Fish &amp Shellfish Immunology (FISH SHELLFISH IMMUN)

Publications in this journal

• Article: Crustin, a WAP domain containing antimicrobial peptide from freshwater prawn Macrobrachium rosenbergii: Immune characterization

  Jesu Q, Arockiaraj, Annie J Gnanam, Dhanaraj Muthukrishnan, Ranganath Gudimella, James Milton, Arun Singh, Saravanan Muthupandian, Marimuthu Kasi, Subha Bhassu
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  ABSTRACT: a b s t r a c t Crustin (MrCrs) was sequenced from a freshwater prawn Macrobrachium rosenbergii. The MrCrs protein contains a signal peptide region at N-terminus between 1 and 22 and a long whey acidic protein domain (WAP domain) at C-terminus between 57 and 110 along with a WAP-type 'four-disulfide core' motif. Phylogenetic results show that MrCrs is clustered together with other crustacean crustin groups. MrCrs showed high sequence similarity (77%) with crustin from Pacific white shrimp Litopenaeus vannamei and Japanese spiny lobster Panulirus japonicas. I-TASSER uses the best structure templates to predict the possible structures of MrCrs along with PDB IDs such as 2RELA and 1FLEI. The gene expressions of MrCrs in both healthy M. rosenbergii and those infected with virus including infectious hypodermal and hematopoietic necrosis virus (IHHNV) and white spot syndrome virus (WSSV) and bacteria Aeromonas hydrophila (Gram-negative) and Enterococcus faecium (Gram-positive) were examined using quantitative real time PCR. To understand its biological activity, the recombinant MrCrs gene was constructed and expressed in Escherichia coli BL21 (DE3). The recombinant MrCrs protein agglutinated with the bacteria considered for analysis at a concentration of 25 mg/ml, except Lactococcus lactis. The bactericidal results showed that the recombinant MrCrs protein destroyed all the bacteria after incubation, even less than 6 h. These results suggest that MrCrs is a potential antimicrobial peptide, which is involved in the defense system of M. rosenbergii against viral and bacterial infections. Q1 Ó 2012 Published by Elsevier Ltd.
  Fish &amp Shellfish Immunology 01/2013; 34(1):109-18.
• Article: The influence of dietary beta-glucan, PAMP exposure and Aeromonas salmonicida on apoptosis modulation in common carp (Cyprinus carpio)

  Joanna Miest, Alberto Falco, Nicolas Pionnier, Patrick Frost, Ilgiz Irnazarow, Gwyn Williams, David Hoole
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  ABSTRACT: The association between b-glucan (MacroGard) supplemented feed and apoptosis in immune-related organs of common carp (Cyprinus carpio) was studied using fluorescence microscopy and real-time PCR. In addition the effect of Aeromonas salmonicida, LPS and Poly(I:C) injections on this relationship was evaluated. Whilst acridine orange staining revealed that apoptosis levels were independent of MacroGard and LPS/Poly(I:C) administration or their combination, it was shown that injection with A. salmonicida increased the percentage of apoptotic cells irrespective of the feeding regime. It was apparent that in all the treatments gene expression profiles displayed organ and time dependency. For example no effect was observed at 7 days of MacroGard administration while 25 days of feeding led to increased iNOS expression and differential up-regulation of anti- or pro-apoptotic genes depending on organ. This may indicate differences in NO sensitivity. MacroGard also led to an elevation of pro- as well as anti-apoptotic genes in LPS or Poly(I:C) injected fish, while LPS/Poly(I:C) alone had little effect. A. salmonicida caused enhanced iNOS expression and it is possible that the type of apoptosis pathway induced is organ dependent as Caspase 9 is induced in mid-gut but not in pronephros. These results indicate that MacroGard feeding alone or in combination with other pathogenic factors did not induce significant apoptosis in immune organs.
  Fish &amp Shellfish Immunology 01/2012; 33:846 - 856.
• Article: Isolation and characterization of major histocompatibility class IIβ genes in an endangered North American cyprinid fish, the Rio Grande silvery minnow (Hybognathus amarus).

