杂色鲍哈维弧茵耐药质粒的鉴定和分析

文章对引起杂色鲍（Hallotis diversicolor）肌肉萎缩症的病原菌哈维弧菌（Vibrio harveyi）质粒上的磺胺耐药基因进行研究,结果显示,该菌株对复方新诺明完全耐药,其质粒上含有sulⅡ耐药基因。接合转化试验显示该质粒具可转移性,可使受体菌对磺胺制剂复方新诺明产生耐药性,鉴定为耐药质粒,并测定了该耐药质粒的全基因组序列,序列总长为10 940 bp,初步分析显示有7个具有一定功能的ORF框。进一步构建重组表达质粒pR-SET-A-sulⅡ,表达了目的蛋白（31 kDa）。     

南蛇藤果实提取液对弧菌的体外抑菌作用

用平板牛津杯法和试管二倍稀释法比较南蛇藤乙酸乙酯相、水相、乙醇相3种提取液对弧菌的抑菌活性和最小抑菌浓度(MIC)。探讨南蛇藤果实提取液对鳗弧菌、副溶血弧菌的体外抑菌作用。结果表明:3种提取液的MIC值分别是145.8、208.32、91.7 mg/mL(鳗弧菌);125.01、87.52、08.33 mg/mL(副溶血弧菌)。南蛇藤果实的3种提取液对鳗弧菌、副溶血弧菌有不同程度的抑制作用,其中乙酸乙酯相提取液抑菌效果最佳,水相和乙醇相次之。     

基于toxR基因的SYBR GreenⅠ实时定量PCR检测病原副溶血弧菌方法的建立

基于副溶血弧菌toxR基因保守序列设计1对特异性引物,建立了SYBR GreenⅠ实时定量PCR检测副溶血弧菌的方法。常规PCR检测,副溶血弧菌扩增出大小为368bp的目的片段,其他4种病原细菌均为阴性;SYBRGreenⅠ实时定量PCR扩增曲线反映了PCR的指数增长阶段和平台阶段;在Tm为85℃,扩增产物的熔解曲线分析只出现1个单特异峰,无引物二聚体;所制作的标准曲线在2.7×108～2.7×102拷贝数之间有较好的线性关系,相关系数为0.998,能对副溶血弧菌进行准确的定量分析;该方法检测时间从核酸抽提到结果分析仅需4～5h,较传统方法敏感、操作简单,耗时短,是副溶血弧菌引起的水产动物疾病的快速诊断及食品中针对副溶血弧菌安全检测的有效手段。     

对虾养殖场底泥中副溶血性弧菌的接合型质粒多样性及其与毒力之间的关系

为了阐明引起急性肝胰腺坏死病(acute hepatopancreatic necrosis disease, AHPND)副溶血性弧菌的接合型质粒在对虾养殖环境中的遗传多样性,实验从中国5个沿海省份的虾场收集了100个底泥样品,以质粒上编码接合转移蛋白的保守基因为目标,利用PCR法检测相关质粒的存在情况,并对质粒进行测序。结果显示,100个样品中有39个样品含有质粒的接合转移蛋白片段。从100个底泥样品中分离出15株副溶血性弧菌,其中13株含有1～2个质粒。质粒序列测序结果显示,这些质粒可分为8种类型/谱型,其中7种不携带pirAB,但均含有编码接合转移的基因簇。根据分离副溶血性弧菌携带质粒的8种谱型,分别选择8株副溶血性弧菌进行凡纳滨对虾攻毒实验,发现这些菌株对凡纳滨对虾的毒性有显著差异,实验虾死亡率为15%～100%。只有pirAB阳性菌株会对实验虾产生AHPND症状,死亡率为100%。对质粒组成进行分析表明,质粒之间遗传物质交换频繁,大部分质粒的遗传组成都来自一个183 kb的超大质粒pVP2HP。综上,本实验通过探究对虾养殖场底泥中结合性质粒的多样性,增强了人们对副溶血性弧菌...     

