The summer occurrence and distribution of vibrios in the fishes and shellfishes in the coasal waters of Hong Kong were investigaed. A total of 69 strains of vibrios were isolated from all samples examined. The strains...The summer occurrence and distribution of vibrios in the fishes and shellfishes in the coasal waters of Hong Kong were investigaed. A total of 69 strains of vibrios were isolated from all samples examined. The strains, along with 10 reference strains were classified with the technique of numerical taxonomy bared on 54 characters and 62 of the 69 strains fell into 5 major phena, identified as V. paraheamolyticus (30 strains), V. alginoloticus (23 strains), V. choloerae (3 strains), V. harveyi (2 strains) and V. fluinalis (4 strains). Among them, V.paraheamolyticus and V. alginolyticus were the predominant species in the fishes, shellfishes and the coastal waters of Hong Kong and comprised 43. 5 % and 33. 3 % of the total Vibrio spp. isolates respectively. Meanwhile, 3 strains of non-Ol V. cholerae were isolated from oyster and it was the first time to record V. cholerae non-Ol in seawater or from shellfishes in Hong Kong. These results highlighted the potential risks of food poisoning associated with raw or undercooked seafood.展开更多
The distribution featires, spotes composition and seasonal variation of halophilic Vibrios in 7 stations ofXiamen Harbor and 9 stations of the Daya Bay were studied. The counts of Vibrios were analyzed with the most p...The distribution featires, spotes composition and seasonal variation of halophilic Vibrios in 7 stations ofXiamen Harbor and 9 stations of the Daya Bay were studied. The counts of Vibrios were analyzed with the most probable number(MPN) technique. The media TCBS was used to isolate Vibrios, and API20E system employed to identifythe Vibrios. The results reveals that the density Of Vibrios in the Daya Bay ranged from 30. 0 x 105 to 2. 4 x 105 celldin-3 and the average density was 6. 61 x 103 cell·din-3, and that in Xiamen Harbor was 2. 3 x 102-2. 4 x 105 cen·din-3 and 7. 8 x 103 cell·din-3 on the average. The number of Vibrios varied seasonally with the water temperature,and was heher in the summer than in the autumn. The Vibrio species in the two bays mainly included Vibrioalginolyticus, V. parahaemolytic-us, V. fluvialis, V. vulnificus, V. mimices and V. metshnikovii and V.alginolyticus was the predominant spotes.展开更多
Turbot Scophthalmus maximus is an important mariculture fish species with high economic value.However,the bacterial diseases caused by Vibrio anguillarum infection bring huge economic losses to the turbot aquaculture ...Turbot Scophthalmus maximus is an important mariculture fish species with high economic value.However,the bacterial diseases caused by Vibrio anguillarum infection bring huge economic losses to the turbot aquaculture industry.To understand the immune response of the turbot against V.anguillarum infection and to explore novel immune-related genes,the transcriptome analysis of turbot spleen and gills were conducted after V.anguillarum infection.Differentially expressed genes(DEGs)were identified in spleen and gill of the turbot amounted to 17261 and 16436,respectively.A large number of immunerelated DEGs were enriched in cytokine-cytokine receptor interaction signaling pathway,and the others by the kyoto encyclopedia of genes and genomes(KEGG)enrichment.The gene ontology(GO)classification analysis revealed that V.anguillarum infection had the greatest effect on biological processes and cellular components.Twelve immune-related DEGs were identified in the spleen(cstl.1,egfl6,lamb21,v2rx4,calcr,and gpr78a)and gills(ghra,sh3gl2a,cst12,inhbaa,cxcl8,and il-1b)by heat map.The proteinprotein interaction(PPI)networks were constructed to analyze the immune mechanism.The results demonstrate that the maturation and antigen processing of major histocompatibility complex(MHC)class II molecule,and calcitonin-or adrenomedullin-regulated physiological activity were important events in the immunity of turbot against V.anguillarum infection.In the gills,the protein interactions in TGF-βsignaling pathway,production of inflammatory factors,and endocytosis regulation were most significant.Our research laid a foundation for discovering novel immune-related genes and enriching the knowledge of immune mechanisms of turbot against V.anguillarum infection.展开更多
Lysozyme(EC3.2.1.17)plays an important role in the immune response;as a nonspecific immune factor,it can resist causative agents.Lysozyme can be divided into c-type and g-type in fish.In a previous study,through genom...Lysozyme(EC3.2.1.17)plays an important role in the immune response;as a nonspecific immune factor,it can resist causative agents.Lysozyme can be divided into c-type and g-type in fish.In a previous study,through genome-wide association analysis,the g-type lysozyme gene,which is named NaLyg in yellow drum(Nibea albiflora),was found to be a key candidate gene for disease resistance in response to Vibrio harveyi infection.The cDNA of NaLyg was 1025 bp,including four exons and three introns,and its open reading frame(ORF)had a full-length of 582 bp,encoding 193 amino acids.NaLyg was found to be conserved during evolution through bioinformatic analyses.The NaLyg protein possessed a sugar binding domain and three catalytic sites,including Glu71,Asp84 and Asp101.Quantitative qRT-PCR results confirmed that NaLyg gene mRNA was visibly increased after V.harveyi infection.The NaLyg protein purified by prokaryotic expression killed some gram-negative bacterial pathogens by inducing cell wall destruction,including V.harveyi,Aeromonas hydrophila and Edwardsiella tarda.Moreover,the NaLyg protein killed two gram-positive bacteria,Bacillus subtilis and Staphylococcus aureus.Taken together,the experimental results suggested that the NaLyg protein of N.albiflora played an important role in fighting bacterial infections.展开更多