  Megan J Osborne, Thomas F Turner
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  ABSTRACT: The major histocompatibility complex (MHC) is a critical component of the adaptive immune response in vertebrates. Due to the role that MHC plays in immunity, absence of variation within these genes may cause species to be vulnerable to emerging diseases. The freshwater fish family Cyprinidae comprises the most diverse and species-rich group of freshwater fish in the world, but some are imperiled. Despite considerable species richness and the long evolutionary history of the family, there are very few reports of MHC sequences (apart from a few model species), and no sequences are reported from endemic North American cyprinids (subfamily Leuciscinae). Here we isolate and characterize the MH Class II beta genes from complementary DNA and genomic DNA of the non-model, endangered Rio Grande silvery minnow (Hybognathus amarus), a North American cyprinid. Phylogenetic reconstruction revealed two groups of divergent MH alleles that are paralogous to previously described loci found in deeply divergent cyprinid taxa including common carp, zebrafish, African large barb and bream. Both groups of alleles were under the influence of diversifying selection yet not all individuals had alleles belonging to both allelic groups. We concluded that the general organization and pattern of variation of MH class II genes in Rio Grande silvery minnow is similar to that identified in other cyprinid fishes studied to date, despite distant evolutionary relationships and evidence of a severe genetic bottleneck.
  Fish &amp Shellfish Immunology 04/2011; 30(6):1275-82.
• Article: Exposure to lipopolysaccharide induces immune genes in cultured preadipocytes of Atlantic salmon.

  Skugor S, Skugor A, Todorcević M, Torgersen J, Ruyter B, Krasnov A
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  ABSTRACT: In addition to its central role of energy storage and release, white adipose tissue (WAT) performs complex endocrine and immune activities. WAT produces physiologically active secretory proteins, including cytokines and complement factors. Furthermore, treatment of mammalian adipocytes with cytokines and inflammatory stimulators induces immune genes, suppresses regulators of adipocyte differentiation and activates lipolysis. Previously we reported up-regulation of immune genes in the course of in vitro development of Atlantic salmon white adipocytes. If WAT is immunoactive tissue in fish, excessive deposition of fat resulting from lipid-rich diets may imply risk for health of farmed fish. In this paper we investigated how lipopolysaccharide (LPS) affects immune activity in the adipose tissue-derived stromo-vascular fraction (aSVF) of Atlantic salmon. Experiments were performed with confluent cultures of proliferating preadipocytes. Exposure to LPS induced expression of immune genes, including TNFalpha and TNF-dependent genes, chemokines and receptors, NFkappaB related genes, matrix metalloproteinases and genes involved in eicosanoid metabolism. LPS decreased expression of adipocyte markers and genes involved in lipid metabolism, however, in parallel, it accelerated a number of transcriptional events that take place during the adipogenic differentiation of aSVF.
  Fish &amp Shellfish Immunology 11/2010; 29(5):817-824.
• Article: In situ localisation of major histocompatibility complex class I and class II and CD8 positive cells in infectious salmon anaemia virus (ISAV)-infected Atlantic salmon

  Dyveke Lem Hetland, Sven Martin Jørgensen, Karsten Skjødt, Ole Bendik Dale, Knut Falk, Cheng Xu, Aase B. Mikalsen, Unni Grimholt, Tor Gjøen, Charles McL. Press
  Fish &amp Shellfish Immunology 01/2010;
• Article: In situ localisation of major histocompatibility complex class I and class II and CD8 positive cells in infectious salmon anaemia virus (ISAV)-infected Atlantic salmon

  Hetland DL, Jørgensen SM, Skjødt K, Dale OB, Falk K, Xu C, Mikalsen AB, Grimholt U, Gjøen T, Press CM
  Fish &amp Shellfish Immunology 01/2010;
• Article: In situ localisation of major histocompatibility complex class I and class II and CD8 positive cells in infectious salmon anaemia virus (ISAV)-infected Atlantic salmon

  Dyveke Lem Hetland a, Sven Martin Jørgensen b, Karsten Skjødt c, Ole Bendik Dale d, Knut Falk d, Cheng Xu a, Aase B. Mikalsen a, Unni Grimholt e, Tor Gjøen f, Charles McL. Press a
  Fish &amp Shellfish Immunology 01/2010;
• Article: Decreased expression of TGF-beta, GILT and T-cell markers in the early stages of soybean enteropathy in Atlantic salmon (Salmo salar L.)

