海洋菌株Aeromonas sp.YS-54来源几丁质酶的制备与性质研究

为实现甲壳类原料的高效降解与N-乙酰氨基寡糖的绿色制备,本研究以青岛近海土壤中筛选的菌株YS-54为材料,使用60%硫酸铵沉淀、Sephadex G-100凝胶过滤层析制备几丁质酶;对几丁质酶的最适温度、最适pH、底物偏好性等性质进行表征;使用质谱技术确认了酶解产物的聚合度。结果表明,菌株YS-54为Aeromonas属,该菌株来源几丁质酶比活力为23.44 U/mg,分子量为63与75 kDa。几丁质酶在40℃和pH为5.0条件下具有良好的稳定性,Ba^(2+)、Co^(2+)、Mg^(2+)与TritonX-100显著促进了几丁质酶活力(P<0.05),而Fe^(3+)、Cu^(2+)、SDS对酶活力起抑制作用。几丁质酶对α-几丁质的比活力为7.99 U/mg,为水解胶体几丁质比活力的34.09%。几丁质酶对胶体几丁质与α-几丁质的酶解产物聚合度分别为1~4与1~3。Aeromonas sp.YS-54来源几丁质酶在较宽的温度与pH范围内保持稳定,同时对α-几丁质具有良好的水解能力,可为甲壳类废弃物的高效综合利用提供技术支持。

Model prediction of inactivation of Aeromonas salmonicida grown on poultry in situ by intense pulsed light

The aim of this study was to evaluate the factors influencing the inactivation effect of intense pulsed light(IPL)on Aeromonas salmonicida grown on chicken meat and skin,and to further develop prediction models of inactivation.In this work,chicken meat and skin inoculated with meat-borne A.salmonicida isolates were subjected to IPL treatments under different conditions.The results showed that IPL had obvious bactericidal effect in the chicken skin and thickness groups when the treatment voltage and time were 7 V combined with 5 s.In addition,the lethality curves of A.salmonicida were fitted under IPL conditions of 3.5-7.5 V.The comparison of statistical parameters revealed that the Weibull model could best fit the mortality curves and could accurately predict the mortality dynamic of A.salmonicida grown on chicken skin.And further a secondary model between the scale factor b and the treatment voltage in Weibull model was established using linear equations,which determined that the secondary model could accurately predict the inactivation of A.salmonicida.This study provides a theoretical basis for future prediction models of Aeromonas,and also provides new ideas for sterilization approaches of meat-borne Aeromonas.

Isolation,identification,and virulence gene analysis of pathogenic Aeromonas dhakensis in Macrobrachium rosenbergii and histopathological observation

To identify the cause of mass mortality of adult Macrobrachium rosenbergii in a farm in Gaoyou City,Jiangsu Province,China,a dominant strain named DKQ-1 was isolated from the hepatopancreas of dying M.rosenbergii and identified as Aeromonas dhakensis by purification culture,biochemical characterization,and 16S rRNA and gyrB gene sequence analysis.The results of the challenge test revealed that the strain was highly pathogenic and the 50%lethal dose(LD_(50))in 72 h to M.rosenbergii was 1.54×10^(5)CFU/mL.The amplification results of virulence genes show that strain DKQ-1 carried 9 virulence genes,including ascV,aexT,aer,act,lip,ompAI,gcaT,acg,and exu,supporting the strong virulence of strain DKQ-1 to M.rosenbergii.Histopathological observation of the hepatopancreas,gills,and intestines indicated that DKQ-1 injection into M.rosenbergii could cause serious tissue damage,which further supported the strong virulence of this strain.In addition,a drug susceptibility test revealed that strain DKQ-1 was sensitive to 16 kinds of antibiotics,resistant to 9 kinds of antibiotics,and had intermediate resistance to spectinomycin and kanamycin.This study is the first report of A.dhakensis isolated from M.rosenbergii and provided a reference for the pathogen identification of bacterial diseases in M.rosenbergii,and for the prevention and treatment caused by A.dhakensis.

