Acute hepatopancreatic necrosis disease (AHPND), caused by marine bacteria Vibrio Parahaemolyticus, is a huge problem in shrimp farms. The V. parahaemolyticus infecting material is contained in a ...plasmid which encodes for the lethal toxins PirABVp, whose primary target tissue is the hepatopancreas, causing sloughing of epithelial cells, necrosis, and massive hemocyte infiltration. To get a better understanding of the hepatopancreas response during AHPND, juvenile shrimp Litopenaeus vannamei were infected by immersion with V. parahaemolyticus. We performed transcriptomic mRNA sequencing of infected shrimp hepatopancreas, at 24 hours post-infection, to identify novel differentially expressed genes a total of 174,098 transcripts were examined of which 915 transcripts were found differentially expressed after comparative transcriptomic analysis: 442 up-regulated and 473 down-regulated transcripts. Gene Ontology term enrichment analysis for up-regulated transcripts includes metabolic process, regulation of programmed cell death, carbohydrate metabolic process, and biological adhesion, whereas for down-regulated transcripts include, microtubule-based process, cell activation, and chitin metabolic process. The analysis of protein- protein network between up and down-regulated genes indicates that the first gene interactions are connected to oxidation-processes and sarcomere organization. Additionally, protein-protein networks analysis identified 20-top highly connected hub nodes. Based on their immunological or metabolic function, ten candidate transcripts were selected to measure their mRNA relative expression levels in AHPND infected shrimp hepatopancreas by RT-qPCR. Our results indicate a close connection between the immune and metabolism systems during AHPND infection. Our RNA-Seq and RT-qPCR data provide the possible immunological and physiological scenario as well as the molecular pathways that take place in the shrimp hepatopancreas in response to an infectious disease.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
The immunostimulatory potential of the marine yeast Yarrowia lipolytica (D1 and N6 strains) administered orally was evaluated in the white shrimp Litopenaeus vannamei. Yeasts and commercial glucans ...were mixed with a commercial feed to formulate diets with a 1.1% concentration of immunostimulants. The shrimp were fed daily for a period of 21 days. Weekly determinations were performed for immunological parameters in hemolymph, such as total hemocyte count (THC), lysozyme activity (LYZ), prophenoloxidase activity, antioxidant enzymatic activities (superoxide dismutase SOD, catalase CAT, and peroxidases), and bactericidal activity against Vibrio parahaemolyticus. Expression profiles of penaeidin (PEN), lysozyme (LYZ), and prophenoloxidase (proPO) immune genes were evaluated in hemocytes. In general, an increase in the immune parameters was observed in shrimp fed yeast diet compared to glucan and the control diets. Yarrowia lipolytica, especially strain N6, provided maximum immunostimulatory effects evidenced by the increase of immune parameters (THC, LYZ, SOD, CAT) and gene expression profile. In conclusion, this study demonstrated that Y. lipolytica had immunostimulatory effects and increased bactericidal activity in L. vannamei hemocytes against V. parahaemolyticus. These findings open the path for the potential application of Y. lipolytica-based immunostimulant for shrimp aquaculture.
•This is the first report on the immunological response to Yarrowia lipolytica in shrimp.•Oral administration of Y. lipolytica stimulates innate immune responses.•Y. lipolytica increased bactericidal activity in shrimp.•Circulating hemocytes and immune-related enzymatic activities increased in shrimp fed Y. lipolytica.•Immune-related gene expression up-regulated in hemocytes upon yeast administration.
The escape response in some decapod crustaceans through tail-flipping represents an excellent model of short-term, exhausting muscular activity to understand the role of energy depletion and ...oxidative stress in muscle fatigue. In this study indicators of oxidative stress and changes in energy metabolism were measured during escape response until fatigue in whiteleg shrimp Litopenaeus vannamei. The escape response of individual shrimp was elicited in a tank by prodding the shrimp with a stick until exhaustion (no further response). A control group consisted of undisturbed shrimp, and a third group was allowed to recover. Arginine phosphate (ArgP) and adenylic energy charge (AEC) were measured by reverse phase HPLC, while fuels, reserves, antioxidant enzyme activities (superoxide dismutase, catalase, glutathione peroxidase-GPx), thiobarbituric acid reactive substances, and protein carbonyl content were analyzed by spectrophotometric methods. Shrimp reached fatigue after 31 tail-flips, which decreased abdominal muscle ArgP by 75% and AEC by 18%. Hemolymph lactate levels increased 5 times and muscle GPx activity by 2.7 times. After 1h of recovery, ArgP and AEC base levels were re-established, while hemolymph lactate concentration and GPx activity were still increased, and glucose hemolymph content increased 1.8 times. The drastic decrease in ArgP levels prevents a drop of AEC to a critical level, in accordance with a situation of non-lethal stress. Although no significant changes were observed in muscle glycogen or lactate concentrations, the increase in hemolymph lactate levels indicates an important contribution of muscle glycolysis to prevent a more pronounced decrease in ATP and AEC. The increase in GPx activity in abdominal muscle suggests an increase in reactive oxygen species production during intensive muscular activity, which was adequately neutralized, for no signs of oxidative damage occurred, as indicated by the lack of lipid and protein oxidation. These responses suggest adequate energy efficiency and prevention of oxidative damage during the escape response, in accordance with the adaptative value of such behavior to avoid instantaneous danger, with a capacity of rapid recovery.