  Lilleeng E, Penn MH, Haugland O, Xu C, Bakke AM, Krogdahl A, Landsverk T, Frøystad-Saugen MK
  Fish &amp Shellfish Immunology 07/2009;
• Article: Decreased expression of TGF-b, GILT and T-cell markers in the early stages of soybean enteropathy in Atlantic salmon (Salmo salar L.)

  Einar Lilleeng, Michael H. Penn, Øyvind Haugland, Cheng Xu, Anne Marie Bakke, Åshild Krogdahl, Thor Landsverk, Marianne K. Frøystad-Saugen
  Fish &amp Shellfish Immunology 05/2009;
• Article: In vitro investigations for the QSAR mechanism of lymphocytes apoptosis induced by substituted aromatic toxicants.

  Hangjun Zhang, Jianying Zhang, Yinmei Zhu
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  ABSTRACT: The objective of this study was to assess the chemicals-induced apoptosis effects on fish lymphocytes and to use the QSAR model to interpret the relationships between apoptotic effects and chemical structures to identify the immune toxicology mechanism. By the use of a simple in vitro toxicological assay, the measured apoptotic parameter (EC50) is used in a QSAR to interpret the apoptotic effects of 25 substituted benzenes at low exposure levels. The apoptotic effects of all tested substituted aromatic chemicals with Carassius auratus lymphocytes were confirmed by DNA ladder and nucleus condensation. For both chlorobenzenes and PCBs, the EC50 values increase with increasing Cl number in the molecule, a result reflecting probably the increased p-pi conjugation of the C-Cl bonds that lowers the molecular reactivity. Furthermore, the apoptotic EC50 data were best correlated with the dipole moment (mu) and the energy of the lowest unoccupied molecular orbital (ELUMO) such that: log(1/EC50)=0.325+0.222 micro-0.163(ELUMO) (with R(2)=0.879). The dependence on the electronic ELUMO factor of the established correlation suggests that during the apoptotic process the ROS (reactive oxygen substance) produced by cells acts as a Lewis base in substituted nucleophilic reactions with toxic chemicals behaving as an electron acceptor. On the basis of the test results, the present toxicological assay offers a rapid tool for assessing the toxic effects of chemicals at low exposure levels.
  Fish &amp Shellfish Immunology 01/2009; 25(6):710-7.
• Article: Development of monoclonal antibodies to rohu [Labeo rohita] immunoglobulins for use in immunoassays.

  Gaurav Rathore, Gokhlesh Kumar, Neeraj Sood, D Kapoor, W S Lakra
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  ABSTRACT: Serum immunoglobulins [Ig] of rohu [Labeo rohita] were purified by affinity chromatography using bovine serum albumin as capture ligand. The purified rohu Ig [r-Ig] had a molecular weight [MW] of 880 kDa as determined with gel filtration chromatography. The heavy chain of r-Ig had an MW of 77.8 kDa and that of light chain was 26.4 kDa in SDS-PAGE. Purified r-Ig was used for the production of two anti-rohu Ig monoclonal antibodies [D7 and H4] that belonged to subclass IgG2b and IgG1, respectively. Both the MAbs were specific to heavy chain of r-Ig as seen in Western blotting. Anti-rohu Ig MAb was used as a diagnostic reagent in ELISA and immunocytochemical assays to demonstrate its application for sero-surveillance and for immunological studies in rohu. A competitive ELISA was used to demonstrate the antigenic relatedness of r-Ig with whole serum Ig of other fish species. Cross reactivity of anti-rohu Ig MAb was observed with serum Ig of Catla catla and Cirrihinus mrigala. No reactivity to serum Ig of Ophiocephalus striatus and Clarias gariepinus was seen. Anti-rohu Ig MAb was found to be suitable for the detection of pathogen specific [Edwardsiella tarda] antibodies in serum of immunized rohu by an indirect ELISA. In flow cytometry using D7 MAb, the mean percentage [+/-SE] of Ig positive cells in spleen and blood of rohu were found to be 64.85% [+/-2.34] and 51.84% [+/-2.55] of gated lymphocytes, respectively. Similarly, D7 MAb also stained 52.84% [+/-1.30] and 10.5% of gated lymphocytes in kidney and thymus, respectively. The anti-rohu Ig MAbs also showed specific staining of Ig bearing cells in spleen sections by the indirect immunoperoxidase test.
  Fish &amp Shellfish Immunology 01/2009; 25(6):761-74.
• Article: Isolation of an Atlantic salmon (Salmo salar) signal transducer and activator of transcription STAT1 gene: kinetics of expression upon ISAV or IPNV infection.

