应用酶联免疫吸附试验检测猪囊虫病的研究

在猪囊虫病的研究中发现在病、健猪血清中除存在着某些抗原性相同的成分引起交叉反应外,在病猪血清中还含有一种特异循环抗原成分。为了弄清囊虫病猪对此循环抗原的免疫应答状况,自囊虫虫浆浸提物(粗     

应用猪囊虫匀浆抗原间接血凝试验检测猪囊虫病抗体的研究

从囊虫匀浆浸提液中分离出一种抗原性与特异循环抗原相同的成分,以这种抗原成分通过间接血凝试验检测病、健猪的血清抗体,结果病猪抗体阳性率为93.2％(96/103),健猪阴性(?)合率为100％。     

酶标葡萄球菌A蛋白染色法用于猪囊尾蚴病诊断的初步研究

用自制的猪囊尾蚴头节“颗粒性抗原”,以HRP—SPA作为第二抗体,进行间接免疫酶染色诊断猪囊尾蚴病。检测61份猪囊尾蚴病血清,阳性59份,检出率为96.6％;健康猪血清69份、细颈囊尾蚴病和肺丝虫病猪血清各10份,均为阴性。此法简便易行,快速、准确,适于基层使用。     

应用McAb—RPHA检测囊虫病猪循环抗原

应用单克隆抗体致敏红细胞,以反向被动血凝试验(RPHA)检测囊虫病猪血清中的循环抗原.囊虫病猪血清的检出率为94.55%(104/110),健猪血清阳性率为8.5%(8/94).假阳性反应主要来自棘球蚴感染.囊虫病猪血清中的循环抗原可耐受100℃30min而仍保持其抗原活性,提示其可能是一种多糖类物质.     

猪衣原体病和布鲁氏菌病血清学调查

应用间接血凝法对来自５个省（湖北、广东、河南、江西、海南）的４４４份猪血清和用虎红平板凝集法对湖北省７个猪场的３４０份猪血清分别血清分别进行衣原体病和布鲁氏菌病血清学调查。结果,衣原体病抗体阳性率平均为１７．０％,各猪场的感染率差异较林;衣原体病血清抗体阳性猪场达到６７．８％;检查了２个猪场不同年龄段的猪,衣原体病抗体阳性率也不同;布鲁氏菌病抗体阳性率平均为９．７％,所检的７个猪场中有４个猪场为阴     

2008～2010年华东部分地区猪圆环病毒2型感染血清学调查

为了解华东地区近三年来猪圆环病毒病（porcine circovirus disease,PCVD）的流行状况,以便更好地控制该病的发生,2008年1月～2010年9月在华东地区5个省、直辖市及河南省的68个规模化猪场（均未免疫商品化猪圆环病毒疫苗）共收集1017份猪血清样品,应用商品化猪圆环病毒（Porcine circovirus,PCV）抗体诊断试剂盒（PCV2-ELISA）,对送检血清样品进行PCV2抗体水平检测。结果显示：送检猪血清中抗体阳性总数为609份,总阳性率为59.88%;2008、2009和2010年送检样品的阳性率分别为53.4%、35.33%和80.63%,PCV血清抗体阳性率呈先下降后上升的趋势;成年猪血清阳性率最高为81.40%,保育猪血清阳性率最低为41.48%,仔猪和育肥猪血清阳性率相近,分别为57.95%和52.20%,说明仔猪断奶后随母源抗体下降其抗体阳性率亦下降,但随猪龄的增长而感染逐渐加重后其抗体阳性率又升高,且不同生长阶段PCV毒抗体阳性率有明显差异。     

甘肃省猪传染性胸膜肺炎病和猪弓形体病的血清学调查

1998～2000年，对全省11个地市22个猪场24批826份猪血清，应用郑州兽医研究所生产的间接血凝诊断液进行了猪传染性胸膜肺炎病的抗体检测。结果表明，抗体阳性率全省平均为87.16％，其中生产母猪群、种公猪群、肥育猪群和后备猪群的抗体阳性率分别为97．40％、85.71％、77．63％和70．09％。同时对全省7个地市140份猪血清进行了猪弓形体病的间接血凝抗体检测，结果在所童猪血清中未见一例阳性。     

