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Fisheries Science

, Volume 85, Issue 1, pp 199–215 | Cite as

Dietary supplementation of Chlorella vulgaris ameliorates chronic sodium arsenite toxicity in Nile tilapia Oreochromis niloticus as revealed by histopathological, biochemical and immune gene expression analysis

  • Eman Zahran
  • Walaa Awadin
  • Engy Risha
  • Asmaa A. Khaled
  • Tiehui Wang
Original Article Aquaculture

Abstract

Arsenic toxicity in an aquatic environment is a major concern, and its elimination has become a global challenge. In the current study, histopathology, serum biomarkers and cytokine gene expression were comparatively examined in fish fed with a control diet or diets containing Chlorella vulgaris (Ch) after exposure to sodium arsenite (NaAsO2) in Nile tilapia (Oreochromis niloticus) with the aim of evaluating the protective role of Ch against arsenite-induced toxicity. Severe histopathological alterations were evident in fish exposed to 7 ppm (parts per million) arsenite for 21 days, compared to unexposed fish. Levels of serum biomarkers ALT, AST, ALP, urea and creatinine were increased, but the levels of Na+, total proteins, albumins and globulins were decreased. Moreover, the expression of all the cytokine genes examined, including IL-1β (7-fold), TNF-α (14-fold) and TGF-β1 (13-fold), were significantly upregulated after arsenite exposure. However, in fish fed with diets containing 5% or 10% Ch, the histopathological alterations in the gills, liver and head kidney were reduced, the biomarkers were stabilized, and the upregulation of cytokine gene expression was lowered, with the high Ch diet (10%) showing more prominent effects. These results suggest the protective and therapeutic roles of Ch as a feed supplement in Nile tilapia against arsenic induced toxicity.

Keywords

Fish Heavy metals Algae Immunity Enzymatic activity Tissue alterations 

Notes

Acknowledgement

This research received no specific grant from any funding agency in the public, commercial, or not-for-profit sectors.