Cloning and Bioinformatics Analysis of hcp Gene in Aeromonas hydrophila

[Objectives]To explore the function of hcp gene in Aeromonas hydrophila.[Methods]A pair of specific primers was designed referring to the hcp gene sequence of A.hydrophila.The hcp gene was amplified by PCR,and performed bioinformatics analysis.[Results]The hcp gene had a total length of 1650 bp and encoded 549 amino acids.The theoretical molecular weight of the protein predicted was about 59476.44 kDa.After predicting the N-terminal signal peptide structure of the amino acid sequence,neither obvious signal peptide cleavage site nor signal peptide was found,and the protein had no transmembrane region.The amino acid sequence had a N-glycosylation site,4 protein kinase C phosphorylation sites,7 casein kinase II phosphorylation sites,9 N-myristoylation sites,4 isoprene binding sites,10 microbody C-terminal target signal sites,and an ATP/GTP binding site motif A(P-ring).The amino acid sequence of hcp gene of A.hydrophila was performed homology analysis with other Aeromonas strains,and it showed higher homology with A.veronii.In the secondary structure,theα-helix,β-sheet,random coil and extended strand accounted for 45.36%,6.01%,37.52%and 11.11%,respectively.The tertiary structure model consisted of 18α-helix and 22β-sheet.Analysis of protein-protein network interaction demonstrated that the proteins interacting with Hcp protein were AHA_3407,nrfA,nirB-1,nirB-2 and AHA_1112.[Conclusions]Through the bioinformatics prediction results,the basic information of hcp gene of A.hydrophila is preliminarily understood,and the possible function of this protein is predicted,in order to provide guidance for subsequent vaccine research.

Isolation,Identification and in vitro Antimicrobial Susceptibility of Pathogenic Aeromonas veronii from Soft-shelled Turtles

In order to effectively control diseases caused by Aeromonas veronii,pathogenic bacteria were isolated and incubated from infected soft-shelled turtles with traditional bacterial isolation method. Four strains of pathogenic bacteria were isolated and identified with traditional biochemical identification method and modern molecular biological identification techniques. According to the results, four strains of pathogenic bacteria were identified as A. veronii biovar sobria. Drug sensitivity test and in vitro antimicrobial test against Bacillus amyloliquefaciens were performed with agar diffusion method. The results showed that B. amyloliquefaciens and several antibiotics such as ofloxacin and gentamicin exerted strong antimicrobial effects on four isolates. B. amyloliquefaciens could be used for the prevention and control of diseases caused by A. veronii in aquaculture.

罗氏沼虾(Macrobrachium rosenbergii)致病性豚鼠气单胞菌(Aeromonas caviae)的分离鉴定及其特性分析

近年来细菌性疾病因病原菌种类繁多、覆盖区域广而影响了其养殖业的健康发展,造成较大的经济损失。从体表具有黑斑、肢体残损症状的罗氏沼虾(Macrobrachium rosenbergii)肝胰腺组织中分离得到1株病原菌,通过形态学观察、生理生化特性鉴定、16S rRNA序列和特定毒力基因检测鉴定,以及人工回归感染试验和药敏试验分析其致病性和耐药性。结果表明,该菌为豚鼠气单胞菌(Aeromonas caviae),含有aer、lip、gcat、ser和OMPA I等毒力基因;药敏试验显示其对头孢曲松、氯霉素等7种抗生素高度敏感,对恩诺沙星、阿米卡星及复方新诺明3种抗生素中度敏感,对四环素、环丙沙星等10种抗生素具有耐药性;人工回归感染实验表明该菌具有较高的致病性,对罗氏沼虾的半致死浓度为6.54×10~5 CFU/mL。豚鼠气单胞菌作为水产养殖中的条件性致病菌,研究结果揭示了该菌株的部分生物学特性,有利于丰富豚鼠气单胞菌属的基础数据,同时也为罗氏沼虾养殖过程中的病害防控提供了一定参考依据。

凡隆气单胞菌(Aeromonas veronii)拮抗菌4-1-3的筛选鉴定及其活性物质初探

凡隆气单胞菌是泥鳅养殖中容易发生而且致死率较高的病原菌之一。为筛选凡隆气单胞菌拮抗菌、研发防治此病害的生物制剂,采集泥鳅养殖池底泥样品,通过滤纸抑菌圈法筛选凡隆气单胞菌拮抗细菌,结合形态观察、生理生化实验和16S r RNA基因序列分析进行菌株鉴定。采用硫酸铵沉淀和离子交换层析方法分离纯化抑菌物质,电泳检测其分子量。结果显示,从样品中分离到17株细菌,其中筛选出的拮抗菌4-1-3,能够高效抑制凡隆气单胞菌,抑菌圈直径达到15.2 mm。经菌株鉴定后命名为Pseudomonas fluorescens 4-1-3。该菌产生的抑菌物质为胞外蛋白,分子量约为100 k Da,推测该物质是荧光假单胞菌产生拮抗物质的一种新类型,有望进一步研制成防治水产病害的生物制剂。

