猪传染性胸膜肺炎重组亚单位疫苗诱导小鼠免疫保护效果的观察

重组表达猪传染性胸膜肺炎放线杆菌6种主要毒力因子基因:apxⅠ、apxⅡ、apxⅢ、apxⅣ、apfa和omp,以重组蛋白rApxⅠ、rApxⅡ、rApxⅢ和rOMP组合免疫小鼠作为试验I组,重组蛋白rApxⅠ、rApxⅡ、rApxⅢr、OMPr、ApxⅣ和rApfa组合免疫小鼠作为试验Ⅱ组,PBS为对照组,分3次免疫小鼠,采用背部皮下多点注射,每次间隔2周,免疫剂量为0.2 mL/只,3免后1周分别以APP1型菌Shope 4074株(5×109cfu)和APP2型菌S1536株(5×1010cfu)进行攻毒试验。通过小鼠保护率与抗体效价的相关性研究、肺部病理变化及肺脏细菌的分布情况等指标进行综合评价。结果显示,试验Ⅰ组4种重组蛋白特异性抗体水平显著高于其他两组(P<0.05),对APP1型菌攻毒的保护率(9/10)明显高于试验Ⅱ组(5/10)和对照组(0/8),小鼠的免疫保护率与抗体效价之间存在显著正相关;且该组对APP2型菌攻毒的保护作用(无肺脏损伤)也明显优于其它两组(典型肺部损伤)。间接免疫荧光试验表明试验Ⅰ组对肺脏细菌的清除效果也明显优于其他两组。本试验揭示试验Ⅰ组对不同血清型APP攻击能够提供很好的交叉保护作用,从而为猪传染性胸膜肺炎新型疫苗的研制提供参考。     

胸膜肺炎放线杆菌Apx ⅢA、Apx ⅣA蛋白表达及重组亚单位疫苗的研究

为研制以胸膜肺炎放线杆菌(APP) 4种外毒素蛋白(ApxⅠA、ApxⅡA、ApxⅢA、ApxⅣA)和外膜蛋白(OMP)为组分的亚单位疫苗,在本研究室已表达纯化的r ApxⅠA、r ApxⅡA、r OMP的基础上,本研究通过PCR扩增得到ApxⅢA、ApxⅣA基因,并分别将其克隆至表达载体p ET-32a中,测序验证后将重组质粒转化入宿主菌BL21 (DE3),经IPTG诱导表达后获得重组蛋白r ApxⅢA和r ApxⅣA。将获得的5种蛋白(rApxⅠA、r ApxⅡA、r ApxⅢA、r ApxⅣA和r OMP)按一定比例混合制备成亚单位疫苗免疫BALB/c雌鼠进行免疫效果测定,设灭活疫苗组(Ⅰ组)、弗氏佐剂乳化的亚单位疫苗组(Ⅱ组)、白油佐剂乳化的亚单位疫苗组(Ⅲ组)、灭活疫苗+白油佐剂乳化的亚单位疫苗组(Ⅳ组)和PBS组(Ⅴ组)。Western blot结果显示,表达了重组蛋白r ApxⅢA和r ApxⅣA且具有较好的反应原性;抗体检测结果显示,加强免疫后,Ⅱ组、Ⅲ组和Ⅳ组的r ApxⅠA、r ApxⅢA、r ApxⅣA和r OMP抗体水平基本一致(p0.05),均显著高于Ⅴ组和Ⅰ组(p0.05);攻毒保护试验结果显示,Ⅱ组与Ⅲ组保护率一致(p0.05),显著高于Ⅴ组(p0.05)。表明本研究中的亚单位疫苗具有一定的免疫保护效果,为猪传染性胸膜肺炎新型疫苗的研制提供参考。     

重组ApxⅡA对胸膜肺炎放线杆菌灭活疫苗免疫效果的影响

将猪传染性胸膜肺炎放线杆菌ApxⅡ毒素的结构基因ApxⅡA克隆到原核表达载体pGEX-6P-1，并在大肠埃希氏菌中进行了表达，表达产物rApxⅡA以包涵体形式存在。为了检测rApxⅡA的免疫原性，分别用rApxⅡA、灭活疫苗以及灭活疫苗添加rApxⅡA对比利时白兔进行了免疫。免疫后进行攻毒，所用菌株分别为同型菌株（APP7型L25—4株）和异型菌株（APP1型4074株）。结果，免疫动物对相同血清型菌株的攻击获得完全保护，且对异型菌的攻击也获得一定保护作用。表明，rApxⅡA蛋白具有良好的免疫原性，作为抗原成分添加到灭活疫苗中后可提高灭活疫苗的免疫效果。     