References

  1. Abdel-Tawwab M, Mousa MA, Ahmad MH, Sakr SF (2007) The use of calcium pre-exposure as a protective agent against environmental copper toxicity for juvenile Nile tilapia, Oreochromis niloticus (L.). Aquaculture 264:236–246CrossRefGoogle Scholar
  2. Ahmed MK, Habibullah-Al-Mamun M, Parvin E et al (2013) Arsenic induced toxicity and histopathological changes in gill and liver tissue of freshwater fish, tilapia (Oreochromis mossambicus). Exp Toxicol Pathol 65:903–909CrossRefGoogle Scholar
  3. Aly HA, El-Shitany NA, El-Beshbishy HA, Ashour OM (2015) Ameliorative effect of lycopene against 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin-induced rat liver microsomal toxicity: an in vitro study. Toxicol Ind Health 31:938–950CrossRefGoogle Scholar
  4. Andrews SR, Sahu NP, Pal AK, Kumar S (2009) Haematological modulation and growth of Labeo rohita fingerlings: effect of dietary mannan oligosaccharide, yeast extract, protein hydrolysate and chlorella. Aquac Res 41:61–69CrossRefGoogle Scholar
  5. Aruljothi B (2014) Effect of arsenic on lipid peroxidation and antioxidants system in fresh water fish, labeo rohita. Int J Mod Res Rev 2:15–19Google Scholar
  6. ATSDR (2002) Toxicological profile for arsenic. agency for toxic substances and disease registry. Washington, DC, SUDHHS, PHSGoogle Scholar
  7. Avigliano E, Schenone NF, Volpedo AV, Goessler W, Cirelli AF (2015) Heavy metals and trace elements in muscle of silverside (Odontesthes bonariensis) and water from different environments (Argentina): aquatic pollution and consumption effect approach. Sci Total Environ 506:102–108CrossRefGoogle Scholar
  8. Bancroft D, Stevens A, Turner R (1996) Theory and practice of histological techniques. Churchill Livingstone, New YorkGoogle Scholar
  9. Bengwayan PT, Laygo JC, Pacio AE, Poyaoan JLZ, Rebugio JF, Yuson ALL (2010) A comparative study on the antioxidant property of Chlorella (Chlorella sp.) tablet and glutathione Tablet. Int Sci Res J 2:25–32Google Scholar
  10. Benhamed S, Guardiola FA, Martínez S et al (2016) Exposure of the gilthead seabream (Sparus aurata) to sediments contaminated with heavy metals down-regulates the gene expression of stress biomarkers. Toxicol Rep 3:364–372CrossRefGoogle Scholar
  11. Benli AÇK, Köksal G, Özkul A (2008) Sublethal ammonia exposure of Nile tilapia (Oreochromis niloticus L.): effects on gill, liver and kidney histology. Chemosphere 72:1355–1358.  https://doi.org/10.1016/j.chemosphere.2008.04.037 CrossRefGoogle Scholar
  12. Camargo MM, Martinez CB (2007) Histopathology of gills, kidney and liver of a Neotropical fish caged in an urban stream. Neotrop Ichthyol 5:327–336CrossRefGoogle Scholar
  13. Capkin E, Birincioglu S, Altinok I (2009) Histopathological changes in rainbow trout (Oncorhynchus mykiss) after exposure to sublethal composite nitrogen fertilizers. Ecotoxicol Environ Saf 72:1999–2004CrossRefGoogle Scholar
  14. Cheng D, Wan Z, Zhang X, Li J, Li H, Wang CJN (2017) Dietary Chlorella vulgaris ameliorates altered immunomodulatory functions in cyclophosphamide-induced immunosuppressive mice. Nutrients 9:708CrossRefGoogle Scholar
  15. Cobbina SJ et al (2015) A multivariate assessment of innate immune-related gene expressions due to exposure to low concentration individual and mixtures of four kinds of heavy metals on zebrafish (Danio rerio) embryos. Fish Shellfish Immunol 47:1032–1042CrossRefGoogle Scholar
  16. Correia TG, Narcizo AdM, Bianchini A, Moreira RG (2010) Aluminum as an endocrine disruptor in female Nile tilapia (Oreochromis niloticus). Comp Biochem Physiol C Toxicol Pharmacol 151:461–466CrossRefGoogle Scholar
  17. Crestani M et al (2007) Effect of clomazone herbicide on biochemical and histological aspects of silver catfish (Rhamdia quelen) and recovery pattern. Chemosphere 67:2305–2311CrossRefGoogle Scholar
  18. Datta S, Saha DR, Ghosh D, Majumdar T, Bhattacharya S, Mazumder S (2007) Sub-lethal concentration of arsenic interferes with the proliferation of hepatocytes and induces in vivo apoptosis in (Clarias batrachus) L. Comp Biochem Physiol C Toxicol Pharmacol 145:339–349CrossRefGoogle Scholar
