Type Ⅳ interferon(IFN-υ)is a recently discovered cytokine crucial for host defense against viral infections.However,the role and mechanisms of IFN-υin bacterial infections remain unexplored.This study investigated ...Type Ⅳ interferon(IFN-υ)is a recently discovered cytokine crucial for host defense against viral infections.However,the role and mechanisms of IFN-υin bacterial infections remain unexplored.This study investigated the antibacterial and antiviral functions and mechanisms of grass carp(Ctenopharyngodon idella)IFN-υ(CiIFN-υ)both in vivo and in vitro.The CiIFN-υgene was first identified and characterized in grass carp.Subsequently,the immune expression of CiIFN-υsignificantly increased following bacterial challenge,indicating its response to bacterial infections.The eukaryotic recombinant expression plasmid of CiIFN-υwas then constructed and transfected into fathead minnow(FHM)cells.Supernatants were collected and incubated with four bacterial strains,followed by plate spreading and colony counting.Results indicated that CiIFN-υexhibited more potent antibacterial activity against gram-negative bacteria compared to gram-positive bacteria and aggregated gram-negative bacteria but not gram-positive bacteria.In vivo experiments further confirmed the antibacterial function,showing high survival rates,low tissue edema and damage,reduced tissue bacterial load,and elevated proinflammatory response at the early stages of bacterial infection.In addition,the antiviral function of CiIFN-υwas confirmed through in vitro and in vivo experiments,including crystal violet staining,survival rates,tissue viral burden,and reverse transcription-quantitative real-time polymerase chain reaction(RT-qPCR).This study highlights the antibacterial function and preliminary mechanism of IFN-υ,demonstrating that IFN-υpossesses dual functions against bacterial and viral infections.展开更多
[Objective] This study aimed to evaluate growth performance of hybrid progenies of 10 grass carp （Ctenopharyngodon idella） populations from different ge- ographical origins （Foshan population, Zhaoqing population, ...[Objective] This study aimed to evaluate growth performance of hybrid progenies of 10 grass carp （Ctenopharyngodon idella） populations from different ge- ographical origins （Foshan population, Zhaoqing population, Jingzhou population 1, Jingzhou population 2, Jingzhou population 3, Ezhou population, Yiyang population, Changsha population 1, Changsha population 2 and Jiangsu population）. [Method] Fifteen hybrid combinations were established with these ten grass carp populations and cultured in the same pond. The body mass of hybrid progenies was measured at the age of 4, 7, 8, 10, 12, 15 and 18 months, respectively. Using body mass at the age of 4 months as a concomitant variable, the growth performance of proge- nies from different hybrid combinations was analyzed by covariance analysis. [Re- suit] The average body mass of hybrid progenies of Jingzhou population 2 c2 x Foshan population $ reached the maximum （1 892.90 g） at the age of 18 months, which was 3.51%-32.36% higher than that of other hybrid progenies. Multiple com- parison analysis showed that average body mass of hybrid progenies of Jingzhou population 2 2 x Foshan population ♂ and Jingzhou population 1 ♀× Foshan population ♂ was significantly higher than that of other hybrid progenies （P〈0.05）. [Conclusion] In the production process, Jingzhou population 2 ♀ × Foshan popula- tion 5 and Jingzhou population 1 ♀×Foshan population ♂ can be used to pro- duce high-quality fries, thereby greatly improving economic benefits. This study laid the foundation for establishment of rapid-growing grass carp core populations.展开更多
