重组牛源荚膜A型多杀性巴氏杆菌脂蛋白E的免疫保护性研究

为鉴定多杀性巴氏杆菌脂蛋白E的免疫保护性,本研究从牛源荚膜A型多杀性巴氏杆菌PM-HLJ株的基因组DNA中扩增出大小为1 011 bp的脂蛋白E编码基因(PlpE),经序列测定显示,其氨基酸序列与A型多杀性巴氏杆菌参考株PlpE序列一致性为95.83%。将PM-HLJ的PlpE基因克隆于表达质粒pET-30a(+)并诱导表达,将表达的重组脂蛋白E纯化后经腹腔免疫小鼠,间接ELISA检测结果显示各免疫组均产生了针对重组脂蛋白E的IgG抗体,免疫保护试验显示,当小鼠免疫剂量为20μg、30μg、40μg和60μg时,保护率分别达到20%、40%、80%和100%。结果表明,牛源荚膜A型多杀性巴氏杆菌重组脂蛋白E具有良好的免疫保护作用。     

天津地区禽多杀性巴氏杆菌分离株血清型鉴定

通过Ｃａｒｔｅｒ氏荚膜型鉴定方法和琼脂扩散试验,对天津地区禽多杀性巴氏杆菌血清型进行鉴定。结果１６株分离菌中１１株为Ａ：１（４）,４株为Ａ：１（３,４）,１株为－：１（４）表明引起天津地区禽霍乱的致病菌与国内报道的以Ａ：１型为主有一定的差异。     

禽多杀性巴氏杆菌ELISA三种抗原比较研究

禽多杀性巴氏杆菌ＥＬＩＳＡ三种抗原比较研究刘云同郑星道金仁哲顾万钧黄伟（吉林省通化县家畜繁改站·１３４１００）（吉林农业大学动物科学系关于检测禽多杀性巴氏杆菌血清抗体的研究,国外学者是应用凝集反应、间接血凝、酶联免疫吸附试验检测禽多杀性巴氏杆菌弱毒免...     

禽多杀性巴氏杆菌保护性荚膜抗原基因PCA_（10）的高效表达及其免疫原性

以ＰｓｔＩ酶切含禽多杀性巴氏杆菌保护性荚膜抗原基因片段ＰＣＡ１０的重组质粒ｒＰＣＡ１０，收集基因片段ＰＣＡ１０，经修饰后依次连接到系列高效表达载体ＰＥＶ—Ｖｒｆ１～３的ＢａｍＨＩ切点上，转化至Ｅ．ｃｏｌｉＲＲＩ（ｐＲＫ２４８ｃＩｔｓ）中。经快速检测，这３种载体的重组子中均有ＰＣＡ１０插入。以间接ＥＬＩｓＡ检测重组子转化菌，只有ＰＥＶ—Ｖｒ１３载体的重组子转化菌与禽巴氏杆菌保护性荚膜抗原（ＰＣＡ）抗血清呈阳性反应，且ＯＤ值为原克隆菌株ｒＰＣＡ１０的３倍，初步证明ＰＣ１０已克隆至高效表达载体ＰＥＶ—Ｖｒｆ３上，而且读码框架正确。用ＥＬＩｓＡ阳性表达克隆株ＰｃＡ７１—Ｖｒｆ３免疫小鼠和鸡，均达到７０％的保护率。     

牛源荚膜血清A型多杀性巴氏杆菌的分离鉴定

从采集于黑龙江、天津的病牛肺组织中分离到2株病原菌,经菌落形态学观察、培养特性、生化反应和小鼠毒力试验,初步鉴定为多杀性巴氏杆菌,分别命名为Pm-HLJ和Pm-TJ.参考多杀性巴氏杆菌种特异性基因kmt1和荚膜血清型特异性基因hyaD-hyaC、bcbD、dcbF、ecbJ和fcbD,合成引物,通过多重PCR扩增2株菌的种特异性基因和荚膜血清型特异性基因.选取Pm-HLJ的目标PCR产物进行克隆、序列测定、Blast搜索同源序列并且比较分析.结果显示,Pm-HLJ的kmt1基因片段全长460 bp,与GenBank中各血清型kmt1基因同源性均在96.6%以上;荚膜血清型A菌株特异性基因同源性为99.9%;而与其他荚膜血清型B、D、E、F的型特异性基因的同源性均低于50%.由此确认,分离的2株多杀性巴氏杆菌均为荚膜血清A型,这是我国A型多杀性巴氏杆菌引发牛出血性败血症的首例报道.     

