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Nitrite stress is a major environmental factor that limits aquatic animal growth, reproduction and survival. Even so, some shrimps still can withstand somewhat high concentrations of nitrite environment. However, few studies have been conducted about the tolerance molecular mechanism of Litopenaeus vannamei in the high concentration nitrite. To identify the genes and pathways involved in the regulation of nitrite tolerance, we performed comparative transcriptomic analysis in the L. vannamei nitrite-tolerant (NT) and nitrite-sensitive (NS) families, and untreated shrimps were used as the control group. After 24 h of nitrite exposure (NaNO 2 , 112.5 mg/L), a total of 1521 and 868 differentially expressed genes (DEGs) were obtained from NT compared with NS and control group, respectively. Functional enrichment analysis revealed that most of these DEGs were involved in immune defense, energy metabolism processes and endoplasmic reticulum (ER) stress. During nitrite stress, energy metabolism in NT was significantly enhanced by activating the related genes expression of oxidative phosphorylation (OXPHOS) pathway and tricarboxylic acid (TCA) cycle. Meanwhile, some DEGs involved in innate immunity- related genes and pathways, and ER stress responses also were highly expressed in NT. Therefore, we speculate that accelerated energy metabolism, higher expression of immunity and ER related genes might be the important adaptive strategies for NT in relative to NS under nitrite stress. These results will provide new insights on the potential tolerant molecular mechanisms and the breeding of new varieties of nitrite tolerant L. vannamei., (Copyright © 2021. Published by Elsevier Inc.)

Maintaining the homeostasis of energy metabolism is crucial for organism's stress tolerance and survival. Acute cold exposure (AC) and waterless duration (WD) represent the two predominate abiotic stressors during waterless live transport of Litopenaeus vannamei. Although previous reports have explored the physiological response of L. vannamei to combined stress AC + WD, the roles of energy metabolism response in regulation of stress tolerance remains unknown. The present study comparatively examined the variations of energy metabolism-related indicators in hemolymph (cortisol, hemocyanin, glucose and lactate), hepatopancreas and muscle tissues (levels of lactate and glycogen, activities of hexokinase (HK), phosphofructokinase (PFK), pyruvate kinase (PK), lactate dehydrogenase (LDH), succinate dehydrogenase (SDH) and adenosine triphosphatase (ATPase), and ATP levels). Combined stress significantly disturbed the homeostasis of energy metabolism with the increase in levels of hemocyanin, glucose and lactate, and decrease in glycogen and ATP content (P < 0.05). In addition, the activities of HK, PFK, PK, and SDH initially elevated and then decreased with the prolongation of combined stress from 3h to 9h duration, while the activity of lactate dehydrogenase (LDH) remained gradual elevation and ATPase activity decreased in a duration time dependent manner throughout the experiment. These alterations revealed that exposure to combined stress could accelerate anaerobic metabolism at initial stage and inhibit aerobic metabolism in a duration time-dependent manner, following with the reduction of energy biosynthesis and the disturbance of energy metabolism equilibrium. On the other hand, the progressive impairment on hepatopancreas tissue was observed under combined stress. In summary, the deficiency of ATP supply and histopathological injures on hepatopancreas tissue might the underlying mechanisms inducing mortality of L. vannamei during live transport., (Copyright © 2021. Published by Elsevier Ltd.)

Water temperature is one of the most common physiological stressors in aquaculture. Previous studies demonstrate that organisms require miRNA activity for survival in various unfavourable environmental conditions. However, the detailed role of miRNA in response to low-temperature stress is still unclear. This study was conducted to construct a comprehensive miRNA dataset for the Penaeus vannamei after low-temperature stress. A total of 329 known miRNAs and 60 putative novel miRNAs were identified. Among them, 17 miRNAs were identified with the most significant differences, and they were found to be involved in stimulation or stress processes. The main enriched target pathways of the 17 miRNAs were the Hippo signalling pathway, autophagy, apoptosis and MAPK signalling. In addition, all the 17 miRNAs identified were up-regulated, suggesting that miRNA by inhibiting the expression of target genes constitutes an effective strategy for Penaeus vannamei to cope with low-temperature stress. The 35-putative target of the 17 miRNAs was related to apoptosis and autophagy-related proteins, such as Pxt, DRAM2, cytochrome c, ATG2B, JNK, ATG4 and API5. The analysis of miRNA expression profiles contributes to the understanding of the molecular mechanisms of low-temperature tolerance in Penaeus vannamei. This study's findings enrich current miRNA resources and offer the possibility to validate the involvement of 17 miRNAs in the response of shrimp to low-temperature stress., (© 2021 John Wiley & Sons Ltd.)

