克隆病与副结核杆菌

分支杆菌、尤其是副结核杆菌长期被疑为克隆病的致病菌。采用聚合酶链反应（ＰＣＲ）技术对手术及内镜活检的７４例石蜡包埋组织（３６例克隆病、１８例溃疡性结肠炎和２０例非炎性肠病对照组织）中的副结核杆菌ＤＮＡ进行检测。扩增的靶ＤＮＡ为副结核杆菌染色体特异重复插入序列ＩＳ９００上４００ｂｐ的片段。其产物特异性通过生物素标记的副结核杆菌全染色体探针Ｓｏｕｔｈｅｒｎ杂交证实。结果显示：４７．２％的克隆病、１１．１％的溃疡性结肠炎和１５．０％的非炎性肠病对照组织中检出了副结核杆菌ＤＮＡ，并发现克隆病组织中副结核杆菌的检出率与其病变部位和是否存在肉芽肿病变无关。提示部分克隆病组织中确有副结核杆菌存在，且后者与克隆病间可能存在特殊相关性。     

奶牛副结核病例的临床诊断

奶牛副结核病不同病例出现的间隔时间较长,临床记录上呈散发性,实际上在养殖场中是一种传播缓慢的流行性传染病。本文就经萋-尼氏抗酸染色和副结核分枝杆菌特异性基因IS900PCR扩增检测确诊的6例荷斯坦泌乳期奶牛副结核临床病例的临床症状进行详细描述,从发病年龄、腹泻症状和体温等方面提出临床诊断依据,以供兽医临床诊断时参考。     

模拟克隆病的肉芽肿性肠炎动物模型的建立

４０只地鼠随机分为两组，实验组：２０只腹腔撞种副结核杆菌；对照组：２０只分别接种生理盐水和死菌（各１０只），８个月后处死动物，作病理学观察。结果：实验组８０％（１６／２０）的地鼠出现肉芽肿性肠炎，而对照组未见肉芽肿性肠炎和溃疡形成。两组肉芽肿性肠炎的发病率有显著性差异（Ｐ〈０．０５）。结果表明：利用副结核杆菌接种地鼠，建立了一个模拟克隆病的肉芽肿性肠炎小动物模型。     

Outcome of Three Commercial Serum ELISAs and Faecal Detection of Mycobacterium avium subsp. paratuberculosis in Consecutive Samples from a Cattle Herd with Low Prevalence of Paratuberculosis (Johne’s Disease)

Paratuberculosis (Johne’s disease) in ruminants is caused by Mycobacterium avium subsp. paratuberculosis (MAP). Owing to the lack of accurate laboratory tests, diagnosis is challenging in subclinically infected cattle. To evaluate the long‐term performance of serum ELISAs for the detection of paratuberculosis in a dairy herd with low MAP‐prevalence, three investigations of all the cows and the consecutive testing of 33 cows suspected to be infected with MAP and 30 cows classified as MAP free were performed over a period of 22 months. Blood samples were tested by three commercial serum ELISAs, MAP shedding was detected by bacteriological culture and polymerase chain reaction (PCR). The ELISA results varied in a wide range in the herd investigations with 1.2% to 18.8% positive samples, the faecal samples were positive for MAP between 1.8% and 4.9% in the three herd investigations. Over the study period, ELISA‐positive serum samples varied between 0.0% and 69.7% in MAP‐suspicious and 0.0% and 17.6% in MAP‐unsuspicious cows with a poor correlation between ELISAs and faecal shedding. The correlation coefficient of the optical density values of the three ELISAs varied between 0.348 and 0.61. Evidence of cow specific variations of residuals was found in all linear models. The linear mixed models showed relevant contribution of cow specific variation in explanation of the residual variances. They also showed significant effects of the explanatory ELISA, the group (MAP‐suspicious or MAP‐unsuspicious) and the time of sampling. It can be concluded that the choice of the laboratory test significantly influences the outcome of the testing for MAP and that none of the three ELISAs can be thoroughly recommended as single test for the early diagnosis of paratuberculosis in cattle. Test results should always be interpreted with caution to avoid erroneous decisions and the disappointment of those engaged in the abatement of paratuberculosis.     

