Diagnosis Of Tuberculosis In Cattle Research Articles

Study on supplemental test to improve the detection of bovine tuberculosis in individual animals and herds

BackgroundBovine tuberculosis (bTB), is a worldwide disease caused by Mycobacterium bovis (M. bovis). The success of bTB eradication and control programs is based on early detection and the removal of reactors from a herd thus routine testing and cull strategy have been applied globally. Since the late nineteenth century, the Tuberculin Skin Test (TST) has been the primary antemortem test available to support bTB eradication campaigns. Due to the TST limitations in terms of Se and Sp, the credibility of the diagnosis is frequently questioned given the occurrence of false-positive and false-negative reactions, therefore, it is necessary to confirm reactive animals using other methods, ensuring the reliability of the diagnosis. The aim of this study was to evaluate the sensitivity and specificity of a multiplex enzyme-linked immunosorbent assay (ELISA) relative to the tuberculin test used for the diagnosis of tuberculosis in cattle in Brazil.ResultsLack of agreement between comparative cervical tuberculin test and ELISA IDEXX TM was observed. The 2 animals positive on the comparative cervical tuberculin test did not react at the ELISA IDEXX TM and 22 negative reactors by comparative cervical tuberculin test were positive by the ELISA IDEXX TM. The ELISA IDEXX TM showed sensitivity that is significantly lower than the official screening test the single cervical tuberculin. ELISA IDEXX TM also detected infected animals and herds undetected by the comparative cervical tuberculin test. The parallel use of comparative cervical tuberculin test and ELISA IDEXX TM increased sensitivity and the feasibility bTB screening.ConclusionsThe results obtained here suggest that the ELISA IDEXX TM may be a supplemental test for the detection of Mycobacterium bovis infection in regions without routine testing and slaughter, where the disease generally progresses to more advanced stages and antibody responses are likely to be more prevalent. Evidence to support the validation of the ELISA IDEXX™ as a supplemental test for bTB eradication programs was provided.

Diagnosis of Bovine Tuberculosis by Comparative Intradermal Tuberculin Test, Interferon Gamma Assay and <i>esxB</i> (CFP-10) PCR in Blood and Lymph Node Aspirates

Bovine tuberculosis (TB) is a chronic debilitating disease of huge economic importance due to loss in production, morbidity and mortality, and has a potential zoonotic threat. TB is endemic in India and has a worldwide prevalence, therefore, needing early diagnostic technique for the eradication of TB globally. Currently, compared to the eradication programme of TB in Medical sector, Veterinary sector is lagging behind though TB is one of the major zoonotic diseases prevalent in dairy animals and wildlife in India. With the “End TB” strategy by WHO in human, parallel measures for early diagnosis and culling has to be followed in case of animals for an overall successful eradication programme. The objective of this study is diagnosis of TB in cattle and buffaloes by using the cell-mediated immune response tests, i.e. Comparative Intradermal Tuberculin Test (CITT) and Interferon gamma (IFN-γ) assay, and Polymerase Chain Reaction (PCR) targeting esxB gene (CFP-10 protein) and to compare their diagnostic capabilities. This study was carried out in 202 dairy cattle and buffaloes from an organized dairy farm, where almost all of the animals appeared clinically healthy. We found that, the combined use of both CITT and IFN-γ assay lead to more accurate diagnosis of TB, although IFN-γ assay was more specific than CITT. However, esxB PCR showed almost similar sensitivity to IFN-γ assay and may be used as a fast alternative method for the diagnosis of bovine TB from blood samples.

Professor Odo Bujwid - from medicine to veterinary medicine

Prof. Odo Bujwid was one of the pioneers of microbiology, prophylaxis and hygiene as well as the precursor of the fight against zoonoses. He was a student of Robert Koch and Luis Pasteur. He propagated the discoveries of these scientists in Poland. In 1885 he established the first bacteriological laboratory in Poland (Warsaw, Wilcza 12). In 1886 he opened a branch of Pasteur’s Institute in Warsaw (the first outside Paris). Following the ideas of Koch he produced a new product, which he called tuberculin and introduced it to the diagnosis of tuberculosis in cattle. He produced many sera and vaccines for medicine and veterinary medicine In the history of medicine and veterinary he enrolled as a pioneer of Polish bacteriology and creator of the biomedical and bioveterinary industries.

Detection of Mycobacterium bovis in whey, by multiplex polymerase chain reaction (PCR) and bacteriological culture