Although Litopenaeus vannamei is a widely studied species, the information on how the organisms respond to natural daily variations of environmental conditions such as temperature and dissolved oxygen, and how such conditions alter the physiological responses, is scarce. In the present work, the strategies used by shrimps to cope with temperature and dissolved oxygen fluctuations during 24 days were investigated through the evaluation of oxygen consumption and heat shock proteins (HSP) gene expression. During daily fluctuations, no change in oxygen consumption in the short-term, but a significant increase in the long-term during hyperthermia conditions was registered, whereas a significant decrease during hypoxia was observed during all the bioassay. On the other hand, HSP70 and HSP90 gene expression increased in gills under thermal stress but was down-regulated under hypoxia, in both the short- and the long-term. This study highlights that to counteract environmental variations of temperature and dissolved oxygen, the shrimps use molecular compensatory mechanisms (HSP gene expression) that are different to those used under constant hypoxic conditions, suggesting that hypoxia can compromise physiological cytoprotection., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Ammonia is an important environmental stress factor in aquaculture. Long-term ammonia stress could affect the normal growth, and also increase the risk for the occurrence of various diseases. In order to learn the mechanism that ammonia caused the outbreak of the shrimp disease, transcriptomics and metabolomics approaches were used to analyze the differential expressions of the genes in hemocytes and different metabolites in the serum of the Pacific white shrimp Litopenaeus vannamei under ammonia exposure. Transcriptional analysis showed that 17 cell apoptosis related genes, seven phagocytosis related genes, 10 immunity related genes and seven cell cycle and lipid metabolism related genes showed differential expressions after ammonia exposure. Metabolomics analysis on the serum showed that 25 differential metabolites were identified in positive and negative ion patterns. They are involved in purine metabolism, amino acids metabolism and lipid metabolism. Injection of two up-regulated metabolites triethanolamine and oxypurinol to normal shrimp could induce apoptosis in normal shrimp. The total hemocytes counts in shrimp showed a significant decrease and the apoptotic cell ratio increased significantly under ammonia exposure. These results suggested that ammonia exposure increased the apoptosis of hemocytes, which affected the immunity of shrimp, and thus caused susceptibility to pathogenic infection. These data will help us understand the mechanism of ammonia stress leading to the immunity decline of shrimp., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

The objective of this study is to explore the immune response of the shrimp Penaeus vannamei to low temperature and air exposure during the mimic waterless transportation. Shrimp were cold shocked at 13 °C for 3 min, then exposed to air at 15 °C for 12 h, and finally revived in water at 25 °C. Hemocyte structure remained intact with only slight distortions of some organelles and nuclear membrane under the stress. Phenoloxidase (PO), lysozyme (Lys) and gamma-glutamyl transferase (GGT) activities, glutamine (Gln) content and relative mRNA expressions of prophenoloxidase (proPO), β-1,3-glucan binding protein (LGBP), ferrin (Fer) and glucose regulated protein 78 (GRP 78) increased and reached peak levels after 3 h-9 h of air exposure, and then decreased to relatively stable levels in the prolonged period of air exposure. The total hemocyte count (THC) and gene expressions of proPO, Fer and LGBP at the end of waterless storage were significantly lower (p < 0.05) than those in fresh shrimp while no significant difference (p ≥ 0.05) was found between revived and fresh shrimp in PO, Lys, GGT activities, Gln content and gene expression level of GRP 78. Of all the hemocytes, the percentage of semi granular cells (SGC) and granular cells (GC) significantly decreased after 6-9 h of storage, while hyaline cells (HC) dramatically increased after 9 h of storage to compensate the loss of SGC and GC. Results suggested that the low temperature and air exposure caused significant immunological response to the shrimp, but the damages to the immune system were partly reparable., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