pH和NaCl质量浓度对发酵鱼糜中腐败菌Aeromonas veronii bv. veronii群体感应的影响

细菌通过群体感应(Quorum sensing,QS)系统来调控自身一些代谢行为,如生物膜的形成、鞭毛运动等,可引起食品腐败和食源性疾病,分析环境因素对细菌QS的影响对控制水产品质量安全具有重要意义。该研究以从发酵鱼糜中分离的腐败菌Aeromonas veronii bv.veronii为研究对象,利用生物报告菌根癌农杆菌及激光共聚焦显微镜等技术分析了pH(6.0、7.0、8.0)和Na Cl(5、10、20 g/L)对细菌N-酰基高丝氨酸内酯(N-acyl homoserine lactone,AHL)类信号分子介导的QS及代谢行为的影响。研究发现,培养基初始pH 8.0或NaCl质量浓度为20g/L时能显著降低细菌产生的AHLs活性、生物膜形成能力及泳动能力(p<0.05)。由此认为,通过控制环境中的pH和Na Cl质量浓度能影响细菌合成AHLs及其QS系统的表达,并调控细菌的代谢行为。

Aeromonas veronii biovar veronii and sepsis-infrequent complication of biliary drainage placement: A case report

BACKGROUND Aeromonas species are uncommon pathogens in biliary sepsis and cause substantial mortality in patients with impaired hepatobiliary function. Asia has the highest incidence of infection from Aeromonas,whereas cases in the west are rare.CASE SUMMARY We report the case of a 64-year-old woman with advanced pancreatic cancer and jaundice who manifested fever,abdominal pain,severe thrombocytopenia,anemia and kidney failure following the insertion of a percutaneous transhepatic biliary drainage. Blood culture results revealed the presence of Aeromonas veronii biovar veronii(A. veronii biovar veronii). After antibiotic therapy and transfusions,the life-threatening clinical conditions of the patient improved and she was discharged.CONCLUSION This was a rare case of infection,probably the first to be reported in West countries,caused by A. veronii biovar veronii following biliary drainage. A finding of Aeromonas must alert clinician to the possibility of severe sepsis.

嗜水气单胞菌(Aeromonas hydrophila)引起斑马鱼肠道生理损伤和肠道菌群失调

研究分析了嗜水气单胞菌(Aeromonas hydrophila)感染斑马鱼后肠道生理健康和肠道微生物的变化。以斑马鱼为研究对象,刮伤皮下真皮,使用10^(5)CFU/mL浓度的嗜水气单胞菌暴露6 h后转入清水,分别在暴露前、暴露后6 h、12 h、24 h取样。使用鱼特异性酶联免疫吸附试验(ELISA)试剂盒,检测肠道中紧密连接蛋白ZO-2(TJP2)含量和乙氧基异戊二烯-O-脱乙基酶(EROD)酶活性,结果表明,嗜水气单胞菌暴露引起了斑马鱼肠道生理损伤,表现为肠道TJP2含量、EROD酶活性在暴露后显著下调。采用16S rRNA高通量测序检测肠道微生物及其菌群结构变化,嗜水气单胞菌暴露斑马鱼后致病菌不动杆菌属(Acinetobacter)明显增加,OTUs数量明显下降,Alpha多样性降低,结果说明嗜水气单胞菌引起了斑马鱼肠道菌群失调。肠道中微生物的多样性与改善肠道上皮和黏膜屏障功能紧密相关,该研究对从肠道健康的角度评价嗜水气单胞菌毒性,探索嗜水气单胞菌的致病机制具有重要意义。

Characterization and Expression of Outer Membrane Protein A I Gene of Aeromonas veronii

The outer membrane protein, ompA, ofAeromonas veronii has a role in the virulence of the organism and is a potential candidate for vaccine development. In this study, ompA I ofAeromonas veronii strain WA106 was cloned and sequenced, then, it was expressed in Escherichia coli BL21. The nucleotide sequence of ompA I gene was 1 023 base pairs （GenBank Accession NO.KC748024）, which showed 100% homology with that of A. veronii （NO.AB290200.1）. This predicted protein was composed of 340 amino acid residues. Its molecular weight was 35.78 ku and isoelectric point was 5.18. The protein was a hydrophilic protein containing alpha helix and random coil with percentage of 35.0% and 49.7%, respectively. The tertiary structure, quaternary structure prediction showed that ompA I protein contained two peptide chains. SDS-PAGE showed that the actual value of the fusion protein was consistent with the expected result. It will facilitate further study of the role of ompA I protein.