  19. Duker AA, Carranza E, Hale M (2005) Arsenic geochemistry and health. Environ Int 31:631–641CrossRefGoogle Scholar
  20. Elgaml SA, Khalil R, Hashish EA, El-Murr A (2015) Protective effects of selenium and alpha-tocopherol against lead-induced hepatic and renal toxicity in Oreochromis Niloticus. J Aquac Res Dev 6:2Google Scholar
  21. Elia AC et al (2018) A comparative study on subacute toxicity of arsenic trioxide and dimethylarsinic acid on antioxidant status in Crandell Rees feline kidney (CRFK), human hepatocellular carcinoma (PLC/PRF/5), and epithelioma papulosum cyprini (EPC) cell lines. J Toxicol Environ Health A 81:333–348CrossRefGoogle Scholar
  22. Figueiredo-Fernandes A, Ferreira-Cardoso JV, Garcia-Santos S, Monteiro SM, Carrola J, Matos P, Fontaínhas-Fernandes A (2007) Histopathological changes in liver and gill epithelium of Nile tilapia, Oreochromis niloticus, exposed to waterborne copper. Pesquisa Veterinária Brasileira 27:103–109CrossRefGoogle Scholar
  23. Fırat Ö, Cogun HY, Yüzereroğlu TA, Gök G, Fırat Ö, Kargin F, Kötemen Y (2011) A comparative study on the effects of a pesticide (cypermethrin) and two metals (copper, lead) to serum biochemistry of Nile tilapia, Oreochromis niloticus. Fish Physiol Biochem 37:657–666CrossRefGoogle Scholar
  24. Geist J, Werner I, Eder KJ, Leutenegger CM (2007) Comparisons of tissue-specific transcription of stress response genes with whole animal endpoints of adverse effect in striped bass (Morone saxatilis) following treatment with copper and esfenvalerate. Aquat Toxicol 85:28–39CrossRefGoogle Scholar
  25. Ghorbel I, Maktouf S, Fendri N, Jamoussi K, Ellouze Chaabouni S, Boudawara T, Zeghal N (2015) Coexposure to aluminum and acrylamide disturbs expression of metallothionein, proinflammatory cytokines and induces genotoxicity: biochemical and histopathological changes in the kidney of adult rats. Environ Toxicol 31:1044–1058CrossRefGoogle Scholar
  26. Gonzalez HO, Roling JA, Baldwin WS, Bain LJ (2006) Physiological changes and differential gene expression in mummichogs (Fundulus heteroclitus) exposed to arsenic. Aquat Toxicol 77:43–52CrossRefGoogle Scholar
  27. Guerriero G, Avino M, Zhou Q, Fugelstad J, Clergeot PH, Bulone V (2010) Chitin synthases from Saprolegnia are involved in tip growth and represent a potential target for anti-oomycete drugs. PLoS Pathog.  https://doi.org/10.1371/journal.ppat.1001070 Google Scholar
  28. Hamias R, Wolak T, Huleihel M, Paran E, Levy-Ontman OJB (2018) Red alga polysaccharides attenuate angiotensin II-induced inflammation in coronary endothelial cells. J Biochem Biophys Res Commun 500:944–951CrossRefGoogle Scholar
  29. Harvey R, Kubena L, Elissalde M (1994) Influence of vitamin E on aflatoxicosis in growing swine. Am J Vet Res 55:572–577Google Scholar
  30. Hasegawa T et al (1997) Effect of hot water extract of Chlorella vulgaris on cytokine expression patterns in mice with murine acquired immunodeficiency syndrome after infection with Listeria monocytogenes. Immunopharmacology 35:273–282CrossRefGoogle Scholar
  31. Hwang P, Tsai Y (1993) Effects of arsenic on osmoregulation in the tilapia Oreochromis mossambicus reared in seawater. Mar Biol 117:551–558CrossRefGoogle Scholar
  32. Jancsó Z, Hermesz E (2015) Impact of acute arsenic and cadmium exposure on the expression of two haeme oxygenase genes and other antioxidant markers in common carp (Cyprinus carpio). J Appl Toxicol 35:310–318CrossRefGoogle Scholar
  33. Jiang G et al (2003) Effect of arsenic trioxide on cytokine expression by acute promyelocytic leukemia cells. Chin Med J 116:1639–1643Google Scholar
  34. Jin Y, Liu Z, Liu F, Ye Y, Peng T, Fu Z (2015) Embryonic exposure to cadmium (II) and chromium (VI) induce behavioral alterations, oxidative stress and immunotoxicity in zebrafish (Danio rerio). Neurotoxicol Teratol 48:9–17CrossRefGoogle Scholar
  35. Kabilan N et al (2013) The combined effects of mercury chloride and cadmium chloride metals on plasma electrolytes of a fish, Lates Calcarifer. Int J Anal Bioanal Chem 3:183–188Google Scholar