PhoR is a histidine kinase in a two-component regulatory system that regulates phosphorus metabolic pathways and undertakes the key mission of information transmission in pathogenic bacteria.The full-length phoR gene ...PhoR is a histidine kinase in a two-component regulatory system that regulates phosphorus metabolic pathways and undertakes the key mission of information transmission in pathogenic bacteria.The full-length phoR gene was successfully cloned from the Vibrio alginolyticus HY9901 strain.A comprehensive analysis of the cloned gene was conducted using bioinformatics.Sequence analysis revealed that the total length of the phoR gene(GenBank accession No.:KJ958404.1)is 1299 bp,with the coding region containing a total of 432 amino acid residues.The phylogenetic tree of PhoR revealed that it belongs to the same subclade as V.diabolicus.The SMART program was employed for the purpose of functional domain prediction,which revealed that PhoR possesses three major functional domains:PAS(amino acids 98-166),HisKA(amino acids 205-272),and HATPase_c(amino acids 317-429).展开更多
Objective:To analyse experimental infection and immune system of shrimp(Penaeus monodon)against Vibrios furnissii(V.furnissii).Methods:Experimental animals were collected and acclimatized by maintaining specific tempe...Objective:To analyse experimental infection and immune system of shrimp(Penaeus monodon)against Vibrios furnissii(V.furnissii).Methods:Experimental animals were collected and acclimatized by maintaining specific temperature,pH and salinity to avoid mortality.Shrimps were experimentally infected with V.furnissii and their immune responses were monitored.After the infection all the shrimps were monitored for any symptoms,death rate in 0,12,24,36,48 h.Then haemolymph were collected and tetrahydrocannabinol,phenol oxidase,nitroblue tetrazolium and lysozyme were monitored in every 12 h at the interval of 48 h.Results:Shrimps infected by live V.furnissii had showed gradual increase in tetrahydrocannabinol,phenol oxidase activity,nitro-blue-tetrazolium and lysozyme activity comparing with the killed and control.Conclusions:The live V.furnissii had showed infection in the shrimp immune system.The live V.furnissii shows infection in experimental shrimps comparing with killed V.furnissii.So the V.furnissii in nature cause the infection in shrimp Penaeus monodon immune system.This report could be applied to control of the infection in shrimp hatchery.展开更多
Objective:To study the ecology of antibiotic resistant bacteria with emphasis on sucrose negative vibrios in water and sediments samples of traditional shrimp farming system(bhery)in West Bengal,India.Methods:The vibr...Objective:To study the ecology of antibiotic resistant bacteria with emphasis on sucrose negative vibrios in water and sediments samples of traditional shrimp farming system(bhery)in West Bengal,India.Methods:The vibrios were isolated from traditional shrimp farm samples on thiosulphate citrate bile salt sucrose agar and sucrose negative bacterial strains were used as biomarkers to assess the frequency of antibiotic resistance.Results:The incoming water brought presumptive vibrios ranging from 5.50×10^(1)to 1.00×10^(3)mL in to the bhery,and there appeared to build up vibrios in the culture system with days of culture,as there was about 9 fold increase in vibrios.The levels of vibrios were observed to be moderately higher in outlet water and ranged between 4.15×10^(2)and 4.15x10^(3)mL.The counts of vibrios in pond sediment was found to be 1.00x10^(2)-4.90×10^(3)g;while in inlet(2.00×10^(2)-4.20×10^(4)g)and outlet(3.00×10^(2)-6.85×10^(3)g)their levels were observed to be higher than the pond sediment.Thirteen different Vibrio species were encountered in traditional shrimp culture system and all vibrios were sensitive to chloramphenicol,followed by ciprofloxacin and gatifloxacin(98.24%),gentamicin(95.61%)and other antibiotics.The multiple antibiotic resistance(MAR),i.e.,resistance to at least two antibiotics,was noticed among 43.85%of the sucrose negative vibrios and 41.86%of the sucrose negative non--vibrios.All vibrios harveyi strains exhibited MAR.Although no antibiotic was used in the bhery,the prevalence of MAR in 44%of the sucrose negative vibrios and nonvibrios is a cause of concern.The MAR index was higher in inlet water and sediment samples.The MAR observed in biomarker strains of pond water and sediment(40%)was comparable to those of inlet samples,thus confirming the fact that incoming water was the major source of antibiotic resistant bacteria.Conclusions:It seems that the shrimp culture in bhery does not favour the proliferation and spread of antibiotic resistant bacteria.展开更多
Objective This study investigated how the natural phytophenol and potent SIRT1 activator resveratrol(RSV)regulate necroptosis during Vibrio vulnificus(V.vulnificus)-induced sepsis and the potential mechanism.Methods T...Objective This study investigated how the natural phytophenol and potent SIRT1 activator resveratrol(RSV)regulate necroptosis during Vibrio vulnificus(V.vulnificus)-induced sepsis and the potential mechanism.Methods The effect of RSV on V.vulnificus cytolysin(VVC)-induced necroptosis was analyzed in vitro using CCK-8 and Western blot assays.Enzyme-linked immunosorbent assays and quantitative real-time polymerase chain reaction,western blot,and immunohistochemistry and survival analyses were performed to elucidate the effect and mechanism of RSV on necroptosis in a V.vulnificus-induced sepsis mouse model.Results RSV relieved necroptosis induced by VVC in RAW264.7 and MLE12 cells.RSV also inhibited the inflammatory response,had a protective effect on histopathological changes,and reduced the expression level of the necroptosis indicator pMLKL in peritoneal macrophages,lung,spleen,and liver tissues of V.vulnificus-induced septic mice in vivo.Pretreatment with RSV downregulated the mRNA of the necroptosis indicator and protein expression in peritoneal macrophages and tissues of V.vulnificusinduced septic mice.RSV also improved the survival of V.vulnificus-induced septic mice.Conclusion Our findings collectively demonstrate that RSV prevented V.vulnificus-induced sepsis by attenuating necroptosis,highlighting its potency in the clinical management of V.vulnificus-induced sepsis.展开更多