Protective efficacy evaluation of immunogenic protein AHA_3793 of Aeromonas hydrophila as vaccine candidate for largemouth bass Micropterus salmoides

Aeromonas hydrophila is a Gram-negative pathogen that can infect various fish,including largemouth bass(Micropterus salmoides),which have caused huge economic losses.In present study,largemouth bass anti-A.hydrophila antibodies were produced,then a highly immunogenic outer membrane proteins,AHA_3793,was identified by combined western blotting and mass spectrometry analysis.Moreover,AHA_3793 was expressed,and its immunogenicity was further verified by western blotting.Subsequently,the protective efficacy of AHA_3793 were evaluated in largemouth bass.The results showed that rAHA_3793 could produce a relative percentage survival(RPS)of 61.76% for largemouth bass against A.hydrophila challenge.ELISA analysis showed the specific serum antibodies of largemouth bass against rAHA_3793 and A.hydrophila in vaccinated group in weeks 4 and 5 after immunization were significantly higher than those in control group,which suggested that rAHA_3793 induced production of specific serum antibodies against rAHA_3793 and A.hydrophila.The qRT-PCR analysis showed that expressions of CD4-2 and MHC Ⅱα were also significantly up-regulated after immunization.These results collectively demonstrated that rAHA_3793 could induce a strong humoral immune response of largemouth bass,and then produce high immune protection ef fects against A.hydrophila infection.

Immuno-Protective Efficiency of the Bivalent Inactivated Vaccine Against Vibrio scophthalmi and Aeromonas salmonicida Infections in Turbot(Scophthalmus maximus L.)

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.

Aeromonas hydrophila infection in acute myeloid leukemia: A case report

Rationale:Aeromonas hydrophila,a bacterium commonly found in a variety of soils,aquatic habitats,and other natural environments,is known for causing infections and hemorrhagic diseases in both aquatic and terrestrial animals.Patients with leukemia,when infected by Aeromonas hydrophila following chemotherapy,face an increased risk of developing necrotizing fasciitis.Patient concerns:A 48-year-old male with a history of acute myeloid leukemia presented with swelling and pain in his right hand after being stabbed by a grass carp two days ago.The injury occurred on the fifth day after his last chemotherapy session when he was accidentally punctured in the right thumb and ring finger by a carp while fishing.Two days following the injury,the skin isurrounding the wound turned black immediately,while the rest of his hand was reddened,swollen,and bleeding.Diagnosis:Aeromonas hydrophila infection in the soft tissue of the right hand.Interventions:Intravenous antibiotics and local debridement.Outcomes:The patient experienced an alleviation of systemic poisoning symptoms,effective control of the local wound infection.Lessons:It is important for physicians to appreciate the potential for highly unusual and life-threatening infections in patients with acute myeloid leukemia.Early diagnosis and prompt treatment can prevent the development of necrotizing fasciitis and save patients’lives.

气单胞菌属低温淀粉酶基因改造与原核表达

【目的】获取低温淀粉酶基因,分析其相关功能,为工业生产上的应用提供参考。【方法】以气单胞菌(Aeromonas)LA77为出发菌株克隆低温α-淀粉酶基因,以BL-21(DE3)为宿主菌进行克隆。分析其它已知气单胞菌属低温α-淀粉酶基因同源性,设计特异引物,PCR扩增获得C13片段,将其克隆到pMAL-2X,转化到BL-21(DE3)中,筛选蓝白斑,验证PCR及EcoRⅠ和HindⅢ的双酶切,获得高效表达生物工程菌株pMAL-2X-C13。设计定点突变引物,以pMAL-2X-C13为模板,获得低温淀粉酶基因突变株三株C19、C29、C43。【结果】表达蛋白的分子大小为114kD,突变菌株C19蛋白表达量均高于pMAL-2X-C13。【结论】低温淀粉酶基因突变株C19比原始菌株pMAL-2X-C13淀粉酶基因蛋白表达量更高。