pH of water environment affects the survival of aquatic animals. Intestine barrier function influences the health of animals, which is related to its mucosa structure, immune components, and microbial communities. In this study, we investigated the histological structure, digestive and metabolic capacity, immune responses, and microbial composition in the intestine of Litopenaeus vanmei under three different conditions: control (pH 8.3), low pH stress (pH 6.9), and high pH stress (pH 9.7) for 72 h. The results showed both low and high pH stress disrupted the intestine morphological structure, and induced variations in the activities of digestive (AMS, LPS, Tryp, and Pep) and metabolic (HK, PK, CCO, and LDH) enzymes. Low and high pH stress also increased oxidative stress (MDA, LPO, PC, and ·O 2 - generation), and decreased the antioxidant enzyme activities (T-AOC, SOD, and GST); shrimp enhanced CAT activity and HSP70, Trx, MT and Fer gene transcripts as defense mechanism. Additionally, Immune confusion was also found in the shrimp intestine in response to low and high pH stress, including the antibacterial ability (T-NOS, PO, proPO, ALF, and Lys), pathogen recognition (TLR and Lec), apoptosis (Casp, IAP and p53), and mucus homeostasis (Muc-1, Muc-2, Muc-5AC, Muc-5B, and Muc-19). pH exposure also decreased the diversity of the intestine bacterial, disturbed the composition of microbiota, and decreased the microbial metabolite SCFA contents. Our results indicated that acute pH stress can impair the intestine barrier function of white shrimp, probably via destroying mucosa structure, confusing digestion and metabolism, inducing oxidative stress, disordering immunity, and disrupting the microbial composition., (Copyright © 2019. Published by Elsevier Ltd.)

Proline (Pro) metabolism is intimately associated with stress adaptation. The catabolism of Pro includes two dehydrogenation reactions catalyzed by proline dehydrogenase (ProDH) and Δ 1 -pyrroline-5-carboxylate dehydrogenase (P5CDh). P5CDh is a mitochondrial matrix NAD + -dependent dehydrogenase that is critical in preventing P5C-Pro intensive cycling and avoiding ROS production from electron run-off. Little is known about the roles of P5CDh in invertebrates, however. We cloned the P5CDh sequence in the Pacific white shrimp, Litopenaeus vannamei, and found that LvP5CDh is expressed predominantly in pleopod, hepatopancreas and gill. Subcellular localization analysis revealed that LvP5CDh protein was mainly found in the cytoplasm. In addition, overexpressing LvP5CDh in cells reduced ROS formation and inhibited apoptosis induced by LC 50 Cd 2+ . Shrimp were exposed to various stress factors including infection with Vibrio alginolyticus, (½ LC 50 and LC 50 ) Cd 2+ , acid (pH 5.6) and alkali stress (pH 9.3). Both biotic and abiotic stress resulted in increased LvP5CDh expression and Pro accumulation; V. alginolyticus infection, pH 9.3 and LC 50 Cd 2+ stress apparently stimulated the Glu pathway of Pro synthesis, while pH 5.6 and ½ LC 50 Cd 2+ stress promoted the Orn pathway of Pro synthesis. Silencing of Lvp53 in shrimp attenuated LvP5CDh expression during Cd 2+ stress, but had no effect on LvP5CDh mRNA levels if no Cd 2+ stress was imposed. Our study contributes to the functional characterization of LvP5CDh in biotic and abiotic stress and reveals it to protect against ROS generation, damage to the cell, including the mitochondria, and apoptosis. Thus, LvP5CDh plays a critical role in immune defense and antioxidant responses., (Copyright © 2018 Elsevier B.V. All rights reserved.)