•Escape response Bioenergetics and oxidative stress of L. vannamei were studied.•Escape response to exhaustion decreased energetic status and phosphoarginine levels.•Escape response to exhaustion increased hemolymph lactate.•After recovery the energetic status was reestablished and glucose hemolymph increased.•Glutathione peroxidase activity increased after exhaustion and after 1h recovery.
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•Three stages of E-RLO infection were identified based on internal lesions.•The intensity of infection differed between tissues.•Damage suggests that the E-RLO has the capacity to ...impairs the C. gigas.
Rickettsia infection in adult Japanese oysters (Crassostrea gigas) was observed in 2015 at San Ignacio Lagoon, Baja California Sur, Mexico and characterized using molecular tools. In the present study, the degree of infection by Eosinophilic Rickettsia-Like Organism (E-RLO), characterized by intracellular inclusions in gill, mantle, labial palps, digestive tract and gonadal ducts, was evaluated using histological methods and was associated with visible injuries on body surface, such as blisters, shell damage and necrosis. Most of the oysters (92.2%) had some type of observable symptoms; 90.2% of all oysters had blisters in the mantle, and all oysters presented with E-RLO in the epithelia of at least one of the analyzed tissues (gills, labial palps, mantle, gonadal ducts and digestive tract). The highest intensity of infection (percent of E-RLO coverage) was observed in the labial palps (18.6%) and the lowest in digestive tract (2.6% of the examined tissue). Oysters with external injuries had higher infection intensities than the oysters without external injuries. Considering the clinical signs (observable symptoms and histological findings), we propose three infective stages of E-RLO disease development, a scale that could be used for early detection.
Biomarkers for monitoring shrimp health have been proposed but scarcely evaluated at the farm level. We analyzed several indicators of energy status in shrimp under farming conditions concerning ...stocking densities (100 m-2 with biofloc, 15 and 7 m-2). The influence of the year's season (temperature) was also analyzed, and, finally, an unfortunate event of White spot syndrome virus (WSSV) infection occurred on the 7 m-2 farms, adding another condition. At shrimp sampling from ponds, the effect of acute handling stress was also analyzed for indicators typically affected by such procedure with a 3- and 10-fold increase in glucose and lactate levels in hemolymph, respectively, regardless of density. This response was partially blunted at lower temperatures and WSSV incidence. Increased levels of protein in the hepatopancreas, adenylic energy charge (AEC) in both hepatopancreas and muscle and phosphagens in muscle were observed in shrimp from the 100 m-2 farms, suggesting a better nutritional and energetic status in shrimp cultured at high density with biofloc technology. Shrimp with WSSV presented lower hemocyanin levels in hemolymph, most likely associated with its role in the immune response. In WSSV-infected shrimp, the stress response regarding glucose increase was blunted, whereas a stress-induced increase in triglycerides (TG) levels in hemolymph was observed only with WSSV. Increased TG levels in those shrimp hepatopancreas could indicate a switch from carbohydrate to lipid-based metabolism associated with the preferential use of carbohydrates (Warburg effect) for virus replication in the early infection state.
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.
•Shrimp's oxygen consumption responses to cyclic hyperthermia and hypoxia are different between short and long-term exposure.•Under oscillating conditions, HSP gene expression in gills is compromised during hypoxia.•Oxygen consumption and HSP gene expression responses during cyclic conditions are different from those at constant hypoxia.
In marine ectotherms, reproduction is an energetically expensive process that affects their thermal window tolerance. For most species, the impacts of hyperthermia during gametogenesis have still not ...been addressed. Our aim was to assess the metabolic response of adult Nodipecten subnodosus scallops to thermal challenges at early development (spring) and advanced gonad maturation (summer). Scallops collected in both seasons were exposed to acute hyperthermia (26 and 30 °C, 24 h), maintaining a group of scallops at acclimation temperature (22 °C) as a control condition. During the summer, relatively low activity of hexokinase (HK), as well as low levels of ATP and GTP were found in the adductor muscle, suggesting a shift in energy investment for reproduction, although arginine phosphate (ArgP) levels were higher in summer scallops. Hyperthermia (30 °C) induced an increased energy expenditure reflected by a transitory enhanced oxygen consumption (VO
) and relatively high activities of HK and arginine kinase (AK). Moreover, a slight decrease in adenylic energy charge (AEC) was partially compensated by a decrease in ArgP. An increase in nucleotide by-products inosine monophosphate (IMP) and hypoxanthine (HX) indicated a thermal stress at 30 °C. Some of the responses to acute hyperthermia were more pronounced at advanced maturation stages (summer scallops), indicating a possible lack of energy balance, with possible implications in animals challenged to global warming scenario.
