Responses Mechanism Of Hepatopancreas And Intestinal Of Penaeus Vannamei To Different Pathogenic Factors

The purpose of this work was to investigate the effect of three prominent pathogenic factors(aflatoxin B1,high rearing density and Vibrio parahaemolyticus)in aquaculture industry on Penaeus vannamei growth,antioxidant,immune and nutrition performance.Meanwhile,the potential response mechanisms and correlations of shrimp hepatopancreas and intestines to different pathogenic factors were systematically studied using transcriptome and microbial 16S sequencing technology.1.Responses mechanism of hepatopancreas and intestinal of P.vannamei fed with aflatoxin B1(AFB1)Healthy P.vannamei with mean weight 2.55±0.08 g were fed with an experimental feedstuff containing 5(Experimental groups)and 0(Control group)p.p.m.AFB1 for 30 consecutive days,respectively.The hepatopancreas and intestine samples of shrimp were measured and sampled at 3,6,12,18,24 and 30 d.The results were as follows:the survival rate and weight gain rate(WG)of shrimp were significantly lower in the experimental group than in the control group(P<0.05);the microstructure of hepatopancreas and intestine in the experimental group was damage severely;the digestive enzymes activities and genes expression level of hepatopancreas and intestine were significantly lower in the experimental group than in the control group and showed a time-dependent decrease(P<0.05);the activities of superoxide dismutase(SOD),catalase(CAT)and glutathione peroxidase(GPX)of hepatopancreas and intestine were significantly higher in the experimental group than in the control group and showed a tendency to increase first and then decrease(P<0.05);the relative m RNA expression of immune genes(Toll,IMD,pro PO,Rab and GST)of the hepatopancreas and intestines were significantly higher in the experimental group than in the control group and showed a tendency to increase first and then decrease(P<0.05);AFB1 induced to12,014 and 1,387 differentially expression genes(DEGs)in the hepatopancreas and intestine,respectively;totally 1,995 and 152 DEGs in hepatopancreas and intestines were mainly annotated and classified into 18 and 7 immune-related pathways or processes,respectively;the relative abundance of Proteobacteria,Firmicutes,Vibrio and Photobacterium in the experimental group increased obviously with the experiment period extension,while the relative abundance of Bacteroidetes,Flavobacterium?sp?M and Tenacibaculum decreased;additionally,the types and amounts of intestinal flora in the control group are more abundant than the experimental group.All above results indicated that challenge 5 p.p.m.AFB1 could lead to damage of antioxidants and immune system,dysregulation of intestinal flora(decreased probiotics and increased pathogenic bacteria),inhibition of digestive system,destruction of histomorphology in P.vannamei,causing growth restriction and dead.Additionally,hepatopancreas of P.vannamei was the main immune organs response to AFB1 challenge and intestine played an important role in metabolism.2.Responses mechanism of hepatopancreas and intestinal of P.vannamei to high rearing density and V.parahaemolyticus E1(VPE1)challenge under the high rearing densityHealthy P.vannamei with mean weight 7.52±0.12 g were conducted two different rearing densities(800-and 400-shrimp/m~3)for 15 consecutive days,respectively,and then challenge VPE1 for 72 consecutive hours.The hepatopancreas and intestine samples of shrimp were measured and sampled at 5,10,15 d(under different rearing density)and 12,24,48,72 h(after VPE1 challenge).The results were as follows:the survival rate and weight gain rate(WG)of shrimp were significantly lower in the high rearing density group than in the low rearing density group(P<0.05);the microstructure of hepatopancreas and intestine in the high rearing density group was damage severely;the digestive enzymes activities and genes expression level of hepatopancreas and intestine were significantly lower in the high rearing density group than in the low rearing density group and showed a time-dependent decrease(P<0.05);the activities of T-SOD,CAT and GPX of hepatopancreas and intestine were significantly higher in the high rearing density group than in the low rearing density group and showed a tendency to increase first and then decrease(P<0.05);the relative m RNA expression of immune genes(Toll,IMD,pro PO,LZM and Relish)of the hepatopancreas and intestines were significantly higher in the experimental group than in the control group at the 15 d and showed a tendency to increase first and then decrease(P<0.05);Rearing shrimp at high density for 15 d resulted in 45 DEGs in the hepatopancreas and mainly related to 4 immune-related pathways or processes;In the intestine,totally 5,470 DEGs were identified and mainly classified into 17 immune-related pathways or processes;challenge to the VPE1 48 h induced 639 DEGs in the hepatopancreas and mainly involved into 14 immune-related pathways or processes;In the intestine,totally 279 DEGs were identified and mainly classified into 14 immune-related pathways or processes;the relative abundance of Planctomycetes,Photobacterium and Vibrio in the experimental group increased obviously in the high rearing density,while the relative abundance of Bacteroidetes,Firmicutes,Pseudoalteromonas and Blastopirellula decreased;All above results showed that high rearing density could inhibit growth,destruct histomorphology,damage antioxidants and immune system,increase pathogenic bacteria and decrease probiotics,resulting in higher mortality due to disease.In addition,the intestine was mainly affected by density stress,and the hepatopancreas of shrimp was the main immune organ in response to the pathogen infection.3.Responses mechanism of hepatopancreas and intestinal of P.vannamei to VPE1 challengeHealthy P.vannamei with mean weight 12.28±0.10 g were randomly divided into two groups for consecutive 72 h VPE1 challenge test,including VPE1 challenge group and control group.The hepatopancreas and intestines samples were collected to transcriptome sequencing after VPE1 challenge 48 h.The results were as follows:the mortality was significantly higher in the VPE1 challenge group than the control group;VPE1 challenge resulted in 2,628(including 2,489 up-regulated and 139 down-regulated)and 1,963(including 1,732 up-regulated and 231 down-regulated)DEGs in the hepatopancreas and intestine,respectively;a total of 27 immune-related pathways or processes were enriched both in the hepatopancreas and intestine.Above results illuminated that VPE1 challenge can induce a large number of differential gene expressions in the hepatopancreas and intestine of shrimp,which led to disturbances in its immune-related pathways and eventually death.Additionally,the hepatopancreas of shrimp was the main immune organ in response to the VPE1 challenge.4.Preliminary study on the potential correlation between hepatopancreas and intestinal of P.vannamei immune response to different pathogenic factorsThe purpose of this study was to analyze the differences and connections between hepatopancreas and intestinal response to three different pathogenic factors,AFB1,density stress and VPE1 at the transcription level using transcriptome and bioinformatics technologies.The results manifested totally 7 common GO terms and18 KEGG pathways were enriched in the hepatopancreas and intestinal of shrimp under three different pathogenic factors conditions,respectively.Among them,5 common KEGG pathways related to immune were identified in the hepatopancreas and intestinal,including apoptosis,phagosome,AMPK signaling pathway,antigen processing and presentation and epstein-bar virus infection.Therefore,it is speculated that the 5common pathways may be the potential pathways connected hepatopancreas with intestinal of shrimp in immune defense function.