Serological and immunological studies of pleuropneumonia of swine caused by Haemophilus parahaemolyticus

The development of circulating antibodies for H. parahaemolyticus was studied in experimentally infected SPF pigs and in-contact SPF pigs. Blood serum titers were determined by a modified complement fixation test with normal SPF swine serum as a source of supplementary complement factor, and by an indirect haemagglutination test.CF and IHA titers became positive within the first 2 weeks following exposure to H. parahaemolyticus, and reached peak values after 2 to 7 weeks (Figs. 1 to 3). The exposed pigs proved immune, in that they showed no clinical symptoms on challenge after resp. 6, 9 and 11 weeks.While distinct titers were thus obtained with both tests in SPF swine experimentally exposed to H. parahaemolyticus, the CF test proved more specific than the IHA test when the 2 tests were compared in a field outbreak of polyserositis (Glässers disease) caused by H. parasuis. The CF test would therefore seem to be preferable to the IHA test in field diagnostic work (Table 1).A noticeable finding was that challenge did not elicit an anamnestic antibody response in any of the immune pigs (Figs. 1 to 3). This fact together with negative bacteriological findings in the animals in question would seem to suggest that the challenge dose was unable to establish a permanent infection in the respiratory tract of the immune pigs.     

养殖半滑舌鳎腹水病的病原分离鉴定及药物敏感性分析

从患腹水病的养殖半滑舌鳎腹水中分离到优势生长的细菌,人工感染试验证明,其中一株细菌(HST2)为致病菌。通过细菌的形态学特征、培养特性、生理生化特性及16S rRNA基因序列分析对病原菌进行鉴定,结果表明,HST2为副溶血弧菌。应用50种抗菌药物对HST2进行药物敏感性分析,结果显示,HST2对氨曲南等43种药物高度敏感,对林可霉素敏感,对青霉素G、氨苄青霉素、羧苄青霉素、苯唑青霉素、杆菌肽、阿洛西林6种药物不敏感。     

Study on the Quantity Dynamic Changes of Heterobacteria and Vibrios in Larvae Industrialized Culture System

[Objective] The dynamic change of heterobacteria and vibrios in larvae industrialized culture system was studied to provide scientific reference for healthy cultivation of shrimp.[Method] The heterobacteria,vibrios and pathogenic vibrio parahaemolyticus were monitored in larvae industrialized culture system.[Result] The heterobacteria,vibrios and pathogenic vibrio parahaemolyticus were the most in fertilized eggs of shrimp but the least in nauplius,then their number would increase with growth. During whole rearing period,both heterobacteria in larvae,vibrios in water would increase by one order of magnitude,while both vibrios in larvae and heterobacteria in water would increase by two orders of magnitude.There were many heterobacteria and vibrios but few vibrio parahaemolyticus in living bait.The correlation coefficients between larvae and heterobacteria and vibrios in water were 0.704 and 0.840 in culture system respectively,while the correlation among heterobacteria,vibrios in living bait and larvae,water were weak or negative.[Conclusion] There was a dynamic relation between water and larvae in rearing period,and restrictly control of culture condition would restrain the occurrence of disease caused by vibrio parahaemolyticus,besides that bacteria number in bait was not obviously correlated with bacteria nubmer in culture system.     

22味中草药对创伤弧菌和灿烂弧菌体外抑菌作用筛选

研究了22味中草药对创伤弧菌(Vibrio vulnificus)和灿烂弧菌(V.splendidus)的体外抑菌作用。结果显示,22味中草药对其均有不同程度的抑制作用;其中,黄连和连翘对创伤弧菌的抑菌和杀菌效果最好,最小抑菌浓度分别为0.49mg/mL和1.95mg/mL,最小杀菌浓度与最小抑菌浓度相同;黄连、黄芩、连翘和黄柏对灿烂弧菌的抑菌和杀菌效果较好,最小抑菌浓度分别为0.98、1.95、1.95和3.90mg/mL,最小杀菌浓度分别为0.98、1.95、1.95和7.81mg/mL。     

大豆异黄酮对鱼类免疫增强作用的初步研究

研究副溶血弧菌对斑马鱼的感染模式。试验以注射、创伤、浸泡等3种不同的感染方式分2次进行,每组10尾斑马鱼,结果显示注射感染只在高浓度菌液时有死亡,48h的LC50是5.63×107CFU·mL-1,而创伤感染偶有死亡,浸泡感染则未见死亡,表明注射感染是斑马鱼副溶血弧菌的重要感染模式。同时注射感染结果也显示低水温的死亡率远远低于高水温的死亡率,表明适宜的温度和弧菌的浓度是其致病的关键。并初步探讨了大豆异黄酮作为饲料添加剂的免疫增强作用,各组试验鱼分别投喂黄酮含量为0.5%、1%、2%的饲料,喂养两周后开始注射3.6×109CFU·mL-1的副溶血弧菌悬液,以投喂1%的试验组的抗感染效果较好,表明其对减少养殖病害的发生有一定的好处,为养殖生产提供一种新的免疫增强剂。