As a unique atypical 2-Cys Peroxiredoxin (Prx) of the Prx-like superfamily, Peroxiredoxin5 (Prx5) possesses special properties, such as its enzymatic mechanism, wide subcellular distribution and high affinity for peroxides and peroxynitrite. Prx5 plays a crucial role in oxidative stress, immune responses, cell apoptosis, proliferation, differentiation, intracellular signaling, the modulation of gene expression, ecdysis, etc. In this paper, we obtained a full-length Prx5 cDNA sequence (designated PmPrx5) from black tiger shrimp (P. monodon). The full-length PmPrx5 cDNA sequence was 1686 bp containing a 5' untranslated region (UTR) of 76 bp with two nucleotide sequences (AAA), a 3' UTR of 1040 bp with a poly (A) tail and two canonical polyadenylation signal sequences (AATAAA), and an open reading frame of 570 bp encoding 189 amino acid residues with a predicted molecular mass of 20 kDa and a theoretical isoelectric point of 6.29. Phylogenetic trees and multiple sequence alignment showed that the PmPrx5 had strong homology with Prx5 proteins from other species, such as similarity with Palaemon carinicauda (69%) and Macrobrachium rosenbergii (69%), containing the highly conserved functional domain. PmPrx5 mRNA was ubiquitously detected in all tested tissues. After P. monodon was exposed to pathogenic bacteria, osmotic pressure, acidity and alkalinity and the heavy metal, the mRNA expression of PmPrx5 in the gills and hepatopancreas was significantly enhanced (P < 0.01) because of the immune response and declined with heavy metal copper and cadmium challenges as time progressed. The recombinant PmPrx5 protein purified in E. coli (DE3) was further confirmed to exhibit antioxidant activity and antibacterial properties to a certain extent using a bacterial growth inhibition test in both liquid and solid cultures in vitro. E. coli transformed with pRSET-PmPrx5 were dramatically protected in response to metal toxicity stress. Thus, PmPrx5 may be developed as a potential therapeutic drug against pathogenic bacteria and as a biomarker for pollutant levels. This work offers useful clues to further explore the functional mechanism of Prx5 in marine shrimp immunity., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Aquatic animals are frequently suffered from starvation due to restricted food availability or deprivation. It is currently known that gut microbiota assists host in nutrient acquisition. Thus, exploring the gut microbiota responses would improve our understanding on physiological adaptation to starvation. To achieve this, we investigated how the gut microbiota and shrimp digestion and immune activities were affected under starvation stress. The results showed that the measured digestion activities in starved shrimp were significantly lower than in normal cohorts; while the measured immune activities exhibited an opposite trend. A structural equation modeling (SEM) revealed that changes in the gut bacterial community were directly related to digestive and immune enzyme activities, which in turn markedly affected shrimp growth traits. Notably, several gut bacterial indicators that characterized the shrimp nutrient status were identified, with more abundant opportunistic pathogens in starved shrimp, although there were no statistical differences in the overall diversity and the structures of gut bacterial communities between starved and normal shrimp. Starved shrimp exhibited less connected and cooperative interspecies interaction as compared with normal cohorts. Additionally, the functional pathways involved in carbohydrate and protein digestion, glycan biosynthesis, lipid and enzyme metabolism remarkably decreased in starved shrimp. These attenuations could increase the susceptibility of starved shrimp to pathogens infection. In summary, this study provides novel insights into the interplay among shrimp digestion, immune activities and gut microbiota in response to starvation stress., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

A 25-days experiment was conducted to evaluate the effect of dietary Haematococcus pluvialis on growth, survival, immune response and stress tolerance ability of post-larval Litopenaeus vannamei. Post-larval white shrimp (mean initial weight 2.1 mg) were fed five isoenergic and isonitrogenous diets containing grade levels of Haematococcus pluvialis (0, 1.7, 3.3, 6.7 and 13.3 g kg -1 diet, respectively). Results indicated that 3.3 g Haematococcus pluvialis kg -1 diet increased the survival rate of post-larval white shrimp. Specific growth rate (SGR) and weight gain (WG) showed no difference among each groups. After the acute salinity stress (salinity decreased rapidly from 28‰ to 5‰), survival of shrimp fed 6.7 g Haematococcus pluvialis kg -1 diet significant higher than the control (P < 0.05), and the total antioxidant capacity (T-AOC) was increased with the increasing dietary Haematococcus pluvialis levels. The malonaldehyde (MDA) contents in whole body decreased with the increasing dietary Haematococcus pluvialis levels before and after the salinity stress. Before the salinity stress, relative mRNA levels of Caspase 3, Rho and Janus kinase (JAK) decreased in shrimp fed diets contain Haematococcus pluvialis. After the salinity stress, relative mRNA levels of anti-oxidative related genes and immune related genes decreased with the dietary Haematococcus pluvialis level increased to 3.3 g kg -1 . Based on the effect of Haematococcus pluvialis on survival, salinity stress tolerance ability and the immune response of post-larval L. vannamei, the optimal level of Haematococcus pluvialis was 3.3-6.7 g kg -1 diet (100-200 mg astaxanthin kg -1 diet)., (Copyright © 2018. Published by Elsevier Ltd.)