D-mannose,essential for protein glycosylation,has been reported to have immunomodulatory effects and to maintain intestinal flora homeostasis.In addition to evaluating growth performance,we examined the impact of D-ma...D-mannose,essential for protein glycosylation,has been reported to have immunomodulatory effects and to maintain intestinal flora homeostasis.In addition to evaluating growth performance,we examined the impact of D-mannose on the structure of epithelial cells and apical junction complexes in the animal intestine.All 1800 grass carp(16.20±0.01 g)were randomly divided into six treatments with six replicates of 50 fish each and fed with six different levels of D-mannose(0.52,1.75,3.02,4.28,5.50 and 6.78 g/kg diet)for 70 d.The study revealed that D-mannose increased feed intake(P 0.05).D-mannose supplementation at 1.75 g/kg increased crude protein content in fish and lipid production value(P<0.05).D-mannose supplementation at 4.28 g/kg increased intestinal length,intestinal weight and fold height of grass carp compared to the control group(P<0.05).This improvement may be attributed to the phosphomannose isomerase(PMI)-mediated enhancement of glycolysis.This study found that D-mannose supplementation at 4.28 or 3.02 g/kg reduced serum diamine oxidase activity or D-lactate content(P<0.05)and improved cellular and intercellular structures for the first time.The improvement of cellular redox homeostasis involves alleviating endoplasmic reticulum(ER)stress through the inositol-requiring enzyme 1(IRE1),RNA-dependent protein kinase-like ER kinase(PERK),and activating transcription factor 6(ATF6)signaling pathways.The alleviation of ER stress may be linked to the phosphomannomutase(PMM)-mediated enhancement of protein glycosylation.In addition,ubiquitin-dependent[PTEN-induced putative kinase 1(PINK1)/Parkin]and ubiquitin-independent[BCL2-interacting protein 3-like(BNIP3L),BCL2-interacting protein 3(BNIP3),and FUN14 domain containing 1(FUNDC1)]mitophagy may play a role in maintaining cellular redox homeostasis.The enhancement of intercellular structures includes enhancing tight junction and adherent junction structures,which may be closely associated with the small Rho GTPase protein(RhoA)/the Rho-associated protein kinase(ROCK)signaling pathway.In conclusion,D-mannose improved intestinal cellular redox homeostasis associated with ER stress and mitophagy pathways,and enhanced intercellular structures related to tight junctions and adherent junctions.Furthermore,quadratic regression analysis of the PWG and intestinal reactive oxygen species content indicated that the optimal addition level of D-mannose for juvenile grass carp was 4.61 and 4.59 g/kg,respectively.展开更多
Bacterial pathogens destroy the structural integrity of functional organs in fish,leading to severe challenges in the aquaculture industry.Vitamin D_(3)(VD_(3))prevents bacterial infections and strengthens immune syst...Bacterial pathogens destroy the structural integrity of functional organs in fish,leading to severe challenges in the aquaculture industry.Vitamin D_(3)(VD_(3))prevents bacterial infections and strengthens immune system function via vitamin D receptor(VDR).However,the correlation between VD_(3)/VDR and the structural integrity of functional organs remains unclarified.This study aimed to investigate the influence of VD_(3)supplementation on histological characteristics,apoptosis,and tight junction characteristics in fish intestine during pathogen infection.A total of 540 healthy grass carp(257.24±0.63 g)were fed different levels of VD_(3)(15.2,364.3,782.5,1,167.9,1,573.8,and 1,980.1 IU/kg)for 70 d.Subsequently,fish were challenged with Aeromonas hydrophila,a pathogen that causes intestinal inflammation.Our present study demonstrated that optimal supplementation with VD_(3)(1)alleviated intestinal structural damage,and inhibited oxidative damage by reducing levels of oxidative stress biomarkers;(2)attenuated excessive apoptosisrelated death receptor and mitochondrial pathway processes in relation to p38 mitogen-activated protein kinase signaling(P<0.05);(3)enhanced tight junction protein expression by inhibiting myosin light chain kinase signaling(P<0.05);and(4)elevated VDR isoform expression in fish intestine(P<0.05).Overall,the results demonstrated that VD_(3)alleviates oxidative injury,apoptosis,and the destruction of tight junction protein under pathogenic infection,thereby strengthening pathogen defenses in the intestine.This finding supports the rationale for VD_(3)intervention as an essential practice in sustainable aquaculture.展开更多
