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1.
Several phenotypic as well as genotypic methods have been published describing the detection of central nervous system (CNS) tissues that are part of the bovine spongiform encephalopathy (BSE) risk material in food products. However, none of these methods is able to differentiate between CNS tissue of the banned ruminant species and tissues of other animal species. A quantitative and species-specific real-time RT-PCR method has been developed that enables the reliable identification of CNS tissues in meat and meat products. This method is based on a messenger (m)RNA assay that uses bovine, ovine and caprine glial fibrillary acidic protein (GFAP) encoding gene sequences as markers. The in-house validation studies evaluated the tissue specificity of up to 15 bovine tissues and the standardization of absolute as well as relative quantitative measurement. The specific amplification of spinal cord and brain tissue GFAP cDNA has been shown previously. In addition, two commercially available ELISA kits were used for the comparative analysis of artificially contaminated minced meat. Small quantities of bovine brain that had been stored over the recommended period of 14 days were examined. The real-time PCR method proved to be suitable for the detection of 0.1% CNS tissue. No false negative results were observed. The quantitative detection of GFAP mRNA using real-time RT-PCR seems a suitable tool in routine diagnostic testing that assesses the illegal use of CNS tissue in meat and meat products. The stability of the selected target region of the GFAP mRNA also allows the detection of CNS tissues after the meat has been processed. 相似文献
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3.
Miyashita M Stierstorfer B Schmahl W 《Journal of veterinary medicine. B, Infectious diseases and veterinary public health》2004,51(5):209-215
The brains of 26 Bavarian bovines clinically suspected of bovine spongiform encephalopathy (BSE) were the subject of a neuropathological evaluation containing histopathology and immunohistochemistry. Six animals tested positive for BSE. In these six brains severe histological lesions that correlated with previous reports from the United Kingdom were observed. Immunohistochemistry with prion protein (PrP(Sc)), glial fibrillary acidic protein (GFAP) and synaptophysin were conducted on the mid-brain containing the red nucleus. All BSE-positive brains stained positively for PrP(Sc), and no plaques were observed. The BSE-affected brains showed a substantially more intense staining pattern for GFAP in comparison with the control groups, some of which were diagnosed with severe neuropathological disorders. Synaptophysin staining on BSE-positive brains was substantially reduced in the neuropil of the mid-brain, especially in the red nucleus. Twenty animals tested negative for BSE. The most common diagnoses were listeriosis, viral infections of unknown aetiology, brain oedema and hypomagnesaemia. These disorders may represent the most important clinical differential diagnoses for BSE in Bavaria. 相似文献
4.
Wemheuer WM Benestad SL Wrede A Wemheuer WE Brenig B Bratberg B Schulz-Schaeffer WJ 《Veterinary research》2011,42(1):32
ABSTRACT: Scrapie in sheep and goats has been known for more than 250 years and belongs nowadays to the so-called prion diseases that also include e.g. bovine spongiform encephalopathy in cattle (BSE) and Creutzfeldt-Jakob disease in humans. According to the prion hypothesis, the pathological isoform (PrPSc) of the cellular prion protein (PrPc) comprises the essential, if not exclusive, component of the transmissible agent. Currently, two types of scrapie disease are known - classical and atypical/Nor98 scrapie. In the present study we examine 24 cases of classical and 25 cases of atypical/Nor98 scrapie with the sensitive PET blot method and validate the results with conventional immunohistochemistry. The sequential detection of PrPSc aggregates in the CNS of classical scrapie sheep implies that after neuroinvasion a spread from spinal cord and obex to the cerebellum, diencephalon and frontal cortex via the rostral brainstem takes place. We categorize the spread of PrPSc into four stages: the CNS entry stage, the brainstem stage, the cruciate sulcus stage and finally the basal ganglia stage. Such a sequential development of PrPSc was not detectable upon analysis of the present atypical/Nor98 scrapie cases. PrPSc distribution in one case of atypical/Nor98 scrapie in a presumably early disease phase suggests that the spread of PrPSc aggregates starts in the di- or telencephalon. In addition to the spontaneous generation of PrPSc, an uptake of the infectious agent into the brain, that bypasses the brainstem and starts its accumulation in the thalamus, needs to be taken into consideration for atypical/Nor98 scrapie. 相似文献
5.
