猪流感抗体间接ELISA检测方法的建立

猪流感病毒A／Swine／Fujian／668／2001（H3N2）株感染的鸡胚尿囊液，经差速离心后，再经蔗糖密度梯度离心，提纯、纯化的猪流感病毒经NP-40处理并反复冻融，作为猪流感间接ELISA抗原，确立了间接ELISA检测方法。对29份HI试验猪流感为阴性的血清进行了检测，经统计学分析，确定间接ELISA判定标准，被检血清OD490nm值≥0．20判定为阳性。该方法对猪瘟等11种猪疫病阳性血清无交叉反应，批内和批间重复试验的吸收变异系数分别在3．34％～8．12％和6．2％～9．04％之间。与HI的符合率达到92．8％，经卡方检验（P〈0．01）比HI试验敏感。为猪流感抗体检测提供了快速、准确、简便的方法。     

模猪场猪瘟野毒感染情况调查

利用ELISA方法对广西6个规模猪场送检的1626份猪血清进行猪瘟野毒感染情况调查。结果6个规模猪场均存在猪瘟野毒感染,其中母猪感染率为7.86%~29.21%,平均为17.52%,种公猪感染率为0%~23.52%,平均为11.83%,育肥猪感染率为5%~22.45%,平均为15.5%,断奶仔猪感染率为8.24%~18.57%,平均为12.69%。此检测结果与猪场临床发病情况基本一致,病猪多表现为繁殖障碍型、温和型的非典型猪瘟。     

Serological survey of exposure to Erysipelothrix rhusiopathiae in poultryin New Zealand

Abstract

AIMS: To estimate the seroprevalence of antibodies to Erysipelothrix rhusiopathiae in chickens in New Zealand, and to estimate the effect of housing type, geographical location and age on seroprevalence.

METHODS: A cross-sectional serological survey of a convenience sample of 545 broiler, breeder, and layer chickens in 55 flocks was conducted in 2010–2011. Birds were aged 5–83 weeks; housing types were free-range, shed, caged, and unknown; and flocks were located in the Auckland, Manawatu, North Canterbury, Otago, Taranaki, Waikato, and Wairarapa regions of New Zealand. An ELISA was used to measure antibodies to E. rhusiopathiae. Samples with an optical density reading ≥1.50 were considered to be positive. Logistic regression analysis was used to model the effect of housing type, geographical location and flock age on the prevalence of samples positive for antibodies to E. rhusiopathiae.

RESULTS: The overall prevalence of samples with antibodies to E. rhusiopathiae was 39.8 (95% CI=35.68–44.06)% for the 545 samples, and 46/55 (84%) farms that were tested had at least one positive sample. Mean seroprevalence for types of housing was 44.2 (95% CI=37.79–50.70)% for free-range (n=240 birds), 23.7 (95% CI=17.83–30.38)% for shed (n=190), 73 (95% CI=56–86)% for caged (n=37) and 50 (95% CI=38–62)% for unknown (n=78). The disease was present in all seven geographical locations from which samples were obtained for this study. Seroprevalence increased with increasing age of birds (p<0.001); for birds ≤12 weeks of age it was 2 (95% CI=0.3–8)% (n=91), 13–24 weeks 29.1 (95% CI=23.34–35.46)% (n=230), 25–36 weeks 47 (95% CI=32–64)% (n=40), 37–48 weeks 75 (95% CI=51–91)% (n=20), >48 weeks 63.8 (95% CI=54.78–72.12)% (n=127). Neither housing type nor geographical location had a significant effect on the likelihood of samples being positive for antibodies to E. rhusiopathiae.

CONCLUSIONS: The results of this study suggest the prevalence of erysipelas under New Zealand field conditions may be higher than expected and that the disease is significantly associated with increasing age. Housing type and geographical location appear to be unrelated to seroprevalence.

CLINICAL RELEVANCE: Further study of the epidemiology of E. rhusiopathiae in chickens in New Zealand should be considered in order to minimise the extent of birds' exposure to the organism. These findings will assist in the design of further studies.     

2011年～2012年我国不同地区猪流感血清学监测

为了解猪流感病毒(SIV)在我国的流行情况,本研究对2011年～2012年从不同省市重点地区屠宰场、规模化猪场和个体养殖户采集猪血清样品13044份,以经典H1N1 (CS)、类禽H1N1 (EA)、甲型H1N1(pdm/09)、H3N2、H5N1、H9N2亚型SIV抗原并采用血凝抑制(HI)方法检测.结果显示,2011年～2012年我国猪血清中类禽H1N1亚型SIV抗体平均阳性率比其他亚型高,分别为42.24％和45.89％,2011年甲型H1N1亚型SIV血清抗体平均滴度(GMT)比其他亚型高,GMT为53.32,2012年经典H1N1 GMT比其他亚型高,GMT为73.2.结果表明2011年～2012年我国猪群中H1N1亚型SIV感染较为普遍.     