应用ELISA间接法检测猪流行性腹泻抗体的研究

应用猪流行性腹泻病毒(PEDV)吉(J)毒株猪胎肠单层细胞培养物,以冻融法制备抗原,建立了酶联免疫吸附试验(ELISA)间接法检测PED抗体的方法。对30头份经直接免疫荧光技术证实的PED病猪群血清测定,97％为阳性;检测32头份无PED猪场猪血清,93％为阴性。对1份PED“华毒株”,1份“川毒株”以及3头份“吉毒株”PED免疫血清测定,均为阳性。对猪传染性胃肠炎血清、猪轮状病毒病血清、疑似猪血球凝集性脑脊髓炎血清、鸡传染性支气管炎血清及猪梭菌性肠炎血清共16头份检测均为阴性,证明PEDV不与TGEV等其它3种冠状病毒血清发生交叉反应。89头份疫区猪血清的区域性试验阳性率为75％(67/89);对82头份屠宰猪血清测定,53％阳性。抗原包被板保存期试验表明,包被板在-20℃可保存2个月。     

ELISA和DIA对猪囊虫病诊断的比较

应用ELISA和DIA两种方法,采用同一抗原,同一结合物,对956份猪血清进行了猪囊虫病的检测对比试验。结果,DIA和ELISA的特异性分别为99.57％与98.88％,敏感性分别为98.39％和97.18％,无明显差异。但在抗体滴度、操作时间、抗体消长规律、诊断液的用量等方面,DIA明显优于ELISA。     

四川省部分种畜(猪)场猪细小病毒病血清抗体的检测

猪细小病毒是引起母猪繁殖障碍的主要病原体之一。以胚胎和胎儿感染及死亡为特征。实践中，本病以预防为主，采用猪细小病毒疫苗免疫。采用乳胶凝集抗原，对我省部分种畜（猪）场的猪血清进行了猪细小病毒病血清抗体检测，结果显示，平均阳性率为56％，其中阳性率最高达93％，最低为14．71％。     

囊虫病猪循环抗原的研究(初报)

用琼脂凝胶双扩散试验发现了囊虫病猪的循环抗原,并用反向间接血凝技术作了检测。103份囊虫病猪血清琼扩试验对循环抗原的检出率为22.33％(23/103);反向间接血凝试验检出率为32.03％(33/103),其余70份呈阴性反应的血清经免疫复合物解离后,又有25份呈阳性反应,总检出率为56.31％(58/103)。     

抗猪囊虫循环抗原杂交瘤细胞系的建立和鉴定

建立了8株抗猪囊虫循环抗原(CA)的杂交病细胞系。其中6F12和2E7为抗囊虫特异McAb;McAb 1C6、1C7、1B5、4D9、2B9和8D8与囊虫、(?)球蚴、细颈囊尾蚴抗原均可发生反应。这些McAb的腹水ELISA效价为105～107,细胞培养上清液效价为102,并均可与病猪血清中的囊虫CA反应形成沉淀线。用1B5、6F12和8D8分别致敏血球,以反向间接血凝试验检测98份囊虫病猪血清,检出率分别为70.41%(69/98)、6.12%(6/98)和7.14%(7/98)。本研究制备的McAb可用于猪囊虫循环抗原检测。     

Effects of sarcoptic mange on serum proteins and immunoglobulin G levels in chamois (Rupicapra pyrenaica) and Spanish ibex (Capra pyrenaica)

Three groups of chamois (Rupicapra pyrenaica) and three groups of Spanish ibex (Capra pyrenaica) were established to study the effects of sarcoptic mange on serum proteins and immunoglobulin G (IgG) levels. The first group of chamois consisted of 22 healthy Pyrenean chamois (R. pyrenaica pyrenaica) from a non-infested area, the second group consisted of 20 healthy Cantabrian chamois (R. p. parva) from an area where sarcoptic mange has been reported since 1994 and the third group consisted of 16 Cantabrian chamois from the same area but naturally infested by Sarcoptes scabiei. The first group of Spanish ibex was 39 healthy animals from a sarcoptic mange non-infested area, the second group was 23 healthy animals from a sarcoptic mange infested area and the third group consisted of 20 animals from the same area but naturally infested with the parasite. Blood samples were taken after killing the animals as part of hunting programmes. Values for total proteins, gamma-globulin and IgG were higher in infested and healthy chamois from the infested area compared to healthy chamois from the non-infested area, and IgG levels were higher in infested chamois compared to healthy-exposed chamois. Values for alpha2-globulin were higher in healthy Cantabrian chamois. In Spanish ibex, albumin, alpha2-globulin and IgG levels were lower in the healthy Spanish ibex from the non-infested area than in healthy animals from an infested area. The differences found in the chamois were indicative of the establishment of a humoral antibody response in the animals in contact with the disease. As the IgG levels were not significantly different between healthy and infested Spanish ibex from the same area, a different pattern of chronic infection with humoral response to the disease was suggested.     

