斑点叉尾鮰败血症3种病原多重PCR检测方法的建立     

刘礼辉  张德锋  李宁求 《西北农林科技大学学报(自然科学版)》2016,44(11):39-46,54

【目的】建立可同时快速检测斑点叉尾鮰败血症3种病原(嗜水气单胞菌、维氏气单胞菌和鮰爱德华菌)的多重PCR检测方法。【方法】根据嗜水气单胞菌溶血素基因(Hly)、维氏气单胞菌脱氧核糖核酸酶Ⅰ基因(DNaseⅠ)以及鮰爱德华菌溶血活化基因(Eha)的保守序列,设计3对特异性引物,优化并建立斑点叉尾鮰败血症3种主要病原菌的多重PCR方法,对其特异性和灵敏度进行考察,并将其用于临床样品的检测。【结果】建立的多重PCR方法,当Hly基因、DNaseⅠ基因以及Eha基因的引物浓度分别为1.0,1.0和0.5μmol/L,退火温度为59.5℃时,各目的片段均可较好地扩增。特异性试验结果表明,建立的多重PCR方法可从嗜水气单胞菌、维氏气单胞菌和鮰爱德华菌分别扩增出1 091,262和450bp的目的片段,从多种细菌DNA的混合种也可扩增出上述目的片段,而对其他对照组的扩增结果均为阴性;敏感性试验结果表明,建立的多重PCR最低可分别检测出200CFU/mL的嗜水气单胞菌、300CFU/mL的维氏气单胞菌和200CFU/mL的鮰爱德华菌;对临床样本的检出率为100%,与传统生物学检测结果的符合率为100%。【结论】建立的多重PCR检测方法特异、灵敏、快速,可单独或同时检测斑点叉尾鮰败血症嗜水气单胞菌、维氏气单胞菌和鮰爱德华菌3种主要病原菌,也可以用于其他水生动物上嗜水气单胞菌、维氏气单胞菌和鮰爱德华菌的检测。  相似文献   

First detection of Edwardsiella ictaluri (Proteobacteria: Enterobacteriaceae) in wild Australian catfish          下载免费PDF全文

E Kelly  P A J Martin  S Gibson‐Kueh  D L Morgan  B C Ebner  J Donaldson  N Buller  D A Crook  S Brooks  A M Davis  M P Hammer  L Foyle  S Hair  A J Lymbery 《Journal of fish diseases》2018,41(2):199-208

The bacterium Edwardsiella ictaluri is considered to be one of the most significant pathogens of farmed catfish in the United States of America and has also caused mortalities in farmed and wild fishes in many other parts of the world. E. ictaluri is not believed to be present in wild fish populations in Australia, although it has previously been detected in imported ornamental fishes held in quarantine facilities. In an attempt to confirm freedom from the bacterium in Australian native fishes, we undertook a risk‐based survey of wild catfishes from 15 sites across northern Australia. E. ictaluri was detected by selective culturing, followed by DNA testing, in Wet Tropics tandan (Tandanus tropicanus) from the Tully River, at a prevalence of 0.40 (95% CI 0.21–0.61). The bacterium was not found in fishes sampled from any of the other 14 sites. This is the first report of E. ictaluri in wild fishes in Australia.  相似文献   

Protection of crucian carp (Carassius auratus Gibelio) against septicaemia caused by Aeromonas hydrophila using specific egg yolk immunoglobulins     

Liji Jin  Xiaoyu Li  Dongliang Zou  Shuying Li  Wenqian Song  Yongping Xu 《Aquaculture Research》2013,44(6):928-936

Egg yolk immunoglobulins (IgY) were obtained from laying hens immunized with inactivated Aeromonas hydrophila. The purified IgY was shown to inhibit the growth of A. hydrophila in vitro and the optimum concentration for inhibition of A. hydrophila‐specific IgY was 75 mg mL^?1. In a subsequent challenge trial, 100 carp (200~250 g) were assigned to one of ten tanks with ten carp per tank. The fish in one tank were unchallenged whereas the remaining 90 fish were injected intraperitoneally with 100 μL of A. hydrophila at a concentration of 10⁸ cfu mL^?1. For the next 21 days, all fish were moved in their respective groups to a clean tank for 20 min day^?1. The fish in four tanks (one unchallenged tank and three challenged tanks) received no treatment whereas the fish in the remaining six tanks were immersed in either 0.5 g L^?1 aqueous nonspecific IgY (N = 3) or 0.5 g L^?1 aqueous specific IgY (N = 3). Haemoglobin concentrations, white and red blood cell numbers as well as the mortality of specific IgY‐treated fish were significantly different from those of the control. These results suggest that passive immunization by immersion with pathogen‐specific IgY may provide a valuable treatment for A. hydrophila infection in carp.  相似文献   

Molecular heterogeneity of plpE gene in Indian isolates of Pasteurella multocida and expression of recombinant PlpE in vaccine strain of P. multocida serotype B: 2     

Ajay Pratap Singh  Satparkash Singh  Rajeev Ranjan  Santosh Kumar Gupta  Vijendra Pal Singh  Bhaskar Sharma 《Journal of veterinary science (Suw?n-si, Korea)》2010,11(3):227-233

