Immune modulation by fish kinetoplastid parasites : a role for nitric oxide

Trypanoplasma borreli and Trypanosoma carassii are kinetoplastid parasites infecting cyprinid fish. We investigated the role of nitric oxide (NO) in immune modulation during T. borreli and T. carassii infection of carp. Phagocytic cells from different organs produced NO and serum nitrate levels increased, demonstrating that T. borreli activates NO production in vivo. In contrast, T. carassii did not induce NO production in vivo and inhibited LPS-induced NO production in vitro. Production of NO was detrimental to the host as T. borreli-infected carp treated with the inducible NO synthase inhibitor aminoguanidine had a higher survival than infected control carp. This detrimental effect can be explained (in part) by the toxicity of NO to cells in vitro as NO inhibited the proliferative response of blood and spleen leukocytes. Head-kidney phagocytes were resistant to the immunosuppressive effects of NO in vitro. The NO-inducing activity of T. borreli may be an adaptation developed to ensure survival and immune evasion in the fish host. Apparently, T. carassii has adopted another strategy by deactivating specific functions of phagocytes. Both strategies may ensure long-term survival of the parasite

Polymorphism of major histocompatibility complex class II B genes in different carp lines of the common carp (Cyprinus carpio L.)

Regular observation of survival of the carp breeding lines constituting a living gene bank at the Institute of Ichthyobiology and Aquaculture in Golysz (Poland) over a period of at least 15 years showed different survival rates for various lines. In this study, we have examined the polymorphism of the major histocompatibility complex (MHC) gene class II B in nine carp lines. The class II B gene encodes for the part of the MHC class II molecule which presents peptides from pathogens and protein antigens that are present in the extracellular milieu and have been taken up into the endocytic vesicles of antigen-presenting cells. Polymerase chain reaction was used to amplify Cyca-DAB gene fragments comprising part of exon 1, complete intron 1 and almost complete exon 2. Exon 2 encodes for the beta(1) domain which is the most polymorphic fragment of MHC class II molecules. Single-strand conformational polymorphism (SSCP) was applied to detect different MHC class II B haplotypes. The analysis revealed the presence of seven different haplotypes occurring with various frequencies. (C) 2003 Editions scientifiques et medicales Elsevier SAS and Ifremer/IRD/Inra/Cemagref. All rights reserved

Immune stimulation in fish and chicken through weak low frequency electromagnetic fields

A hypothesis is proposed how Low Frequency Electromagnetic Field (LF EMF) exposure can stimulate an immune response, based on recent insights in immunology. We hypothesize that the Immunent EMF treatment induces mild stress to cells, which then produce cytokines that function as alarms or so called danger signals for the immune system. In this way EMF treatment takes the place of multiplying pathogens, and the damage these cause, in the triggering of an immune response. In a first series of experiments in¿vitro common carp head kidney-derived phagocytes were used to determine ROS production as a measure for immune activation. Exposure to LF EMF signals (200¿5,000 Hz) at 5 ¿T or 1.5 mT led to 42 or 33% increase in immune activity, respectively, compared to negative control values. EMF could also additionally stimulate chemically pre-stimulated samples up to 18% (5 ¿T) or 22% (1.5 mT). Significance of increase in ROS production in the total series was: p <0.0001. In a second series of experiments in¿vitro commercial goldfish were used. Groups of fish were housed under equal conditions in at least four control tanks and 8¿16 EMF-exposed tanks. Exposure was done with a predominantly vertical field at field strengths (rms) between 0.15 and 50 ¿T. Without treatment mortality was about 50% after 18 days, while the treatment at 5 ¿T reduced it to 20% on average. At field strengths 0.15, 0.5, 1.5, 5, 15 and 50 ¿T an equally strong effect was found. Reducing the field strength to 0.05, 0.06, 0.01 and 0.003 ¿T showed a gradually decreasing effect, which only at 0.003 ¿T is no longer statistically significant. Finally, in¿vitro experiments were done with 560 commercial broiler chickens exposed to infection pressure from coccidiosis. EMF exposure at 6.5 ¿T reduced intestinal lesions by 40% and improved feed conversion by 8%

Transcriptome sequencing supports a conservation of macrophage polarization in fish

Mammalian macrophages can adopt polarization states that, depending on the exact stimuli present in their extracellular environment, can lead to very different functions. Although these different polarization states have been shown primarily for macrophages of humans and mice, it is likely that polarized macrophages with corresponding phenotypes exist across mammals. Evidence of functional conservation in macrophages from teleost fish suggests that the same, or at least comparable polarization states should also be present in teleosts. However, corresponding transcriptional profiles of marker genes have not been reported thus far. In this study we confirm that macrophages from common carp can polarize into M1- and M2 phenotypes with conserved functions and corresponding transcriptional profiles compared to mammalian macrophages. Carp M1 macrophages show increased production of nitric oxide and a transcriptional profile with increased pro-inflammatory cytokines and mediators, including il6, il12 and saa. Carp M2 macrophages show increased arginase activity and a transcriptional profile with increased anti-inflammatory mediators, including cyr61, timp2b and tgm2b. Our RNA sequencing approach allowed us to list, in an unbiased manner, markers discriminating between M1 and M2 macrophages of teleost fish. We discuss the importance of our findings for the evaluation of immunostimulants for aquaculture and for the identification of gene targets to generate transgenic zebrafish for detailed studies on M1 and M2 macrophages. Above all, we discuss the striking degree of evolutionary conservation of macrophage polarization in a lower vertebrate.Animal science

Immunogenetics of disease resistance in fish

The aim of the work described in this thesis is to investigate the possibility to select for immune responsiveness, and subsequently produce isogenic carp ( Cyprinus carpio L.) lines, via gynogenesis, that express the trait under selection. If possible, this would allow the repeated production of numerous isogenic fish lines, all selected for different immune parameters, that can be used for immunogenetic studies on disease resistance. As a model, we chose a defined antigenic determinant (dinitrophenyl: DNP), coupled to a carrier molecule (keyhole limpet haemocyanin: KLH) in combination with a reliable read-out system (enzymelinked immunosorbent assay: ELISA), to divergently select carp for the magnitude of their primary antibody response. The possibility to reproduce both homozygous gynogenetic females and functional males with a high or a low antibody response, resulted in the establishment of a number of F 1 hybrid crosses with high or low immune responsiveness to DNP-KLH. Typically, these isogenic lines showed no within-line genetic variation. Between-line genetic variation in susceptibility to infection with Trypanoplasma borreli, a haemoflagellate parasite of carp, was dependent upon the immune response type of the carp lines. Major histocompatibility complex (Mhc) class II&nbsp;β-chain polymorphism could be associated with the immune response types described and, in retrospect, may have contributed to the observed differences in magnitude of immune responsiveness to DNP-KLH, and possibly T.borreli