  Bertrand Collet, Nicola Bain, Sylviane Prevost, Guillaume Besinque, Alastair McBeath, Mike Snow, Catherine Collins
  Fish &amp Shellfish Immunology 01/2009; 25(6):861-7.
• Article: Oral delivery of live yeast Debaryomyces hansenii modulates the main innate immune parameters and the expression of immune-relevant genes in the gilthead seabream (Sparus aurata L.).

  Martha Reyes-Becerril, Irene Salinas, Alberto Cuesta, José Meseguer, Dariel Tovar-Ramirez, Felipe Ascencio-Valle, Maria Angeles Esteban
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  ABSTRACT: Microorganisms isolated from fish can be used as prophylactic tools for aquaculture in the form of probiotic preparations. The purpose of this study was to evaluate the effects of dietary administration of the live yeast Debaryomyces hansenii CBS 8339 on the gilthead seabream (Sparus aurata L.) innate immune responses. Seabream were fed control or D. hansenii-supplemented diets (10(6) colony forming units, CFU g(-1)) for 4 weeks. Humoral (seric alternative complement and peroxidase activities), and cellular (peroxidase, phagocytic, respiratory burst and cytotoxic activities) innate immune parameters and antioxidant enzymes (superoxide dismutase (SOD) and catalase (CAT)) were measured from serum, head-kidney leucocytes and liver, respectively, after 2 and 4 weeks of feeding. Expression levels of immune-associated genes, Hep, IgM, TCR-beta, NCCRP-1, MHC-II alpha, CSF-1R, C3, TNF-alpha and IL-1 beta, were also evaluated by real-time PCR in head-kidney, liver and intestine. Humoral immune parameters were not significantly affected by the dietary supplementation of yeast at any time of the experiment. On the other hand, D. hansenii administration significantly enhanced leucocyte peroxidase and respiratory burst activity at week 4. Phagocytic and cytotoxic activities had significantly increased by week 2 of feeding yeast but unchanged by week 4. A significant increase in liver SOD activity was observed at week 2 of feeding with the supplemented diet; however CAT activity was not affected by the dietary yeast supplement at any time of the experiment. Finally, the yeast supplemented diet down-regulated the expression of most seabream genes, except C3, in liver and intestine and up-regulated all of them in the head-kidney. These results strongly support the idea that live yeast Debaryomyces hansenii strain CBS 8339 can stimulate the innate immune parameters in seabream, especially at cellular level.
  Fish &amp Shellfish Immunology 01/2009; 25(6):731-9.
• Article: Mj-DWD, a double WAP domain-containing protein with antiviral relevance in Marsupenaeus japonicus.

  Dandan Chen, Nanhai He, Xun Xu
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  ABSTRACT: The Mj-DWD (Marsupenaeus japonicus' double-WAP domains) gene was originally found up-regulated in virus-resistant shrimp M. japonicus by suppression subtractive hybridization (SSH). The full-length cDNA of Mj-DWD encodes a novel protein containing a KGD (Lys-Gly-Asp) motif and double WAP domains. Performed by quantitative real-time PCR, the expression level of Mj-DWD gene was consistently maintained at a high level in the newly prepared virus-resistant shrimp compared to the normal one. In addition, the Mj-DWD gene was also found to be rapidly up-regulated by WSSV infection during the early phase. Furthermore, the recombinant Mj-DWD, expressed by Pichia pastoris, showed specific protease inhibitory activity on Bacillus subtilis. These findings suggest that Mj-DWD plays an important role in the host defence system against WSSV infection in M. japonicus, possibly through its protease inhibitory activity.
  Fish &amp Shellfish Immunology 01/2009; 25(6):775-81.
• Article: The immune response of white shrimp Litopenaeus vannamei and its susceptibility to Vibrio alginolyticus under low and high pH stress.