Biedermann W Lücker E Hensel A 《Berliner und Münchener tier?rztliche Wochenschrift》2002,115(3-4):131-133
Determination of specified risk material (SRM) in processed meat products was performed by quantification of brain specific fatty acids using gas chromatography-mass spectrometry (GC-MS). Results from SMP (internal standardised meat products) based analyses showed that absolute concentrations of CNS are correlated (r2 = > 0.97) with the contents of the CNS typical fatty acids docosahexaenoic acid (C 22:6), nervonic acid (C 24:1), lignoceric acid (C 24) and cerebronic acid (C 24oh). GC-MS detection limits were measured at 0.01% CNS. The cut off value was calculated at 0.39% (w/w) CNS in SMP. In a controlled blindfold experiment we were able to identify correctly all positive and negative SMP samples, respectively. Our results indicate that GC-MS based SRM detection may serve as a reference method for immunochemical and immunohistochemical determination of SRM in processed meat products. 相似文献
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Addition of blood plasma to meat products is not permitted in the FRG unless these products are heat processed using an internal temperature of 80 degrees C (German regulation of meat and meat products: "Verordnung für Fleisch und Fleischerzeugnisse"). Such heat process may have an unfavourable effect on the detectability of blood plasma. Since blood plasma or dried plasma may originate from different animal species (porcine or bovine) two different anti dried blood plasma-sera (porcine and bovine) are required for immunochemical analysis. The varying quality of these sera has to be considered when interpreting the results. Seven M urea extract turned out to be suitable for detection of dried plasma additives and proved to be highly effective particularly when examining heated samples. Both the gel-diffusion and the electro immuno assay proved useful for the detection of dried blood plasma, provided the examined extracts had been adequately diluted. Immunochemical reactivity was hampered by the heat process which was given to the sample. Accordingly, the concentration of the plasma in a particular sample cannot be determined unless the time/temperature data of the process applied to the sample were given and model samples were tested for comparison. 相似文献
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Transmissible spongiform encephalopathies are degenerative disorders affecting the central nervous system (CNS) occurring in a variety of species. The causative agent is thought to be composed of an abnormal form of the host encoded prion protein (PrPC), termed PrPSc. The conformational change of PrPC into PrPSc can occur spontaneously, however, it can also be induced by PrPSc. Prion diseases such as bovine spongiform encephalopathy (BSE), scrapie and variant Creutzfeldt-Jakob-Disease (vCJD) are most likely caused by peripheral uptake of prions. The process by which prions proceed to the CNS following peripheral uptake is referred to as neuroinvasion. Infection with prions is thought to occur in two phases: After ingestion prions first replicate in lymphatic tissue and then gain access to the CNS via peripheral nerves. Studies looking at the biochemical and clinical characteristics of BSE and vCJD demonstrated that BSE is most likely responsible for vCJD in humans. 相似文献
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Sponge samples were taken from the carcases, meat, personnel and surfaces involved in stunning, slaughter and dressing/boning activities at three abattoirs, and from retail beef products. The samples were examined for the presence of central nervous system (CNS)-specific proteins (syntaxin 1B and/or glial fibrillary acidic protein (GFAP), as indicators of contamination with CNS tissue. Syntaxin 1B and GFAP were detected in many of the sponge samples taken along the slaughter line and in the chill rooms of all three abattoirs; GFAP was also detected in one sample of longissimus muscle (striploin) taken in the boning hall of one of the abattoirs but not in the other two abattoirs or in retail meats. 相似文献
9.
