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1.
An indirect enzyme linked immunosorbent assay (ELISA) using an unpurified antigen was assessed for its accuracy in detecting Mycobacterium bovis infected cattle. The ELISA test recorded sensitivities of 88.7% and 63%, respectively, for infected cattle tuberculin tested positive and for infected cattle never tuberculin tested. Specificity was determined at 52.6% for cattle from confirmed free herds which had never been tuberculin tested. Significant differences in the mean ELISA values were recorded between the 3 groups. No evidence was found for long term effects of tuberculin testing upon the titre of antibodies detected by the ELISA in unaffected cattle. The indirect ELISA using the unpurified antigen of this assay was considered to be unsuitable as an alternative to tuberculin testing for the detection of M. bovis infected animals.  相似文献   

2.
OBJECTIVE: To determine whether cattle testing positive for Mycobacterium avium subsp paratuberculosis as determined by microbial culture of feces or antibody ELISA were more likely to have false-positive responses on the caudal fold tuberculin (CFT) test or interferon-gamma (IFN-gamma) assay for Mycobacterium bovis than cattle testing negative for M paratuberculosis. ANIMALS: 1043 cattle from 10 herds in Michigan. PROCEDURE: Feces and blood samples for plasma were collected from cattle > or =24 months old on the day the CFT test was read. Fecal samples were submitted for microbial culture for M paratuberculosis. Plasma samples were tested for antibody against M paratuberculosis, and IFN-gamma after stimulation with purified protein derivative tuberculin from M bovis or M avium. RESULTS: Of 1043 cattle, 180 (17.3%) had positive CFT test results (suspects) and 8 (0.8%) had positive IFN-gamma assay results after stimulation with purified protein derivative tuberculin from M bovis. Forty-five (4.3%) and 115 (11.0%) cattle tested positive for M paratuberculosis as determined by microbial culture of feces and antibody ELISA, respectively. Cattle with positive responses for M paratuberculosis appeared to have an increased likelihood of false-positive results on the CFT test, although this association was not significant. CONCLUSIONS AND CLINICAL RELEVANCE: No significant association was detected among cattle testing positive for M paratuberculosis as determined by microbial culture of feces and antibody ELISA and positive CFT test and IFN-gamma assay results for M bovis.  相似文献   

3.
A fluorescence polarization assay (FPA) utilizing fluorescein-labelled MPB70 protein as the antigen was developed and evaluated for its ability to detect antibodies to Mycobacterium bovis in cattle sera. Three panels of sera were examined in this study. These included: (A) sera (n=28) obtained from cattle from which M. bovis was cultured; (B) sera (n=5666) from Canadian field cattle which were presumed to be free from M. bovis; (C) sera (n=10) from cattle infected with Mycobacterium paratuberculosis and known to contain antibodies to this organism. Receiver operating characteristic (ROC) curve analysis of the results of panels A and B yielded an area under the curve value of 0.975 (95% confidence interval=0.971-0.979), which indicated that this FPA is an accurate indicator of M. bovis infection. At the cut-off point recommended by the ROC curve analysis, the FPA sensitivity and specificity estimates were 92.9% (95% confidence interval=76.5-98.9%) and 98.3% (95% confidence interval=97.9-98.6%) respectively. The FPA results were compared to the results of the single intradermal (SID) test for the 28 infected cattle. Fifteen of these animals were scored positive with the SID test (sensitivity=53.6%). The FPA detected 15/15 (100%) of the SID test-positive animals and 11/13 (84.6%) of the SID test-negative animals. Two of the culture-positive cattle were not detected by either test. None of the sera that were obtained from the M. paratuberculosis-infected animals cross-reacted in this assay.  相似文献   