Experimental sarcoptes scabiei infestation in pigs: (2) Effects on production.

A study on the development and effect of experimental Sarcoptes scabiei var suis infestations in growing pigs is described. Pigs were infested at either weekly or fortnightly intervals throughout each experimental and individual growth rates and feed conversion ratios were determined. The animals were fed diets which contained either optimal or sub-optimal levels of protein. They were housed either intensively or extensively. In all experiments the majority of infested animals developed a generalised hypersensitivity to sarcoptes mites and performed significantly less efficiently than non-infested littermates. Mean growth rates were depressed from 9.2 to 12.5 per cent and feed conversion efficiencies by a similar margin. Well fed, intensively housed pigs developed a more severe hypersensitivity reaction than poorly fed, extensively housed pigs.     

Antigens from Rhipicephalus sanguineus ticks elicit potent cell-mediated immune responses in resistant but not in susceptible animals

In the present study we compared the immunological reactions between Rhipicephalus sanguineus tick-infested susceptible (dogs and mice) and tick-resistant hosts (guinea pigs), elucidating some of the components of efficient protective responses against ticks. We found that T-cells from guinea pigs infested with adult ticks proliferate vigorously in the presence of concanavalin A (ConA), whereas ConA-induced cell proliferation of tick-infested mice and dogs was significantly decreased at 43.1 and 94.0%, respectively, compared to non-infested controls. Moreover, cells from mice and dogs submitted to one or three successive infestations did not exhibit a T-cell proliferative response to tick antigens, whilst cells from thrice tick-infested guinea pigs, when cultured with either a tick extract or tick saliva, displayed a significant increase in cell proliferation. Also, we evaluated the response of tick-infested mice to a cutaneous hypersensitivity test induced by a tick extract. Tick-infested mice developed a significant immediate reaction, whereby a 29.9% increase in the footpad thickness was observed. No delayed-type hypersensitivity (DTH) reaction was detected. Finally, the differential cell count at the tick attachment site in repeatedly infested mice exhibited a 6.6- and 4.1-fold increase in the percentage of eosinophils and neutrophils, respectively, compared to non-infested animals, while a decrease of 77.0-40.9 in the percentage of mononuclear cells was observed. The results of the cutaneous hypersensitivity test and the cellular counts at the tick feeding site for mice support the view that tick-infested mice develop an immune response to R. sanguineus ticks very similar to dogs, the natural host of this species of tick, but very different from guinea pigs (resistant host), which develop a DTH reaction in addition to a basophil and mononuclear cell infiltration at the tick-attachment site. In conclusion, saliva introduced during tick infestations reduces the ability of a susceptible animal host to respond to tick antigens that could stimulate a protective immune response. As a consequence, the animals present a lack of DTH response and disturbed cellular migration to tick feeding site, which can represent a deficient response against ticks.     

Validation of ELISAs for the detection of antibodies to Sarcoptes scabiei in pigs

An Enzyme-linked ImmunoSorbent Assay (ELISA) was developed for the detection of antibodies to Sarcoptes scabiei. This 'Animal Health Service'-ELISA (AHS-ELISA) was compared with a commercial test (Checkit(R) Sarcoptest) using experimental and field sera. The experimental study was a contact infestation experiment. Eighty piglets were randomly divided between the experimental and control group. After introduction of three Sarcoptes scabiei var. suis infested pigs in the experimental group, both groups were monitored by determining scratching indices, taking ear scrapings and blood samples in Weeks 0, 2, 4, 6, 8, 12 and 16. Four pigs in the control group were immunised with either Dermatophagoides pteronyssinus (Dp) antigens (n=2), or Acarus siro (As) antigens (n=2). In the control group all (non-immunised) pigs were negative in all tests. In the experimental group only slightly elevated scratching indices were observed, with a maximum in Week 8. After 2 weeks for the first time an ear scraping was positive (2.5%). In Week 8 the highest number of positive ear scrapings were found (25.0%). Positive results in the Sarcoptest were first obtained in Week 12 (10.5% positive), while eventually 29.0% of the finishing pigs were positive after 16 weeks. The AHS-ELISA first detected a serological response after 6 weeks (5. 0% positives), increasing until after 16 weeks a large proportion (74.2%) of the finishing pigs were seropositive, making the AHS-ELISA the most sensitive test. In the AHS-ELISA one As-immunised pig remained seronegative, but the other hyper-immunised pigs crossreacted. In the Sarcoptest, only Dp-immunised pigs had elevated Optical Densities (OD's) albeit below the cut-off level. Although hyper-immunisation is not a representation of field conditions, it cannot be excluded that the AHS-ELISA is not 100% specific.Field samples were taken from 20 sows in 30 herds, classified as mange-free, suspect, or infested. On a herd level there was high agreement among the ELISAs. Both serological tests were suitable to distinguish mange-free herds from infested herds. In one infested herd the decline of maternal antibody in piglets was studied by sampling 40 piglets from 20 different litters. The lowest average OD using the AHS-ELISA was found at 5 weeks of age, followed by a significant increase at 7 weeks. The average OD with the Sarcoptest was at a minimum level at 3 weeks, but no increase was found later. For screening of herds, interference of maternal antibodies is avoided by sampling at an age of 7 weeks or older.     

