猪胸膜肺炎放线杆菌的血清型分型及毒力因子研究进展

根据荚膜多糖和脂多糖抗原表位的不同，猪胸膜肺炎放线杆菌分为15个血清型，某些血清型问存在交叉反应，不同血清型甚至同一血清型不同菌株间的毒力大小不一样，致病性也有强弱之别。该菌的致病性与多种毒力因子密切相关，主要有外毒素、外膜蛋白、脂多糖、英膜多糖、转铁蛋白、脲酶等。本文综述了国内外对胸膜肺炎放线杆菌的几种主要毒力因子及血清型的研究进展。     

An abattoir survey of pneumonia and pleuritis in slaughter weight swine from 9 selected herds. III. Serological findings and their relationship to pathomorphological and microbiological findings

Blood samples from 777 pigs, originating from 9 different herds, were collected at slaughter and examined for antibodies to Mycoplasma hyopneumoniae and Actinobacillus (Haemophilus) pleuropneumoniae by the indirect hemagglutination assay (IHA) and the complement fixation (CF) test, respectively. Results were compared to pathological and microbiological findings. Antibodies to M. hyopneumoniae in positive titers of 1/80 or higher were found in 62% of the samples. The relationship between positive IHA titers to M. hyopneumoniae and gross findings indicative of enzootic pneumonia of pigs (EPP), histological findings indicative of EPP, the isolation of M. hyopneumoniae and the demonstration of M. hyopneumoniae by indirect immunofluorescent testing ranged from 64% to 68%. No correlation was noted between positive IHA titers and the isolation of Mycoplasma flocculare. Positive antibody titers to A. pleuropneumoniae of 1/10 or higher were detected in 5% to 85% of the samples from individual herds. Positive titers to A. pleuropneumoniae serotype 2 were found in 71% to 79% of the sampled animals from herds with high frequencies of pneumonic lesions indicative of pleuropneumonia. In herds with low frequencies of pleuropneumonia, positive titers were recorded in from 0 to 4% of the tested pigs. However, no statistical association was found between pleuropneumonia and positive titers to A. pleuropneumoniae serotype 2 in individual animals. Twenty-one per cent of samples with positive CF titers to A. pleuropneumoniae showed antibodies to more than one serotype.     

A longitudinal study of serological patterns of respiratory infections in nine infected Danish swine herds

Sixteen litters of seven pigs from each of nine Danish farrow-to-finish herds were followed to investigate the serological patterns caused by natural infection with Mycoplasma hyopneumoniae, Pasteurella multocida toxin and Actinobacillus pleuropneumoniae serotypes 2, 5–7, 12. In seven of the herds, pigs were followed as two separate cohorts started 4 weeks apart, and in two herds only one cohort was followed.

A total of 999 pigs were included in the study. The pigs were blood sampled at weaning and subsequently every fourth week until slaughter. All pigs were examined for antibodies against M. hyopneumoniae (enzyme-linked immunosorbent assay), P. multocida toxin (enzyme-linked immunosorbent assay) and A. pleuropneumoniae serotypes 2, 5–7, 12 (complement-fixation tests). The most-common pattern (28%) of seroconversion was that of pigs first seroconverting to A. pleuropneumoniae serotype 2, followed by seroconversion to M. hyopneumoniae. Each herd had a dominant serotype of A. pleuropneumoniae to which most pigs seroconverted. Seroconversion to the respiratory pathogens occurred mainly in the growing-to-finishing units (8–24 weeks). The risk of seroconversion to the P. multocida toxin was very low (<20%) and occurred late.

None, four and seven herds tested seropositive to PRRS and to swine influenza virus subtypes H3N2 and H1N1, respectively, when testing 10 pigs per herd (selected randomly among the study pigs) at the age of 20 weeks.     

不同NAD含量条件下培养的猪胸膜肺炎放线杆菌对小白鼠致病性和免疫保护性的影响

将在含体积分数为0.050%、0.001%的尼克酰胺腺嘌呤二核苷酸(NAD)的LB培养基中培养的血清5型猪胸膜肺炎放线杆菌(APP),分别以3×109CFU.只-1的菌量对30日龄小白鼠进行接种,观察不同NAD含量条件下培养的APP对小白鼠致病性的影响.结果表明:用0.001%NAD培养的血清5型APP对小白鼠的致病性较强,小白鼠接种APP后,多数表现肺脏严重出血;而用0.050%NAD培养的血清5型APP接种小白鼠后,只有少数小白鼠肺脏出血严重,多数表现为轻度出血.将上述条件培养的APP按常规方法制备油乳剂灭活疫苗,分别在第3周龄和第6周龄对小白鼠进行免疫接种,在2免后3周对免疫的小白鼠进行同源攻击,并对免疫前后小白鼠的抗体效价进行检测.结果表明:分别用0.050%、0.001%NAD培养的血清5型APP制备的灭活疫苗,对小白鼠的保护率差异不显著;用0.001%NAD培养的血清5型APP制备的灭活疫苗免疫小白鼠,用血清5型APP同源攻击后,发病的小白鼠肺脏出血较轻,而用0.050%NAD培养的疫苗免疫小白鼠攻毒后,发病的小白鼠肺脏出血严重.可见在不同NAD含量条件下培养的APP对小白鼠的致病性和免疫保护力不同.     