4.
An ELISA for the detection of anergic tuberculous cattle   总被引：6，自引：0，他引：6  
An enzyme-linked immunosorbent assay (ELISA) for bovine antibody to antigens in unheated Mycobacterium bovis culture filtrate was standardised against a reference serum from an experimentally infected cow. Two Northern Territory herds with a total of 561 cattle were tested. All cattle reacting in the caudal fold tuberculin test, those giving strong reactions in the ELISA and those with visible lesions of tuberculosis were subjected to a detailed bacteriological examination. Of the 19 cattle which yielded isolates of M. bovis, only 4 were positive to the tuberculin test. Serum samples from 5 cattle gave ELISA values greater than 7.0 units. None of these 5 reacted in the tuberculin-test and 2 had no visible lesions. Of the 10 remaining cattle from which M. bovis was isolated, 3 had ELISA values between 6.5 and 7.0 units and were also without visible lesions. The ELISA values for the remaining 7 infected cattle ranged down to 4.6 units. Forty cattle yielded no M. bovis on culture of their tissues. They included 7 which were reactors in the tuberculin test and 23 with ELISA values of 7.0 units or more. The evident low specificity and sensitivity of the ELISA make it of little value as an alternative to the tuberculin test, but it can detect some anergic cattle at the cost of increasing the number of false positive reactors. This may be acceptable in some circumstances and would justify the use of the ELISA as a complement to the tuberculin test or to an in vitro assay of T-cell immunity. In the 2 Northern Territory herds described, the removal of 5 of the anergic cattle would have required a cull of 28 animals of 5% of the total. A cut off value of 6.5 units would have eliminated 3 more, but at the cost of culling 80 animals or nearly 15% of the cattle. Even so, 7 cattle from which M. bovis was isolated would have remained undetected by either test.  相似文献   

5.
A total of 23 (15.3 per cent) of 150 cattle infected with Mycobacterium bovis and which had never been tuberculin tested showed specific antibody responses to M bovis. Their sera may be important keys to the identification of unique M bovis antigens for use in specific serodiagnostic tests. Assessment of specific and non-specific responses was done by screening sera in six indirect anti-IgG enzyme-linked immunosorbent assays using whole cell sonicates of M bovis and five members of the Mycobacterium avium-intracellulare-scrofulaceum complex as respective antigens. Sera from 16 infected cattle that had been tuberculin tested positive and nine uninfected cattle (never tuberculin tested) were also assayed for specific and non-specific responses. Three other findings emerged. First, 43 of the 150 infected animals (28.7 per cent) showed no antibody responses to any of the mycobacterial antigens used. Secondly, the cattle showing the highest antibody levels were associated with the greatest cross reactivity. Lastly, the results indicated that tuberculin injections may increase antibody responses to shared, rather than specific, M bovis antigens in infected cattle.  相似文献   

6.
In the UK there has been a sharp rise in the incidence of bovine tuberculosis since the early 1990s and the badger has been identified as an important wildlife reservoir for this infection. Infected badgers can excrete Mycobacterium bovis, putting other badgers and cattle at risk of becoming infected. Vaccination has been proposed as an approach to reducing the excretion of M. bovis by tuberculous badgers. In order to evaluate the efficacy of a badger vaccine it will be necessary to accurately determine the number of badgers excreting M. bovis without removing them for post-mortem evaluation. The existing live tests for tuberculosis in the badger (culture, indirect ELISA, Western blot) have not been assessed for their ability to detect badgers excreting M. bovis. Over the past 18 years, badgers from 31 social groups have been trapped and sampled in a study area of the Cotswold escarpment. We have examined the serological responses of 128 badgers trapped between 1985 and 1998 from social groups where M. bovis infection was endemic. These responses were compared with culture from faeces, urine, tracheal aspirates and bite wound swabs taken from these animals while alive. ELISA was found to be more sensitive than Western blot in detecting badgers excreting M. bovis. The majority of culture-positive badgers excreted M. bovis intermittently over the period of study. As a result, there was only a 27.5% chance of sampling a badger for culture when it was excreting M. bovis. In contrast, a positive ELISA result correctly predicted 68.2% of badgers with a history of excreting M. bovis. In the absence of alternative live tests for the badger, the Brock Test indirect ELISA appears to be more valuable than culture for measuring the effect of vaccination on reducing the number of badgers at risk of transmitting tuberculosis.  相似文献   