维氏气单胞菌拮抗菌株J2-2的筛选、鉴定与效果评价

为筛选对维氏气单胞菌(Aeromonas veronii)有潜在抑菌作用的菌株,本研究从湖北公安县泥鳅养殖池塘底泥中分离一株细菌J2-2,对其特性进行了研究,并对抑菌活性物质进行了初步分析。结果显示:菌株J2-2为解淀粉芽孢杆菌(Bacillus amyloliquefaciens),具有抑菌广谱性,对米尔伊丽莎白菌(Elizabethella milieri)、嗜水气单胞菌(A.hydrophila)等6种水产病原菌的抑菌直径在9.6~29.5 mm。菌株J2-2的抑菌物质具有较好的酸碱稳定性、热稳定性和蛋白酶K处理的稳定性。电镜结果显示菌株J2-2的发酵产物对维氏气单胞菌的细胞膜结构具有破坏作用。通过LC-MC分析该拮抗细菌可产生至少15种不同的脂肽类化合物如杆菌霉素等。药敏试验结果显示,菌株J2-2对氨苄西林、青霉素具有耐药性,对克林霉素中度敏感,对大多数抗生素高度敏感。

斑鳜肠道组织对嗜水气单胞菌感染早期的免疫响应

【目的】嗜水气单胞菌是危害斑鳜养殖生产的主要病原之一。研究斑鳜感染嗜水气单胞菌后,其肠道组织基因表达水平的变化,解析鱼体对细菌性感染的免疫应答分子机制,为防控淡水鱼类细菌性疾病提供依据。【方法】用嗜水气单胞菌感染斑鳜6 h后,利用Illumina Hiseq 6000测序平台对斑鳜肠道组织进行RNA-seq测序,并利用生物信息学方法对数据进行分析。【结果】感染组和未感染组斑鳜肠道组织存在1077个差异表达基因(DEGs),包括669个上调DEGs和408个下调DEGs。其中,与免疫相关的上调DEGs主要有基质金属蛋白酶3(MMP3)、白细胞介素-1β(IL-1β)、白细胞介素-17F(IL-17F)、正五聚蛋白3(PTX3)、核苷酸结合寡聚化结构域样受体(NLR)含pyrin结构域蛋白6(NLRP6)、C型凝集素域家族4E(CLEC4E)、胆固醇-25-羟化酶(CH25H)和激活转录因子1(ATF1)等。基因本体(GO)富集分析显示,DEGs主要富集在免疫反应激活信号通路、免疫应答调节细胞表面受体信号通路、T细胞活化的正调控过程以及细胞修复和调节过程上。京都基因与基因组百科全书(KEGG)富集分析显示,上调DEGs主要富集在多种疾病发生、发展过程和免疫代谢相关信号通路上。实时荧光定量PCR验证结果表明,7个随机选取的与免疫相关的DEGs的表达量与RNA-seq测序结果存在一致性。【结论】感染组与未感染组斑鳜肠道组织MMP3、IL-1β、PTX3、NLRP6、CLEC4E表达量的差异较为显著,可能是参与斑鳜肠炎发生的关键候选基因。

ExsA对维氏气单胞菌致病性的影响

维氏气单胞菌是一种重要的人鱼共患病原菌,可以引起多种水产生物的大量死亡,对水产养殖业以及公共卫生安全有重大危害。Ⅲ型分泌系统(typeⅢsecretion system,T3SS)是决定气单胞菌属致病性最关键的毒力机制之一,AraC家族转录因子ExsA作为主调控因子,响应环境条件变化,激活T3SS组装及效应蛋白分泌。为了探究ExsA对维氏气单胞菌致病性的影响,通过同源重组策略构建exsA基因敲除菌株,并检测其生物膜形成能力、黏附性及培养后上清的细胞毒性。结果显示,维氏气单胞菌中exsA基因缺失导致其生物膜形成能力、对宿主离体组织的黏附能力以及培养后上清液的细胞毒性显著下降。与野生型相比,exsA敲除株形成生物膜的能力降低1.5倍,对上皮细胞的黏附能力无显著性变化,但对小鼠离体肠道组织的黏附能力降低3.4倍,细菌培养后上清的细胞毒性下降1.2倍。研究结果表明ExsA对于维氏气单胞菌的毒力机制有重要调控作用,为进一步阐明T3SS对于病原菌的致病机制以及指导水生生物微生物感染防治策略奠定坚实基础。