育肥猪传染性胸膜肺炎的诊断及分离菌毒力基因检测

<正>猪传染性胸膜肺炎(Porcine contagious pleuropneumonia,PCP)是由胸膜肺炎放线杆菌(Actinobacillus pleuropneumoniae,APP)感染所引起的一种高度接触性、致死性呼吸系统疾病,以急性纤维素性胸膜肺炎、出血坏死性肺炎或慢性局灶性坏死性肺炎为特征~([1])。目前研究发现,APP有4种外毒素,即ApxⅠ、ApxⅡ、ApxⅢ、ApxⅣ,是APP的主要毒力因子~([2])。     

猪胸膜肺炎放线杆菌Apx毒素的分子生物学与致病机理

猪传染性胸膜肺炎是由胸膜肺炎放线杆菌(App)引起的猪的一种呼吸道传染病.到目前为止,APP已经被分离鉴定出了15种血清型(1～15).15种血清型的APP能产生属于RTX毒素家族的4种Apx毒素,分别为ApxⅠ、ApxⅡ、ApxⅢ和ApxⅣ.ApxⅠ操纵子包含有ApxⅠ C、A、B、D 4个基因;ApxⅠ毒素含有13个富含甘氨酸的重复区.ApxⅡ操纵子仅包括结构基因ApxⅡA和激活基因ApxⅡC,而无分泌基因;ApxⅡ蛋白含有8个甘氨酸的序列重复.ApxⅢ操纵子与ApxⅠ操纵子相同,具有一个完整的操纵子;ApxⅢ在N～末端有3个疏水区,在C末端部分有13个富含甘氨酸的区域.Apx毒素的致病机理为先由没有活性的前体蛋白转变为有活性的ApxA蛋白,再由有活性的ApxA蛋白侵害细胞.     

猪传染性胸膜肺炎新型亚单位菌苗对小鼠的效力研究

利用大肠杆菌表达猪胸膜肺炎放线杆菌的分泌毒素ApxⅠ、ApxⅡ、ApxⅢ，提取表达产物包涵体，再加入我国流行的猪胸膜肺炎放线杆菌7型菌，和等量弗氏佐剂乳化后制成亚单位菌苗，免疫BALB／c小鼠，间隔2周加强免疫1次。每次免疫后采血检测毒素I、Ⅱ、Ⅲ的EuSA抗体和7型菌的血凝抗体，第2次免疫2周后用5LDso的猪胸膜肺炎放线杆菌1型、7型菌株进行攻毒。结果发现第2次免疫后抗体水平显著升高，用2种血清型进行攻毒后其保护力分别为83．3％和91．7％，从死亡小鼠的体内分离到了攻毒菌株，初步的动物试验表明此种新型亚单位菌苗对同型和异型的APP菌株具有较好的保护力，为进一步研制高效的亚单位菌苗打下基础。     

不同IBD疫苗对IBDV超强毒株NN1172感染的免疫保护试验

为了评估不同传染性法氏囊病疫苗对广西地方品种鸡的免疫保护效果,试验将1日龄后备种鸡分成4组,分别用A、B、C三种商品疫苗对Ⅰ、Ⅱ、Ⅲ组鸡接种免疫,Ⅳ组鸡在C疫苗免疫的基础上于14日龄用D疫苗进行第二次免疫。在免疫前(1日龄)及免疫后的第7,14,21,28,35,42,49,56日龄每组分别采集血清30份,进行传染性法氏囊病病毒(IBDV)抗体的测定;Ⅰ、Ⅱ、Ⅲ组鸡在21日龄,Ⅳ组鸡于28日龄分别用IBDV超强毒地方分离株NN1172进行攻毒,并分别在攻毒后的3,7,10天观察各组雏鸡的临床症状、死亡情况、发病鸡剖检病变,统计免疫保护率。结果表明:各免疫组雏鸡抗体滴度均呈现先降再升的趋势,其中Ⅳ组在免疫后期呈现比较平稳的上升趋势;免疫攻毒保护率结果表明,Ⅱ组的保护率最低(73.3%,11/15),Ⅰ、Ⅲ组均为80%(12/15),Ⅳ组保护率最好(86.7%,13/15)。说明不同疫苗对广西地方品种鸡免疫效果存在一定差异,而二次免疫可以对雏鸡提供更好的免疫保护。     