抗伊氏锥虫表膜单克隆抗体的研究 Ⅱ.用抗伊氏锥虫表膜抗原McAb检测循环抗原

利用抗伊氏锥虫表膜抗原的McAb双抗体夹心法,检测了48匹健康马(骡)、5匹人工接种伊氏锥虫马(骡),9匹先用锥虫弱毒株免疫后用强毒株攻毒马(骡)共278份血清的循环抗原(CA),同时用常规间接ELISA检测了相应的循环抗体(CAb)。结果: 278份血清的CA、CAb阳性率无显著差异,但其阳性检出时机不完全吻合;人工接种马(骡)的CA阳性检出率高于CAb,于接虫后经1-2个CA高峰而死,濒死前4/5动物的CA呈现阳性,免疫马(骡)在攻虫前CA即可呈现阳性,多经1-3个CA高峰后逐渐转阴;人工接种马(骡)的CA水玉高于免疫马(骡),二次攻虫后的CA水平高于首次。     

大庆市及周边地区非免疫猪群猪圆环病毒感染情况调查

为了解大庆市及周边地区猪群中PCV的感染情况,应用ELISA和PCR技术对采自大庆市5区4县及周边地区55个村屯/猪场未免疫PCV2疫苗猪血清样本649份,5家病猪场和6家屠宰场组织样本448套,进行血清学和分子生物学检查,并对检测结果进行了比较分析。结果显示,大庆市及周边地区未免疫PCV2疫苗猪血清样品PCV2抗体平均阳性率为54.70%,其中断奶仔猪样品阳性率为50.68%,育肥猪样品为70.00%,繁育母猪样品为54.70%。组织样品PCV1阳性率为4.46%,PCV2阳性率为22.32%。调查结果可为确定地区流行毒株序列提供原始材料和制定科学防控措施提供依据。     

Experimental quantification of the transmission of Sarcoptes scabiei var. suis among finishing pigs

In this study, the rate of S. scabiei var. suis transmission among finishing pigs was quantified in a contact transmission experiment. Forty piglets originating from a mange free farrow-to-finish herd were randomly allocated to three groups and one S. scabiei var. suis infested finishing pig was subsequently added to each of these groups. After 35 days, the three seeder pigs were removed from the groups and the remaining 40 pigs were re-allocated to five pens. Ear scrapings, to be examined for mites, were collected from each pig on days 1, 14, 28, 42, 56, and 84 of the experiment. Blood samples, to be tested for antibodies against S. scabiei, were collected from each pig on days 0, 14, 28, 42, 56, 70, 84 and 112 after the introduction of the seeder pigs.From the results of the ear scrapings and the blood samples the number of susceptible (not infested) and infested pigs was derived at the time of each sample collection and the number of new infestations in the intervals between the sample collections. From these data the infestation rate parameter beta (average number of new infestations per infested pig per day) was estimated by use of a Generalised Linear Model (GLM) and accordingly, beta was estimated at 0.056 (95% CI: 0.037-0.085) infestations per infested pig per day.Next, by use of beta, the transmission of S. scabiei was simulated in a population of 100 finishing pigs for 100 days after the introduction of a single infested pig. For this purpose, 500 simulations were done. The 90% confidence interval of the number of infested pigs at day 100 ranged from 12 to 88 (median: 63). It was concluded that transmission of S. scabiei among finishing pigs is slow. Due to the presumed lower contact rate between sows as compared to finishing pigs, it is anticipated that transmission of S. scabiei among sows will even be slower than among finishers These findings are of particular interest for the development of surveillance programmes for S. scabiei free herds.