副猪嗜血杆菌病原夹心ELISA检测方法的建立及应用

为了建立一种准确、快速的副猪嗜血杆菌病原夹心ELISA检测方法,以副猪嗜血杆菌外膜蛋白P5的特异性单克隆抗体1E2作为捕获抗体,兔抗副猪嗜血杆菌多克隆抗体作为检测抗体,通过方阵滴定优化夹心ELISA检测方法最佳反应条件,并对该检测方法进行特异性、敏感性验证以及临床样品的检测应用。结果显示,该检测方法中单克隆抗体1E2最佳包被浓度为3.434μg/m L,兔抗副猪嗜血杆菌多抗的最佳稀释浓度为3.350μg/m L,用10 mg/m L明胶37℃封闭1 h优于其他封闭液,辣根过氧化物酶标记的山羊抗兔多体最佳使用浓度为1∶4 000。该检测方法的特异性试验结果显示可检测出15个血清型的副猪嗜血杆菌病原,而与其他病原菌的检测结果均为阴性。敏感性试验结果显示该方法能够检测出副猪嗜血杆菌的最低菌落浓度为1×106cfu/m L。利用该检测方法对临床115份副猪嗜血杆菌可疑病料进行检测结果显示可检出83份阳性样品,检出的阳性样品数高于细菌分离及PCR鉴定方法。上述结果表明所建立的副猪嗜血杆菌病原夹心ELISA检测方法特异性强、敏感性好并可应用到临床样品的检测。     

副猪嗜血杆菌间接血凝诊断液的制备

【目的】建立一种便于检测副猪嗜血杆菌抗体的方法。【方法】将分离纯化的副猪嗜血杆菌5型超声裂解后,致敏双醛处理的健康绵羊红细胞,制成副猪嗜血杆菌5型间接血凝诊断液,检测该诊断液的活力及特异性。【结果】该诊断液特异性强,仅与副猪嗜血杆菌5型阳性血清发生凝集反应。活力也比较高,达到1:512以上,可以用其进行副猪嗜血杆菌疫苗免疫后抗体检测。【结论】该间接血凝诊断液成功制备为广大兽医化验人员提供一种简单、快速、经济的检测副猪嗜血杆菌抗体的方法,为副猪嗜血杆菌病的诊断和防治提供一种有效的手段     

副猪嗜血杆菌外膜蛋白 P5的分段表达及免疫学性质分析

为获得副猪嗜血杆菌外膜蛋白P5的分段表达产物,并对各片段的免疫学性质进行鉴定。采用生物信息学软件分析Omp P5全长基因,设计3对分段引物,PCR分别从HPS汕头分离株（H0801）Omp P5中扩增出P5F1、P5F2和P5F3基因片段,构建于pET-32a（＋）原核表达载体上,并将重组质粒转化大肠杆菌BL21（DE3）,用IPTG诱导表达,利用SDS-PAGE分析蛋白表达情况。将蛋白纯化后,利用ELISA及Western Blot分别鉴定其免疫原性及抗原性。结果表明,扩增得到的Omp P5的3个基因片段与预期大小相符合,基因测序结果与GenBank 公布的序列一致,经SDS-PAGE分析,3个蛋白表达相对分子质量大小都与预期相符。 ELISA和Western Blot分析显示,表达的3个蛋白表现出良好的抗原性和免疫原性。 HPS Omp P5的分段表达为进一步研究HPS保护性表位及疫苗鉴定奠定了新的基础。     

青海省部分地区副猪嗜血杆菌流行病学研究

为了明确青海省4个地区12个猪场副猪嗜血杆菌病的流行情况,应用间接血凝对不同猪场639份血清进行了检测;对疑似副猪嗜血杆菌病的15份病料进行了病原分离鉴定,应用琼脂扩散试验对分离菌进行了分型.结果4个地区副猪嗜血杆菌最高阳性率为58.73%,最低阳性率为0,平均阳性率为25.19%;从疑似副猪嗜血杆菌病患病猪的肺脏和腹膜中分离到2株革兰氏阴性细小杆菌,经生化试验和PER鉴定,确定分离菌为副猪嗜血杆菌,分型结果为副猪嗜血杆菌血清5型(QH0804)和7型(QH0811).结果表明,副猪嗜血杆菌病在4个地区一些猪场有不同程度的流行和发生.这一结果为该地区有效控制该病防制提供了依据.     