Objective This study aimed to investigate whether the VCA0560 gene acts as an active diguanylate cyclase(DGC)in Vibrio cholerae and how its transcription is regulated by Fur and Hap R.Methods The roles of VCA0560 was ...Objective This study aimed to investigate whether the VCA0560 gene acts as an active diguanylate cyclase(DGC)in Vibrio cholerae and how its transcription is regulated by Fur and Hap R.Methods The roles of VCA0560 was investigated by utilizing various phenotypic assays,including colony morphological characterization,crystal violet staining,Cyclic di-GMP(c-di-GMP)quantification,and swimming motility assay.The regulation of the VCA0560 gene by Fur and Hap R was analyzed by luminescence assay,electrophoretic mobility shift assay,and DNase I footprinting.Results VCA0560 gene mutation did not affect biofilm formation,motility,and c-di-GMP synthesis in V.cholerae,and its overexpression remarkably enhanced biofilm formation and intracellular c-di-GMP level but reduced motility capacity.The transcription of the VCA0560 gene was directly repressed by Fur and the master quorum sensing regulator Hap R.Conclusion Overexpressed VCA0560 functions as an active DGC in V.cholerae,and its transcription is repressed by Fur and Hap R.展开更多
Vibrio scophthalmi and Aeromonas salmonicida can cause high turbot mortality and huge economic losses.Presently,vaccination is the most promising method for preventing communicable diseases.In this study,we used forma...Vibrio scophthalmi and Aeromonas salmonicida can cause high turbot mortality and huge economic losses.Presently,vaccination is the most promising method for preventing communicable diseases.In this study,we used formalin to kill V.scophthalmi and A.salmonicida cells,and mixed with the mineralized oil adjuvant(Montanide^(TM)ISA 763 AVG)to prepare the bivalent inactivated vaccine.The results showed that turbot inoculated with the bivalent inactivated vaccine exhibited strong tolerance to the infection of V.scophthalmi and A.salmonicida,and no obvious clinical symptoms and pathological changes were observed.The activities of enzymes lysozyme,acid phosphatase and complement C3 had significantly increased after the vaccination.The antibody titer response of vaccinated turbot was greatly boosted,which was positively connected with the immunological impact according to ELISA results.Simultaneously,the expression levels of immune-related genes such as MHC-IIα,MHC-IIβ,CD4,CD8,TNF-αand IL^(-1)βwere up-regulated,demonstrating that it might stimulate humoral and cellular immunological response in turbot.These findings highlight the potential of the bivalent inactivated vaccine for controlling V.scophthalmi and A.salmonicida infections in turbot.展开更多
The caspase gene family is a crucial gene cluster that regulates apoptosis which contribute to programmed cell death,cell proliferation and differentiation,and several immune responses.In our study,a complete set of 1...The caspase gene family is a crucial gene cluster that regulates apoptosis which contribute to programmed cell death,cell proliferation and differentiation,and several immune responses.In our study,a complete set of 12 caspase genes were identified in spotted sea bass Lateolabrax maculatus.These genes were divided into three subfamilies:2 inflammatory caspases(casp-1 and casp-14-like),5 apoptosis initiators(casp-2,casp-8a,casp-8b,casp-9,and casp-10),and 5 apoptosis executioners(casp-3a,casp-3b,casp-3-like,casp-6,and casp-7).Their phylogenetic relationships,synteny and gene structures were systematically analyzed.Furthermore,the relative expression profiles of the caspase family members in the liver,intestine,head kidney,and spleen were measured by q PCR after infection with Vibrio harveyi.The results showed that the overall mRNA levels of the caspase genes were dramatically increased after V.harveyi infection,and the expression patterns varied among genes and tissues.More caspase genes underwent pronounced expression changes in the head kidney and spleen than in the liver or intestine,mainly after 48 h of the challenge.Specifically,casp-3a,casp-3b,casp-3-like,casp-6,casp-7,casp-8a,casp-8b,casp-10,and casp-14-like in the head kidney,and casp-3-like,casp-6,casp-7,and casp-14-like in the spleen,were the most responsive caspase genes which may contribute significantly to immune regulation in spotted sea bass.Additionally,the apoptosis level in head kidney and spleen after infection were examined using the Caspase assay.Our study provides a systemic overview of the caspase gene family in spotted sea bass after V.harveyi infection and lays a foundation for further deciphering the biological roles of these caspase genes.展开更多
The appropriate reference gene is a prerequisite for accurate normalization of gene expression level,and research on suitable reference genes in clam Cyclina sinensis is scarce.To improve the situation,we selected fiv...The appropriate reference gene is a prerequisite for accurate normalization of gene expression level,and research on suitable reference genes in clam Cyclina sinensis is scarce.To improve the situation,we selected five commonly used housekeeping genes,including β-actin,Elongation factor 1-α(EF1-α),Glyceraldehyde-3-pho sphate dehydrogenase(GAPDH),40S ribosomal protein S18(RPS18),and Tubulin a(TUB-α),then evaluated their expression stability in different adult tissues and under different experimental treatments(salinity stress and Vibrio parahaemolyticus infection).Their expression stability was analyzed by three frequently used programs,geNorm,NormFinder,and BestKeeper.This analysis indicated that multiple genes should be used for normalization,and we concluded that the reference gene combination GAPDH-RPS18-β-actin,should be used for qRT-PCR analysis in different tissues of C.sinensis under normal physiological conditions.For the clams under salinity stress and Vibrio infection,EF1-α-GAPDHRPS18 was recommended as the gene combination for qRT-PCR normalization.TUB-αwas generally poorly ranked by all programs,and should not be used in future studies.This study should provide fundamental support for accurate quantitative gene expression analysis of this species.展开更多