7.
An enzyme-linked immunosorbent assay (ELISA) for the detection of antibodies against Mycobacterium paratuberculosis in cattle was evaluated in three herds known to have Johne's disease. Prior to testing, the plasma was absorbed with dried Mycobacterium phlei in order to remove cross-reacting antibody specificities. The sensitivity and specificity of the ELISA were calculated after repeatedly testing 327 cattle in the infected herds. Of these, 53 animals had one or more positive faecal cultures or had post-mortem histopathological evidence of infection. The other 274 had three or more negative faecal culture results, and were regarded as non-infected for the purpose of evaluation of the ELISA. Using these criteria for the presence or absence of infection, the M. phlei-absorbed ELISA under field conditions had a 57% sensitivity and a 98.9% specificity. The sensitivity of the absorbed ELISA depended on the stage of disease of the animal under test. In general, it appeared that animals in the more advanced stages of disease were absorbed ELISA positive, whereas those in the early stages of infection were not detected. These results indicate that the M. phlei-absorbed ELISA has an important role as a test for the diagnosis and control of Johne's disease in cattle.  相似文献   

8.
Vaccination of cattle against Mycobacterium avium subsp. paratuberculosis (MAP) provides partial protection by delayed shedding of MAP and reduced numbers of clinically affected animals. The duration of vaccine induced immune response is not known. The primary objective of this study was therefore to characterize the long-term effect of whole-cell based vaccination against MAP on the immune response. A secondary objective was to evaluate whether immunodiagnosis of MAP and Mycobacterium bovis infections is affected by MAP vaccination. Two studies were performed: (1) A retrospective longitudinal study including 895 vaccinated and 2526 non-vaccinated dairy cows in 9 Danish dairy herds aiming at characterizing the long-term antibody-response to vaccination; and (2) a cross-sectional study of responses in the IFN-γ assay carried out in 140 vaccinated animals in two herds to evaluate the effect of vaccination on the cell-mediated immune response and to evaluate a possible interference with the diagnosis of M. bovis infections. The results showed that 37% of samples from vaccinated animals and 5% of samples from non-vaccinated animals, respectively, were test positive in the milk antibody ELISA. The prevalence of antibody responses of the vaccinated animals was relatively constant from 2 to 6 years of age, but decreased in older animals. Among the 140 vaccinated animals 88% tested positive with the IFN-γ test to johnin PPD and 50% responded to PPDb with IFN-γ production above a similar cut-off. Although Denmark is free of M. bovis, two of the vaccinated animals responded with higher IFN-γ levels when cultured with PPDb compared to PPDa. In conclusion, immunization with whole-cell MAP vaccines elicits both humoral and cell-mediated immune reactions, which may interfere with surveillance and diagnosis of both MAP and M. bovis infections using currently available tests.  相似文献   

9.
Skin test negative cattle from a herd containing an unusually high proportion (194/382) of tuberculin skin test positive cattle were investigated for remaining Mycobacterium bovis infected animals. Blood samples from the skin test negative cattle, analysed by an antibody ELISA and an interferon-gamma assay, were mostly test negative for M. bovis. Radiometric culture of nasal mucus samples from 48 of the cattle yielded 22 culture positives with acid-fast bacilli and cording in 6 of these. Subculture on solid media was successful for 7, including 2 with cording of the 22 radiometric culture positives. Mycobacterium tuberculosis complex DNA probe testing using the Accuprobe (Gen-Probe, Inc.) and M. tuberculosis complex-specific PCR amplification, performed on the solid media subcultures, were negative. 16S rRNA PCR and sequence analysis were successful for 6 of the 7 solid media subcultures obtained and revealed the presence of Mycobacterium nonchromogenicum in all 6 subcultures. This is the first report of M. nonchromogenicum in nasal mucus of cattle. The observation highlights the importance of integrating definitive tests such as the PCR for diagnosis of bovine tuberculosis and indicates a possible zoonotic risk.  相似文献   