副猪嗜血杆菌病的防治

副猪嗜血杆菌病是由副猪嗜血杆菌引起的一种世界范围内流行的重要疾病。由于我国的饲养管理条件较差，该病流行和危害日渐严重，应引起广大养殖户足够的重视。一、流行情况副猪嗜血杆菌只感染猪，主要存在于猪的上呼吸道（鼻腔、扁桃体和气管前段等）内，通过猪群间的接触和空气飞沫以及污染物而传播，带菌猪和慢性感染猪为本病的传染源。     

副猪嗜血杆菌中毒素蛋白基因hipA的生物信息学分析

为预测毒素蛋白HipA的二级结构和B细胞抗原表位,首先从GenBank中查找副猪嗜血杆菌毒素蛋白基因hipA的序列,应用生物信息学方法,对副猪嗜血杆菌毒素蛋白HipA的理化性质、二级结构和B细胞抗原表位进行预测。结果显示,HipA蛋白的理论等电点为7.23,前50个氨基酸为信号肽,二级结构的预测结果显示该酶主要以α螺旋、无规则卷曲和延伸链为主,只有少数的β转角;推测该蛋白有17个B细胞优势抗原表位区域。该研究为进一步研究分析副猪嗜血杆菌毒素蛋白HipA的功能奠定理论基础。     

猪圆环病毒和副猪嗜血杆菌混合感染病例的诊断与治疗

<正>猪圆环病毒感染,又称断奶仔猪多系统衰竭综合征,是由猪圆环病毒引起的一种传染病,临床以仔猪消瘦、腹泻、呼吸困难和体质下降为特征。副猪嗜血杆菌病是由猪副嗜血杆菌引起的泛嗜性细菌性传染病,仔猪和架子猪的感染性较强,该病多发生于运输疲劳时或应激而引起,临床以肺浆膜和心包以及腹腔浆膜和四肢关节浆膜的纤维素性炎为特征。副猪嗜血杆菌病是世界性分布的条件性疾病,往往并发感染于蓝耳病毒、圆环病毒等病毒性疾病,由猪圆环病毒并发副猪嗜血杆菌的案例较为常见。一、典型病例     

副猪嗜血杆菌鉴定分型方法研究现状

随着养殖水平的提高,近年来,副猪嗜血杆菌(HPS)在各猪场引起多发性浆膜炎和关节炎的报道屡见不鲜,已从多个省市地区分离出副猪嗜血杆菌,HPS的血清型复杂,用传统的方法往往不能达到对其快速准确鉴定的目的。大量研究表明不同血清型HPS毒株其毒力和致病力存在差异,因此,找出一种敏感性、特异性与重复性都非常良好的分型方法,对副猪嗜血杆菌病的防控有着重要的意义。     

基于猪囊尾蚴M13h蛋白的ELISA方法的优化

利用纯化的猪囊尾蚴重组蛋白M13h作为抗原包被酶标板,通过棋盘滴定方法筛选最佳反应条件,初步建立了猪囊尾蚴病间接ELISA检测方法.确定最佳包被量为1 μg/mL,待检血清最佳稀释倍数为1∶100,酶标二抗的稀释倍数为1∶1000,二者的作用时间均为45 min；与猪弓形虫病、猪旋毛虫病、细颈囊尾蚴病、猪蛔虫病、包虫病...     

副猪嗜血杆菌hhdB基因的鉴定和分析

为了明确副猪嗜血杆菌致病菌株的毒力相关因子,从广东省不同地区发病猪场送检病猪中分离到9株细菌(H1~H9),并对其hhdB基因进行了鉴定和分析。结果显示,所分离的9株细菌经形态观察、生化特性鉴定和PCR鉴定为副猪嗜血杆菌阳性,并具有卫星现象和不溶血特征。用hhdB基因引物从分离菌株中扩增到了特异性片段,序列分析表明,所获得的hhdB基因序列与GenBank中副猪嗜血杆菌SH0165的hhdB基因同源性为99%,分离菌株hh-dB基因之间序列同源性为97.5%~99.8%,推导的氨基酸序列与少数放线杆菌、杜克雷嗜血杆菌的溶血素激活蛋白具有一定的相似性。RT-PCR扩增到了hhdB基因的目的条带,表明该基因在分离菌株中得到了相应的表达。这将为副猪嗜血杆菌致病菌株毒力特征基因的鉴定以及副猪嗜血杆菌致病机理的研究奠定基础。     

Location of a high-lysine gene and the DDT-resistance gene on barley chromosome 7