猪源荚膜血清F型多杀性巴氏杆菌分离鉴定

本研究对从病死猪肺脏分离出的1株菌株,经菌落形态观察、培养特性、生化反应、药敏试验、动物致病性试验,并采用PCR对该分离菌株进行菌种及荚膜血清型鉴定。结果显示,该分离菌株为较少见的荚膜血清F型多杀性巴氏杆菌,对小鼠有较强致病性,且对不同药物的敏感性不同,为指导临床科学用药提供依据。     

4株荚膜A型多杀性巴氏杆菌的分离与鉴定

2011年7月,从高邮某养殖户两次送检病死麻鸭的肝脏、心脏中分离出4株细菌,对纯化的菌株进行形态学和培养特性的观察、生化反应、血清型PCR鉴定及16SrDNA基因序列分析,确定4株分离茵均为荚膜A型多杀性鸭巴氏杆菌,鸭群所患疾病为该致病菌引起的鸭霍乱。4株分离茵的药敏结果表明,其对氯霉素、强力霉素、萘啶酸等药物多重耐药,而对左氟沙星等药物敏感。     

重组H5N1亚型禽流感疫苗免疫孔雀后的抗体消长规律及免疫程序的研究

为了探讨孔雀对H5高致病性禽流感灭活疫苗的免疫原性,做好孔雀禽流感的防控,采用哈尔滨兽医研究所研制的重组H5N1亚型高致病性禽流感疫苗,不同剂量免疫孔雀后,跟踪其抗体消长规律。检测结果显示:小孔雀母源抗体能维持约6周;首免0.5 mL/羽颈部皮下注射,有效抗体能维持18周以上;1.0,1.5,2.0 mL/羽3个剂量组肌肉注射进行二免,以1.5 mL/羽组效果最好,有效抗体能维持10个月左右;成年孔雀1.5 mL/羽进行三免,有效抗体能维持1年以上。而且孔雀产生的抗体均一性良好,抗体阳性率均在75%以上。根据抗体消长规律,推荐了孔雀的免疫程序。     

H5N1亚型禽流感灭活疫苗免疫鸵鸟后抗体消长规律及免疫程序的研究

为了探讨鸵鸟对H5亚型禽流感灭活疫苗的免疫原性,做好鸵鸟禽流感的防控工作,采用哈尔滨兽医研究所研制的重组H5N1亚型禽流感疫苗,不同剂量免疫鸵鸟,用HI方法检测母源抗体和免疫抗体,根据母源抗体的衰减和免疫抗体的消长规律确定首免和再免日龄.结果表明,雏鸵鸟母源抗体能维持约3周～8周;8周龄时分组首免,C1组产生的免疫反应优于C2组,抗体峰值能达到7.50 log2,维持时间为9周左右;17周龄时C1组以3.0 mL/羽进行二免,2周后抗体达到7.40log2,3周～7周抗体维持在高峰值,最高达8.80log2,以后逐渐下降,有效抗体水平能维持至接种后25周左右.二免后25周进行三免,以5 mL/羽三免后2周抗体可达到9.80log2,2周～4周抗体维持在最高峰,以后缓慢下降,期间抗体时有起伏,但有效抗体水平约可维持1年时间.三免后45周内抗体水平合格率均在70%以上.根据抗体消长规律,初步推荐了鸵鸟的免疫程序.     