The sodium bicarbonate cotransporter (NBC) is an integral membrane ion transporter that can transport HCO 3 - (or a related species, such as CO 3 2- ) across the plasma membrane. Previous researches revealed that NBC might play an important role in the regulation of intracellular pH in vertebrates. In the present study, an NBC cDNA was identified from Pacific white shrimp (Litopenaeus vannamei) and designated as Lv-NBC. The full-length Lv-NBC cDNA is 4479 bp in size, containing a 5'-untranslated region (UTR) of 59 bp, a 3'-UTR of 835 bp and an open reading frame (ORF) of 3585 bp that encodes a protein of 1194 amino acids with a deduced molecular weight of 134.34 kDa. The Lv-NBC protein contains two functional domains (Band&#95;3&#95;cyto and HCO3&#95;cotransp) and twelve transmembrane (TM) domains. Expression of the Lv-NBC mRNA was ubiquitously detected in all selected tissues, with the highest level in the gill. By in situ hybridization (ISH) with Digoxigenin-labeled probe, the Lv-NBC positive cells were shown mainly located in the secondary gill filaments. After low or high pH challenge, the transcript levels of Lv-NBC in the gill were found to be up-regulated. After knockdown of the Lv-NBC level by siRNA, the mortality of shrimp significantly increased under pH stress. Our study, as a whole, may provide evidences for the role of NBC in shrimp responding to pH stress, and give a new insight of the acid/base homeostasis mechanism in crustaceans., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

In the practical farming of Litopenaeus vannamei, the intensive culture system and environmental pollution usually results in a high concentration of ammonia, which usually brings large detrimental effects to shrimp, such as increasing the susceptibility to pathogens, reducing growth, decreasing osmoregulatory capacity, increasing the molting frequency, and even causing high mortality. However, little information is available on the molecular mechanisms of the detrimental effects of ammonia stress in shrimp. In this study, we performed comparative transcriptome analysis between ammonia-challenged and control groups from the same family of L. vannamei to identify the key genes and pathways response to ammonia stress. The comparative transcriptome analysis identified 136 significantly differentially expressed genes that have high homologies with the known proteins in aquatic species, among which 94 genes are reported potentially related to immune function, and the rest of the genes are involved in apoptosis, growth, molting, and osmoregulation. Fourteen GO terms and 6 KEGG pathways were identified to be significantly changed by ammonia stress. In these GO terms, 13 genes have been studied in aquatic species, and 11 of them were reported potentially involved in immune defense and two genes were related to molting. In the significantly changed KEGG pathways, all the 7 significantly changed genes have been reported in shrimp, and four of them were potentially involved in immune defense and the other three were related to molting, defending toxicity, and osmoregulation, respectively. In addition, majority of the significantly changed genes involved in nitrogen metabolisms that play an important role in reducing ammonia toxicity failed to perform the protection function. The present results have supplied molecular level support for the previous founding of the detrimental effects of ammonia stress in shrimp, which is a prerequisite for better understanding the molecular mechanism of the immunosuppression from ammonia stress., Competing Interests: The authors have declared that no competing interests exist.