Neuronal nitric oxide synthase(nNOS)was the producer of nitric oxide(NO)which played important gas messenger molecules in biological process.It also can take effect as immune regulation molecule in organism.Black rock...Neuronal nitric oxide synthase(nNOS)was the producer of nitric oxide(NO)which played important gas messenger molecules in biological process.It also can take effect as immune regulation molecule in organism.Black rockfish(Sebastes schlegelii)is an important economic fish which were widely farmed in East Asia countries.Meanwhile,the pathogenic bacteria such as the Edwardsiella tarda and Vibrio anguillarum in seawater always brought serious obstacles to their healthy growth.In order to explore the expression pattern of n NOS gene under the pathogen stimulation and predict its immune function,the n NOS gene in black rockfish named Ssn NOS was identified.It was 3780 bp in length,located on chromosome 6,and contained 27 coding domain sequence(CDs).According to the phylogenetic analysis,the Ssn NOS showed closest relative to the counterpart gene of swamp eel(Monopterus albus).Meanwhile,analysis of Ssn NOS expression in various healthy tissues showed that Ssn NOS expression level was highest in healthy brain tissues,followed by intestinal tissues.In addition,Ssn NOS showed significant expression changes in response to stimulation by two pathogens.Particular in gill,the expression of Ssn NOS after pathogenic stimulation increased significantly.The Elisa analysis showed the Ssn NOS content in gills was much higher than that in other tissues at all time points.Moreover,the expression patterns of Ssn NOS in brain,intestine and kidney after stimulation by pathogens showed a distinct expression pattern which first down-regulated and then up-regulated.Therefore,the Ssn NOS may be an important signaling molecule for fish to respond rapidly in immune stimulation.展开更多
Vibrio splendidus is an important opportunistic pathogen ubiquitously present in the marine environment,exhibiting virulence to a variety of cultured animals.The extracellular products secreted by V.splendidus are cru...Vibrio splendidus is an important opportunistic pathogen ubiquitously present in the marine environment,exhibiting virulence to a variety of cultured animals.The extracellular products secreted by V.splendidus are crucial to bacterial survival and virulence.In this study,the secretion of outer membrane vesicles(OMVs)by V.splendidus was determined,purified,and morphologically characterized.The protein composition of OMVs was analyzed by proteomic analysis.The results showed that approximately 120 proteins were contained in these OMVs,including outer membrane proteins,flagellins,ABC transporters,protease,and iron regulation proteins,etc.,which were involved in bacterial motility,formation of biofilms and the cell membrane components,and cellular localization based on their structural molecule activity,passive transmembrane transporter activity,channel activity,neurotransmitter receptor activity,extracellular ligand-gated ion channel activity,glutamate receptor activity,ligand-gated ion channel activity,and transmembrane signaling receptor activity.To explore the biological functions of OMVs in V.splendidus,the effects of OMVs on the bacterial adaption to iron limitation,antibiotic,and the coelomic fluid of the Apostichopus japonicus were confirmed.This study is the first time to show that V.splendidus secretes OMVs,and OMVs carry functional proteins that enhance bacterial survival under various stresses.展开更多
Leopard coral groupers(Plectropomus leopardus),commercially bred in South China,are a significant economical fish species.In this study,by means of quantitative real-time PCR(qRT-PCR)technology,we assessed the stabili...Leopard coral groupers(Plectropomus leopardus),commercially bred in South China,are a significant economical fish species.In this study,by means of quantitative real-time PCR(qRT-PCR)technology,we assessed the stability of six common reference genes expression and selected the appropriate reference genes in leopard coral groupers with or without Vibrio harveyi stimulation at different time points.These data produced by qRT-PCR was handled via the geNorm,NormFinder,and BestKeeper software.The results revealed all the examined reference genes had manifest tissue-specific expression in different tissues.Prior to V.harveyi stimulation,RPL13 gene was the appropriate reference gene among eleven tissue types(blood,spleen,hepatopancreas,kidney,stomach,gill,heart,skin,muscle,intestine,brain)in leopard coral groupers.Under V.harveyi stimulation,the most reliable reference genes varied from tissue to tissue and were closely hinged upon different time points.At 6-h post-bacterial injection,the appropriate reference genes in hepatopancreas,spleen,kidney,and gill were Actin,B2M,UBCE,and Actin,respectively.At 9-and 12-h post-bacterial injection,the appropriate reference genes in hepatopancreas,spleen,kidney,and gill were RPL13,Actin,Actin,and Actin,respectively.If one reference gene was preferable,RPL13,Actin,Actin,and Actin could be selected as the reference gene in hepatopancreas,spleen,kidney,and gill of leopard coral groupers after V.harveyi infection,respectively.Expression profiles of two target genes(IL-6 and NK-lysin)were used to further validate reliability of these selected appropriate candidates.This study will lay a solid foundation for the future research on qRTPCR analysis of gene expression in leopard coral groupers.展开更多
文摘The summer occurrence and distribution of vibrios in the fishes and shellfishes in the coasal waters of Hong Kong were investigaed. A total of 69 strains of vibrios were isolated from all samples examined. The strains, along with 10 reference strains were classified with the technique of numerical taxonomy bared on 54 characters and 62 of the 69 strains fell into 5 major phena, identified as V. paraheamolyticus (30 strains), V. alginoloticus (23 strains), V. choloerae (3 strains), V. harveyi (2 strains) and V. fluinalis (4 strains). Among them, V.paraheamolyticus and V. alginolyticus were the predominant species in the fishes, shellfishes and the coastal waters of Hong Kong and comprised 43. 5 % and 33. 3 % of the total Vibrio spp. isolates respectively. Meanwhile, 3 strains of non-Ol V. cholerae were isolated from oyster and it was the first time to record V. cholerae non-Ol in seawater or from shellfishes in Hong Kong. These results highlighted the potential risks of food poisoning associated with raw or undercooked seafood.
