Osmoregulation,growth and moulting cycles of the giant freshwater prawn (Macrobrachium rosenbergii) at different salinities

The giant freshwater prawn, Macrobrachium rosenbergii, is a species with a high commercial value in aquaculture. Two experiments were performed to determine the effects of salinities on the osmoregulation, growth and molting cycles of M. rosenbergii during growout. The first experiment was designed to determine whether these animals are capable of adapting to the changes in salinity seen in salinity intrusions in tropical deltas, with an incremental increase in salinity of 3‰ per day from 0‰ to 30‰ Haemolymph osmolality was rapidly regulated up to salinities of 15‰ , whereas animals conformed at higher salinities. The second experiment determined the growth, moulting cycle, osmolality, muscle water content and mortality during a 4‐month experiment at 0‰, 15‰ or 25‰ salinity. The weight gains in 0‰ and 15‰ were not significantly different and were comparable to the growth rates achieved in production farms with body mass increases of 2.6 and 2.3‐fold their initial body mass, respectively, after 4 months. The 25‰ group suffered from low growth, high mortality and a significantly lower moulting frequency. These data show that this species can be reared in brackish water up to 15‰, allowing for farming in the large areas impacted by salt water intrusions in tropical deltas.     

外源Fe3+诱导罗氏沼虾铁蛋白mRNA的表达

探讨铁蛋白ferritin基因在罗氏沼虾中的组织表达分布及外源Fe3+对ferritin mRNA表达的影响。通过RT-PCR表明,ferritin mRNA在罗氏沼虾精巢、卵巢、肌肉、心脏、中肠腺和血淋巴中均有表达;荧光定量PCR结果证实了ferritin mRNA在中肠腺中表达量最高,其次是精巢。对罗氏沼虾进行FeCl3溶液肌肉注射后,荧光定量PCR结果表明,肌肉中ferritin mRNA表达量在注射后3 h增加了61倍,高水平的表达一直维持到8 h,随后在12 h表达量急剧地下降;卵巢中ferritin mRNA表达量在3 h有近4倍显著增加,在注射后48 h内一直维持在较高水平;心脏ferritin mRNA表达量在24 h后增加了8倍,48 h后表达量开始下降;在血淋巴、中肠腺和精巢注射前后均未发现有显著性差异(P>0.05)。以上实验结果表明,外源Fe3+可以诱导罗氏沼虾铁蛋白mRNA的表达。     

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.     

Growth and carcass composition of giant freshwater prawn, Macrobrachium rosenbergii (De Man), fed different isonitrogenous and isocaloric diets

A feeding experiment was conducted for 135 days to observe the effect of different isonitrogenous (35% crude protein) and isocaloric (385 kcal) diets on the growth and carcass composition of giant freshwater prawn, Macrobrachium rosenbergii ( De Man 1879 ). Three experimental diets (ED1, ED2 and ED3) were prepared using locally available ingredients. These diets differed mainly in terms of percent contribution of major protein sources such as fish meal, soybean meal, groundnut oil cake and mustard oil cake. Experimental diets were evaluated against a commercial diet, which served as the control (CD). Juveniles 1.87–2.44 g in size were stocked at a population density of 40 000 ha⁻¹ and fed thrice daily at 10% in the beginning and reducing gradually to 7% and 5% of the body weight during the experimental period. No significant differences (P>0.05) in the growth performance were observed; however, a significantly (P<0.05) higher yield (721.9 kg ha⁻¹ 135 days⁻¹) was recorded for prawn fed with control diet, followed by experimental diet 2 (676.5 kg ha⁻¹ 135 days⁻¹, having soybean meal as a major protein source). The survival ranged between 63.8% and 77.7%, with different diets showing significantly higher survival. The apparent feed conversion ratio values of diets ranged between 3.15 and 3.49, with experimental and control diets showing non‐significantly lower AFCR values. At the end of the experiment, representative specimens from each treatment were collected and their carcass composition was measured. Analysis of variance showed that carcass protein and total carbohydrate contents were significantly (P<0.05) higher in prawns fed on a fish–soybean meal‐based diet (ED3) and a control diet. The total lipid contents of prawns, however, did not differ significantly among the various dietary treatments. The results of our study suggest that the experimental diets could be used effectively for M. rosenbergii without compromising growth and flesh quality.     

Engulfed pathogen-induced apoptosis in haemocytes of giant freshwater prawn, Macrobrachium rosenbergii

Haemocytes of the giant freshwater prawn, Macrobrachium rosenbergii, were investigated for the induction of apoptosis after phagocytosis of pathogenic yeasts, bacteria and non-pathogenic latex beads in vitro. Isolated haemocytes of M. rosenbergii were cultured at a ratio of 1:50 haemocytes to pathogen with the yeast Debaryomyces hansenii, the bacteria Aeromonas hydrophila or Enterococcus faecium, or with latex beads at 25 degrees C for 2 h, followed by washing to remove free particles. At least 200 haemocytes were counted to determine the phagocytosis rate, and the results showed that haemocytes engulfed latex beads at a higher rate than the aquatic pathogens. By transmission electron microscopy, the yeast- or bacterium-engulfing haemocytes displayed morphological changes characteristic of apoptosis, including formation of cytoplasmic vacuoles, chromatin condensation and fragmentation of nuclei. This pathogen-induced apoptosis was further confirmed by DNA laddering and TUNEL (terminal deoxynucleotidyl transferase-mediated deoxy-UTP nick-end-labelling) assays. Neither haemocytes treated with latex beads nor uninfected haemocytes (control group) showed signs of apoptosis after 48 h in culture.     

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.     

