Shrimp Hemolymph Extraction

Research on an innate immune system of penaeid shrimp is greatly motivated by economical requirements, because their culture is limited by the development of infectious diseases. As invertebrates, shrimp’s natural immunity acts as a fast and efficient defence mechanism against the pathogens. Their immune system involve hemocytes (for encapsulation, nodule formation and phagocytosis), several plasma components (antimicrobial peptides, histones, lysosomal enzymes, lipopolysaccharide, -1,3-glucan binding proteins, and recognition molecules), and multimeric systems (clotting protein cascade, prophenoloxidase system). When these defense mechanisms fail to protect the shrimp against bacteria, viruses, fungi, protozoa and their products, disease develops and a negative impact takes place in the shrimp culture system. Studying the shrimp immune system is attractive for the advancement of a basic knowledge on invertebrate and vertebrate general immunity, because it offers various possible alternatives for disease management in shrimp aquaculture. The aim of this document is to present the general status of the shrimp defense system, to help in the development of strategies that favour the control and prevention of disease.

2016

Research on an innate immune system of penaeid shrimp is greatly motivated by economical requirements, because their culture is limited by the development of infectious diseases. As invertebrates, shrimp’s natural immunity acts as a fast and efficient defence mechanism against the pathogens. Their immune system involve hemocytes (for encapsulation, nodule formation and phagocytosis), several plasma components (antimicrobial peptides, histones, lysosomal enzymes, lipopolysaccharide, β-1,3-glucan binding proteins, and recognition molecules), and multimeric systems (clotting protein cascade, prophenoloxidase system). When these defense mechanisms fail to protect the shrimp against bacteria, viruses, fungi, protozoa and their products, disease develops and a negative impact takes place in the shrimp culture system. Studying the shrimp immune system is attractive for the advancement of a basic knowledge on invertebrate and vertebrate general immunity, because it offers various possible a...

Fish & Shellfish Immunology, 1995

In order to ensure shrimp aquaculture production, immunology has become a priority in terms of disease control and prevention. Research must be devoted to the identification of anti-infectious effectors at the cellular and molecular levels. In this review, recent results obtained in the shrimp Penaeus japonicus are presented and discussed with reference to the Crustacea and other arthropods. Various cellular and plasma haemolymph components were characterised both antigenically and functionally. Haemocyte subpopulations, separated by isopycnic centrifugation on Percoll were discriminated using two monoclonal antibodies, 40E2 and 40El0. The granular cells were recognised by the antibody 40E2 specific for a plasmatic protein. By comparison, the hyaline and semi-granular cells were recognised by the antibody 40El0, which immunoprecipitated a plasmatic protein identified as an agglutinin. Moreover, an a2-macroglobulin-like protein present both in plasma and in haemocytes was antigenically characterized in P. japonicus as well as a plasma clotting factor. Functional studies have been undertaken by considering antimicrobial effectors and haemocyte-mediated oxidative defence mechanisms studied by chemiluminescence. Prospects on findings in shrimp immunology are discussed with reference to their application to zoosanitary prophylaxis and disease prevention. ~$)

Fish & Shellfish Immunology, 1995

In order to ensure shrimp aquaculture production, immunology has become a priority in terms of disease control and prevention. Research must be devoted to the identification of anti-infectious effectors at the cellular and molecular levels. In this review, recent results obtained in the shrimp Penaeus japonicus are presented and discussed with reference to the Crustacea and other arthropods. Various cellular and plasma haemolymph components were characterised both antigenically and functionally. Haemocyte subpopulations, separated by isopycnic centrifugation on Percoll were discriminated using two monoclonal antibodies, 40E2 and 40E10. The granular cells were recognised by the antibody 40E2 specific for a plasmatic protein. By comparison, the hyaline and semi-granular cells were recognised by the antibody 40E10, which immunoprecipitated a plasmatic protein identified as an agglutinin. Moreover, an α2-macroglobulin-like protein present both in plasma and in haemocytes was antigenically characterized in P. japonicus as well as a plasma clotting factor. Functional studies have been undertaken by considering antimicrobial effectors and haemocyte-mediated oxidative defence mechanisms studied by chemiluminescence. Prospects on findings in shrimp immunology are discussed with reference to their application to zoosanitary prophylaxis and disease prevention.

