Prokaryotic Expression and Polyclonal Antibody Preparation of African Swine Fever Virus Georgia 2007/1 Strain CD2v Protein

The study was aimed to express the EP402R gene of African swine fever virus (ASFV) Georgia 2007/1 strain via prokaryotic expression system,obtain the recombinant CD2v protein,and prepare polyclonal antibodies against the purified recombinant CD2v protein.After codon optimization,ASFV EP402R full-length gene was linked into pET-28a(+) expression vector to construct prokaryotic recombinant expression plasmid.After induction by 1 mmol/L IPTG at 16 ℃ for 12 h,the recombinant protein was identified by SDS-PAGE and Western blotting.The purified recombinant CD2v protein was used as immunogen to prepare mouse anti-CD2v polyclonal antibodies.The antibody titer was measured by indirect ELISA and the specificity was further analyzed by indirect immunofluorescence assay (IFA) and Western blotting.The results showed that ASFV EP402R gene was successfully cloned into pET-28a(+),and pET-28a-EP402R was obtained.The recombinant plasmid was transformed into E.coli BL21(DE3) for expression,the recombinant protein was expressed mainly in the form of inclusion bodies,with molecular mass at about 47 ku,while some of the recombinant protein could also exist in a soluble form.Western blotting results showed that the purified protein had good immunoreactivity.The indirect ELISA result showed that the polyclonal antibodies had a high titer of 1:512 000,IFA and Western blotting results indicated that it could specifically recognize recombinant CD2v protein.These results confirmed the recombinant CD2v protein expressed via prokaryotic system had good immunogenicity,and the prepared polyclonal antibodies had high titer and specificity.This research provided technical support for further study of ASFV EP402R biological function,as well as its gene-deletion based vaccine development.     

猪流行性腹泻IgA、IgG抗体与PEDV抗原相关性研究

为了研究仔猪所吮乳汁中IgA抗体水平与乳汁中PEDV感染情况,探讨母猪免疫猪流行性腹泻疫苗后乳汁与血清中IgA、IgG抗体水平的相关性,临床采集多家规模化猪场母猪群乳汁、血清样品以及发病仔猪所吮母猪乳汁样品。采用实验室已经建立的IgA、IgG抗体ELISA方法,检测临床上猪流行性腹泻疫苗免疫后IgA抗体与IgG抗体的相关性,同时采用RT-PCR方法检测发病仔猪所吮乳汁中PEDV感染情况。结果表明,免疫猪流行性腹泻活疫苗后母猪乳汁中IgA抗体水平与血清中IgA、IgG抗体水平呈现很好的正相关性;当发病仔猪所吮母猪乳汁IgA抗体检测结果为阴性时,其PEDV抗原检测结果为阳性;乳汁IgA抗体检测结果趋于阳性样品临界值时,其PEDV抗原检测结果亦为阳性。结论:乳汁中低水平的IgA抗体很难有效地保护仔猪抵御PEDV感染;乳汁中IgA抗体与血清IgA、IgG抗体呈现很好的相关性,且乳汁中IgA抗体水平远远高于血清;通过IgG抗体水平可以间接反映乳汁中的IgA抗体水平,在初乳样品采集难度较大的情况下,可用于间接评估猪流行性腹泻免疫保护水平。     

Maternal behaviour and performance in first-parity outdoor sows

When sows and their litters are kept outdoors, much of the responsibility for the litter is shifted from the herdsman to the sow compared to when the sows are kept indoors. Therefore, the maternal ability, including maternal behaviour, is believed to be important in outdoor piglet production. The aim of this study was to describe maternal traits in conventionally bred first-parity sows kept outdoors. Nursing behaviour, sow activity, sow body reserves, litter size and piglet growth were studied in 40 first-parity sows during a seven week long lactation. The sows were kept in groups outdoors. Sow activity (lying down or active) and nursing behaviour (nursing frequency, nursing duration and nursing terminator) were recorded on videotapes at four days post partum (pp) and directly observed at four and six weeks pp. Cross suckling was observed at four and six weeks pp. Sows were weighed and measured for backfat depth five days before farrowing, two weeks pp and at weaning seven weeks pp. Piglets were weighed at four days pp, at two weeks pp and at weaning. We concluded that sow nursing behaviour and activity are individual characteristics repeatable within sows' late lactation. Less active sows are more available for suckling. Light and thin sows have an earlier and more progressed weaning process than heavier, fatter sows. A significant positive relation was found between sow appetite in early lactation and piglet growth until weaning. Sow nursing behaviour is not important for piglet growth when sows and piglets are held in groups, piglets have access to sow feed and piglets are weaned as late as at seven weeks of age.     

Genetic parameters for maternal behaviour traits in sows

Data from 32 nucleus and multiplier herds in Germany was used to estimate variance components and breeding values for five maternal behaviour traits in sows. The estimation was performed univariately using an animal threshold model. About 31,000 farrowings recorded from December 2003 until July 2005 were included. The heritability coefficients were 0.07 (0.06) for group behaviour, 0.06 (0.03) for attitude to people, 0.05 (0.01) for maternal ability, 0.03 (0.01) for crushing of piglets and 0.02 (0.02) for savaging of piglets. Additionally, genetic correlations between the behaviour traits and between the behaviour traits and litter size, respectively, were estimated multivariately by REML with a linear model. Low heritability and weak genetic correlation to litter size at birth indicate that it may be difficult to genetically improve the maternal behaviour, and that selection for better mothering ability is not necessarily accompanied by reduced litter size at birth.     