Copper(Cu)is a trace element,essential for fish growth.In the current study,in addition to growth performance,we first explored the effects of Cu on collagen synthesis and myofiber growth and development in juvenile g...Copper(Cu)is a trace element,essential for fish growth.In the current study,in addition to growth performance,we first explored the effects of Cu on collagen synthesis and myofiber growth and development in juvenile grass carp(Ctenopharyngodon idella).A total of 1080 fish(11.16±0.01 g)were randomly divided into 6 treatments(3 replicates per treatment)to receive five doses of organic Cu which were Cu citrate(CuCit)at 0.99(basal diet),2.19,4.06,6.15,and 8.07 mg/kg,and one dose o inorganic Cu(CuSO_(4)·5H_(2)O at 3.15 mg/kg),for 9 weeks.The results showed appropriate Cu level(4.06 mg kg)enhanced growth performance,improved nutritional Cu status,and downregulated Cu-transporting ATPase 1 mRNA levels in the hepatopancreas,intestine,and muscle of juvenile grass carp.Meanwhile collagen content in fish muscle was increased after Cu intake,which was probably due to the following pathways:(1)activating CTGF/TGF-β1/Smads signaling pathway to regulate collagen transcription;(2upregulating of La ribonucleoprotein domain family 6(LARP6)mRNA levels to regulate translation initiation;(3)increasing proline hydroxylase,lysine hydroxylase,and lysine oxidase activities to regulate posttranslational modifications.In addition,optimal Cu group increased myofiber diameters and the frequency of myofibers with diameter>50μm,which might be associated with upregulation of cyclin B cyclin D,cyclin E,proliferating cell nuclear antigen,myogenic determining factor(MyoD),myogenic factor 5,myogenin(MyoG),myogenic regulatory factor 4 and myosin heavy chain(MyHC)and down regulation of myostatin mRNA levels,increasing protein levels of MyoD,MyoG and MyHC in fish muscle Finally,based on percentage weight gain(PWG),serum ceruloplasmin(Cp)activity and collagen conten in fish muscle,Cu requirements were determined as 4.74,4.37 and 4.62 mg/kg diet(CuCit as Cu source of juvenile grass carp,respectively.Based on PWG and Cp activity,compared to CuSO_(4)·5H_(2)O,the efficacy of CuCit were 131.80%and 115.38%,respectively.Our findings provide new insights into Cu supple mentation to promote muscle growth in fish,and help improve the overall productivity of aquaculture.展开更多
[ Objective] This study aimed to investigate the biochemical and genetic characteristics of F1 hybrids of Ctenopharyngodon idellus ♀× Elopichthys bambusa ♂ - [ Method ] By using vertical polyacrylamide gel elec...[ Objective] This study aimed to investigate the biochemical and genetic characteristics of F1 hybrids of Ctenopharyngodon idellus ♀× Elopichthys bambusa ♂ - [ Method ] By using vertical polyacrylamide gel electrophoresis technology, three isozymes （LDH, EST, MDH） in nine tissues （heart, brain, eyes, liver, kidney, spleen, fin, muscle, plasma） of F1 hybrids of C. ideUus and parents were compared. [Result] Three isozymes in F1 hybrids of ♀× E. bambusa ♀ were analyzed; differences in isozyme and proteins between F1 hybrids idellus ♀× E. bambusa ♀ exhibited varying degrees of tissue-specificity; isozymes and proteins in F, hybrids were signiticandy different from the parents. [ Conclusion] These nine differences could be used as indicators to identify F1 hybrids and parents.展开更多