猪胸膜肺炎放线杆菌溶血外毒素研究进展

从分子质量、基因结构及致病机理等方面概述了猪传染性胸膜肺炎放线杆菌溶血外毒素ApxⅠ,ApxⅡ,ApxⅢ和ApxⅣ的基本特点和研究进展.比较并归纳了这4种溶血外毒素的异同点及在15种血清型中存在的特点.     

猪胸膜肺炎放线杆菌ApxⅠA蛋白的原核表达及单克隆抗体的制备

以猪胸膜肺炎放线杆菌(APP)血清10型标准株ApxⅠ天然外毒素为免疫抗原,接种Balb/c小鼠；设计编码ApxⅠN-端具有保护性抗原表位apxⅠA基因(1149 bp)的引物,构建原核表达载体pET-32a-apxⅠA,转化至E.coli BL21,获得分子质量约为41 ku的rApxⅠA作为包被抗原,用间接ELIS...     

胸膜肺炎放线杆菌RTX毒素抗原优势决定簇的筛选和融合表达

为了制备胸膜肺炎放线杆菌(Actinobacillus pleuropneumoniae,APP)亚单位疫苗和建立配套的诊断方法，本研究在对APP的RTX毒素(ApxⅠ～Ⅲ)生物学信息分析的基础上，以APP血清5型和8型的DNA为模板对ApxⅠ～Ⅲ分段扩增后进行截短表达，获得12个截短表达蛋白。利用Western blot对12个表达蛋白进行抗原性分析，确定ApxⅠ～Ⅲ的优势抗原决定簇分别为蛋白AⅠ2、AⅡ3和AⅢ2。以蛋白AⅡ3、AⅢ2、AⅠ2的顺序排列并在蛋白间加入GPGPG氨基酸序列，无缝克隆3个蛋白片段基因，通过原核表达获得融合蛋白A231。该蛋白可与临床APP阳性猪血清特异性结合，具有良好的免疫反应性。该研究的成功开展可为研制具有交叉保护力的亚单位疫苗及建立配套ELISA检测方法奠定基础。     

白术、微米白术和白术多糖对断奶仔猪生长性能和肠道形态及微生态区系的影响

试验选用96头平均体重14.82 kg左右的杜×长×大断奶仔猪,随机分成4组,每组3栏,每栏8头(公母各半)。对照组饲喂基础日粮,试验1、2、3组分别添加1%80目白术、0.2%白术多糖和1%微米白术。试验期30 d。结果表明:在生长性能方面,与对照组相比,1%微米白术添加组可显著提高日增重(P0.05)、降低饲料增重比和腹泻率,而且效果优于1%80目白术组和0.2%白术多糖组,在肠道形态和肠道微生态区系方面,与对照组相比,日粮添加1%80目白术、0.2%白术多糖、1%微米白术均可不同程度的提高十二指肠和空肠的绒毛高度,加深十二指肠和空肠的隐窝深度,并且增加肠道微生态区系的多样性,其中以1%微米白术添加组的效果最佳。     

Feline intracranial neoplasia: retrospective review of 160 cases (1985-2001)