Outer membrane proteins of Pasteurella (P.) multocida have been known to be protective immunogens. Pasteurella lipoprotein E (PlpE) has been reported to be an important cross reactive outer membrane protein in P. multocida. The gene encoding the PlpE of P. multocida serotypes A: 3, B: 2 and D: 1 was amplified from the genomic DNA. The amplified products were cloned and the nucleotide sequence was determined. Sequence analysis of the recombinant clones revealed a single open reading frame of 1,011 bp, 1,008 bp and 1,017 bp encoding a protein with a calculated molecular mass of 37.829 kDa, 37.389 kDa and 37.965 kDa for serotypes A: 3, B: 2 and D: 1 respectively. The comparison of the plpE sequence in different capsular types revealed a high degree (>90%) of homology. Furthermore, the plpE gene of Haemorhhagic septicaemia causing serotype (B: 2) was expressed in E. coli and recombinant PlpE was strongly immunostained by antiserum against whole cell antigen, indicating that the protein is expressed in vivo.  相似文献   

中华鲟源维氏气单胞菌的分离鉴定及其药敏特性   总被引：1，自引：0，他引：1  

田甜  张建明  杜合军 《安徽农业大学学报》2017,44(6):1010

以自然患细菌性败血症的子二代中华鲟(Acipenser sinensis)幼鱼为研究对象,无菌操作从濒死病鲟体内分离到1株致病菌(zhx20120301)。结合传统细菌学鉴定及分子生物学鉴定方法,对其形态、生理生化特性、16S r DNA序列及系统发育分析等方面进行研究,研究结果显示这株细菌为维氏气单胞菌(Aeromomas veronii)。回归感染试验证实分离菌株具有较强的致病性,能导致健康杂交鲟(西伯利亚鲟Acipenser baerii♂×施氏鲟Acipenser schrenckii♀)死亡,并呈现与自然条件下相似的临床症状,且再分离菌株各种生理生化特性与原分离菌株相同。药敏试验结果表明该菌株对头孢氨苄、头孢唑啉、庆大霉素、四环素、强力霉素、氟苯尼考、多粘霉素B、恩诺沙星、氟哌酸和左氧氟沙星敏感。  相似文献   

The potential of waste items in aquatic environments to act as fomites for viral haemorrhagic septicaemia virus     

Pham PH  Jung J  Lumsden JS  Dixon B  Bols NC 《Journal of fish diseases》2012,35(1):73-77

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大菱鲆(Scophthalmus maximus L.)鳗利斯顿氏菌(Listonella anguillarum)感染症及其病原特性   总被引：1，自引：0，他引：1  

房海  陈翠珍  张晓君  葛慕湘  何振平  巩元芳 《中国兽医学报》2006,26(2):144-146

对一养殖场所发生的大菱鲆（Scophthalmus maximus L．）病害进行了发病情况、临床表现、病理变化及病原等方面的检验，结果表明为细菌性败血感染症。通过对15尾病（死）大菱鲆病变组织中细菌检查、细菌分离与鉴定，以及人工感染试验的致病作用等，表明所检出的细菌为利斯顿氏菌属（Listonella MaeDonell and Colwell 1986）的鳗利斯顿氏菌[L．anguillarum（Bergeman 1909）MacDonell and Colwell 1986]，系该病例的病原菌。  相似文献   

Specific Antibodies of Pasteurella multocida in Newborn Calves of Vaccinated Dams     

el-Eragi AM  Mukhtar MM  Babiker SH 《Tropical animal health and production》2001,33(4):275-283

Twenty-five newborn Holstein Friesian calves, from dams vaccinated against haemorrhagic septicaemia (HS), were tested repeatedly over the first 6 months of life to monitor the transferred antibody levels against HS. Enzyme-linked immunosorbent assays were used to measure the specific HS antibodies with antigens from Pasteurella multocida strains B:2 and E:2. There was a significant curvilinear relationship between the monitored IgG response and the age of the calves. Peak serum IgG levels were obtained during the period from 8 to 16 weeks of age. Beyond this age, the concentration of IgG in the serum fell away.  相似文献   

Epidemiological aspects of viral haemorrhagic septicaemia virus genotype II isolated from Baltic herring, Clupea harengus membras L     

Gadd T  Jakava-Viljanen M  Tapiovaara H  Koski P  Sihvonen L 《Journal of fish diseases》2011,34(7):517-529

This study was carried out to clarify the role of wild fish, especially Baltic herring, Clupea harengus membras L., in the epidemiology of viral haemorrhagic septicaemia virus (VHSV) in brackish water in Finland. Baltic herring with no visible signs of disease were collected from the Archipelago Sea, the Gulf of Bothnia and the eastern Gulf of Finland. In total, 7580 herring were examined by virus isolation as 758 pooled samples and 3029 wild salmonid broodfish as pooled samples during 2004-2006. VHSV was isolated from 51 pooled herring samples in bluegill fibroblast-2 cells, but not in epithelioma papulosum cyprini cells. The majority of isolations were from the coastal archipelago and from fish caught during the spawning season. Based on glycoprotein (G) gene sequences, the virus was classified as a member of genotype II of VHSV. Pairwise comparisons of the G gene regions of herring isolates revealed that all the isolates were closely related, with 98.8-100% nucleotide homology. Phylogenetic analyses revealed that they were closely related to the strains isolated previously from herring and sprat, Sprattus sprattus (L.), in Gotland and to the VHSV isolates from European river lamprey, Lampetra fluviatilis (L.), in the rivers that flow into the Bothnian Bay. The infection in Baltic herring is likely to be independent of the VHSV Id epidemic in farmed rainbow trout, Oncorhynchus mykiss (Walbaum).  相似文献   

Temporal trends in in vitro antimicrobial susceptibility patterns of bacteria isolated from foals with sepsis: 1979–2010     

M. J. P. Theelen  W. D. Wilson  J. M. Edman  K. G. Magdesian  P. H. Kass 《Equine veterinary journal》2014,46(2):161-168

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