Summary The high-lysine gene in Risø mutant 1508 conditions an increased lysine content in the endosperm via a changed protein composition, a decreased seed size, and several other characters of the seed. The designation lys3a, lys3b, and lys3c, is proposed for the allelic high-lysine genes in three Risø mutants, nos 1508, 18, and 19. Linkage studies with translocations locate the lys3 locus in the centromere region of chromosome 7. A linkage study involving the loci lys3 and ddt (resistance to DDT) together with the marker loci fs (fragile stem), s (short rachilla hairs), and r (smooth awn) show that the order of the five loci on chromosome 7 from the long to the short chromosome arm is r, s, fs, lys3, ddt. The distance from locus r to locus ddt is about 100 centimorgans.     

A note on the origin of Kalanchoë x vadensis Boom & Zeilinga (Crassulaceae)

Summary K x vadensis is a hybrid of K. blossfeldiana and K. marmorata obtained after doubling the number of chromosomes.     

Simultaneous Determination of Four Phenolic Acids in Radix Salviae Miltiorrhizae Formula Granules by QAMS

[Objectives]This study aimed to establish a QAMS(quantitative analysis of multi-components by single-marker)method for simultaneous determination of four phenol...     

Avoidance of leaf rust fungi in wild relatives of cultivated cereals

Summary Avoidance of rust fungi that was based on poor appressorium induction was previously found in Hordeum chilense. In the present study 95 accessions of Triticeae were screened for avoidance of Puccinia hordei. The percentage of appressorium formation per germinated spore ranged from 6 to 90%. On none of the 41 accessions of Aegilops, Agropyron, Elymus, Secale, Thinopyrum or Triticum studied was the rate of appressorium formation lower than 25%. Lower rates of appressorium formation were, however, found on accessions of wild barley species Hordeum brachyantherum, H. marinum, H. parodii and H. secalinum. Its implications in cereal breeding are discussed.     

Present status and future strategy in breeding faba beans (Vicia Faba L.) for resistance to biotic and abiotic stresses

Progress is being made, mainly by ICARDA but also elsewhere, in breeding for resistance to Botrytis, AScochyta, Uromyces, and Orobanche; and some lines have resistance to more than one pathogen. The strategy is to extend multiple resistance but also to seek new and durable forms of resistance. Internationally coordinated programs are needed to maintain the momentum of this work.Tolerance of abiotic stresses leads to types suited to dry or cold environments rather than broad adaptability, but in this cross-pollinated species, the more hybrid vigor expressed by a cultivar, the more it is likely to tolerate various stresses.     

Water Extraction Process of Chinese Herbal Compound Man Gan Ning

[Objectives]To optimize the water extraction process of Chinese Herbal Compound Man Gan Ning and establish a method for its extraction and content determination...     

Pollen and pollination experiments. III. The viability of apple and pear pollen as affected by irradiation and storage

Summary Apple and pear pollen was irradiated with doses of 0, 50, 100, 250 and 500 krad (gamma rays) and stored at 4°C and 0–10% r.h. From the in-vitro germination percentages an average LD 50 dose of about 220 krad was estimated. For both irradiated and untreated pollen a close and corresponding lineair relationship existed between germination percentage and pollen tube growth.Irradiated pollen was much more sensitive to dry storage conditions than untreated pollen, resulting in less germination and more bursting. Apparently, irradiation caused the pollen cell membrane to lose its flexibility faster than normal. Rehydration of dry-stored, irradiated pollen in water-saturated air restored germination percentages up to their initial levels. The importance of this procedure in germination trials is stressed.     

Cytoplasmic male sterility,resistance to gall mite and mildew,and season of leafing out in black currants

