared in rounded fibre glass tanks (1.3 m, diameter). Pre‐mating moult occurs in prawns once the ovaries ripen in their carapace cavity so as to transform the prawn into berried stage. Recorded incubation period ranged from 18 to 24 days. The number of eggs ranged from 2050 to 150 500 and the fecundity ranged from 435.2 to 3849.1 eggs. Number of hatched larvae ranged from 1825 to 123 410 larvae for females of 4.71 to 39.1 g respectively. The number of eggs carried by female prawn was directly proportional to its body weight (no. of eggs = 3441.3 wt. of female – 32 292, r² = 0.819). (fecundity = 82.066 wt. of female ? 235.04, r² = 0.7779; fecundity = 317.86 length of female ? 2651, r² = 0.833). Hatching rate ranged from 65% to 91%, and there was a correlation between number of hatched larvae and size of female parent (no. of larvae = 10 369 length of female – 102 965, r² = 0.8159; no. of larvae = 2792.9 wt. of female – 26 268, r² = 0.829). This study can greatly help in the management strategies of prawn hatcheries and improve its hatching technology.     

Genetic diversity of cultured and wild populations of the giant freshwater prawn Macrobrachium rosenbergii (de Man, 1879) based on microsatellite analysis

Freshwater prawn (Macrobrachium rosenbergii) culture in the Western Hemisphere is primarily, if not entirely, derived from 36 individual prawns originally introduced to Hawaii from Malaysia in 1965 and 1966. Little information is available regarding genetic variation within and among cultured prawn stocks worldwide. The goal of the current study was to characterize genetic diversity in various prawn populations with emphasis on those cultured in North America. Five microsatellite loci were screened to estimate genetic diversity in two wild (Myanmar and India‐wild) and seven cultured (Hawaii‐1, Hawaii‐2, India‐cultured, Israel, Kentucky, Mississippi and Texas) populations. Average allelic richness ranged from 3.96 (Israel) to 20.45 (Myanmar). Average expected heterozygosity ranged from 0.580 (Israel) to 0.935 (Myanmar). Many of the cultured populations exhibited reduced genetic diversity when compared with the Myanmar and the India‐cultured populations. Significant deficiency in heterozygotes was detected in the India‐cultured, Mississippi and Kentucky populations (overall F_is estimated of 0.053, 0.067 and 0.108 respectively) reflecting moderate levels of inbreeding. Overall estimate of fixation index (F_st = 0.1569) revealed moderately high levels of differentiation among the populations. Outcome of this study provide a baseline assessment of genetic diversity in some available strains that will be useful for the development of breeding programmes.     

人工诱导罗氏沼虾同步产卵与卵巢组织学研究

运用保幼激素类似物－ＺＲ５１５诱导罗氏沼虾同步产卵进行了生产性规模试验，每性雌虾体钙点滴４０μｌＪＨＡ－ＺＲ５１５，通过体表渗稼促进卵巢成熟和产卵。药物处理后１６－１７天，产卵率为４１．８１％，较常规生产的自然产卵率提高５０．２３％。罗氏沼虾卵巢生物学和组织滂研究表明，性成熟后的任何发育阶段，卵巢中均存在由卵原细胞组成的生发带和卵黄发生卵母细胞。     

A sandwich enzyme linked immunosorbent assay (S-ELISA) for detection of MrNV in the giant freshwater prawn,Macrobrachium rosenbergii (de Man)

A sandwich enzyme-linked immunosorbent assay (S-ELISA) was developed to improve diagnosis of white tail disease of the giant freshwater prawn, Macrobrachium rosenbergii, caused by the nodavirus, MrNV. Polyclonal antibodies were produced by immunization of Balb/C mice using a purified suspension of the virus and IgG anti-MrNV were purified from ascitic fluid. A sandwich method was successfully developed, coating first with unlabelled antibody and detecting trapped antigens with a second biotinylated antibody. Reaction was demonstrated using an avidin-peroxidase conjugate. Tissue extracts from M. rosenbergii infected with MrNV or purified viral extracts (control) were successfully identified in an individual ELISA, thus confirming the validity of the method. This S-ELISA should be the technique of choice for epidemiological studies of this disease and is a rapid and inexpensive assay with high specificity and sensitivity.     

Immunomodulatory effect of Cynodon dactylon against white tail disease of giant freshwater prawn,Macrobrachium rosenbergii (de Man, 1879)

The giant freshwater prawn, Macrobrachium rosenbergii, is an economically important and extensively cultured crustacean worldwide. The viral pathogens, Macrobrachium rosenbergii nodavirus (MrNV) and extra small virus (XSV) are responsible for causing severe mortalities in the hatchery and nursery phases. This study investigates the protection of postlarvae of freshwater against white tail disease (WTD) using plant extract derived from Cyanodon dactylon and the modulation of the prawn non‐specific immunity. To determine the immunomodulatory effect of C. dactylon extract, the prawn was injected with plant extract and various immunological parameters were estimated. The immunological parameters such as proPO, SOD, THC and clotting time were found to be significantly higher in the plant extract‐injected prawn when compared with control groups. The results of real time PCR analysis revealed up regulation on the expression proPO, SOD and lysozyme genes in MrNV and XSV challenged prawn postlarvae treated with C. dactylon extract. Infectivity experiment showed high relative per cent survival in MrNV and XSV‐challenged prawn postlarvae treated with C. dactylon extract. These results strongly indicate that the administration of C. dactylon plant extract enhances immunity of the prawn. Based on the results, this study recommends that the immersion of postlarvae in C. dactylon plant extract is a potential prophylactic agent against WTD.     