Effects of salinity on standard metabolic rate and critical oxygen tension in the giant freshwater prawn (Macrobrachium rosenbergii)

The extensively farmed giant freshwater shrimp, Macrobrachium rosenbergii, can survive salinities up to 26 g L^?1, but the commercially important grow‐out occurs exclusively in freshwater areas. Recent studies suggest the shrimp equally capable of growing in brackish as fresh water and a better understanding of how this species responds to changing salinity could significantly impact freshwater prawn farming in deltas and coastal areas. Here, the effect of salinity (0 and 15 g L^?1) on standard metabolic rate (SMR) and critical oxygen tension (P_crit) was measured in adult M. rosenbergii using intermittent closed respirometry. SMR was 79.8 ± 3.1 and 72.7 ± 2.9 μmol kg^?1 min^?1 in fresh and brackish water, respectively, with no significant difference between the two salinities (P = 0.122). During hypoxia M. rosenbergii maintained oxygen uptake down to a P_crit of 26.3 ± 1.4 mmHg in fresh and 27.2 ± 2.0 mmHg in brackish water (P = 0.682), showing that salinity had no overall effect on oxygen conductance in the animals. These findings are in agreement with recent growth studies and provide further evidence that grow‐out phase could be accomplished in brackish water areas. Thus, the predicted intrusions of brackish water in tropical deltas as a consequence of future global warming may not impact this important production.     

Immunomodulatory effect of recombinant RNA‐dependent RNA polymerase protein of Macrobrachium rosenbergii nodavirus in giant freshwater prawn M. rosenbergii

The present study evaluated the role of recombinant RNA‐dependent RNA polymerase (RdRp) protein of Macrobrachium rosenbergii nodavirus (MrNV) in modulating the immune response and in reducing MrNV load in infected prawn. In the first experiment, prawns (25–30 g) were injected with recombinant RdRp protein (RP) at a concentration of 0, 1.0 and 10 μg, and immune parameters and expression of some immune‐related genes were measured up to 14 days post injection (p.i.). In the second experiment, early juveniles were injected with a similar dose of RdRp and animals were challenged by immersion with MrNV. The infection status was detected in muscles by nested RT‐PCR up to 21 days post challenge. Prawn injected with higher concentration of RP showed significantly higher total haemocyte count at different period post injection. Significant up‐regulation of immune‐related genes was observed within 24 h in prawn treated with lower dose of RP and after 7 days p.i. at higher level of RP injection compared with adult control. Most of the tested samples (63%) were found to be RT‐PCR positive for MrNV at 48 h of post‐immersion challenge. After 14 days, MrNV was detected only in control prawn, while both RP‐injected groups were MrNV negative. This study elucidated the potential viral load reduction role played by RdRP in MrNV‐infected prawn.     

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.     

Experimental susceptibility of different life-stages of the giant freshwater prawn, Macrobrachium rosenbergii (de Man), to white spot syndrome virus (WSSV)

Studies were conducted by injecting/feeding white spot syndrome virus (WSSV) derived from infected shrimp, Penaeus monodon (Fabricius), to different life-stages, namely post-larvae, juveniles, sub-adults and adults of Macrobrachium rosenbergii (de Man). The disease was also induced in brood stock, and the eggs and larvae derived from these animals were subsequently tested for WSSV infection. All the stages except egg used for the experiment were found WSSV positive in histopathology, cross infection bioassay and polymerase chain reaction (PCR) analysis. Experimentally infected post-larvae and juveniles showed a high percentage of mortality and an increased rate of cannibalism. The cumulative mortality in post-larvae was up to 28%; with 28–40% cannibalism resulting in a maximum loss of up to 68%. In juveniles, observed mortality and cannibalism were 10–20% and 6.7–30.0%, respectively, and the maximum loss recorded was 50%. In sub-adults, mortality ranged from 2.8 to 6.7%, cannibalism was up to 20% and the total loss was up to 26.7%. Sub-adults and adults were found to be more tolerant to the infection as evidenced by the mortality pattern. A nested (two-step) PCR resulted in a 570-bp product specific to WSSV in all stages, except the eggs.     

Dietary supplementation of host‐associated lactic acid bacteria modulates growth,metabolic activities,and immune‐related gene expression in giant freshwater prawn,Macrobrachium rosenbergii

The present study was carried out to evaluate the dietary effects of host‐associated lactic acid bacteria on growth performance, metabolic enzyme activities, and immune response of Macrobrachium rosenbergii juveniles. To formulate the test diets, a control (Con) diet was supplemented with a commercial probiotic and three host‐derived bacteria Enterococcus faecalis (EC), Lactococcus lactis I (LC‐I), and L. lactis II (LC‐II), which were previously isolated from the gastrointestinal tract of adult individuals of M. rosenbergii. Juvenile M. rosenbergii (0.65 ± 0.008 g) were randomly stocked at 20 individuals/100 L of fiberglass tanks with three replications for each test diet. After 50 days, juveniles fed the diets LC‐I and LC‐II showed significantly higher (p < .05) weight gain, specific growth rate, and the lowest feed conversion ratio. The analyses of glutamic oxaloacetate transaminase and glutamic pyruvate transaminase in muscle and hepatopancreas revealed significantly (p < .05) reduced values in LC‐I fed juveniles. The total hemocyte count and phenoloxidase activity were significantly increased (p < .05) in LC‐I and LC‐II fed juveniles. The relative mRNA expression patterns of immune‐related α2‐M, LGBP, ProPO, Cu, Zn‐SOD, TG, PE, AKP, and ACP genes were significantly (p < .05) upregulated in juveniles fed with LC‐I followed by the diet LC‐II. Finally, the study suggests that the growth performance and immune response of M. rosenbergii can be improved through supplementation of host‐associated L. lactis bacteria for its higher production.     

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.