The crustacean hyperglycemic hormone (CHH) family is an important group of neuropeptides involved in controlling growth, reproduction, and stress response in decapod species. In this study, a new ...gene containing 4 exons-3 introns flanked by canonical 5′-GT-AG-3′ intron splice-site junctions was isolated from Litopenaeus vannamei. Two full length transcripts of this CHH were isolated from eyestalk and pericardial tissue of males and females using rapid amplification of cDNA ends (RACE). Transcripts sequences were 1578bp in length in males pericardial tissues and in males and females eyestalk with 100% identity, but the transcript isolated from females pericardial tissues was shorter (974bp). The differences in transcripts length is a result of two polyadenylation sites present in the 3'UTR resulting in two transcription termination signals. Transcript sequences encoded one unique protein that can be classified as type I CHH subfamily because of the 4 exons and 3 introns structure, although the CPRP region is not-well conserved and there is no amidation in the C-terminal of the deduced amino acid sequence. Furthermore, there is a glycine inserted in the mature peptide not at position 12 as in type II CHHs but after amino acid 31 and the phylogenetic analysis did not group the peptide within type I, but closer to type II CHHs. We demonstrated by endpoint-PCR, qPCR, and in situ hybridization (ISH), that this gene is expressed in neuroendocrine organs known to express CHHs in penaeid shrimp, including X-organ and optic nerve in eyestalk, supraesophageal ganglion (SoG), but it is also expressed in other organs as gill, gut, pericardial cavity, as well as in terminal ampoule or spermatophore and vas deferens of males.
•We isolated a novel CHH gene from the Pacific White shrimp.•Phylogenetic analysis indicated it is not a strict type I or type II CHH.•Expressed in eyestalk, brain, heart, gills, gut, and males ductus deferens and spermatophore•It might function in osmoregulation, ion exchange and water uptake
Understanding the nature and origins of food sources supporting coastal lagoon-inhabiting organisms is necessary to evaluate the ecological status of such ecosystems. The trophic ecology of a bivalve ...species Spondylus crassisquama was studied in the Ojo de Liebre lagoon (Baja California, Mexico), combining stable isotope (SI), fatty acid (FA) and sterol analyses along a transect under oceanic influences. The second objective of the study was to investigate if sterol compositions give complementary information to those obtained from FA and SI. Temporal and spatial patterns of the three biomarkers in bivalve tissues suggest oceanic inputs at the mouth of the lagoon, while the inner station was characterized by a contribution of local sources including an important role of micro heterotrophs. This study revealed that the association of lipid biomarkers provide higher taxonomic resolution of the spatiotemporal heterogeneity in nutrient sources influencing the trophic functioning of a large coastal lagoon.
•Stable isotopes fatty acids and sterols were used to characterize the diet of bivalves.•A transect was sampled in a coastal lagoon with increasing distance from the mouth.•Bivalves at the entrance of the lagoon relied on oceanic phytoplankton inputs.•Bivalves in the inner lagoon relied on local microbial food web.•Lipid biomarkers and SI were complementary to distinguish the food sources origins.
High temperature increases energy demand in ectotherms, limiting their physiological capability to cope with hypoxic events. The present study aimed to assess the metabolic tolerance of juvenile ...Nodipecten subnodosus scallops to acute hyperthermia combined with moderate hypoxia. A previous study showed that juveniles exhibited a high upper temperature limit (32 °C), but the responses of juveniles to combined hyperthermia and low dissolved oxygen are unknown. Scallops were exposed to control conditions (treatment C: 22 °C, ∼7.1 mg O2 L−1 or PO2 156.9 mmHg), acute hyperthermia under normoxia (treatment T: 30 °C, ∼6.0 mg O2 L−1 or PO2 150.9 mmHg) or acute hyperthermia plus hypoxia (treatment TH: 30 °C, ∼2.5 mg O2 L−1 or PO2 62.5 mmHg) for 18 h. In T, juveniles exhibited an enhanced oxygen consumption, together with a decrease in adenylate energy charge (AEC) and arginine phosphate (ArgP), and with no changes in metabolic enzyme activity in the muscle. In TH, scallops maintained similar AEC and ArgP levels in muscle as those observed in T treatment. This response occurred along with the accumulation of inosine monophosphate and hypoxanthine. Besides, reduced citrate synthase and pyruvate kinase activities, enhanced hexokinase activity, and a higher octopine dehydrogenase/lactate dehydrogenase ratio in the mantle indicated the onset of anaerobiosis in TH. These responses indicate that juvenile scallops showed tissue-specific compensatory responses regarding their energy balance under moderate hypoxia at high temperatures. Our results give an insight into the tolerance limit of this species to combined hyperthermia and hypoxia in its northern limit of distribution.
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•Oxygen consumption increases at high temperatures, a response affected by hypoxia.•Adenylate energy charge decreases, buffered by phosphagen hydrolysis and adenylate kinase.•Tissue-specific responses of metabolic enzyme activity under hyperthermia and hypoxia.
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