10.
The upper respiratory tract surfaces, the palatine and pharyngeal tonsils and associated lymph nodes of 32 tuberculin reactor cattle were examined pathologically and bacteriologically. Tuberculous lesions were observed histologically in the palatine tonsils of five animals and in both the palatine and pharyngeal tonsils of a sixth. Mycobacterium bovis was cultured from the tonsils of four of these animals and from the palatine or pharyngeal tonsils of a further eight cattle in which no lesions were observed. The upper respiratory tract surfaces of 10 animals were M bovis-positive.  相似文献   

11.
The pneumonic lungs of 42 cattle from 26 feedlots were examined for the presence of mycoplasma, pathogenic bacteria and viruses. Four animals representative of two lots failed to yield mycoplasma. One of these yielded the virus of infectious bovine rhinotracheitis and Pasteurella hemolytica, the other yielded only P. P. multocida. Nine animals in eight lots yielded Mycoplasma sp.: five of these were M. bovirhinis, two were M. arginini and two were untypable. All of these animals yielded one or more of P. hemolytica, P. multiocida, infectious bovine rhinotracheitis virus or bovine virus diarrhea virus. Twenty-five of 29 animals in 16 lots yieled M. agalactiae subsp. bovis from lung tissues. The same organism was recovered from the arthritic joints of 12 of these animals. Eight of the 25 animals yielded no other pathogen and all of these had not received any treatment. Nine of the 25 M. agalactiae subsp. bovis positive animals also yielded one or more of P. hemolytica, P. multocida, Corynebacterium pyogenes or infectious bovine rhinotracheitis virus. Bacteriological and virological studies were not completed for the remaining eight of the 25 positive animals. In five lots of cattle which had not received medication for pneumonia and for arthritis only M. agalactiae subsp. bovis was recovered. Twenty-five grossly normal lungs obtained from normal cattle at the time of slaughter were cultured and all were negative. The possible role of M. agalactiae subsp. bovis in pneumonia and arthritis was discussed.  相似文献   

12.
OBJECTIVE: To determine necropsy and Mycobacterium bovis culture results in cattle from herds with tuberculosis, the role of the bovine NRAMP1 gene in resistance and susceptibility to infection with M bovis, and the association between magnitude of the tuberculous lesions and various types of M bovis isolates. ANIMALS: 61 cattle from herds with tuberculosis in Texas and Mexico. PROCEDURE: 61 cattle were evaluated by necropsy; 59 had positive and 2 had negative caudal fold tuberculin intradermal test (CFT) results. Thirty-three cattle with positive CFT results were genotyped to evaluate polymorphism of the 3' untranslated region of the bovine NRAMP1 gene, using single-stranded conformational analysis, 9 were resistant to M bovis with no tuberculous lesions and negative M bovis culture results, and 24 were susceptible with tuberculous lesions and positive M bovis culture results. Isolates of M bovis were analyzed by restriction fragment length polymorphism (RFLP) on the basis of IS6110 sequences and direct-repeat fingerprinting patterns. RESULTS: 21 (35.6%; 21/59) cattle with positive CFT results had tuberculous lesions or positive culture results; in addition, 1 of 2 cattle with negative CFT results had tuberculous lesions and positive culture results. Tuberculous lesions were most common in the thorax (35/63; 55.5%) and lymphoid tissues of the head (10/63; 15.9%). Tuberculous lesions varied from 1 to 11/animal; 8 of 21 (38.1%) had solitary lesions. Associations were not found between resistance or susceptibility to infection with M bovis and polymorphism in the NRAMP1 gene or between the magnitude of the lesions and various RFLP types of M bovis isolates. CONCLUSIONS AND CLINICAL RELEVANCE: The NRAMP1 gene does not determine resistance and susceptibility to infection with M bovis in cattle.  相似文献   