AMP-activated protein kinase (AMPK) serves as a major regulator of cellular energy metabolism by activating ATP production pathways and blocking ATP consumption. However, information on AMPK genes in aquatic animals is limited. In this study, three subunits of AMPK were cloned from the Pacific white shrimp Litopenaeus vannamei. The full-length cDNAs of the α, β and γ subunits were 1617, 1243 and 3467bp long, respectively, with open reading frames of 1566, 873 and 2988bp encoding for 521, 290 and 996 amino acids, respectively. Amino acid sequence alignments of the three subunits showed that the functional domains in the L. vannamei proteins retained the highest similarity with those of other animals, at 89%, 58%, and 75%, respectively. The expression levels of the three subunits were higher in the muscle and gills than in the eyestalk and hepatopancreas. The mRNA levels of AMPK-α and AMPK-β were up-regulated in the hepatopancreas and muscle after acute low-salinity stress at 3psu for 6h compared with control salinity at 20psu. After 8-week salinity stress at 3psu, AMPK-α and AMPK-β mRNA levels in the hepatopancreas were significantly higher than those of the control at 30psu. However, in the muscle only AMPK-γ mRNA was significantly up-regulated at low salinity relative to controls. Muscle and hepatopancreas showed increases in AMPK protein after 6h exposure to low salinity, but there were no differences seen after long term acclimation. The change patterns of protein were slightly differing from the mRNA patterns due to the distinguishing function of individual subunits of AMPK. These findings confirm that three AMPK subunits are present in L. vannamei and that all encode proteins with conserved functional domains. The three AMPK subunits are all regulated at the transcriptional and protein levels to manage excess energy expenditure during salinity stress., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Cytosolic manganese superoxide dismutase (cMnSOD) is an important antioxidant enzyme which catalyzes the conversion of superoxides to oxygen and hydrogen peroxide in several organisms. In the Pacific white shrimp, Litopenaeus vannamei, three cMnSOD genes (LvcMnSOD1-3) have previously been characterized. Here, the genomic structure of LvcMnSOD2 and its mRNA expression in response to thermal stress was examined. Analysis of the nucleotide sequence demonstrated that LvcMnSOD2 is comprised of 2392 bp spanning from the ATG translation start site to the stop codon and contains six exons interrupted by five introns. The 5' region upstream of the LvcMnSOD2 gene contains several putative regulatory elements but lacks the accepted TATA sequence. The putative transcription factor binding elements that may be involved in LvcMnSOD2 mRNA expression level include activator protein-1 (AP-1), cAMP response element binding protein (CREB), upstream stimulatory factor (USF), CAAT-enhancer binding protein (C/EBP), nuclear factor-κB (NF-κB) and heat shock regulatory element (HSE). In addition, we compared the 5' upstream sequences of the LvcMnSOD2 gene between two shrimp strains that are resistant or susceptible to Taura syndrome virus (TSV), respectively, which revealed the absence of the USF and C/EBP elements at positions -2125 and -1986, respectively, in the TSV-susceptible shrimp line. Moreover, genomic variations between the two shrimp strains were detected in some of the putative C/EBP, USF, HSE and NF-κB transcription factor binding elements. That these genomic variations might be involved in the TSV resistance as well as in stress responses remains to be evaluated. The presence of 15 putative HSEs suggests that the expression of LvcMnSOD2 is regulated under thermal stress. Here, we found that in response to a 1 or 3 h thermal stress (35 °C), the mRNA expression levels of LvcMnSOD2 were significantly increased and then gradually decreased in the recovering phase at room temperature (25 °C) to control levels by 3 h after the heat shock. Thus, the antioxidant system may be induced to protect cells from the oxidative damage caused by thermal stress. The genomic organization of LvcMnSOD2 likely provides a clue to the mechanisms that might regulate the antioxidant defense pathway in shrimps and so potentially in marine invertebrates., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

V-H ATPase and NaK ATPase are important classes of ATP-driven proton pumps that are present in the intracellular and plasma membranes of eukaryotic cells and play diverse roles in both normal and abnormal cellular processes. Among the subunits of the V-H ATPase complex, subunit a is a transmembrane glycoprotein that plays crucial roles in metabolism, growth, survival and cellular immunity. NaK ATPase subunit beta is thought to participate in the proper folding and movement of the NaK ATPase enzyme and may also aid cation transport. In this study, we analyzed the functions of V-H ATPase subunit a and NaK ATPase subunit beta from the Pacific white shrimp, Litopenaeus vannamei. Full-length cDNAs of the genes corresponding to V-H ATPase subunit a and NaK ATPase subunit beta were obtained, which were 2654 and 2055 bp long, with open reading frames encoding 830 and 313 amino acids, respectively. RT-PCR analysis indicated that mRNA transcripts were strongly (but differentially) expressed in the gills and hepatopancreas, and at lower levels in other shrimp tissues. In this study, for the first time, the gene expression of V-H ATPase subunit a and NaK ATPase beta of white shrimp Litopenaeus vannamei were analysed by quantitative real-time PCR after exposure to five kinds of environmental stresses (bacteria, pH, Cd, salinity and low temperature). The results demonstrate that both of the two genes are sensitive and involved in all different stress responses and are more sensitive to salinity than other stresses. And they may have relationship with the anti-stress mechanism induced by environment stress in shrimp.