根据牛特异性线粒体DNA片段,设计合成1对引物,以生、熟牛肉为材料,建立了肉制品中牛源性成分的PCR检测方法,并用该法对市售的67份牛肉制品进行检测。结果显示,所检牛源性成分在271 bp处出现预期的条带,扩增片段经Sau3AⅠ酶切分析确认,获得的214和57 bp片段与预期一致;运用该引物均可扩增出水牛肉、牦牛肉、奶牛肉、黄牛肉单一的相同大小的DNA条带,而对羊、马、狗、驴、兔和鸭等14种动物肉的DNA扩增则呈阴性,其检测灵敏度达到53.2 fg/μL DNA;利用该法对67份牛肉制品进行检测,检出率为100％。结果表明,该法快速简便,且具有较高的特异性和敏感性,可用于市售牛肉制品中牛源性成分的鉴定。 相似文献
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参考Saborio和Soto创建的名为蛋白质错误折叠循环扩增(protein misfolding cyclic amplification,PMCA)的方法建立一种22L毒株系朊蛋白错误折叠循环扩增方法,以便研究22L毒株系异常朊蛋白(scrapie PrP,PrPSc)的合成机制。在阴离子去污剂十二烷基硫酸钠(SDS)存在的条件下,这种方法可以通过超声降解从而重复扩增产物,并在仓鼠脑匀浆中快速有效的模仿体内朊病毒PrPSc的复制,因此可以在几个小时内高浓度扩增PrPSc。已有试验结果发现,PrPSc体外转化率在使用粗提的脑组织匀浆时比使用纯化重组PrP更高,这个发现暗示着高效率的PrPSc转化可能还需要探索其他未知的宿主因素。研究结果表明,建立的这种改良的PMCA技术可用于研究PrPSc合成的生物化学机制,以此来进行诊断与治疗方法的研究。 相似文献
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Okada H Iwamaru Y Imamura M Masujin K Matsuura Y Shimizu Y Kasai K Mohri S Yokoyama T Czub S 《Veterinary research》2011,42(1):79
ABSTRACT: Atypical bovine spongiform encephalopathy (BSE) has recently been identified in Europe, North America, and Japan. It is classified as H-type and L-type BSE according to the molecular mass of the disease-associated prion protein (PrPSc). To investigate the topographical distribution and deposition patterns of immunolabeled PrPSc, H-type BSE isolate was inoculated intracerebrally into cattle. H-type BSE was successfully transmitted to 3 calves, with incubation periods between 500 and 600 days. Moderate to severe spongiform changes were detected in the cerebral and cerebellar cortices, basal ganglia, thalamus, and brainstem. H-type BSE was characterized by the presence of PrP-immunopositive amyloid plaques in the white matter of the cerebrum, basal ganglia, and thalamus. Moreover, intraglial-type immunolabeled PrPSc was prominent throughout the brain. Stellate-type immunolabeled PrPSc was conspicuous in the gray matter of the cerebral cortex, basal ganglia, and thalamus, but not in the brainstem. In addition, PrPSc accumulation was detected in the peripheral nervous tissues, such as trigeminal ganglia, dorsal root ganglia, optic nerve, retina, and neurohypophysis. Cattle are susceptible to H-type BSE with a shorter incubation period, showing distinct and distinguishable phenotypes of PrPSc accumulation. 相似文献
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动物产品中牛、羊源性成分多重PCR检测方法的建立 总被引：14，自引：2，他引：14
以肉骨粉、鱼粉、猪肉干和鱼肉干为研究对象,异硫氰酸胍法提取总DNA,18S rDNA片段的扩增结果表明提取到的DNA中不存在抑制PCR的物质。应用梯度PCR技术对牛、羊源性成分检测的退火温度进行了优化,在单一PCR检测技术的基础上分别进行了18S rDNA片段和牛、羊源性成分的多重PCR分析,得到了预期的结果。试验表明,本文建立的多重PCR方法具有快速、简便、准确等特点,对动物产品牛、羊源性成分检测具有重要意义。 相似文献
13.