广东省部分地区规模化猪场猪流感病毒感染的血清学调查与分析

为了解广东省规模化猪场猪流感病毒(SIV)感染情况,本研究采用ELISA方法和血凝抑制试验(HI),对2011年～2012年采集的1 050份血清样品进行SIV血清学检测.两种检测方法结果显示1 050份血清样品中SIV抗体阳性率分别为50.4％(ELISA)和50.2％(HI).其中珠三角地区和粤东地区的感染率高于粤西地区.SIV亚型调查结果显示该地区流行的SIV主要为H1和H3亚型,抗体阳性率分别为39.2％和18.2％(ELISA).部分猪场存在H9亚型SIV抗体.部分猪群中同时存在H1和H3两种亚型SIV抗体,表明猪群中存在不同亚型SIV混合感染.本研究为广东省猪流感的预防提供参考依据.     

1株猪源H9N2亚型流感病毒的分子特征

旨在进一步了解山东省猪流感的流行情况及其病原特征,笔者于2019年春季,在山东省泰安某屠宰场采集130份猪鼻拭子,进行病毒分离鉴定;并对分离病毒进行全基因组测序和分子特征分析;用禽H9N2亚型标准抗原联合血凝抑制方法检测2018—2019年从山东省8个地区猪场采集的1 527份猪血清样品中的猪流感病毒抗体。结果显示:分离到1株H9N2亚型流感病毒,命名为A/swine/Shandong/TA009/2019(H9N2)。分离病毒与A/environment-air/Kunshan/NIOSH-BL20/2018(H9N2)和A/environment-air/Kunshan/NIOSH-BL25/2018(H9N2)遗传关系最近,其基因片段的核苷酸相似性均在99.5%以上。分离病毒的HA和NA基因属于Y280-like分支,PB2和M基因属于G1-like分支,PB1、PA、NP和NS基因属于SH/F98-like分支。分离病毒HA蛋白裂解位点处的氨基酸序列为\"PSRSSR/GL\",符合低致病性禽流感病毒的分子生物学特性。HA蛋白的216位为L,具有结合人源唾液酸α2,6-Gal的能力。血清学分析结果显示,9份血清中H9N2抗体呈阳性,其总阳性率为0.59%。综上:本研究分离到1株猪源H9N2亚型流感病毒,并在猪血清中检测到H9N2抗体,提示应加强对猪流感的流行情况及其病原特征的持续监测。     

Detection of Taenia solium Cysticercosis in Pigs by ELISA with an Excretory-Secretory Antigen

Serodiagnosis of Taenia solium cysticercosis in pigs was conducted by an enzyme-linked immunosorbent assay with an excretory-secretory antigen. The antigen was obtained by in vitro cultivation of the cysticerci in a synthetic medium RPMI 1640. The sensitivity and specificity of the ELISA in detecting infection in pigs reared on free range was 92% and 100%, respectively. In addition, 33.33% of pigs in which infection could not be detected at meat inspection were found positive by ELISA. However, none of the sera from a group of farm-reared pigs were positive. No cross reactions were observed in pigs that contained either the cysticerci of Taenia hydatigena or hydatid cysts.     

H3亚型猪流感病毒抗原捕捉ELISA检测方法的建立

为快速检测H3亚型猪流感病毒(SIV),本研究将一株H3N2亚型SIV特异性单克隆抗体(MAb)进行纯化,建立了以多抗作为包被抗体,MAb作为检测抗体的H3亚型SIV抗原捕捉ELISA (AC-ELISA)方法.并对该方法的特异性和敏感性进行了分析,其敏感性是血凝试验的4倍,与猪繁殖与综合征病毒、猪圆环病毒、猪瘟病毒、猪细小病毒以及H1、H5、H9亚型SIV均没有明显的交叉性反应,批间和批内重复试验变异系数均小于10％.应用建立的AC-ELISA方法对84份H3亚型SIV实验室感染样品进行检测与鸡胚滴定病毒结果符合率达90.47％.本实验建立的AC-ELISA方法具有良好的特异性和敏感性,可用于H3亚型SIV的临床检测.     

四川农村猪圈舍改造方法

本文针对四川农村猪圈舍存在的阴暗、无仔猪保温设施、母猪配种难等实际问题,力求经济实用、科学合理等原则,初步提出以种猪圈舍为重点的四川农村猪圈舍改造意见。     

三种禽流感灭活疫苗对农村散养鸭的免疫效果比较

采用重组禽流感灭活疫苗(H5N1亚型,Re-1株)、禽流感灭活疫苗(H5亚型,N28株)、禽流感H5-H9二价灭活疫苗三种疫苗免疫农村散养鸭,结果表明,在一次免疫和加强免疫中,重组禽流感灭活疫苗、禽流感灭活疫苗的抗体效价都分别明显高于H5-H9二价灭活疫苗同期的H5抗体效价,因此对水禽免疫宜使用重组禽流感灭活疫苗或禽流感灭活疫苗;加强免疫明显比一次免疫效果好,应用禽流感灭活苗对散养鸭免疫,必须加强方可达到理想的免疫效果。