文摘The distribution featires, spotes composition and seasonal variation of halophilic Vibrios in 7 stations ofXiamen Harbor and 9 stations of the Daya Bay were studied. The counts of Vibrios were analyzed with the most probable number(MPN) technique. The media TCBS was used to isolate Vibrios, and API20E system employed to identifythe Vibrios. The results reveals that the density Of Vibrios in the Daya Bay ranged from 30. 0 x 105 to 2. 4 x 105 celldin-3 and the average density was 6. 61 x 103 cell·din-3, and that in Xiamen Harbor was 2. 3 x 102-2. 4 x 105 cen·din-3 and 7. 8 x 103 cell·din-3 on the average. The number of Vibrios varied seasonally with the water temperature,and was heher in the summer than in the autumn. The Vibrio species in the two bays mainly included Vibrioalginolyticus, V. parahaemolytic-us, V. fluvialis, V. vulnificus, V. mimices and V. metshnikovii and V.alginolyticus was the predominant spotes.
基金the National Key Research and Development Program of the Ministry of Science and Technology(CN)(No.2022YFD2400401)the Key Research and Development Plan of Shandong Province(CN)(for Academician Team in Shandong)(No.2023ZLYS02)+1 种基金the Fundamental Research Funds for the Central Universities(No.202261029)the Enterprise Authorized Project(No.20200025)。
文摘Turbot Scophthalmus maximus is an important mariculture fish species with high economic value.However,the bacterial diseases caused by Vibrio anguillarum infection bring huge economic losses to the turbot aquaculture industry.To understand the immune response of the turbot against V.anguillarum infection and to explore novel immune-related genes,the transcriptome analysis of turbot spleen and gills were conducted after V.anguillarum infection.Differentially expressed genes(DEGs)were identified in spleen and gill of the turbot amounted to 17261 and 16436,respectively.A large number of immunerelated DEGs were enriched in cytokine-cytokine receptor interaction signaling pathway,and the others by the kyoto encyclopedia of genes and genomes(KEGG)enrichment.The gene ontology(GO)classification analysis revealed that V.anguillarum infection had the greatest effect on biological processes and cellular components.Twelve immune-related DEGs were identified in the spleen(cstl.1,egfl6,lamb21,v2rx4,calcr,and gpr78a)and gills(ghra,sh3gl2a,cst12,inhbaa,cxcl8,and il-1b)by heat map.The proteinprotein interaction(PPI)networks were constructed to analyze the immune mechanism.The results demonstrate that the maturation and antigen processing of major histocompatibility complex(MHC)class II molecule,and calcitonin-or adrenomedullin-regulated physiological activity were important events in the immunity of turbot against V.anguillarum infection.In the gills,the protein interactions in TGF-βsignaling pathway,production of inflammatory factors,and endocytosis regulation were most significant.Our research laid a foundation for discovering novel immune-related genes and enriching the knowledge of immune mechanisms of turbot against V.anguillarum infection.
基金supported by the National Natural Science Foundation of China(No.32072969)the National Key R&D Program of China(No.2022YFD2401002)+1 种基金the Natural Science Foundation of Fujian Province(No.2022 J01325)the Open Research Fund Program of Fujian Provincial Key Laboratory of Marine Fishery Resources and Eco-Environment(No.Z822280).
文摘Lysozyme(EC3.2.1.17)plays an important role in the immune response;as a nonspecific immune factor,it can resist causative agents.Lysozyme can be divided into c-type and g-type in fish.In a previous study,through genome-wide association analysis,the g-type lysozyme gene,which is named NaLyg in yellow drum(Nibea albiflora),was found to be a key candidate gene for disease resistance in response to Vibrio harveyi infection.The cDNA of NaLyg was 1025 bp,including four exons and three introns,and its open reading frame(ORF)had a full-length of 582 bp,encoding 193 amino acids.NaLyg was found to be conserved during evolution through bioinformatic analyses.The NaLyg protein possessed a sugar binding domain and three catalytic sites,including Glu71,Asp84 and Asp101.Quantitative qRT-PCR results confirmed that NaLyg gene mRNA was visibly increased after V.harveyi infection.The NaLyg protein purified by prokaryotic expression killed some gram-negative bacterial pathogens by inducing cell wall destruction,including V.harveyi,Aeromonas hydrophila and Edwardsiella tarda.Moreover,the NaLyg protein killed two gram-positive bacteria,Bacillus subtilis and Staphylococcus aureus.Taken together,the experimental results suggested that the NaLyg protein of N.albiflora played an important role in fighting bacterial infections.
基金Supported by Outstanding Graduate Entering Laboratory Project of College of Fisheries,Guangdong Ocean UniversityNational Natural Science Foundation of China(32073015)+1 种基金Undergraduate Innovation Team of Guangdong Ocean University(CCTD201802)Undergraduate Innovation and Entrepreneurship Training Program of Guangdong Ocean University(CXXL2024007).