Grass carp hemorrhagic disease caused by grass carp reovirus(GCRV)results in significant economic losses to the global grass carp aquaculture industry.Oral vaccination is an ideal choice for disease precaution in aqua...Grass carp hemorrhagic disease caused by grass carp reovirus(GCRV)results in significant economic losses to the global grass carp aquaculture industry.Oral vaccination is an ideal choice for disease precaution in aquaculture.However,oral vaccine can be degraded in the gut.Therefore,the selection of loading materials is essential.In this study,the S6 and S7 fragments(encoding the outer capsid protein VP4 and fibronectin VP56 of GCRV)and grass carp interferons(IFNs),including IFN1,IFN3,and IFNγ2 were used to create DNA vaccines and adjuvants based on pcDNA3.1,respectively.The oral DNA vaccine was encapsulated in poly(lactic-co-glycolic acid)(PLGA)and polyvinyl alcohol(PVA)with IFNs.The PLGA-PVA(PP)nano-microspheres were prepared by double emulsionsolvent evaporation technique.Using transmission electron microscopy and dynamic light scattering assays,it was determined the vaccines had a spherical structure with uniform particle size(643.5±35.3 nm).The nanomicrospheres possessed excellent encapsulation efficiency(81.6±2.6%)and loading rate(0.54±0.02%),and simultaneously exhibited negligible hemolytic activity and cell toxicity.The protection rate and tissue viral loads post-GCRV challenge in grass carp were assessed.The oral PP nano-microsphere with pVP4 t pIFN1(PP41)vaccine increased protection rate by 44%compared with the control group and was correlated with relatively low viral loads in the spleen,head kidney,and hindgut.Further,three crucial serum biochemical indexes,total superoxide dismutase(TSOD),complement C3(C3),and lysozyme(LZM),were also dramatically increased.Furthermore,mRNA expressions of representative immune-related genes(IgM,IFN1,IFNγ2,MHC-I,and CD8α)in the head kidney and spleen were significantly enhanced.In addition,mRNA expression of IgT was significantly boosted in the hindgut.The results indicate that DNA vaccine capsulated with PP is effective against GCRV infection.The present study provides insights into a prospective strategy for oral vaccine development in aquaculture.展开更多
基金supported by the Biological Breeding-Major Projects(2023ZD04065)。
文摘Type Ⅳ interferon(IFN-υ)is a recently discovered cytokine crucial for host defense against viral infections.However,the role and mechanisms of IFN-υin bacterial infections remain unexplored.This study investigated the antibacterial and antiviral functions and mechanisms of grass carp(Ctenopharyngodon idella)IFN-υ(CiIFN-υ)both in vivo and in vitro.The CiIFN-υgene was first identified and characterized in grass carp.Subsequently,the immune expression of CiIFN-υsignificantly increased following bacterial challenge,indicating its response to bacterial infections.The eukaryotic recombinant expression plasmid of CiIFN-υwas then constructed and transfected into fathead minnow(FHM)cells.Supernatants were collected and incubated with four bacterial strains,followed by plate spreading and colony counting.Results indicated that CiIFN-υexhibited more potent antibacterial activity against gram-negative bacteria compared to gram-positive bacteria and aggregated gram-negative bacteria but not gram-positive bacteria.In vivo experiments further confirmed the antibacterial function,showing high survival rates,low tissue edema and damage,reduced tissue bacterial load,and elevated proinflammatory response at the early stages of bacterial infection.In addition,the antiviral function of CiIFN-υwas confirmed through in vitro and in vivo experiments,including crystal violet staining,survival rates,tissue viral burden,and reverse transcription-quantitative real-time polymerase chain reaction(RT-qPCR).This study highlights the antibacterial function and preliminary mechanism of IFN-υ,demonstrating that IFN-υpossesses dual functions against bacterial and viral infections.