响应面法优化产琥珀酸发酵培养基

[目的]提高琥珀酸产量。[方法]在单因素试验确定显著因素的基础上,采用响应面法优化琥珀酸放线杆菌JNUBE0709的发酵培养基,利用Design-Expert软件对优化的结果进行二次回归分析。[结果]单因素试验确定碳源麦芽糖的浓度为50 g/L,氮源酵母膏的浓度为25 g/L,酸碱中和剂MgCO3的浓度为40 g/L。响应面法优化得麦芽糖、酵母膏、MgCO33个显著因素的最佳浓度分别为51.1、24.5、40.4g/L,此条件下琥珀酸的产量达33.45 g/L,比优化前提高了14.28 g/L,与模型预测值33.68 g/L基本吻合。麦芽糖与酵母膏的交互作用不显著(P>0.05),MgCO3与麦芽糖、酵母膏的交互作用均显著(P<0.05)。[结论]响应面法优化琥珀酸放线杆菌发酵培养基后的琥珀酸产量比优化前提高了74.49%。     

用PCR检测猪胸膜肺炎放线杆菌

为了快速诊断猪的传染性胸膜肺炎，试验设计合成了1对引物，建立了检测猪胸膜炎肺放线杆菌的PCR方法，猪胸膜炎肺放线杆菌Ⅰ型菌血清2、5、6、9、10型均能扩增出预期片段；大肠杆菌，猪痢疾蛇形螺旋体、多杀性巴氏杆菌、鸡葡萄球菌和支气管败血性波氏杆菌的扩增结果均为阴性，用该方法检测了自猪肺脏分离的30个菌株，有11株为阳性，19株为阴性，结果证实，本方法可以用于猪胸膜肺炎的快速诊断。     

Immunological study of an attenuated Salmonella Typhimurium expressing ApxIA,ApxIIA, ApxIIIA and OmpA of Actinobacillus pleuropneumoniae in a mouse model

Salmonella Typhimurium strain expressing the Actinobacillus pleuropneumoniae antigens, ApxIA, ApxIIA, ApxIIIA and OmpA, was previously constructed as a vaccine candidate for porcine pleuropneumonia. This strain was a live attenuated (∆lon∆cpxR∆asd)Salmonella as a delivery host and contained a vector containing asd. An immunological study of lymphocyte proliferation, T-lymphocyte subsets and cytokines in the splenocytes of a mouse model was carried out after stimulation with the candidate Salmonella Typhimurium by intranasal inoculation. The splenic lymphocyte proliferation and the levels of IL-4, IL-6 and IL-12 of the inoculated mice were significantly increased, and the T- and B-cell populations were also elevated. Collectively, the candidate may efficiently induce the Th1- and Th2-type immune responses.     

猪传染性胸膜肺炎的研究进展

猪传染性胸膜肺炎是一种严重的呼吸道传染病,在世界范围内发病率较高。近年来,国内外学者在该病的病原、流行病学、诊断、防治措施以及免疫预防等方面进行了深入的研究。本文就该病的研究进展进行了综述。     

二温式PCR检测猪传染性胸膜肺炎放线杆菌方法的建立与应用

为了探索简便、快速确诊猪传染性胸膜肺炎的方法,根据猪传染性胸膜肺炎放线杆菌(APP)apx IV基因序列,设计合成1对特异性引物,建立聚合酶链反应(PCR)检测APP的方法。采用该方法对APP和其他6种猪病病原核酸进行检测,结果只对APP扩增出与预期大小相符的422bp DNA片段,而对其他6种猪病病原核酸的扩增结果为阴性。该PCR最低可检出10pg的APP DNA。对送检的46份可疑病猪组织进行检测,结果有19份样品为阳性,27份为其他病原感染。     

猪鼻腔棉拭子中胸膜肺炎放线杆菌检测与分离方法的优化

为提高APP亚临床感染的核酸检出率和细菌分离率,通过检测3种PCR的敏感性和特异性,运用3种PCR方法分别检测母猪鼻腔棉拭子中APP核酸;并同时使用4种选择性培养基分离母猪鼻腔棉拭子中APP。结果显示,3种PCR的最低检测限度均为102 CFU,且除APP外与其他细菌不发生反应;dsb E-PCR方法的App核酸检出率(23.7%)显著高于oml A-PCR(8.1%)和Apx IVA-PCR(14.8%),三者符合率较高,分别为84.4%(dsb E/oml A-PCR)、91.1%(dsb E/Apx IVA-PCR)和93.3%(oml A/Apx IVA-PCR);m PPLO的APP分离率(11.9%)显著高于TSA(0.7%)、CA(5.2%)和BA(2.2%),四者符合率较高,分别为88.9%(m PPLO/TSA)、93.3%(m PPLO/CA)、90.37%(m PPLO/BA)、95.6%(TSA/CA)、98.5%(TSA/BA)和97.0%(CA/BA);dsb E-PCR检测结果与m PPLO分离结果的符合率较高(88.1%)。说明dsb E-PCR和m PPLO的检测分离效率最高,可综合2种方法,完善APP检测分离操作程序,为APP亚临床感染的诊断和治疗以及疫苗免疫提供科学依据。