The purpose of this study was to determine the frequency of different tumor types within a large cohort of cats with intracranial neoplasia and to attempt to correlate signalment, tumor size and location, and survival time for each tumor. Medical records of 160 cats with confirmed intracranial neoplasia evaluated between 1985 and 2001 were reviewed. Parameters evaluated included age, sex, breed, FeLV/FIV status, clinical signs, duration of signs, number of tumors, tumor location(s), imaging results, treatment, survival times, and histopathologic diagnosis. Most of the cats were older (11.3 +/- 3.8 years). Primary tumors accounted for 70.6% of cases. Metastasis and direct extension of secondary tumors accounted for only 5.6 and 3.8% of cases, respectively. Twelve cats (7.5%) had 2 or more discrete tumors of the same type, whereas 16 cats (10.0%) had 2 different types of intracranial tumors. The most common tumor types were meningioma (n = 93, 58.1%), lymphoma (n = 23, 14.4%), pituitary tumors (n = 14, 8.8%), and gliomas (n = 12, 7.5%). The most common neurological signs were altered consciousness (n = 42, 26.2%), circling (n = 36, 22.5%), and seizures (n = 36, 22.5%). Cats without specific neurological signs were common (n = 34, 21.2%). The tumor was considered an incidental finding in 30 (18.8%) cats. In addition to expected relationships (eg, meninges and meningioma, pituitary and pituitary tumors), we found that lesion location was predictive of tumor type with diffuse cerebral or brainstem involvement predictive of lymphoma and third ventricle involvement predictive of meningioma.     

Prevalence and seasonal incidence of nematode parasites and fluke infections of sheep and goats in eastern Ethiopia

Sissay, M.M., Uggla, A. and Waller, P.J., XXXX. Prevalence and seasonal incidence of nematode parasites and fluke infections of sheep and goats in eastern Ethiopia. Tropical Animal Health and Production, XXXX. A 2-year abattoir survey was carried out to determine the prevalence, abundance and seasonal incidence of gastro-intestinal (GI) nematodes and trematodes (flukes) of sheep and goats in the semi-arid zone of eastern Ethiopia. During May 2003 to April 2005, viscera including liver, lungs and GI tracts were collected from 655 sheep and 632 goats slaughtered at 4 abattoirs located in the towns of Haramaya, Harar, Dire Dawa and Jijiga in eastern Ethiopia. All animals were raised in the farming areas located within the community boundaries for each town. Collected materials were transported within 24 h to the parasitology laboratory of Haramaya University for immediate processing. Thirteen species belonging to 9 genera of GI nematodes (Haemonchus contortus, Trichostrongylus axei, T. colubriformis, T. vitrinus, Nematodirus filicollis, N. spathiger, Oesophagostomum columbianum, O. venulosum, Strongyloides papillosus, Bunostomum trigonocephalum, Trichuris ovis, Cooperia curticei and Chabertia ovina), and 4 species belonging to 3 genera of trematodes (Fasciola hepatica, F. gigantica, Paramphistomum {Calicohoron} microbothrium and Dicrocoelium dendriticum) were recorded in both sheep and goats. All animals in this investigation were infected with multiple species to varying degrees. The mean burdens of adult nematodes were generally moderate in both sheep and goats and showed patterns of seasonal abundance that corresponded with the bi-modal annual rainfall pattern, with highest burdens around the middle of the rainy season. In both sheep and goats there were significant differences in the mean worm burdens and abundance of the different nematode species between the four geographic locations, with worm burdens in the Haramaya and Harar areas greater than those observed in the Dire Dawa and Jijiga locations. Similar seasonal variations were also observed in the prevalence of flukes. But there were no significant differences in the prevalence of each fluke species between the four locations. Overall, the results showed that Haemonchus, Trichostrongylus, Nematodirus, Oesophagostomum, Fasciola and Paramphistomum species were the most abundant helminth parasites of sheep and goats in eastern Ethiopia.     

Y‐chromosomal variation of local goat breeds of Turkey close to the domestication centre

Genetic variations in chromosome Y are enabling researchers to identify paternal lineages, which are informative for introgressions and migrations. In this study, the male‐specific region markers, sex‐determining region‐Y (SRY), amelogenin (AMELY) and zinc finger (ZFY) were analysed in seven Turkish native goat breeds, Angora, Kilis, Hair, Honaml?, Norduz, Gürcü and Abaza. A SNP in the ZFY gene defined a new haplotype Y2C. All domestic haplogroups originate from Capra aegagrus, while the finding of Y1A, Y1B, Y2A and Y2C in 32, 4, 126 and 2 Turkish domestic goats, respectively, appears to indicate a predomestic origin of the major haplotypes. The occurrence of four haplotypes in the Hair goat and, in contrast, a frequency of 96% of Y1A in the Kilis breed illustrate that Y‐chromosomal variants have a more breed‐dependent distribution than mitochondrial or autosomal DNA. This probably reflects male founder effects, but a role in adaptation cannot be excluded.     