Changes in the risk management of Salmonella enterica subspecies diarizonae serovar 61:(k):1, 5, (7) in Swedish sheep herds and sheep meat due to the results of a prevalence study 2012

Background

The prevalence of Salmonella in food producing animals is very low in Sweden due to rigorous control programmes. However, no active surveillance is in place in sheep. The authorities decided to perform a prevalence study in sheep herds because findings at slaughter indicated that sheep associated S. diarizonae (S. enterica subspecies diarizonae serovar 61:(k):1, 5, (7)) might be common in sheep. Sampling was stratified by herd size in two groups, small herds with ≤ 30 animals and large herds with > 30 animals. In each stratum, 237 herds were selected at random. Faecal samples received from 244 out of the 474 randomly selected herds were analysed.

Results

A total of 40 of 100 (40%) of large herds and 17 of 144 (12%) of small herds were positive. The overall adjusted prevalence was 17.6% (95% CI, 12.9-22.2). Sheep associated S. diarizonae was detected in all counties (n = 21). Scientific opinions and an evaluation of on-farm control measures performed concluded that the impact of sheep associated S. diarizonae on human health is very low, and that risk management measures applied in response to findings of sheep associated S. diarizonae in sheep or sheep meat can be expected to have very little impact on reducing risks to human health. As a result, Swedish authorities decided to make an exemption for sheep associated Salmonella diarizonae in sheep and sheep meat in the current Salmonella control measures.

Conclusions

Sheep associated S. diarizonae is endemic in Swedish sheep herds. It is more common in large herds and not limited to certain parts of the country. The responsible authorities concluded that current risk management actions regarding sheep associated S. diarizonae in sheep and sheep meat are not proportional to the risk. This is the first time in the history of the Swedish Salmonella control programme that an exemption from the legislation has been made for a specific serovar. If there is any future indication of an increasing risk, due to e.g. change in the pathogenicity or development of antimicrobial resistance, the risk assessment will be re-evaluated and control measures reinforced if needed.     

Interaction of bovine immunoglobulin gamma 2 (IgG2) with staphylococcal protein A: evidence for IgG2a and IgG2b sub-subclasses in the bovine

The reactivity of bovine IgG with protein A is confusing with respect to which of the bovine IgG class and subclasses are reactive. We have, therefore, re-examined the interaction of bovine immunoglobulins with protein A. The results presented in this paper indicated that at pH 8.0 protein A binds only immunoglobulin of the IgG2 subclass. The bound IgG2 can be readily recovered from an immobilized protein A column at pH 5.0. Furthermore, the antigenic IgG2 eluted demonstrated two charged species which could readily be separated by ion-exchange chromatography. These results indicate that IgG2 in the bovine exists in two sub-subclasses, IgG2a and IgG2b. The two sub-subclasses of IgG2 could be rapidly isolated with a good yield in two-steps namely protein A affinity chromatography followed by ion exchange chromatography.     

The indirect immunofluorescence assay using cardiac tissue from chickens, quails and ducks for identification of influenza A virus during an outbreak of highly pathogenic avian influenza virus (H5N1): a rapid and simple screening tool for limited resource settings

Here we describe the diagnostic utility of the indirect immunofluorescence assay (IFA) during a recent outbreak of highly pathogenic avian influenza (HPAI) subtype H5N1 virus in southern Thailand and demonstrate the usefulness of the cardiac tissue from infected chickens, quail, and ducks for diagnosis. The most reliable sample for IFA diagnosis of influenza A virus was cardiac tissue (83.0%; 44/53) which when divided by species (chicken, quail and duck cardiac tissues) gave respective positivity rates of 88% (22/25), 88.9% (16/18) and 60.0% (6/10). Cardiac tissue also gave the highest IFA intensity for the three species. We believe that the IFA method has wide applicability in developing countries or remote settings where clinically similar avian diseases with high morbidity and mortality such as Newcastle disease and fowl cholera are common and could be rapidly excluded thereby conserving valuable reference laboratory capacity for true HPAI outbreaks.