  Chang-Che Li, Jiann-Chu Chen
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  ABSTRACT: White shrimp Litopenaeus vannamei (also known as Penaeus vannamei) held in 34 per thousand seawater at pH 8.2 were injected with tryptic soy broth (TSB)-grown Vibrio alginolyticus at 8 x 10(5) colony-forming units (cfu) shrimp(-1), and then transferred to tanks at pH 6.5, 8.2 (control) and 10.1, respectively. After 24-168 h, the mortality of V. alginolyticus-injected shrimp that were transferred to pH 6.5 and pH 10.1 tanks was significantly higher than that of V. alginolyticus-injected shrimp held at pH 8.2. In another experiment, L. vannamei held at pH 8.2 following transfer to pH 6.5, 8.2 (control) and 10.1 for 6, 12, 24, 72 and 120 h were examined for immune parameters, phagocytic activity, and the clearance efficiency of shrimp against V. alginolyticus. The results indicated that the shrimp that were transferred to pH 6.5 and 10.1 showed significantly decreased phenoloxidase (PO) activity, respiratory burst, phagocytic activity, and clearance efficiency against V. alginolyticus over 6-72 h; significantly decreased superoxide dismutase (SOD) activity over 6-24h; and decreased total haemocyte count (THC) over 12-72 h. Shrimp transferred to pH 10.1 showed significantly decreased granular cell counts, and THC after 6h, and decreased SOD activity after 72 h. The immune parameters of shrimp transferred to pH 6.5 and 10.1 returned to the original values after 120 h. However, shrimp transferred to pH 6.5 still maintained lower phagocytic activity, and clearance efficiency against V. alginolyticus, and shrimp transferred to pH 10.1 still maintained lower clearance efficiency against V. alginolyticus. It was therefore concluded that low pH and high pH stress decrease the resistance of white shrimp L. vannamei against V. alginolyticus and decrease several parameters of the immune response.
  Fish &amp Shellfish Immunology 01/2009; 25(6):701-9.
• Article: Soybean meal induces intestinal inflammation in common carp (Cyprinus carpio L.).

  P A Urán, A A Gonçalves, J J Taverne-Thiele, J W Schrama, J A J Verreth, J H W M Rombout
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  ABSTRACT: The development of soybean meal (SBM) induced enteritis in the hindgut of the omnivorous common carp (Cyprinus carpio L.). The developed condition was assessed when carp, continuously fed on animal protein, were transferred to a diet in which 20% of the protein was replaced by SBM. After week 1, most of the inflammation parameters were already present, but at week 3, a strong aggravation of the condition was observed which included a shortening of the mucosal folds, the disappearance of the supranuclear vacuoles, an increased number of goblet cells, a thickened lamina propria and sub-epithelial mucosa with increased numbers of basophilic granulocytes as well as a decreased uptake capacity of enterocytes (impaired endocytosis and microvilli). Contrary to previous observations made with respect to Atlantic salmon, common carp start to recover from the fourth to the fifth week after switching to SBM feeding. At this stage, the supranuclear vacuoles refill and most of the parameters revert to basal levels. During the enteritis process, a real-time quantitative PCR analysis was conducted to measure the expression of inflammatory and anti-inflammatory cytokine genes in the isolated intraepithelial lymphocytes (IEL). The pro-inflammatory interleukin 1 beta (IL-1 beta) and tumour necrosis factor alpha1 (TNF-alpha1) genes were up-regulated during the inflammation process while the anti-inflammatory interleukin 10 (IL-10) was down-regulated after an initial up-regulation at week 1. Transforming growth factor beta (TGF-beta) expression showed an up-regulation from week 3 onwards despite the high Ct value and the low primer efficiency shown. This study confirms the contribution of IEL (mainly T-like cells) and basophils in the enteritis process. In addition, the results show a clear involvement of up- and down-regulated cytokine genes in both the onset and recovery of the SBM-induced enteritis in the hindgut of carp.
  Fish &amp Shellfish Immunology 01/2009; 25(6):751-60.