13.
A study to determine the presence and prevalence of bovine tuberculosis in slaughter cattle in Kenya was carried out in two abattoirs from July to November 2009. Routine postmortem meat inspection was performed on a subpopulation of 929 cattle selected randomly from among 4,984. Carcases were inspected for gross tuberculous lesions which were then examined for acid-fast bacilli, (AFB), cultured for isolation of mycobacteria and the isolates characterised by DNA molecular analysis. Of the carcases examined, 176 (18.95?%, 95?% CI) had lesions suggestive of tuberculosis. AFB were observed in 63/176 of the lesioned cattle and mycobacteria were isolated from 64 of them. The isolates were identified as Mycobacterium bovis (19/64), Mycobacterium tuberculosis, (2/64) and mycobacteria other than tuberculosis (43/64). The prevalence of M. bovis by molecular analysis was 2.05?% (95?% CI). This study documents for the first time the presence of bovine tuberculosis among slaughter cattle in Kenya. There is therefore a need to formulate and implement control programmes in order to minimise transmission among animals and to humans. Isolation of M. tuberculosis from cattle underscores the risk tuberculous humans pose to animals.  相似文献   

14.
为建立快速检测牛分枝杆菌(M.bovis)的TaqMan荧光定量PCR方法,本研究以GenBank登录的M.bovis特有229 bp基因为研究对象,设计并合成引物及探针。该方法具有较好的特异性,与标准质控菌株呈阳性反应,与其他微生物样品呈阴性反应;灵敏性最低检测值可达1 pg/mL;对20阳性临床样品进行荧光定量PCR检测,均为阳性;而对培养为阴性的20份临床样品进行检测,6份为阳性。该研究结果表明,建立的方法特异性强,敏感性高,稳定性好,能够用于M.bovis的鉴别检测,对牛分枝杆菌病的快速检测和早期诊断具有重要意义。  相似文献   

15.
Mycobacterium bovis is the cause of bovine tuberculosis (bovine Tb) in animals and is considered to be zoonotic and accordingly it infects humans, although cattle are the main host. Buffalo can also be infected and develop bovine Tb. In Iran, almost half a million buffaloes are farmed, mainly in three provinces. In West Azerbaijan, which has the largest numbers of buffaloes, cattle and buffalo are often farmed together. According to the reports of the Iranian Veterinary Organization over the last 25 years, there have been no reports of bovine Tb in buffalo, although the disease is often reported in cattle in this province. Eighteen and 140 pathology specimens from cattle and buffalo, respectively, collected from West Azerbaijani abattoirs were cultured. From one buffalo specimen out of 140, M. bovis was recovered, whereas the pathogen was isolated from 13 cattle specimens. Spoligotyping showed a relatively higher polymorphism within these isolates compared with M. bovis isolated from other Iranian provinces.  相似文献   

16.
本文报道应用两种活体诊断方法检测牛结核病的试验结果。2010～2011年度,应用PPD皮内变态反应方法分3批次对5156头奶牛和奶水牛实施监测,监测阳性数57头,阳性率1.11%。采集该57头牛抗凝全血,应用γ-干扰素酶联免疫吸附试验进行检测,检出阳性样品3份,阳性率5.26%。对14头第一批PPD皮内变态反应阳性牛进行病理检验和进行细菌分离培养和PCR检测,结果3份样品分离培养呈阳性,其中1份PCR鉴定为结核分枝杆菌,2份为非分枝杆菌。PCR方法检测的14份组织样品中,2份为结核分枝杆菌阳性,1份为牛结核分枝杆菌阳性。结合病理剖检和病原PCR诊断,分析比较PPD皮内变态反应试验和γ-干扰素酶联免疫吸附试验的敏感性和特异性,由于PPD纯度、非特异性反应、结果判定的主观性等因素,导致PPD皮内变态反应监测检出假阳性率较高。  相似文献   