基金Supported by National Science and Technology Project in Rural Areas during the"12th Five-Year Plan"Bulk Breeding of New Varieties of Major Freshwater Fishes"(2012BAD26B02)Construction of National Bulk Freshwater Fish industry Technology System"Selective Breeding and Molecular-assisted Breeding of Grass Carp in South China"(CARS-46-03)~~
文摘[Objective] This study aimed to evaluate growth performance of hybrid progenies of 10 grass carp （Ctenopharyngodon idella） populations from different ge- ographical origins （Foshan population, Zhaoqing population, Jingzhou population 1, Jingzhou population 2, Jingzhou population 3, Ezhou population, Yiyang population, Changsha population 1, Changsha population 2 and Jiangsu population）. [Method] Fifteen hybrid combinations were established with these ten grass carp populations and cultured in the same pond. The body mass of hybrid progenies was measured at the age of 4, 7, 8, 10, 12, 15 and 18 months, respectively. Using body mass at the age of 4 months as a concomitant variable, the growth performance of proge- nies from different hybrid combinations was analyzed by covariance analysis. [Re- suit] The average body mass of hybrid progenies of Jingzhou population 2 c2 x Foshan population $ reached the maximum （1 892.90 g） at the age of 18 months, which was 3.51%-32.36% higher than that of other hybrid progenies. Multiple com- parison analysis showed that average body mass of hybrid progenies of Jingzhou population 2 2 x Foshan population ♂ and Jingzhou population 1 ♀× Foshan population ♂ was significantly higher than that of other hybrid progenies （P〈0.05）. [Conclusion] In the production process, Jingzhou population 2 ♀ × Foshan popula- tion 5 and Jingzhou population 1 ♀×Foshan population ♂ can be used to pro- duce high-quality fries, thereby greatly improving economic benefits. This study laid the foundation for establishment of rapid-growing grass carp core populations.
基金supported by the earmarked fund for CARS (CARS-45),and the National Key R&D Program of China (2019YFD0900200).
文摘D-mannose,essential for protein glycosylation,has been reported to have immunomodulatory effects and to maintain intestinal flora homeostasis.In addition to evaluating growth performance,we examined the impact of D-mannose on the structure of epithelial cells and apical junction complexes in the animal intestine.All 1800 grass carp(16.20±0.01 g)were randomly divided into six treatments with six replicates of 50 fish each and fed with six different levels of D-mannose(0.52,1.75,3.02,4.28,5.50 and 6.78 g/kg diet)for 70 d.The study revealed that D-mannose increased feed intake(P 0.05).D-mannose supplementation at 1.75 g/kg increased crude protein content in fish and lipid production value(P<0.05).D-mannose supplementation at 4.28 g/kg increased intestinal length,intestinal weight and fold height of grass carp compared to the control group(P<0.05).This improvement may be attributed to the phosphomannose isomerase(PMI)-mediated enhancement of glycolysis.This study found that D-mannose supplementation at 4.28 or 3.02 g/kg reduced serum diamine oxidase activity or D-lactate content(P<0.05)and improved cellular and intercellular structures for the first time.The improvement of cellular redox homeostasis involves alleviating endoplasmic reticulum(ER)stress through the inositol-requiring enzyme 1(IRE1),RNA-dependent protein kinase-like ER kinase(PERK),and activating transcription factor 6(ATF6)signaling pathways.The alleviation of ER stress may be linked to the phosphomannomutase(PMM)-mediated enhancement of protein glycosylation.In addition,ubiquitin-dependent[PTEN-induced putative kinase 1(PINK1)/Parkin]and ubiquitin-independent[BCL2-interacting protein 3-like(BNIP3L),BCL2-interacting protein 3(BNIP3),and FUN14 domain containing 1(FUNDC1)]mitophagy may play a role in maintaining cellular redox homeostasis.The enhancement of intercellular structures includes enhancing tight junction and adherent junction structures,which may be closely associated with the small Rho GTPase protein(RhoA)/the Rho-associated protein kinase(ROCK)signaling pathway.In conclusion,D-mannose improved intestinal cellular redox homeostasis associated with ER stress and mitophagy pathways,and enhanced intercellular structures related to tight junctions and adherent junctions.Furthermore,quadratic regression analysis of the PWG and intestinal reactive oxygen species content indicated that the optimal addition level of D-mannose for juvenile grass carp was 4.61 and 4.59 g/kg,respectively.