The Prevalence and Intensity of Helminth and Coccidial Infections in Dairy Cattle in Central Kenya

A survey of gastrointestinal parasite infections of young (<6 months old), immature (6–12 months old) and adult (>12 months old) dairy cattle on 16 farms in Kiambu District, Kenya was conducted during a dry season (September 1991 to January 1992) and during a wet season (March to July 1992). The survey was based on monthly coproparasitological examination of cohorts and worm counts in tracer calves. The effects of age, sex, farm and season on the prevalence and intensity of helminth and coccidial infections were determined. Faecal egg and oocyst counts revealed that the overall prevalences were: strongyles (including trichostrongyles) (85.5%), liver flukes (Fasciola gigantica) (34.0%), coccidia (30.9%) and tapeworms (9.6%). Eight species of the protozoan Eimeria were identified, the most prevalent species being E. bovis and E. zuernii. The most prevalent nematode genera were Haemonchus, Cooperia, Oesophagostomum and Trichostrongylus. Season, farm and age of the animals had a significant (p<0.05) influence on the intensity of infection with strongyles, liver flukes and coccidia, whereas the sex of the animals had no significant (p>0.05) effect on the prevalence or intensity of infections. A higher intensity of infection with strongyles and coccidia was found in the wet season than in the dry season (p<0.05). The age-specific intensity was in the following order: for strongyles, immature animals of 6–12 months of age had the highest egg counts, followed by young calves and adults. Calves had significantly (p<0.05) higher oocyst counts than immatures or adults. Liver fluke egg counts did not differ significant (p>0.05) between immatures and adult cattle.     

A brief history of the prevention of infectious diseases by immunisations

Infectious diseases have always been a terrible scourge for humans. The appearance of these plagues, as they were called without distinction, was generally connected to various conditions: asters, climatic changes or religious reasons. The concept of contagious, and then infectious, diseases came slowly. Variolation, i.e. transmission of ‘virulent’ matter to induce a natural disease and the immunity against it, was brought from Constantinople to England by Lady Montague, in 1721. This ‘variolation’ technique was also often performed in veterinary medicine against diseases like sheep-pox or pleuropneumonia. As ‘vaccination’ is the term generally accepted for ‘immunisation’, variolation can be the word designating such a technique. The second period of the history of immunisation began, in 1880, with the studies of Pasteur and his collaborators. A great number of bacterial vaccines were developed: dead, live but attenuated or only parts of pathogens. The viruses were produced in animals, then in eggs and at last, in tissue cultures. Second generation vaccines appeared with genetic engineering: recombinant vaccines, vector vaccines, nucleic acids vaccines, and markers vaccines, among others. These novel technologies can permit the development of new ones and improve the quality of the vaccines already existing.     

Micronutrient Content of Certain Tropical Conventional and Unconventional Feed Resources of Southern India