17.
The MacKenzie Basin, an area of about 5150 km2 in the South Island of New Zealand, was free of bovine tuberculosis prior to 1980. During the next 13 years, the majority of the cattle and deer herds in this area became infected with Mycobacterium bovis. The history of infection in the MacKenzie Basin has all the characteristics of a newly developed region of endemic tuberculosis with a wildlife reservoir of M. bovis. Tuberculous possums and ferrets were found in the MacKenzie Basin and both may have been a source of infection for domestic animals. DNA fingerprinting of 125 isolates of M. bovis from domestic animals and wildlife by restriction endonuclease analysis revealed two major groups of isolates. The same groups were identified using IS6110 as a DNA probe. Restriction endonuclease analysis enabled one group to be subdivided into seven restriction types and the other group into eight types. Mycobacterium bovis isolates with the most common restriction types were present in both domestic animals and wildlife, indicating that infection had spread between these two groups of animals. DNA fingerprinting also revealed that M. bovis was introduced into the MacKenzie Basin from at least two distinct sources. Furthermore, DNA finger-printing was able to identify probable sources of infection.  相似文献   

18.
The sensitivity and specificity of an ELISA for the detection of bovine IgG anti-Mycobacterium bovis antibodies were 73.6% and 94.1%, respectively, as determined in 53 bacteriologically confirmed tuberculous cattle and 101 healthy cattle from a tuberculosis-free area. In addition, the results of ELISA and tuberculin tests in 149 cattle were compared with those of subsequent necropsy studies. Both tests failed to detect 2 animals with tuberculous lesions and positive culture; 3/12 cattle with M. bovis isolation and no lesions, and 2/7 with atypical mycobacterial infection reacted to tuberculin, but none had antibodies; in 128 cattle with neither lesions nor mycobacterial isolation, 6 were tuberculin reactors and 7 others had antibodies. Negative results were obtained by ELISA in 21/22 paratuberculous cattle. Antibodies were not detected in 88.9% to 96.4% of 697 cattle from two tuberculin negative herds of an endemic area. In a herd with proved M. bovis infection, distribution of seropositive animals in tuberculin and non-tuberculin reactors was similar. Antibody responses to cutaneous tuberculin stimuli were observed in 4 experimentally infected cattle, but only in 2/10 healthy controls after repeated PPD stimuli. Nine controls which had either received a single tuberculin dose or none showed no increase in antibody levels. The low sensitivity of this ELISA limits its usefulness as a diagnostic tool for bovine tuberculosis eradication campaigns. However, it could be helpful in epidemiological surveillance if its efficiency to identify infected herds is demonstrated.  相似文献   

19.
Tuberculosis, caused by Mycobacterium bovis is emerging as the most important disease affecting cattle. Furthermore, it results in a major public health problem when transmitted to humans. Due to its difficult and non-specific diagnosis, M. bovis has been declared to be one of the etiologic agents causing significant economic loss in the cattle industry. Our group evaluated a more rapid and specific method, based on a new polymerase chain reaction species-specific primers, which amplifies a 470-base pair fragment of the M. bovis genome. A total of 275 milk-producing cows were studied by intradermal tuberculin test (ITT) which gave 184 positive and 91 negative cases. From them, 50 animals were taken from a cattle ranch free of tuberculosis. Three different samples were collected from each animal (blood, nasal mucus, and milk). Positive results were obtained from 26 animals by PCR (11.4%), 1 by bacteriological culturing (0.4%) and 1 by bacilloscopy (0.4%). This finding suggests, as in previous reports, that ITT, normally used for bovine tuberculosis detection, has the inconvenience of having a broad range of specificity and sensitivity, and the PCR technique is a more specific and sensitive test to detect infection associated with M. bovis. Therefore, we propose this PCR assay as a useful tool in the epidemiological characterization of infected animals in areas considered to be at high risk of transmission.  相似文献   

20.
On a cattle farm latently infected by M. bovis, field studies aiming at the formation of a mycoplasma free herd, were carried out with a group of newborn female calves. These calves were strictly separated from their dams and any other cattle immediately after parturition. Intensive investigations for mycoplasmas were made over 30 months (mycoplasma isolation from nasal swabs, antibody detection by means of indirect hemagglutination test and ELISA technique). M. bovis could never be isolated from the samples. Also, there were no antibodies to M. bovis. In some animals antibody titers to M. bovis occurred after having contact with cattle infected with M. bovis. The results demonstrate a practicable way to establish cattle herds free from M. bovis infection.  相似文献   

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