  36. Kamat JP, Boloor KK, Devasagayam TP (2000) Chlorophyllin as an effective antioxidant against membrane damage in vitro and ex vivo. Mol Cell Biol Lipids 1487:113–127CrossRefGoogle Scholar
  37. Kan Y, Cengiz EI, Ugurlu P, Yanar M (2012) The protective role of vitamin E on gill and liver tissue histopathology and micronucleus frequencies in peripheral erythrocytes of Oreochromis niloticus exposed to deltamethrin. Environ Toxicol Pharmacol 34:170–179CrossRefGoogle Scholar
  38. Karan V, Vitorović S, Tutundžić V, Poleksić V (1998) Functional enzymes activity and gill histology of carp after copper sulfate exposure and recovery. Ecotoxicol Environ Saf 40:49–55CrossRefGoogle Scholar
  39. Kavitha C, Malarvizhi A, Senthil Kumaran S, Ramesh M (2010) Toxicological effects of arsenate exposure on hematological, biochemical and liver transaminases activity in an Indian major carp, Catla catla. Food Chem Toxicol 48:2848–2854.  https://doi.org/10.1016/j.fct.2010.07.017 CrossRefGoogle Scholar
  40. Kumar M, Jeon J, Choi J, Kim S-R (2018) Rapid and efficient genetic transformation of the green microalga Chlorella vulgaris. J Appl Phycol 30:1735–1745CrossRefGoogle Scholar
  41. Lakra WS, Nagpure NS (2009) Genotoxicological studies in fishes: a review. Indian J Anim Sci 79:93–97Google Scholar
  42. Lantz RC, Hays AM (2006) Role of oxidative stress in arsenic-induced toxicity. Drug Metab Rev 38:791–804CrossRefGoogle Scholar
  43. Lavanya S, Ramesh M, Kavitha C, Malarvizhi A (2011) Hematological, biochemical and ionoregulatory responses of Indian major carp (Catla catla) during chronic sublethal exposure to inorganic arsenic. Chemosphere 82:977–985CrossRefGoogle Scholar
  44. Liu C, Leung M, Koon J, Zhu L, Hui Y, Yu B, Fung K (2006) Macrophage activation by polysaccharide biological response modifier isolated from Aloe vera L. var. i (Haw.) Berg. Int Immunopharmacol 6:1634–1641CrossRefGoogle Scholar
  45. Martinez C, Cólus I (2002) Biomarcadores em peixes neotropicais para o monitoramento da poluição aquática na bacia do rio Tibagi A bacia do Rio Tibagi Editora dos Editores, Londrina, PR, Brazil: 551–577Google Scholar
  46. Mason R (2001) Chlorella and Spirulina: green supplements for balancing the body. Altern Complement Ther 7:161–165CrossRefGoogle Scholar
  47. Mazon A, Cerqueira C, Fernandes M (2002) Gill cellular changes induced by copper exposure in the South American tropical freshwater fish Prochilodus scrofa. Environ Res 88:52–63CrossRefGoogle Scholar
  48. Mazumder DG (2005) Effect of chronic intake of arsenic-contaminated water on liver. Toxicol Appl Pharmacol 206:169–175CrossRefGoogle Scholar
  49. McCollum CW, Hans C, Shah S, Merchant FA, Gustafsson J-Å, Bondesson M (2014) Embryonic exposure to sodium arsenite perturbs vascular development in zebrafish. Aquat Toxicol 152:152–163CrossRefGoogle Scholar
  50. Moe SM (2008) Disorders involving calcium, phosphorus, and magnesium. Prim Care Clin Office Pract 35:215–237CrossRefGoogle Scholar
  51. Möller A-M, Korytář T, Köllner B, Schmidt-Posthaus H, Segner H (2014) The teleostean liver as an immunological organ: intrahepatic immune cells (IHICs) in healthy and benzo [a] pyrene challenged rainbow trout (Oncorhynchus mykiss). Dev Comp Immunol 46:518–529CrossRefGoogle Scholar
  52. Nayak AS, Lage CR, Kim CH (2007) Effects of low concentrations of arsenic on the innate immune system of the zebrafish (Danio rerio). Toxicol Sci 98:118–124CrossRefGoogle Scholar
  53. Özkan-Yılmaz F, Özlüer-Hunt A, Gündüz SG et al (2014) Effects of dietary selenium of organic form against lead toxicity on the antioxidant system in Cyprinus carpio. Fish Physiol Biochem 40:355–363CrossRefGoogle Scholar
  54. Ozmen M, Güngördü A, Kucukbay FZ, Güler RE (2006) Monitoring the effects of water pollution on Cyprinus carpio in Karakaya Dam Lake, Turkey. Ecotoxicology 15:157–169CrossRefGoogle Scholar
  55. Pacheco M, Santos M (2002) Naphthalene and β-naphthoflavone effects on (Anguilla anguilla) L. hepatic metabolism and erythrocytic nuclear abnormalities. Environ Int 28:285–293CrossRefGoogle Scholar