Botteron C 《Schweizer Archiv für Tierheilkunde》2002,144(12):639-644
Histopathological examination of the central nervous system is essential for the confirmation of a TSE diagnosis. Typical lesions are spongiform changes of the grey matter, intraneuronal vacuoles in particular nuclei of the brain stem, gliosis and neuronal degeneration. The nature of the lesions is similar between species. However, the variation in the distribution and severity of the changes is striking. Even more reliable than histopathology is the detection of disease-specific protease-resistant prion protein (PrPSc) using immunohistochemistry. The so-called "rapid tests" allow detection of PrPSc in unfixed tissues and are mostly used for the screening of risk populations and slaughtered animals. 相似文献
14.
疯牛病不仅给全球畜牧经济造成重大损失,而且还严重危害着人类的健康,而造成疯牛病传播的主要原因则是由于携带有致病因子的牛羊肉骨粉饲料及牛羊制品在各国之间的贸易往来。因而加强对进口饲料产品中牛羊源成分的检测是防止疯牛病流行的重要措施。本文就疯牛病的流行病学、病原学、病理变化与诊断以及牛羊源成分检测方法的研究进展作以简述。 相似文献
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Shibuya M Matsuki N Fujiwara K Imajoh-Ohmi S Fukuda H Pham NT Tamahara S Ono K 《The Journal of veterinary medical science / the Japanese Society of Veterinary Science》2007,69(3):241-245
Cerebrospinal fluids (CSFs) from 9 Pug dogs with necrotizing meningoencephalitis (NME: Pug dog encephalitis) were examined to identify the antigens for anti-astrocyte autoantibodies. Each CSF exhibited a positive reaction to the cytoplasm of cultured canine astrocytes by an indirect fluorescent antibody test. In an immunoblotting analysis on normal canine brain proteins, eight of 9 CSFs showed a common band of 52 kDa, corresponding to glial fibrillary acidic protein (GFAP), and all of 9 CSFs reacted with purified bovine GFAP. From these results, GFAP is one of the common autoantigens in Pug dogs with NME. On the other hand, the reactivity of CSFs to chymotrypsin-digested bovine GFAP fragments were variable among dogs, indicating that the antibodies in the CSFs recognized different epitopes on GFAP. 相似文献
16.
为鉴定和区分饲料及动物产品中牛、山羊、绵羊源性成分,根据线粒体DNA(mitochondrial DNA,mtDNA)种间保守序列,设计合成了3对特异性引物与TaqMan探针,通过对荧光PCR反应体系和反应条件的优化筛选,建立了三重荧光PCR方法,在同一个荧光PCR反应中完成3种动物源性成分的检测。用该方法对16种不同源性的动物DNA进行检测,结果表明能特异地鉴别检测出牛、山羊和绵羊源性成分,且敏感性比现行国标PCR法高100倍。该方法适用于饲料、肉制品、奶制品等动物源性产品的检测。 相似文献
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Stevenson MA Wilesmith JW Ryan JB Morris RS Lawson AB Pfeiffer DU Lin D 《The Veterinary record》2000,147(14):379-384
This was a spatial analysis of the epidemic of bovine spongiform encephalopathy (BSE) in Great Britain, based on agricultural census data collected between 1986 and 1996 and BSE case data collected up to June 1997. Kernel smoothing techniques were used to plot the distribution of BSE-positive cattle holdings per 100 holdings per square kilometre and the distribution of confirmed BSE cases per 100 head of cattle per square kilometre. In the early stages of the epidemic reported BSE cases were scattered widely throughout Great Britain, with no clearly identifiable focus. By June 1997, a statistically significant cluster of BSE-positive holdings was identifiable in the eastern part of the South west region of England. During the epidemic the highest densities of confirmed BSE cases per 100 cattle per square kilometre occurred in the greater part of the South west region of England and within Dyfed in the south west of Wales. In Wales, a small number of holdings experienced large numbers of confirmed BSE cases. In the South west region of England a large number of holdings experienced small numbers of confirmed cases. By June 1997, the distribution of BSE-positive holdings across Great Britain was largely determined by factors that influenced the amount of recycled infectious material they were exposed to. 相似文献