文摘PhoR is a histidine kinase in a two-component regulatory system that regulates phosphorus metabolic pathways and undertakes the key mission of information transmission in pathogenic bacteria.The full-length phoR gene was successfully cloned from the Vibrio alginolyticus HY9901 strain.A comprehensive analysis of the cloned gene was conducted using bioinformatics.Sequence analysis revealed that the total length of the phoR gene(GenBank accession No.:KJ958404.1)is 1299 bp,with the coding region containing a total of 432 amino acid residues.The phylogenetic tree of PhoR revealed that it belongs to the same subclade as V.diabolicus.The SMART program was employed for the purpose of functional domain prediction,which revealed that PhoR possesses three major functional domains:PAS(amino acids 98-166),HisKA(amino acids 205-272),and HATPase_c(amino acids 317-429).
基金Supported by University Grant Commission(UGC),New Delhi,India.
文摘Objective:To analyse experimental infection and immune system of shrimp(Penaeus monodon)against Vibrios furnissii(V.furnissii).Methods:Experimental animals were collected and acclimatized by maintaining specific temperature,pH and salinity to avoid mortality.Shrimps were experimentally infected with V.furnissii and their immune responses were monitored.After the infection all the shrimps were monitored for any symptoms,death rate in 0,12,24,36,48 h.Then haemolymph were collected and tetrahydrocannabinol,phenol oxidase,nitroblue tetrazolium and lysozyme were monitored in every 12 h at the interval of 48 h.Results:Shrimps infected by live V.furnissii had showed gradual increase in tetrahydrocannabinol,phenol oxidase activity,nitro-blue-tetrazolium and lysozyme activity comparing with the killed and control.Conclusions:The live V.furnissii had showed infection in the shrimp immune system.The live V.furnissii shows infection in experimental shrimps comparing with killed V.furnissii.So the V.furnissii in nature cause the infection in shrimp Penaeus monodon immune system.This report could be applied to control of the infection in shrimp hatchery.
基金supported by Department of Aquatic Animal Health,Faculty of Fishery Sciences,West Bengal University of Animal and Fishery Sciences,West Bengal,India.(FFS/Adm-21/323)
文摘Objective:To study the ecology of antibiotic resistant bacteria with emphasis on sucrose negative vibrios in water and sediments samples of traditional shrimp farming system(bhery)in West Bengal,India.Methods:The vibrios were isolated from traditional shrimp farm samples on thiosulphate citrate bile salt sucrose agar and sucrose negative bacterial strains were used as biomarkers to assess the frequency of antibiotic resistance.Results:The incoming water brought presumptive vibrios ranging from 5.50×10^(1)to 1.00×10^(3)mL in to the bhery,and there appeared to build up vibrios in the culture system with days of culture,as there was about 9 fold increase in vibrios.The levels of vibrios were observed to be moderately higher in outlet water and ranged between 4.15×10^(2)and 4.15x10^(3)mL.The counts of vibrios in pond sediment was found to be 1.00x10^(2)-4.90×10^(3)g;while in inlet(2.00×10^(2)-4.20×10^(4)g)and outlet(3.00×10^(2)-6.85×10^(3)g)their levels were observed to be higher than the pond sediment.Thirteen different Vibrio species were encountered in traditional shrimp culture system and all vibrios were sensitive to chloramphenicol,followed by ciprofloxacin and gatifloxacin(98.24%),gentamicin(95.61%)and other antibiotics.The multiple antibiotic resistance(MAR),i.e.,resistance to at least two antibiotics,was noticed among 43.85%of the sucrose negative vibrios and 41.86%of the sucrose negative non--vibrios.All vibrios harveyi strains exhibited MAR.Although no antibiotic was used in the bhery,the prevalence of MAR in 44%of the sucrose negative vibrios and nonvibrios is a cause of concern.The MAR index was higher in inlet water and sediment samples.The MAR observed in biomarker strains of pond water and sediment(40%)was comparable to those of inlet samples,thus confirming the fact that incoming water was the major source of antibiotic resistant bacteria.Conclusions:It seems that the shrimp culture in bhery does not favour the proliferation and spread of antibiotic resistant bacteria.
基金supported by the Beijing Municipal Natural Science Foundation[7204314]the Medical Innovation Project Foundation[CX19027]the Young Researcher Supporting Program[QNF19066]of the Chinese PLA General Hospital.
文摘Objective This study investigated how the natural phytophenol and potent SIRT1 activator resveratrol(RSV)regulate necroptosis during Vibrio vulnificus(V.vulnificus)-induced sepsis and the potential mechanism.Methods The effect of RSV on V.vulnificus cytolysin(VVC)-induced necroptosis was analyzed in vitro using CCK-8 and Western blot assays.Enzyme-linked immunosorbent assays and quantitative real-time polymerase chain reaction,western blot,and immunohistochemistry and survival analyses were performed to elucidate the effect and mechanism of RSV on necroptosis in a V.vulnificus-induced sepsis mouse model.Results RSV relieved necroptosis induced by VVC in RAW264.7 and MLE12 cells.RSV also inhibited the inflammatory response,had a protective effect on histopathological changes,and reduced the expression level of the necroptosis indicator pMLKL in peritoneal macrophages,lung,spleen,and liver tissues of V.vulnificus-induced septic mice in vivo.Pretreatment with RSV downregulated the mRNA of the necroptosis indicator and protein expression in peritoneal macrophages and tissues of V.vulnificusinduced septic mice.RSV also improved the survival of V.vulnificus-induced septic mice.Conclusion Our findings collectively demonstrate that RSV prevented V.vulnificus-induced sepsis by attenuating necroptosis,highlighting its potency in the clinical management of V.vulnificus-induced sepsis.