  56. Palaniappan PR, Vijayasundaram V (2009) The effect of arsenic exposure and the efficacy of DMSA on the proteins and lipids of the gill tissues of Labeo rohita. Food Chem Toxicol 47:1752–1759CrossRefGoogle Scholar
  57. Palipoch S, Jiraungkoorskul W, Tansatit T, Preyavichyapugdee N, Jaikua W, Kosai P (2011) Protective efficiency of Thunbergia laurifolia leaf extract against lead (II) nitrate-induced toxicity in Oreochromis niloticus. J Med Plants Res 5:719–728Google Scholar
  58. Patel M, Rogers JT, Pane EF, Wood CM (2006) Renal responses to acute lead waterborne exposure in the freshwater rainbow trout (Oncorhynchus mykiss). Aquat Toxicol 80:362–371CrossRefGoogle Scholar
  59. Prieto-Álamo M-J, Abril N, Osuna-Jiménez I, Pueyo C (2009) Solea senegalensis genes responding to lipopolysaccharide and copper sulphate challenges: large-scale identification by suppression subtractive hybridization and absolute quantification of transcriptional profiles by real-time RT-PCR. Aquat Toxicol 91:312–319CrossRefGoogle Scholar
  60. Pugazhendy K (2012) Protective role of spirulina on the variation of haematological parameter induced by herbicide atrazine in the fresh water fish Cyprinus carpio (Linn). Int J Pharm Biol Arch 3:249–254Google Scholar
  61. Puntoriero ML, Cirelli AF, Volpedo AV (2018) Histopathological changes in liver and gills of Odontesthes bonariensis inhabiting a lake with high concentrations of arsenic and fluoride (Chasicó Lake, Buenos Aires province). J Revista Internacional de Contaminación Ambiental 34:69–77CrossRefGoogle Scholar
  62. Queiroz ML, Rodrigues AP, Bincoletto C, Figueirêdo CA, Malacrida S (2003) Protective effects of Chlorella vulgaris in lead-exposed mice infected with Listeria monocytogenes. Int Immunopharmacol 3:889–900CrossRefGoogle Scholar
  63. Reyes-Becerril M, Guardiola F, Rojas M, Ascencio-Valle F, Esteban MÁ (2013) Dietary administration of microalgae Navicula sp. affects immune status and gene expression of gilthead seabream (Sparus aurata). Fish Shellfish Immunol 35:883–889CrossRefGoogle Scholar
  64. Saïdi SA, Azaza MS, Windmolders P et al (2013) Cytotoxicity evaluation and antioxidant enzyme expression related to heavy metals found in tuna by-products meal: an in vitro study in human and rat liver cell lines. Exp Toxicol Pathol 65:1025–1033CrossRefGoogle Scholar
  65. Samuel S, Kathirvel R, Jayavelu T, Chinnakkannu P (2005) Protein oxidative damage in arsenic induced rat brain: influence of DL-α-lipoic acid. Toxicol Lett 155:27–34CrossRefGoogle Scholar
  66. Sharma KP, Upreti N, Sharma S, Sharma S (2012) Protective effect of Spirulina and tamarind fruit pulp diet supplement in fish (Gambusia affinis Baird & Girard) exposed to sublethal concentration of fluoride, aluminum and aluminum fluoride. Ind J Exp Biol 50:897–903Google Scholar
  67. Silva AG, Martinez CB (2007) Morphological changes in the kidney of a fish living in an urban stream. Environ Toxicol Pharmacol 23:185–192CrossRefGoogle Scholar
  68. Simonato JD, Guedes CL, Martinez CB (2008) Biochemical, physiological, and histological changes in the neotropical fish Prochilodus lineatus exposed to diesel oil. Ecotoxicol Environ Saf 69:112–120CrossRefGoogle Scholar
  69. Sirakov I, Velichkova K, Stoyanova S, Staykov Y (2015) The importance of microalgae for aquaculture industry. Review. Int J Fish Aquatic Stud 2:81–84Google Scholar
  70. Soetaert A, Vandenbrouck T, van der Ven K, Maras M, van Remortel P, Blust R, De Coen WM (2007) Molecular responses during cadmium-induced stress in Daphnia magna: integration of differential gene expression with higher-level effects. Aquat Toxicol 83:212–222CrossRefGoogle Scholar
  71. Soontornchaiboon W, Joo SS, Kim SM (2012) Anti-inflammatory effects of violaxanthin isolated from microalga Chlorella ellipsoidea in RAW 264.7 macrophages. Biol Pharm Bull 35:1137–1144CrossRefGoogle Scholar
  72. Suhendrayatna Ohki A, Kuroiwa T, Maeda S (1999) Arsenic compounds in the freshwater green microalga Chlorella vulgaris after exposure to arsenite. Appl Organomet Chem 13:127–133CrossRefGoogle Scholar