Two isoforms of 14-3-3 protein, namely 14-3-3A and 14-3-3B, from the shrimp Penaeus monodon were investigated for their potential role in adaptation to salinity stress. Transcripts of 14-3-3A were found in various shrimp tissues whilst expression of 14-3-3B transcripts was more specific being observed in the osmoregulatory tissues, that is in the gills and epipodites. In shrimp gills, the 14-3-3A transcript levels slightly changed after transfer of the shrimp from 3 to 25 or to 40 ppt. In contrast, significant change in the mRNA levels in response to salinity stress was detected for 14-3-3B, where a significant decrease of 14-3-3B transcript was observed in gills of shrimp transferred from hypo-osmotic (3 ppt) salinity to iso-osmotic or hyper-osmotic (25 and 40ppt, respectively) salinity. On the other hand, shrimp transferred from 40 ppt to 3 ppt showed a strong induction of 14-3-3B mRNA expression in the gills. These transcript expression analyses suggest that 14-3-3B is likely to be involved in the hyper-osmotic regulation in P. monodon. In addition, 14-3-3B appeared to regulate ATPase function since suppression of the gene by RNA interference resulted in a significant decrease in the total ATPase activity., (Copyright © 2011 Elsevier Inc. All rights reserved.)

To evaluate the genotoxic, physiological and immunological effects of short-term acute low temperature stress on the Pacific white shrimp, Litopenaeus vannamei, we rapidly transferred shrimp from tanks at 23±2 °C to aquaria at the same temperature (controls) or 12±2 °C for 12 h. Changes in the shrimp hemocyte respiratory burst activity and DNA damage were examined during and after exposure to the temperature stress using flow cytometry and the comet assay, respectively. We also monitored changes in the total hemocyte count, malondialdehyde levels, total protein concentration and osmolality in shrimp plasma. The results show that hemocyte respiratory burst activity, malondialdehydes levels and hemocyte DNA damage in the plasma all increased significantly after exposure to 12±2 °C for 3 h. In contrast, total hemocyte count, total protein concentration and osmolality in the plasma decreased compared to the controls. We conclude that acute low temperature can induce oxidative stress, DNA damage, lipid peroxidation and changes in osmolality in L. vannamei., (Copyright © 2011 Elsevier Inc. All rights reserved.)

In this study, the probiotic, Bacillus subtilis E20, isolated from the human health food, natto, was used for white shrimp, Litopenaeus vannamei, larvae breeding to improve the larval survival rate and development by adding probiotic to the rearing water at (control), 10(8), and 10(9) cfu L(-1) salt water once every 3 days during the 14 days of breeding experiment. Thereafter, stress tolerance and immune status of postlarvae were evaluated. Shrimp larval development was significantly accelerated after adding the probiotic to the larval rearing water at a level of 10(9) cfu L(-1). The survival rate of larvae was significantly higher in the treatment with 10(9) cfu L(-1) compared to the control and the treatment with 10(8) cfu L(-1) after all larvae had metamorphosed to postlarvae. Adding the probiotic to the shrimp larvae rearing water produced a weak inhibition of bacterial growth by an analysis of the total bacterial count and presumptive Vibrio count. For stress tests, no postlarvae died when they were reared in water in which the temperature was decreased from 30 to 2 degrees C at a rate of 0.1 degrees C min(-1). Postlarvae had significantly lower cumulate mortality in the treatments with 10(8) and 10(9) cfu L(-1) compared to the control when they were suddenly exposed to fresh water and 60 per thousand salt water. A significant decrease in the cumulative mortality of postlarvae treated with the probiotic at a level of 10(9) cfu L(-1) was recorded after the sudden transfer to 300 mg L(-1) nitrite-N compared to the control and treatment with 10(8) cfu L(-1). The analysis of immune-related gene expressions showed that the gene expression of prophenoloxidase I, prophenoloxidase II, and lysozyme of larvae were significantly increased after being reared in probiotic-containing water at the levels of 10(8) and 10(9) cfu L(-1). However, no significant difference in serine proteinase or glutathione peroxidase gene expressions was recorded in this study. It is therefore suggested that 10(9) cfu L(-1) of probiotic, B. subtilis E20 adding to rearing water for shrimp larva breeding., (2010 Elsevier Ltd. All rights reserved.)