基金financially supported by the earmarked fund for CARS(CARS-45)National Natural Science Foundation of China for Outstanding Youth Science Foundation(31922086)+1 种基金the Young Top-Notch Talent Support Programand the 111 project(D17015)
文摘Bacterial pathogens destroy the structural integrity of functional organs in fish,leading to severe challenges in the aquaculture industry.Vitamin D_(3)(VD_(3))prevents bacterial infections and strengthens immune system function via vitamin D receptor(VDR).However,the correlation between VD_(3)/VDR and the structural integrity of functional organs remains unclarified.This study aimed to investigate the influence of VD_(3)supplementation on histological characteristics,apoptosis,and tight junction characteristics in fish intestine during pathogen infection.A total of 540 healthy grass carp(257.24±0.63 g)were fed different levels of VD_(3)(15.2,364.3,782.5,1,167.9,1,573.8,and 1,980.1 IU/kg)for 70 d.Subsequently,fish were challenged with Aeromonas hydrophila,a pathogen that causes intestinal inflammation.Our present study demonstrated that optimal supplementation with VD_(3)(1)alleviated intestinal structural damage,and inhibited oxidative damage by reducing levels of oxidative stress biomarkers;(2)attenuated excessive apoptosisrelated death receptor and mitochondrial pathway processes in relation to p38 mitogen-activated protein kinase signaling(P<0.05);(3)enhanced tight junction protein expression by inhibiting myosin light chain kinase signaling(P<0.05);and(4)elevated VDR isoform expression in fish intestine(P<0.05).Overall,the results demonstrated that VD_(3)alleviates oxidative injury,apoptosis,and the destruction of tight junction protein under pathogenic infection,thereby strengthening pathogen defenses in the intestine.This finding supports the rationale for VD_(3)intervention as an essential practice in sustainable aquaculture.
基金This research was financially supported by National Natural Science Foundation of China(U21A20266)National Key R&D Program of China(2018YFD0900400,2019YFD0900200)+1 种基金National Natural Science Foundation of China for Outstanding Youth Science Foundation(31922086)supported by the earmarked fund for CARS(CARS-45).
文摘Copper(Cu)is a trace element,essential for fish growth.In the current study,in addition to growth performance,we first explored the effects of Cu on collagen synthesis and myofiber growth and development in juvenile grass carp(Ctenopharyngodon idella).A total of 1080 fish(11.16±0.01 g)were randomly divided into 6 treatments(3 replicates per treatment)to receive five doses of organic Cu which were Cu citrate(CuCit)at 0.99(basal diet),2.19,4.06,6.15,and 8.07 mg/kg,and one dose o inorganic Cu(CuSO_(4)·5H_(2)O at 3.15 mg/kg),for 9 weeks.The results showed appropriate Cu level(4.06 mg kg)enhanced growth performance,improved nutritional Cu status,and downregulated Cu-transporting ATPase 1 mRNA levels in the hepatopancreas,intestine,and muscle of juvenile grass carp.Meanwhile collagen content in fish muscle was increased after Cu intake,which was probably due to the following pathways:(1)activating CTGF/TGF-β1/Smads signaling pathway to regulate collagen transcription;(2upregulating of La ribonucleoprotein domain family 6(LARP6)mRNA levels to regulate translation initiation;(3)increasing proline hydroxylase,lysine hydroxylase,and lysine oxidase activities to regulate posttranslational modifications.In addition,optimal Cu group increased myofiber diameters and the frequency of myofibers with diameter>50μm,which might be associated with upregulation of cyclin B cyclin D,cyclin E,proliferating cell nuclear antigen,myogenic determining factor(MyoD),myogenic factor 5,myogenin(MyoG),myogenic regulatory factor 4 and myosin heavy chain(MyHC)and down regulation of myostatin mRNA levels,increasing protein levels of MyoD,MyoG and MyHC in fish muscle Finally,based on percentage weight gain(PWG),serum ceruloplasmin(Cp)activity and collagen conten in fish muscle,Cu requirements were determined as 4.74,4.37 and 4.62 mg/kg diet(CuCit as Cu source of juvenile grass carp,respectively.Based on PWG and Cp activity,compared to CuSO_(4)·5H_(2)O,the efficacy of CuCit were 131.80%and 115.38%,respectively.Our findings provide new insights into Cu supple mentation to promote muscle growth in fish,and help improve the overall productivity of aquaculture.