Diagnostic potential of the peptide‐mediated magnetic separation (PMS)‐phage assay and PMS‐culture to detect Mycobacterium avium subsp. paratuberculosis in bovine milk samples

Controlling the spread of Johne's disease, caused by Mycobacterium avium subsp. paratuberculosis (MAP ), in domestic livestock is challenging. Current diagnostic methods lack sufficient sensitivity to detect subclinically infected animals, and thus, better diagnostic methods are needed. This study was carried out to investigate the diagnostic potential of two novel peptide‐mediated magnetic separation (PMS )‐based tests—a PMS ‐phage assay and PMS ‐culture—both of which have been developed and optimized to detect viable MAP cells in bovine milk. Individual milk samples (50 ml) were obtained from 105 “non‐infected” and 40 “MAP ‐infected” animals (classified as such on the basis of prior faecal culture and serum‐ELISA results) in three dairy herds and tested in parallel by the PMS ‐phage assay and PMS ‐culture. Diagnostic sensitivity (DS e) and specificity (DS p) of the PMS ‐phage and PMS ‐culture methods were determined relative to the MAP infection status of the animal contributing the milk sample. The PMS ‐based tests applied individually showed moderate DS e (PMS ‐culture 0.250 and PMS ‐phage assay 0.325) and high DS p (0.962 and 1.000, respectively). When results of the two PMS ‐based tests were combined, DS e increased substantially to 0.525, and the DS p was calculated to be 0.962. It was concluded that combined application of the PMS ‐phage assay and PMS ‐culture provided the most complete picture regarding the presence of viable MAP in bovine milk samples. A comprehensive validation of the PMS ‐based assays relative to currently used diagnostic methods (faecal culture and serum‐ELISA ) would be the next step in assessment of the diagnostic potential of these novel PMS ‐based methods.     

Evaluation of two in-house immunoenzymatic tests to serodiagnose subclinical paratuberculisis in a sheep flock in Mexicali valley,Mexico

Paratuberculosis (PTB) or Johne’s disease is a common ruminant infectious disease caused by Mycobacterium avium subsp. paratuberculosis (MAP). In this study, two MAP antigens were compared for their diagnostic utility to detect subclinical PTB in a sheep flock in Mexicali, Mexico. Sheep (n = 31) without clinical signs but positive on a direct fecal-polymerase chain reaction were tested with two preabsorbed in-house enzyme linked immunosorbent assays (ELISAs) using: (1) an ethanol-extracted surface lipid antigen (EVELISA) and (2) a protoplasmic antigen (ELISA-PPA). Sensitivities of the EVELISA and ELISA-PPA were 84% (95% CI; 66–95%) and 29% (95% CI; 14–48%), respectively. The EVELISA test could be a fast and effective way to identify subclinical ovine PTB for severely affected flocks.     

Mycobacterium avium subspecies paratuberculosis and its relationship with Crohn's disease

The hypothesis postulating that Mycobacterium avium paratuberculosis (MAP) is the cause of Crohn's disease (CD) has been circulating for many years. Advances in molecular techniques, such as polymerase chain reaction and culture methods, have enabled researchers to demonstrate that there is an association between MAP and CD. Recently, genome-wide association studies have identified novel susceptibility genes for CD, which are critical for generation of an adaptive immune response that is protective against intracellular pathogens,including M. tuberculosis infection. However,the role of MAP as a cause of CD suffered a setbackwith the report that administration of antimycobacterial therapy failed to lead to a sustained response in CD patients. Accordingly, this review sought neither to confirm nor refute this, but instead to survey recent literature on the role of MAP in CD.     

Histopathological classification of lesions observed in natural cases of paratuberculosis in free-ranging fallow deer (Dama dama)

Ninety-five adult fallow deer, legally hunted in the Regional Hunting Reserve of El Sueve (Northern Spain), were subjected to a post-mortem examination for paratuberculosis, samples being taken from the proximal and distal jejunum, proximal and distal ileum, ileocaecal valve and associated lymph nodes. The lesions were divided into four categories. Focal lesions (n=19 cases) consisted of small granulomas, mainly in the jejunal and ileal lymph nodes. Multifocal lesions (n=4) consisted of well-demarcated granulomas in the intestinal lymphoid tissue and also in the intestinal lamina propria. Diffuse multibacillary lesions (n=2) were characterized by a severe granulomatous enteritis and lymphadenitis. Macrophages and numerous Langhans giant cells containing many mycobacteria were present, resulting in macroscopical changes in the normal gut morphology. These changes were found from the proximal jejunum to the ileocaecal valve, but lesions were always particularly severe in the distal jejunum. In diffuse intermediate (multibacillary-lymphocytic) lesions (n=3) the infiltrate consisted of lymphocytes, macrophages and Langhans giant cells, with small numbers of mycobacteria. Mycobacterium avium subspecies paratuberculosis was identified by a polymerase chain reaction technique. The widespread occurrence of paratuberculosis in fallow deer in this Reserve represents a potential source of infection for other susceptible species.