Detection and recovery of Rhizoctonia solani in naturally infested glasshouse soils using a combined baiting, double monoclonal antibody ELISA

A combined baiting, double monoclonal antibody immunoassay was developed that allows specific and sensitive detection of the economically important soil-borne plant pathogen Rhizoctonia solani in naturally infested soils. The assay is quick, taking only three days to complete from receipt of soil samples and the immunoassay format allows recovery of Rhizoctonia isolates from colonized baits for determination of anastomosis group (AG) affiliation and pathogenicity. The assay was tested on naturally infested soils from commercial glasshouses used to grow lettuce. Using the immunoassay, conventional anastomosis tests against known AG isolates, and pathogenicity tests, it was shown that R. solani isolates recovered from soil samples were pathogenic towards lettuce and belonged to AG4. Furthermore, those isolates that exhibited strong pathogenicity towards lettuce were recovered from sites that had experienced severe Rhizoctonia damage in previous lettuce crops. The possibility of developing a preplanting test to predict damage to specific crop plants due to the presence of particular AGs in the soil is discussed.     

狐阴道加德纳氏菌疫苗免疫狐血清抗体消长规律

应用ＤｏｔＥＬＩＳＡ检测狐阴道加德纳氏菌疫苗免疫狐的抗体水平及其哺乳仔狐的母源抗体水平。结果表明,接苗后４天开始产生抗体,７天阳转率７２７％,１０天时免疫狐１００％抗体阳转,２１～３０天抗体效价达高峰。接种后６个月血清抗体仍在１∶６４０以上,哺乳仔狐母源抗体随日龄增长而逐渐降低。３０日龄前,平均抗体效价（ＧＭＴ）为１∶２２６,以后逐渐降低。５０日龄抗体效价只有１∶１２。通过免疫抗体与攻毒保护相关性测试,抗体效价在１∶３２０以上,能抵抗１００个ＩＤ５０狐阴道加德纳氏菌强毒攻击。     

植物精油对广西灵山土鸡生长性能、养分消化率和血清抗体指标的影响

试验旨在研究饲粮中添加植物精油对广西灵山土鸡生长性能、养分表观消化率、血清抗体指标的影响.将1800只30日龄、体重(284.09±1.18)g、体况良好的广西灵山土鸡,随机分为3组,每组3个重复,每个重复200只鸡,空白对照组饲喂基础饲粮,混合植物精油组、抗生素组分别在基础饲粮中添加100 mg/kg混合植物精油、3...     

三甲氧苄氨嘧啶单克隆抗体制备以及酶联免疫试剂盒的研究

[目的]探讨系统地检测水质中三甲氧苄氨嘧啶残留量的方法.[方法]通过三甲氧苄氨嘧啶与马来酸酐反应,得到三甲氧苄氨嘧啶半抗原,再通过免疫动物得到抗三甲氧苄氨嘧啶的单克隆抗体,并将其应用于能够检测水质中三甲氧苄氨嘧啶残留量的ELISA试剂盒.[结果]该试剂盒对水质中三甲氧苄氨嘧啶的检测限为2.34 μg/kg,IC50(50％抑制浓度)为4.8 μg/L,回收率为60.5％～79.7％,试剂盒的标准曲线范围为0～80 μg/L,批内、批间的相对标准偏差均小于10％.三甲氧苄氨嘧啶单克隆抗体与二甲氧苄氨嘧啶的交叉反应率小于1％.4℃下能够保存12个月,稳定性较好.[结论]研究结果为监管三甲氧苄氨嘧啶的滥用提供参考依据.     

猪圆环病毒2型Cap蛋白单克隆抗体的制备

本研究利用已构建的基因工程重组菌表达猪圆环病毒2型(porcine circovirus type 2,PCV2)ORF2编码的Cap蛋白,纯化后作为免疫原,免疫8周龄BALB/c小鼠,取其脾细胞与SP2/0骨髓瘤细胞融合。经间接免疫荧光试验(IFA)筛选及有限稀释法进行3次亚克隆后,本试验最终获得5株稳定分泌抗PCV2 Cap蛋白单克隆抗体的杂交瘤细胞,分别命名为3D12、4D5、4B9、4C9和4G10。其中4D5为IgG2b亚型,其余4株单克隆抗体均为IgG1亚型,轻链类型均为κ。Western blotting鉴定结果表明获得的5株单克隆抗体均不能特异性的识别47 ku重组PCV2 Cap蛋白;对病毒感染细胞进行IFA试验,结果显示5株单克隆抗体均特异性的识别病毒抗原,表明5株单克隆抗体识别的抗原表位均为构象表位。中和试验结果表明,5株单克隆抗体均有中和活性。本试验结果为进一步探索ORF2基因的结构、功能及建立快速准确的诊断方法奠定了基础。