Summary Cytoplasmic male sterility (cms) is described in the F₁ hybrids Ribes × carrierei (R. glutinosum albidum × R. nigrum) and R. sanguineum × R. nigrum. In backcrosses to R. nigrum, progenies with R. glutinosum cytoplasm were either all male sterile, or segregated for full male fertility (F) and complete (S) and partial (I) male sterility. Ratios of F:I+S suggested that two linked genes controlled cms, F plants being dominant for one (Rf ₁) and recessive for the other (Rf ₂).Segregation for cms in relation to three linded genes, Ce (resistance to the gall mite, Cecidophyopsis ribes), Sph ₃(resistance to American gooseberry mildew, Sphaerotheca mors-uvae) and Lf ₁(one of two dominant additive genes controlling early season leafing out) indicated that Rf ₁and Rf ₂were in this linkage group. The gene order and approximate crossover values appeared to be: % MathType!MTEF!2!1!+-% feaafiart1ev1aaatCvAUfeBSjuyZL2yd9gzLbvyNv2CaerbuLwBLn% hiov2DGi1BTfMBaeXafv3ySLgzGmvETj2BSbqef0uAJj3BZ9Mz0bYu% H52CGmvzYLMzaerbd9wDYLwzYbItLDharqqr1ngBPrgifHhDYfgasa% acOqpw0xe9v8qqaqFD0xXdHaVhbbf9v8qqaqFr0xc9pk0xbba9q8Wq% Ffea0-yr0RYxir-Jbba9q8aq0-yq-He9q8qqQ8frFve9Fve9Ff0dme% aabaqaciGacaGaamqadaabaeaafaaakeaacaWGdbGaamyzamaamaaa% baGaaiiiaiaacccacaGGWaGaaiOlaiaacgdacaGG0aGaaiiiaiaacc% caaaGaaiiiaiaacccacaGGGaGaamOuaiaadAgaliaaigdakmaamaaa% baGaaiiiaiaacccacaGGGaGaaiiiaiaaccdacaGGUaGaaiOmaiaacs% dacaGGGaGaaiiiaiaacccacaGGGaGaaiiiaaaacaWGsbGaamOzaSGa% aGOmaOWaaWaaaeaacaGGGaGaaiiiaiaacccacaGGGaGaaiiiaiaacc% cacaGGGaGaaiiiaiaacccaaaGaamitaiaadAgaliaaigdakmaamaaa% baGaaiiiaiaacccacaGGGaGaaiiiaiaacccacaGGGaGaaiiiaiaacc% cacaGGGaGaaiiiaiaacccacaGGGaaaaiaadofacaWGWbGaamiAaSGa% aG4maaaa!6E4D!\[Ce\underline { 0.14 } Rf1\underline { 0.24 } Rf2\underline { } Lf1\underline { } Sph3\]. Crossover values of 0.36 for Ce-Lf ₁, and 0.15 for Lf ₁-Sph ₃were estimated from the relative mean differences in season of leafing out between seedlings dominant and recessive for Ce and Sph ₃.It is suggested that competitive disadvantage of lf ₁-carrying gametes and/or zygotes at low temperatures may be implicated in the almost invariable deficit of plants dominant for the closely linked mildew resistance allele Sph ₃. Poor performance of lf ₁- (and possibly lf ₂-) carrying gametes and young zygotes during periods of low temperature at flowering might also account for the liability of some late season cultivars and selections to premature fruit drop (running off).     

Optimization of Extraction Technology and Determination of Content of Total Phenols of Leaves in Acanthopanax giraldii Harms

[Objectives] To determine the optimum extraction technology for total phenols of leaves in Acanthopanax giraldii Harms.[Methods]The single factor test and ortho...     

Screening techniques and sources of resistance to parasitic angiosperms

Parasitic angiosperms cause great losses in many important crops under different climatic conditions and soil types. The most widespread and important parasitic angiosperms belong to the genera Orobanche, Striga, and Cuscuta. The most important economical hosts belong to the Poaceae, Asteraceae, Solanaceae, Cucurbitaceae, and Fabaceae. Although some resistant cultivars have been identified in several crops, great gaps exist in our knowledge of the parasites and the genetic basis of the resistance, as well as the availability of in vitro screening techniques. Screening techniques are based on reactions of the host root or foliage. In vitro or greenhouse screening methods based on the reaction of root and/or foliar tissues are usually superior to field screenings and can be used with many species. To utilize them in plant breeding, it is necessary to demonstrate a strong correlation between in vitro and field data. The correlation should be calculated for every environment in which selection is practiced. Using biochemical analysis as a screening technique has had limited success. The reason seems to be the complex host-parasite interactions which lead to germination, rhizotropism, infection, and growth of the parasite. Germination results from chemicals produced by the host. Resistance is only available in a small group of crops. Resistance has been found in cultivated, primitive and wild forms, depending on the specific host-parasite system. An additional problem is the existence of pathotypes in the parasites. Inheritance of host resistance is usually polygenic and its transfer is slow and tedious. Molecular techniques have yet to be used to locate resistance to parasitic angiosperms. While intensifying the search for genes that control resistance to specific parasitic angiosperms, the best strategy to screen for resistance is to improve the already existing in vitro or greenhouse screening techniques.