七彩神仙鱼BTN1A1基因克隆及其在嗜水气单胞菌胁迫下的表达分析

【目的】克隆七彩神仙鱼(Symphysodon aequifasciatus)嗜乳脂蛋白1A1(Butyrophilin 1A1,BTN1A1)基因,分析其在病原刺激下的表达模式,为了解七彩神仙鱼BTN1A1基因功能提供依据。【方法】利用生物信息学对七彩神仙鱼2个BTN1A1基因(BTN1A1-1和BTN1A1-2)进行结构和进化分析。根据前期获得的七彩神仙鱼皮肤转录组数据,选取BTN1A1-1和BTN1A1-2基因的CDS区设计引物进行克隆。采用qRT-PCR分析BTN1A1在各组织中的表达及嗜水气单胞菌刺激下的表达模式。【结果】BTN1A1-1和BTN1A1-2的ORF序列长度为894、1275 bp,分别编码298、424个氨基酸。BTN1A1-1和BTN1A1-2蛋白均具有1个信号肽、1个Ig结构域、1个lg_like结构域和1个跨膜结构域的经典结构,BTN1A1-2还具有1个胞质结构域。七彩神仙鱼BTN1A1-1与慈鲷科其他鱼类相似性较高,与尼加拉瓜湖始丽鱼(Archocentrus centrarchus)BTNIAI对应氨基酸序列同源性最高、为94.14%。但BTN1A1-2与其他鱼类发生分离,形成独特分支。BTN1A1-1和BTN1A1-2在七彩神仙鱼各组织中均有表达,但在鳃、肠道、皮肤等与免疫相关的组织中表达量相对较高。七彩神仙鱼BTN1A1-1在嗜水气单胞菌胁迫后表达量先显著下降后上升,而BTN1A1-2表达量则先显著上升后下降。【结论】七彩神仙鱼BTN1A1-1基因相对保守,而BTN1A1-2基因在进化上较为特殊。二者在免疫刺激下的差异表达暗示其可能在七彩神仙鱼免疫防御中发生功能分化。

维氏气单胞菌弱毒株FS12001及其疫苗对异育银鲫的免疫效果

为探究由维氏气单胞菌(Av)弱毒株FS12001制备的疫苗免疫效果,将Av弱毒株FS12001制备活疫苗、灭活疫苗、ISA763A-灭活疫苗和蜂胶-灭活疫苗,腹腔注射免疫异育银鲫,同时设ISA763A佐剂、蜂胶佐剂及0.65%NaCl作为对照组。于免疫后1、3、5、7和14 d经实时荧光定量PCR(qRT-PCR)检测脾脏组织中IgM、IL-1β及LZM的基因表达量;于免疫后7、14、21、28、42和48 d尾静脉采血分离血清,检测IgM抗体水平,测定SOD和LZM酶活性;于免疫28 d后用2株Av强毒株分别攻毒各组实验鱼,连续观察记录7 d,计算各免疫组的相对保护率。结果显示,各免疫组脾脏组织中IgM、IL-1β及LZM的基因表达量高于对照组,且ISA763A-灭活疫苗组和蜂胶-灭活疫苗组的表达量显著高于其他各实验组。各疫苗免疫组血清特异性抗体水平均显著高于0.65%NaCl对照组和佐剂对照组。各疫苗免疫组的LZM活性均在28 d最高,灭活疫苗组、ISA763A-灭活疫苗组及蜂胶-灭活疫苗组SOD活性均在7 d最高。菌株AVCA07攻毒后,活疫苗组、灭活疫苗组、ISA763A-灭活疫苗组和蜂胶-灭活疫苗组的RPS分别为63%、56%、100%和56%;菌株YC170511攻毒后,以上4个免疫组RPS分别为59%、55%、93%和72%。研究表明,用弱毒株FS12001制备疫苗,注射免疫异育银鲫后均可增加脾脏中IgM、IL-1β及LZM的基因表达量,提高血清抗体水平,增强SOD和LZM酶活性,增强其抵抗Av感染的能力。