基金Supported by Project of Project of Science and Technology of Wuhan City(2013021001010464)
文摘[ Objective] This study aimed to investigate the biochemical and genetic characteristics of F1 hybrids of Ctenopharyngodon idellus ♀× Elopichthys bambusa ♂ - [ Method ] By using vertical polyacrylamide gel electrophoresis technology, three isozymes （LDH, EST, MDH） in nine tissues （heart, brain, eyes, liver, kidney, spleen, fin, muscle, plasma） of F1 hybrids of C. ideUus and parents were compared. [Result] Three isozymes in F1 hybrids of ♀× E. bambusa ♀ were analyzed; differences in isozyme and proteins between F1 hybrids idellus ♀× E. bambusa ♀ exhibited varying degrees of tissue-specificity; isozymes and proteins in F, hybrids were signiticandy different from the parents. [ Conclusion] These nine differences could be used as indicators to identify F1 hybrids and parents.
基金supported by the National Key R&D Program of China(2022YFF1000302)Major Project of Hubei Hongshan Laboratory(2022hszd011).
文摘Grass carp hemorrhagic disease caused by grass carp reovirus(GCRV)results in significant economic losses to the global grass carp aquaculture industry.Oral vaccination is an ideal choice for disease precaution in aquaculture.However,oral vaccine can be degraded in the gut.Therefore,the selection of loading materials is essential.In this study,the S6 and S7 fragments(encoding the outer capsid protein VP4 and fibronectin VP56 of GCRV)and grass carp interferons(IFNs),including IFN1,IFN3,and IFNγ2 were used to create DNA vaccines and adjuvants based on pcDNA3.1,respectively.The oral DNA vaccine was encapsulated in poly(lactic-co-glycolic acid)(PLGA)and polyvinyl alcohol(PVA)with IFNs.The PLGA-PVA(PP)nano-microspheres were prepared by double emulsionsolvent evaporation technique.Using transmission electron microscopy and dynamic light scattering assays,it was determined the vaccines had a spherical structure with uniform particle size(643.5±35.3 nm).The nanomicrospheres possessed excellent encapsulation efficiency(81.6±2.6%)and loading rate(0.54±0.02%),and simultaneously exhibited negligible hemolytic activity and cell toxicity.The protection rate and tissue viral loads post-GCRV challenge in grass carp were assessed.The oral PP nano-microsphere with pVP4 t pIFN1(PP41)vaccine increased protection rate by 44%compared with the control group and was correlated with relatively low viral loads in the spleen,head kidney,and hindgut.Further,three crucial serum biochemical indexes,total superoxide dismutase(TSOD),complement C3(C3),and lysozyme(LZM),were also dramatically increased.Furthermore,mRNA expressions of representative immune-related genes(IgM,IFN1,IFNγ2,MHC-I,and CD8α)in the head kidney and spleen were significantly enhanced.In addition,mRNA expression of IgT was significantly boosted in the hindgut.The results indicate that DNA vaccine capsulated with PP is effective against GCRV infection.The present study provides insights into a prospective strategy for oral vaccine development in aquaculture.