基金supported by grants from the National Natural Science Foundation of China[Grant No.81471917]the National Basic Research Priorities Program[Grant 2015CB554201]the Science Foundation for the State Key Laboratory for Infectious Disease Prevention and Control of China[Grant No.2015SKLID509]
文摘Objective This study aimed to investigate whether the VCA0560 gene acts as an active diguanylate cyclase(DGC)in Vibrio cholerae and how its transcription is regulated by Fur and Hap R.Methods The roles of VCA0560 was investigated by utilizing various phenotypic assays,including colony morphological characterization,crystal violet staining,Cyclic di-GMP(c-di-GMP)quantification,and swimming motility assay.The regulation of the VCA0560 gene by Fur and Hap R was analyzed by luminescence assay,electrophoretic mobility shift assay,and DNase I footprinting.Results VCA0560 gene mutation did not affect biofilm formation,motility,and c-di-GMP synthesis in V.cholerae,and its overexpression remarkably enhanced biofilm formation and intracellular c-di-GMP level but reduced motility capacity.The transcription of the VCA0560 gene was directly repressed by Fur and the master quorum sensing regulator Hap R.Conclusion Overexpressed VCA0560 functions as an active DGC in V.cholerae,and its transcription is repressed by Fur and Hap R.
基金supported by the Fish Innovation Team of Shandong Agriculture Research System (No. SDAIT-1206)the Aquatic Animal Immunologic Agents Engineering Research Center of Shandong Province, the Qingdao Agricultural University Doctoral Start-Up Fund (6631122030)+5 种基金the National Natural Science Foundation of China (No. 32002421)the Advanced Talents Foundation of QAU (No. 6651118016)the Natural Science Foundation of Shandong Province (No. ZR2019BC009)the ‘First-Class Fishery Discipline’ program of Shandong Province, the special top talent plan ‘One Thing One Decision (Yi Shi Yi Yi)’the Key Research and Development Program in Shandong Province (No. 2018YFJH0703)Breeding Plan of Shandong Provincial Qingchuang Research Team (2019)
文摘Vibrio scophthalmi and Aeromonas salmonicida can cause high turbot mortality and huge economic losses.Presently,vaccination is the most promising method for preventing communicable diseases.In this study,we used formalin to kill V.scophthalmi and A.salmonicida cells,and mixed with the mineralized oil adjuvant(Montanide^(TM)ISA 763 AVG)to prepare the bivalent inactivated vaccine.The results showed that turbot inoculated with the bivalent inactivated vaccine exhibited strong tolerance to the infection of V.scophthalmi and A.salmonicida,and no obvious clinical symptoms and pathological changes were observed.The activities of enzymes lysozyme,acid phosphatase and complement C3 had significantly increased after the vaccination.The antibody titer response of vaccinated turbot was greatly boosted,which was positively connected with the immunological impact according to ELISA results.Simultaneously,the expression levels of immune-related genes such as MHC-IIα,MHC-IIβ,CD4,CD8,TNF-αand IL^(-1)βwere up-regulated,demonstrating that it might stimulate humoral and cellular immunological response in turbot.These findings highlight the potential of the bivalent inactivated vaccine for controlling V.scophthalmi and A.salmonicida infections in turbot.
基金the National Key R&D Program of China(No.2020YFD0900204)the National Natural Science Foundation of China(No.32072947)+1 种基金the China Agriculture Research System of MOF and MARA(No.CARS-47)the KU-OUC Dual Master’s Program and Ocean University of China Scholarship Council。
文摘The caspase gene family is a crucial gene cluster that regulates apoptosis which contribute to programmed cell death,cell proliferation and differentiation,and several immune responses.In our study,a complete set of 12 caspase genes were identified in spotted sea bass Lateolabrax maculatus.These genes were divided into three subfamilies:2 inflammatory caspases(casp-1 and casp-14-like),5 apoptosis initiators(casp-2,casp-8a,casp-8b,casp-9,and casp-10),and 5 apoptosis executioners(casp-3a,casp-3b,casp-3-like,casp-6,and casp-7).Their phylogenetic relationships,synteny and gene structures were systematically analyzed.Furthermore,the relative expression profiles of the caspase family members in the liver,intestine,head kidney,and spleen were measured by q PCR after infection with Vibrio harveyi.The results showed that the overall mRNA levels of the caspase genes were dramatically increased after V.harveyi infection,and the expression patterns varied among genes and tissues.More caspase genes underwent pronounced expression changes in the head kidney and spleen than in the liver or intestine,mainly after 48 h of the challenge.Specifically,casp-3a,casp-3b,casp-3-like,casp-6,casp-7,casp-8a,casp-8b,casp-10,and casp-14-like in the head kidney,and casp-3-like,casp-6,casp-7,and casp-14-like in the spleen,were the most responsive caspase genes which may contribute significantly to immune regulation in spotted sea bass.Additionally,the apoptosis level in head kidney and spleen after infection were examined using the Caspase assay.Our study provides a systemic overview of the caspase gene family in spotted sea bass after V.harveyi infection and lays a foundation for further deciphering the biological roles of these caspase genes.
基金Supported by the funding for school-level research projects of Yancheng Institute of Technology(No.xjr2019047)the National Natural Science Foundation of China(No.31902362)。
文摘The appropriate reference gene is a prerequisite for accurate normalization of gene expression level,and research on suitable reference genes in clam Cyclina sinensis is scarce.To improve the situation,we selected five commonly used housekeeping genes,including β-actin,Elongation factor 1-α(EF1-α),Glyceraldehyde-3-pho sphate dehydrogenase(GAPDH),40S ribosomal protein S18(RPS18),and Tubulin a(TUB-α),then evaluated their expression stability in different adult tissues and under different experimental treatments(salinity stress and Vibrio parahaemolyticus infection).Their expression stability was analyzed by three frequently used programs,geNorm,NormFinder,and BestKeeper.This analysis indicated that multiple genes should be used for normalization,and we concluded that the reference gene combination GAPDH-RPS18-β-actin,should be used for qRT-PCR analysis in different tissues of C.sinensis under normal physiological conditions.For the clams under salinity stress and Vibrio infection,EF1-α-GAPDHRPS18 was recommended as the gene combination for qRT-PCR normalization.TUB-αwas generally poorly ranked by all programs,and should not be used in future studies.This study should provide fundamental support for accurate quantitative gene expression analysis of this species.
