| Much research has been devoted to exploring the physiological and biochemical changes of Nile tilapia Oreochromis niloticus during salinity acclimation.However,little work has been done to reveal the effects of hyperosmotic stress on the immunity system of Nile tilapia.And there are few reports regarding histologic and transcriptomics stress responses of immune organs to hyperosmotic conditions.In this paper,an eight-week growth trial was conducted to explore the effects of long-term hyperosmotic stress on the two immune organs(spleen and head kidney)of Nile tilapia.The freshwater group was used as a control group to assess long-term effects of hyperosmotic stress(16 psu)on the growth,histology,hematology,and transcriptomics of the two immune organs(spleen and head kidney).The results indicate that no significant difference was observed on the weight gain and specific growth rate of Nile tilapia between the fresh water group and the hyperosmotic stress group.However,hyperosmotic stress(16 psu)led to congestive splenomegaly,and that the expression of platelet-and coagulation-related genes were significantly reduced,and that the variation coefficient of red blood cell distribution width and mean corpuscular hemoglobin were significantly increased.Meanwhile,a large number of macrophage centers and pigmentation were observed in tissue sections of spleen and head kidney of Nile tilapia.The results of transmission electron microscopy show that the hyperosmotic stress(16psu)group had more abnormal macrophages containing sedimentary particles and more neutrophils in the spleen and head kidney than the control group.In the transcriptomics of the spleen and head kidney,772 and 502 genes were marked as significantly differentially expressed respectively,corresponding to 5 and 1 pathways which can significantly change the immune system,and the spleen complementary pathway was significantly reduced in the hyperosmotic stress(16psu)group.This study shows that Nile tilapia exposed to hyperosmotic environment for a long period of time can induce splenomegaly and coagulation function decrease,as well as the improvement of spleen phagocytic activity and the reduction of complementary pathway.Subsequently,further studies on hyperosmotic stress relief experiments were conducted to investigate the role of β-glucan in the immunonutrition regulation to hyperosmotic stress,and to evaluate the effect of in-diet β-glucan on the injury alleviation of the immune organs of Nile tilapia caused by hyperosmotic stress in terms of growth,histology,hematology,expression of immune inflammatory growth factors,and intestinal microbiology.Three diets containing equal levels of nitrogen and energy but Opsu(control feed),2 psu and 4 psu of β-glucan were prepared to feed fresh water group and hyperosmotic stress groups(16 psu)for 8 weeks.The results show that,under hyperosmotic stress,the growth and survival levels of Nile tilapia with different levels of β-glucan were higher than those without addition,which has no significant difference compared with the control diet freshwater group.Under hyperosmotic stress,splenomegaly,spleen coefficient increase(P<0.05)and the significant increase of the number of red blood cells in the spleen were observed.But the 0.2%β-glucan can alleviate splenomegaly,significantly decrease the spleen coefficient and there was no significant difference of spleen coefficient compared to freshwater control diet group.Under hyperosmotic stress,the intestinal tissues appears to be disrupted,intestinal villi in the midgut is broken and the goblet cells are few.However,in the addition group with different levels of added β-glucan,the intestinal villi are not broken but long,the goblet cells are numerous,and the villi are arranged closely,which is similar to the integrity of intestinal tissue in the freshwater group.In hematology,under hyperosmotic stress,red blood cell RBC,leukocyte WBC,hematocrit HCT,mean red blood cell hemoglobin concentration MCHC,red blood cell distribution width variation coefficient RDW,PLT platelet count and platelet pressure product PCT were all elevated compared with freshwater group.But the addition of β-glucan in diet can reduce these blood parameters.At the same time,with the influence of salinity on the growth and histological structure of tilapia,the corresponding changes were also observed in the immune growth factor.The expressions of macrophage migration inhibitory factor MIF-2,transformant factor TGF-β1,TNF-α,and IL-1 were increased in the spleen of hyperosmotic group.However,β-glucan in diets can significantly reduce the expression of MIF-2,TGF-β1,and TNF-α,while significantly increase the expression of IL-1.In the intestine,addition of β-glucan significantly reduced the expression of MIF-2 and IL-1β under hyperosmotic stress,and increased the expression of TNF-a;and 0.4%β-glucan addition under hyperosmotic stress also significantly elevated TGF-β1.In head kidney,dietary β-glucan significantly increased the expression of TNF-α under hyperosmotic stress conditions(P<0.05)and 0.4%β-glucan significantly reduced the expression of IL-1(P<0.05).The gut microbiota as a health indicators provides a new direction for immunization research of tilapia.The results show that salinity stress can change the structure and richness of intestinal flora.Under the salinity stress,in terms of genus,the abundance of some pathogenic bacteria increased,such as:Veillonella,Rubrobacter,and Caldilinea.Some stress-stressed bacteria have also increased significantly,such as Gracilibacter;Perlucidibaca.However,the abundance of some dominant endogenous bacteria significantly reduced,such as Acinetobacter.Dietary addition of different concentrations of β-glucan can significantly improve the structure of the bacteria.Some conditional pathogenic bacteria significantly reduced,such as:Pseudomonas,Staphylococcus,Pseudomonas,Corynebacterium;some antagonistic bacteria increased significantly,such as Dokdonella,Thermus,Sphingopyxis and Salinococcus. |
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