  73. Suvetha L, Ramesh M, Saravanan M (2010) Influence of cypermethrin toxicity on ionic regulation and gill Na+/K+-ATPase activity of a freshwater teleost fish Cyprinus carpio. Environ Toxicol Pharmacol 29:44–49.  https://doi.org/10.1016/j.etap.2009.09.005 CrossRefGoogle Scholar
  74. Suzuki T, Moribe M, Okabe Y, Niinae M (2013) A mechanistic study of arsenate removal from artificially contaminated clay soils by electrokinetic remediation. J Hazard Mater 254:310–317CrossRefGoogle Scholar
  75. Teles M, Mackenzie S, Boltana S, Callol A, Tort L (2011) Gene expression and TNF-alpha secretion profile in rainbow trout macrophages following exposures to copper and bacterial lipopolysaccharide. Fish Shellfish Immunol 30:340–346CrossRefGoogle Scholar
  76. Thévenod F (2009) Cadmium and cellular signaling cascades: to be or not to be? Toxicol Appl Pharmacol 238:221–239CrossRefGoogle Scholar
  77. Torres-Perez J, Gerente C, Andres Y (2012) Conversion of agricultural residues into activated carbons for water purification: application to arsenate removal. J Environ Sci Health A 47:1173–1185CrossRefGoogle Scholar
  78. Tripathi S, Kumar A (2011) Effect of acute and chronic exposure of sodium arsenite (Na3AsO3) on total protein, albumin, and globulin in serum of Oryctolagus cuniculus L. Toxicol Environ Chem 93:307–313CrossRefGoogle Scholar
  79. Upasani C, Balaraman R (2003) Protective effect of Spirulina on lead induced deleterious changes in the lipid peroxidation and endogenous antioxidants in rats. Phytother Res 17:330–334CrossRefGoogle Scholar
  80. Vaglio A, Landriscina C (1999) Changes in liver enzyme activity in the teleost Sparus auratain response to cadmium intoxication. Ecotoxicol Environ Saf 43:111–116CrossRefGoogle Scholar
  81. van Heerden D, Vosloo A, Nikinmaa M (2004) Effects of short-term copper exposure on gill structure, metallothionein and hypoxia-inducible factor-1α (HIF-1α) levels in rainbow trout (Oncorhynchus mykiss). Aquat Toxicol 69:271–280CrossRefGoogle Scholar
  82. Vetrivel C, Pugazhendy K, Prabakaran S (2014) Protective effect of spirulina against the lead acetate induced ALP and ACP activity in the liver tissue of fresh water fish, Labeo rohita. Int J Modn Res Revs 2:226–228Google Scholar
  83. Wang X et al (2012) Arsenic and chromium in drinking water promote tumorigenesis in a mouse colitis-associated colorectal cancer model and the potential mechanism is ROS-mediated Wnt/β-catenin signaling pathway. Toxicol Appl Pharmacol 262:11–21CrossRefGoogle Scholar
  84. Wangkahart E, Scott C, Secombes CJ, Wang T (2016) Re-examination of the rainbow trout (Oncorhynchus mykiss) immune response to flagellin: Yersinia ruckeri flagellin is a potent activator of acute phase proteins, anti-microbial peptides and pro-inflammatory cytokines in vitro. Dev Comp Immunol 57:75–87CrossRefGoogle Scholar
  85. Webster TMU, Williams TD, Katsiadaki I et al (2017) Hepatic transcriptional responses to copper in the three-spined stickleback are affected by their pollution exposure history. Aquat Toxicol 184:26–36CrossRefGoogle Scholar
  86. Yu H-S, Liao W-T, Chang K-L, Yu C-L, Chen G-S (2002) Arsenic induces tumor necrosis factor α; Release and tumor necrosis factor receptor 1 signaling in T helper cell apoptosis. J Invest Dermatol 119:812–819CrossRefGoogle Scholar
  87. Zahran E, Risha E (2014) Modulatory role of dietary Chlorella vulgaris powder against arsenic-induced immunotoxicity and oxidative stress in Nile tilapia (Oreochromis niloticus). Fish Shellfish Immunol 41:654–662CrossRefGoogle Scholar

Copyright information

© Japanese Society of Fisheries Science 2018

Authors and Affiliations

  1. 1.Department of Internal Medicine, Infectious and Fish Diseases, Faculty of Veterinary MedicineMansoura UniversityMansouraEgypt
  2. 2.Department of Pathology, Faculty of Veterinary MedicineMansoura UniversityMansouraEgypt
  3. 3.Department of Clinical Pathology, Faculty of Veterinary MedicineMansoura UniversityMansouraEgypt
  4. 4.Animal and Fish Production Department, Faculty of Agriculture Saba BashaAlexandria UniversityAlexandriaEgypt
  5. 5.Scottish Fish Immunology Research Centre, School of Biological SciencesUniversity of AberdeenAberdeenUK

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