罗氏沼虾核型及长臂虾亚科核型演化关系的探讨

以略作改进的空气干燥法制备染色体标本，研究罗氏沼虾精巢细胞染色体数目，首次报道了其有丝分裂中期的详细核型。罗氏沼虾染色体数目ｎ＝５９，２ｎ＝１１８，依着线点位置及相对长度不同共划分为Ａ、Ｂ、Ｃ三个染色体组。     

The androgenic gland and monosex culture of freshwater prawn Macrobrachium rosenbergii (De Man): a biotechnological perspective

Males of the freshwater prawn Macrobrachium rosenbergii (De Man) grow faster and reach a larger size at harvest than females of the species. It is thus obvious that culture of monosex all‐male populations would be economically advantageous. Sexual differentiation in crustaceans is regulated by the androgenic gland (AG), which plays a pivotal role in the regulation of male differentiation and in the inhibition of female differentiation. In M. rosenbergii, AG removal from immature males resulted in sex reversal, with complete female differentiation. Similarly, AG implantations into immature females lead to the development of the male reproductive system. Sex‐reversed M. rosenbergii animals were capable of mating with normal specimens to produce offspring. Early attempts in Israel and more recently, attempts in other countries to establish all‐male populations through manual segregation showed that for the production of monosex prawn populations to be economically feasible, intervention via the AG is probably required. However, a suitable biotechnology is still to be developed, and an androgenic hormone has yet to be identified in decapods. Three lines of aquacultural and biotechnological research and development are proposed for the future: (1) Establishment of monosex cultures through manual segregation, together with the application of selective harvesting and claw ablation, as well as examination of different monosex culture strategies under a variety of economic conditions. (2) Microsurgical intervention in the AG, leading to the development of functional neo‐females, which would subsequently be mated with normal males to produce all‐male progeny. (3) Elucidation of AG bioactive products to enable biochemical or molecular manipulation of sex differentiation.     

Economic analysis of monosex culture of giant freshwater prawn (Macrobrachium rosenbergii De Man): a case study

All‐male monosex culture of Macrobrachium rosenbergii (De Man) has emerged as a popular practice in India, especially in the state of Andhra Pradesh. A study was conducted to compare the economics of all‐male, mixed and all‐female culture in 15 adjacent, rectangular ponds of 4000 m² each by stocking juveniles previously reared in a nursery for 60 days. The experiment was conducted using a completely randomized design with three treatments; T₁ (all male), T₂ (mixed) and T₃ (all female), and five replicates for a period of 5 months after the nursery phase. Statistical analysis showed highly significant (P<0.01) differences among the three types of culture. The cost of production was estimated and the economic feasibility of the culture methods was evaluated by cost‐return and partial budgeting analysis. The average weight, productivity and specific growth rate were the highest for all male culture, being 80.92±2.41 g, 1532 kg ha^?1 and 1.97±0.02 respectively. All‐female culture registered significantly higher survival (89.16±0.77%) and the best apparent feed conversion ratio of 1.26±0.02. The economic analysis revealed that all‐male monosex culture of M. rosenbergii was 63.13% and 60.20% more profitable than mixed and all‐female cultures respectively.     

Effect of cold shock and hormone on growth and male morphotypes of freshwater prawn,Macrobrachium rosenbergii (de Man)

A study was carried out to determine the effects of cold shock and 17α‐methyl testosterone hormone on growth in male morphotypes of giant freshwater prawn Macrobrachium rosenbergii. Juvenile males were exposed at 18 ± 0.25°C for 24 h with normal feed and cultured at 27 ± 0.7°C for 70 days as well as administrated feed with hormone separately at 400 mg kg^?1 feed for the first 30 days of culture period (70 days). The growth (Mean ± SD) of male juveniles at harvest was closely homogenous in the cold shock (CLS) treated group; while it was heterogeneous in hormone‐treated (MH) and control (CO) groups. Although total body weight of blue claw (BC) males was not significantly different in the controls and the treatments, the 2nd pereiopods (claw) weight and length of the BC was significantly lower in CLS (0.52 ± 0.05 g, 10.02 ± 0.65 cm) than that in the CO (0.71 ± 0.16 g, 12.53 ± 0.57 cm) and MH (0.75 ± 0.08 g, 12.85 ± 1.18 cm) treatments. Survival of prawns was similar in treatments. The morphotypes of male prawn were significantly different with 59% of BC males with small‐sized claws in CLS and, 45% and 30% of BC males with large‐sized claws in MH and CO respectively. Cold shock adversely affected male morphotypes of the prawn so that numbers of BC males with small‐sized claw was the highest in CLS tank. However, the CLS treatment resulted in more uniform marketable prawns and suggested a high production of BC males in an all‐male culture system.