Fish Pathology, 2003

Morphology and immunological roles of the three distinct types of hemocytes, hyaline, small granular and large granular cells in black tiger shrimp Penaeus monodon were stud ied. The study of functions of these hemocytes in the elimination of injected yeast (Saccharomy ces cerevisiae) or Vibrio harveyi as foreign bodies showed a rapid response against those particles. Together with fixed phagocytes, the blood cells removed the yeast through the process of phagocy tosis, nodule formation and encapsulation, which occurred at almost all parts of the body. The injection of the yeast caused a marked reduction in the blood cell counts in the hemolymph during the first 1 h. V. harveyi was efficiently removed within 3 h after injection. The granular cells (small granular and large granular hemocytes) were the major blood cells that are associated with phenoloxidase activity. The role of hemocytes and fixed phagocytes on defense mechanism in the shrimp were discussed.

Comparative Biochemistry and Physiology Part A: Physiology, 1993

Natural haemolytic activity in brown shrimp (Penaeus californiensis) haemolymph was detected using mouse erythrocytes as target cells. This activity is unrelated to agglutinating and phenoloxidase activity, but it is another probable component of the shrimp defence system. 2. The haemolytic reaction is time and dose dependent, and a serine-protease is involved. 3. The haemolytic factor is thermolabile and has an apparent molecular weight of 23.5 kDa.

Aquaculture, 2000

Journal of Cell Science, 1995

Various haemolymph components of the shrimp Penaeus japonicus were identified and characterised by monoclonal antibodies. Three groups of monoclonal antibodies were raised. Their reactivity to haemocyte types and/or secreted molecules was determined by immunofluorescence and the molecular masses of the antigens were analysed by western-blotting. A 170 kDa protein, in reducing conditions, was recognized by four panhaemocytic monoclonal antibodies from group 1. This protein was present both in the plasma and in the haemocytes from which it appears to be secreted. The shrimp haemocytes were separated by isopycnic centrifugation on a Percoll gradient and the different subpopulations were antigenically analysed using the two monoclonal antibodies, 40E2-2A and 40E10-2B, from group 2. The granular cells were labelled by 40E2-2A which was specific for a protein of 142 kDa also present in plasma. By comparison, the 40E10-2B monoclonal antibody was assumed to be the marker for small hyaline a...

The molecular mechanism of cellular immunity in arthropods has until recently been largely unknown, but with the d~.velopment of a technique to isolate and handle the different blood cell types of crustaceans and with the purification of several proteins associated with the so-called proPO system of freshwater crayfish the processes have now begun to be better understood. In this article Mats ff ohansson and Kenneth Srderh~ill discuss the function of the proPO system in cellular immune reactions in crustaceans and in particular the role of a protein with a molecular mass of 76 kDa, which has been shown to be involved in the communication between the different blood cell types of crayfish.

PLoS ONE, 2011

It has long been viewed that invertebrates rely exclusively upon a wide variety of innate mechanisms for protection from disease and parasite invasion and lack any specific acquired immune mechanisms comparable to those of vertebrates. Recent findings, however, suggest certain invertebrates may be able to mount some form of specific immunity, termed 'specific immune priming', although the mechanism of this is not fully understood (see Textbox S1). In our initial experiments, either formalin-inactivated Vibrio harveyi or sterile saline were injected into the main body cavity (haemocoel) of juvenile shrimp (Litopenaeus vannamei). Haemocytes (blood cells) from V. harveyi-injected shrimp were collected 7 days later and incubated with a 1:1 mix of V. harveyi and an unrelated Gram positive bacterium, Bacillus subtilis. Haemocytes from 'vaccinated' shrimp showed elevated levels of phagocytosis of V. harveyi, but not B. subtilis, compared with those from saline-injected (non-immunised) animals. The increased phagocytic activity was characterised by a significant increase in the percentage of phagocytic cells. When shrimp were injected with B. subtilis rather than vibrio, there was no significant increase in the phagocytic activity of haemocytes from these animals in comparison to the non-immunised (saline injected) controls. Whole haemolymph (blood) from either 'immunised' or non-immunised' shrimp was shown to display innate humoral antibacterial activity against V. harveyi that was absent against B. subtilis. However, there was no difference in the potency of antibacterial activity between V. harveyi-injected shrimp and control (saline injected) animals showing that 'vaccination' has no effect on this component of the shrimp's immune system. These results imply that the cellular immune system of shrimp, particularly phagocytosis, is capable of a degree of specificity and shows the phenomenon of 'immune priming' reported by other workers. However, in agreement with other studies, this phenomenon is not universal to all potential pathogens.