The ash, silica and certain important micronutrients were estimated in conventional and unconventional feed and fodder resources available in Southern India. Commonly used dry roughages, such as paddy straw, ragi straw, maize kadbi/stalk, jowar kadbi/stalk, bajra stalk and wheat straw, were high in ash (9.9% +/- 0.77%) and silica (6.4% +/- 0.65%) and low in most of the other micronutrients, except iron, with paddy straw containing most silica (>9%). Cultivated non-leguminous (maize, jowar) and improved green crops (hybrid napier, guinea, green panic, NB-21, CO-1) were also high in ash (10.5% +/- 0.60% and 12.5% +/- 0.51%) but were moderate sources of P, Mg and Cu and good sources of Zn (98 +/- 13.8 ppm and 55 +/- 6.7 ppm). Leguminous green fodders (stylosanthus, lucerne, cow pea, soyabean) were excellent sources of Ca (1.9% +/- 0.16%), Mg (0.40% +/- 0.05%), Cu (30 +/- 5.2 ppm), Zn (121 +/- 14.7 ppm) and Fe (1234 +/- 166 ppm) and moderate sources of P. Mixed local grasses and weeds were high in silica (6.9% +/- 1.00%) but were good sources of Cu, Zn and Fe. Cereal grains (maize, wheat, rice, ragi) were low in ash (2.9% +/- 0.33%) and were relatively poor sources of Ca (0.22% +/- 0.03%), Mg (0.19% +/- 0.03%) and Cu (13 +/- 3.1 ppm). Pulses were low to medium sources of most minerals and good sources of Fe (1230 +/- 293 ppm). Oil seed cake/extractions (groundnut cake, cotton seed cake, soyabean meal, sunflower cake, safflower cake) and cereal by-products (rice polish, rice bran, wheat bran) were excellent sources of P (1.1% +/- 0.47% and 2.3% +/- 0.19%) and good sources of Zn (65 +/- 3.9 ppm and 66 +/- 10.7 ppm) and Fe (938 +/- 130 ppm and 662 +/- 126 ppm). Among the unconventional feeds screened, orange peel, sunflower heads, meat meal, rubber seed cake, spirulina algae and sea weeds contained plentiful Ca, Zn and Fe: tree leaves/top feeds (mulberry, erythrina, glyricidia, banana, subabul, groundnut haulms) were excellent sources of Ca (1.5% +/- 0.13%), Zn (120 +/- 22.9 ppm) and Fe (1033 +/- 133 ppm) but relatively poor sources of P. Soyabean husk, cocoa seed husk, rubber seed cake and meat meal were moderate to good sources of P (1.0% and 0.33%). The high Zn and Fe values of most feeds/fodders were probably due to soil contamination. This account of the micronutrient content of feed/fodder resources should help in strategic supplementation intended to alleviate local deficiencies.     

Bovine Mastitis in Selected Areas of Southern Ethiopia

A study on bovine mastitis, designed to determine the causal agents, prevalence of infection and impact of risk factors in three cattle breeds, was conducted in selected areas of southern Ethiopia. A total of 307 lactating and non-lactating cows, of which 162 were indigenous Zebu, 85 Jersey and 60 Holstein-Friesian, were examined by clinical examination and the California mastitis (CMT) test. Of these, 40.4% were positive by CMT and bacteriology for clinical or subclinical mastitis, with prevalence rates of 37.1% and 62.9%, respectively. Out of 1133 quarters examined, 212 (18.7%) were found to be infected, 83 (39.2%) clinically and 129 (60.8%) subclinically. The prevalence of mastitis was significantly higher in Holstein-Friesian than in indigenous Zebu, in non-lactating cows than in lactating cows, in the early lactation stage than in the mid-lactation stage, in cows with lesions and/or tick infestation on skin of udder and/or teats than in cows without this factor, and in the wet season than in the dry season. Mastitis increased with parity number (R = 0.9). Of 248 CMT and clinically positive udder quarter samples analysed microbiologically, 212 were culturally positive for known mastitis pathogens and 36 were negative. Of the 199 positive samples, Staphylococcus accounted for 39.2%, Streptococcus for 23.6%, coliforms for 14.1%, Micrococcus and Bacillus species for 8.0% each and Actinomyces or Arcanobacterium (Corynebacterium) for 7.0%. It was concluded that there was a high prevalence of clinical and subclinical mastitis, mainly caused by Staphylococcus aureus, Streptococcus agalactiae and Escherichia coli, in this study area.     

不同日粮氮水平对山羊氮代谢和微生物蛋白质合成的影响

本试验旨在探讨不同日粮氮水平对内蒙古白绒山羊氮代谢和微生物蛋白质合成(MCP)的影响,以达到提高氮利用率,减少环境污染和饲料资源的浪费。试验选用9只体况良好、体重为(43.83±2.95)kg,装有瘤胃瘘管的内蒙古白绒山羊,按体重随机分为3组,每组3只。日粮分为低氮7.5%、中氮10.5%、高氮13.5%3个氮水平。试验使用常规试验方法分析测定粪尿血液等生化指标,通过全收尿法和尿嘌呤衍生物法估测MCP。结果表明:随着氮水平增加,氨氮(NH3-N)浓度、氮摄入量、尿氮排出、尿中尿素氮(UUN)和MCP显著增加(P0.05),干物质采食量(DMI)显著降低(P0.01);沉积氮占总摄入氮的比例在中氮组最高,MCP在高氮组最高。综上所述,适当降低日粮氮水平可减少粪尿氮排放,提高反刍动物氮利用率。