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All sheep older than 1 year of age from a flock of the Rygja breed in which clinical scrapie was detected for the first time in two animals (4%) were examined for accumulation of pathogenic prion protein (PrPSc) by immunohistochemistry in the obex, the cerebellum, and the medial retrophayngeal lymph node. In addition, six lambs, 2-3 months old, all offspring of PrPSc-positive dams, were examined for PrPSc in the ileal Peyers' patch (IPP), the distal jejunal lymph node, the spleen, and the medial retropharyngeal lymph node (RPLN). In this flock, 35% (17/48) of the adult sheep showed accumulation of PrPSc, an eightfold increase compared with clinical disease. All positives carried susceptible PrP genotypes. Three sheep had deposits of PrPSc in the RPLN and not in the brain, suggesting that this organ, easily accessible at slaughter, is suitable for screening purposes. Two 7-year-old clinically healthy homozygous V136Q171 ewes showed sparse immunostaining in the central nervous system and may have been infected as adults. Further, two littermates, 86-days-old, showed PrPSc in the IPP. Interestingly, one of these lambs had the intermediate susceptible PrP genotype, VA136QR171. In addition to early immunolabeling in the dorsal motor nucleus of the vagal nerve, a few of the sheep had early involvement of the cerebellum. In fact, a 2-year-old sheep had sparse deposits of PrPSc in the cerebellum only. Because experimental bovine spongiform encephalopathy (BSE) in sheep seems to behave in a similar manner as natural scrapie, these results, particularly regarding spread of infectivity, may have implications for the handling of BSE should it be diagnosed in sheep. 相似文献
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Tanaka M Inoue A Yamamoto K Tamahara S Matsuki N 《The Journal of veterinary medical science / the Japanese Society of Veterinary Science》2012,74(6):733-737
Necrotizing meningoencephalitis (NME), necrotizing leukoencephalitis (NLE) and granulomatous meningoencephalomyelitis (GME) are common idiopathic inflammatory central nervous system (CNS) diseases with unknown etiology in dogs. We previously showed that IgG autoantibodies in the cerebrospinal fluid (CSF) of NME cases reacted to unknown brain proteins as well as to glial fibrillary acidic protein (GFAP). In the present report, we evaluated the autoantibodies against transglutaminase2 (TG2) in the canine CNS diseases. CSF samples obtained from dogs with NME (n=19), NLE (n=7), GME (n=11) and miscellaneous CNS diseases (n=12) were subjected. CSFs from 20 healthy dogs were used as controls. Indirect fluorescent antibody test on the canine cerebrum revealed astrocyte-binding IgG in the CSF of NME. After absorption of the CSF with bovine GFAP, the CSF still possessed the reactivity to astrocytes. Double-color staining showed clear colocalization of the autoantibodies and anti-human TG2 rabbit polyclonal IgG. An immunoblot assay against human recombinant TG2 revealed anti-TG2 IgG in the CSF from dogs with NME, NLE and GME. The CSF of canine idiopathic encephalitis cases, notably of NME, tended to show high ELISA OD values against human recombinant TG2 compared to healthy controls. The presence of anti-TG2 autoantibodies in the CSF may contribute to the elucidation of the etiology of canine NME, NLE and GME. 相似文献
20.
文章论述了"调猪"向"调肉"转变的历史背景,进行了冷却肉代替常温鲜肉的可行性论证,加强对冷却肉营养价值高,口味几乎与常温鲜肉媲美等关于冷却肉的知识宣传,让冷却肉成为肉类消费主流产品,是破解"调猪"向"调肉"困境的主要方法。尖锐指出目前排名前十的猪企纷纷布局的,大城市大型屠宰和肉联厂项目,是走"调猪"的老路和死胡同,会造成极大的资源浪费。提出在生猪主产区建立肉联厂,充分利用和整合现有肉联厂资源,配齐官方兽医,加强产地检疫和屠宰检疫,加快非洲猪瘟快速检测试纸的研发,加强冷却肉加工、质量控制技术研究和国家标准的制订,中国肉类加工企业协会,承接国家主管部门移交的肉类加工社会管理职能等建议,供同行参考。 相似文献