基金supported by the Natural Science Foundation of Shandong Province(No.ZR2020QC214)the Young Experts of Taishan Scholars(No.tsqn201909130)+3 种基金the Science and Technology Support Plan for Youth Innovation of Colleges and Universities in Shandong Province(No.2019KJF003)the‘First Class Fishery Discipline’Programme in Shandong Provincea special talent programme‘One Thing One Decision(YishiYiyi)’Programme in Shandong Province,Chinathe Breeding Plan of Shandong Provincial Qingchuang Research Team(2019)。
文摘Neuronal nitric oxide synthase(nNOS)was the producer of nitric oxide(NO)which played important gas messenger molecules in biological process.It also can take effect as immune regulation molecule in organism.Black rockfish(Sebastes schlegelii)is an important economic fish which were widely farmed in East Asia countries.Meanwhile,the pathogenic bacteria such as the Edwardsiella tarda and Vibrio anguillarum in seawater always brought serious obstacles to their healthy growth.In order to explore the expression pattern of n NOS gene under the pathogen stimulation and predict its immune function,the n NOS gene in black rockfish named Ssn NOS was identified.It was 3780 bp in length,located on chromosome 6,and contained 27 coding domain sequence(CDs).According to the phylogenetic analysis,the Ssn NOS showed closest relative to the counterpart gene of swamp eel(Monopterus albus).Meanwhile,analysis of Ssn NOS expression in various healthy tissues showed that Ssn NOS expression level was highest in healthy brain tissues,followed by intestinal tissues.In addition,Ssn NOS showed significant expression changes in response to stimulation by two pathogens.Particular in gill,the expression of Ssn NOS after pathogenic stimulation increased significantly.The Elisa analysis showed the Ssn NOS content in gills was much higher than that in other tissues at all time points.Moreover,the expression patterns of Ssn NOS in brain,intestine and kidney after stimulation by pathogens showed a distinct expression pattern which first down-regulated and then up-regulated.Therefore,the Ssn NOS may be an important signaling molecule for fish to respond rapidly in immune stimulation.
基金the Zhejiang Provincial Natural Science Foundation for Distinguished Young Scholars(No.LR20C190001)the National Natural Science Foundation of China(No.31972833)+1 种基金the Fundamental Research Funds for the Provincial Universities of Zhejiang(No.SJ LZ2020001)the K.C.Wong Magna Fund at Ningbo University。
文摘Vibrio splendidus is an important opportunistic pathogen ubiquitously present in the marine environment,exhibiting virulence to a variety of cultured animals.The extracellular products secreted by V.splendidus are crucial to bacterial survival and virulence.In this study,the secretion of outer membrane vesicles(OMVs)by V.splendidus was determined,purified,and morphologically characterized.The protein composition of OMVs was analyzed by proteomic analysis.The results showed that approximately 120 proteins were contained in these OMVs,including outer membrane proteins,flagellins,ABC transporters,protease,and iron regulation proteins,etc.,which were involved in bacterial motility,formation of biofilms and the cell membrane components,and cellular localization based on their structural molecule activity,passive transmembrane transporter activity,channel activity,neurotransmitter receptor activity,extracellular ligand-gated ion channel activity,glutamate receptor activity,ligand-gated ion channel activity,and transmembrane signaling receptor activity.To explore the biological functions of OMVs in V.splendidus,the effects of OMVs on the bacterial adaption to iron limitation,antibiotic,and the coelomic fluid of the Apostichopus japonicus were confirmed.This study is the first time to show that V.splendidus secretes OMVs,and OMVs carry functional proteins that enhance bacterial survival under various stresses.
基金Supported by the Natural Science Foundation of Hainan Province (Nos. 320QN212, 2019RC078)the Key Research Project of Hainan Province (No. ZDKJ2019011)
文摘Leopard coral groupers(Plectropomus leopardus),commercially bred in South China,are a significant economical fish species.In this study,by means of quantitative real-time PCR(qRT-PCR)technology,we assessed the stability of six common reference genes expression and selected the appropriate reference genes in leopard coral groupers with or without Vibrio harveyi stimulation at different time points.These data produced by qRT-PCR was handled via the geNorm,NormFinder,and BestKeeper software.The results revealed all the examined reference genes had manifest tissue-specific expression in different tissues.Prior to V.harveyi stimulation,RPL13 gene was the appropriate reference gene among eleven tissue types(blood,spleen,hepatopancreas,kidney,stomach,gill,heart,skin,muscle,intestine,brain)in leopard coral groupers.Under V.harveyi stimulation,the most reliable reference genes varied from tissue to tissue and were closely hinged upon different time points.At 6-h post-bacterial injection,the appropriate reference genes in hepatopancreas,spleen,kidney,and gill were Actin,B2M,UBCE,and Actin,respectively.At 9-and 12-h post-bacterial injection,the appropriate reference genes in hepatopancreas,spleen,kidney,and gill were RPL13,Actin,Actin,and Actin,respectively.If one reference gene was preferable,RPL13,Actin,Actin,and Actin could be selected as the reference gene in hepatopancreas,spleen,kidney,and gill of leopard coral groupers after V.harveyi infection,respectively.Expression profiles of two target genes(IL-6 and NK-lysin)were used to further validate reliability of these selected appropriate candidates.This study will lay a solid foundation for the future research on qRTPCR analysis of gene expression in leopard coral groupers.