The objective of this study was to detect the presence of Mycobacterium bovis (M. bovis) in whey. A total of 233 cow milk samples were analyzed together with 26 tank milk samples that came from dairy herds of several states of the Mexican Republic (Queretaro, San Luis Potosi, Guanajuato, Hidalgo, Coahuila). DNA was obtained from whey and used for polymerase chain reaction-multiplex (PCR-M). Tuberculosis complex was first identified through the detection of gene RD1. Positive samples were subjected to a second PCR-M with the primers for gene RD9 to identify M. bovis. Samples were bacteriologically cultured using conventional techniques for the isolation of mycobacteria. Cohen’s Kappa test (k) and Pearson’s Chi2 were carried out for statistical analysis. A 150 bp amplification product of the RD1 region was obtained, which corresponds to the tuberculosis complex, in 34/233 (14.59%) of the individual milk samples and in 4/26 (15.38%) of the tank milk samples. PCR-M with primer RD9, of the 34 individual samples and the 4 tank milk samples, gave an amplification product of 200 pb, which is the expected product for M. bovis. By bacteriological culture, six isolates were obtained; four in individual whey samples and two from tank milk samples, which were then classified by biochemical tests as M. bovis. The concordance between RD1, RD9 PCR-M and bacteriological culture was low, but there was a significant difference between diagnostic techniques with a P = 0.000. The results showed the potential of the PCR-M as a confirmatory test for the diagnosis of tuberculosis in cattle, as well as the advantage of using whey samples, that may be a possible source of infection for the herd and/or humans. Key words: Bovine tuberculosis, whey, polymerase chain reaction-multiplex (PCR-M), RD1, RD9, milk, isolation.

Cross reactive immune responses in cattle arising from exposure to Mycobacterium bovis and non-tuberculous mycobacteria

Accurate diagnosis of tuberculosis in cattle may be compromised in areas where there are high rates of exposure to environmental/non-tuberculous mycobacteria (NTM). This cross reaction of immune responses to Mycobacterium bovis antigens shared with NTMs can result in reduced specificity of commonly used diagnostic tests including tuberculin skin tests and the interferon gamma assay (IFN-ɣ). In this study we assessed the cross-reactive immune responses of M. bovis (infected) and NTM exposed animals to M. bovis and M. avium tuberculin, the ESAT6/CFP10 cocktail antigen, tuberculin derived from cultures of selected NTMs, and a panel of recombinant mycobacterium tuberculosis complex (MTBC) antigens sharing homology with orthologues in NTM. Gamma interferon (IFN-ɣ) responses were measured in whole blood cultures using the IFN-ɣ assay and the IFN-ɣ elispot assay on purified peripheral blood mononuclear cells (PBMC).We observed the expected strong IFN-ɣ response to PPD-B in the M. bovis infected animals that distinguished this group from non-infected NTM exposed cattle. The IFN-ɣ responses to PPD-N (M. nonchromogenicum), were relatively high in both infected and non-infected NTM exposed cattle, but were not significantly different to classify the true infection status of each group. The results indicated that the cross-reactive responses to PPD-B and/or PPD-A with PPD-N, likely arose from prior exposure to environmental non-tuberculous mycobacteria. The IFN-ɣ immune responses to the 10 R-Mag measured by the IFN-ɣ elispot assay revealed that three of the selected antigens, Rv3615 (ESpC), Rv0287 (esxG) and the ESAT6/CFP10, were immunogenic in the infected cattle, and distinguished the infected cattle from the non-infected NTM exposed animals. The combined data of PPDs and R-Mags derived from NTM mycobacteria may prove useful in future development of novel bTB diagnostic tests.

Sensibilidad y especificidad de PCR anidada y Spoligotyping como pruebas rápidas de diagnóstico de tuberculosis bovina en tejido fresco

Quick diagnosis of tuberculosis in cattle is critical to decide whether or not to quarantine or depopulate a herd. Currently, decisions are based on culture, which takes between four and eight weeks to accomplish. Therefore, the purpose of this study was to evaluate relative sensitivity and specificity of two PCR´s (MPB70 and IS6110) and spoligotyping as quick diagnostic tests for cattle tuberculosis in fresh tissue.

Advances in ante-mortem diagnosis of tuberculosis in cattle

The tuberculin skin test is effective in the early detection of pre-clinical cases of Mycobacterium bovis infection in cattle. This allows the rapid removal of infected animals, thus limiting transmission of the disease, and has resulted in the eradication of bovine tuberculosis (Tb) from many countries. This test is very likely to remain the primary screening test for M. bovis infection in cattle as it is a simple, robust and inexpensive test. However, a number of ancillary tests are being used, or are currently being validated. These ancillary tests are likely to provide a more accurate diagnosis following skin-testing. The blood-based BOVIGAM interferon-γ (IFN-γ) test is a cellular immune assay which can detect early infection, and has become the main ancillary test in New Zealand. It can be used for re-testing skin test-positive animals, to improve specificity and minimise wastage from slaughtering animals with false-positive tests. Alternatively, it can be used in locations of increased risk of infection in parallel with skin-testing, for examining skin test-negative animals for pre-movement testing or in problem herds to identify M. bovis-infected animals that do not respond to the skin test. Several modifications of the test are now being used to improve specificity by altering the cut-off or using specific antigens present in virulent mycobacteria such as the 6 kDa early secreted antigenic target (ESAT 6) and 10 kDa culture filtrate protein (CFP-10). While antibody based tests generally lack sensitivity, as high levels of antibodies tend to occur late in the disease process, they may have unique desirable properties such as the ability to be used as a cow-side test. The use of these new ancillary tests in association with skin-testing will improve the detection of M. bovis-infected cattle and reduce the unnecessary slaughter of false-positive reactors.

Antibody to ovalbumin and delayed-type hypersensitivity to Candida albicans and mycobacteria in lactating Holstein cows using Quil A or Freund's complete adjuvant

Antibody-mediated immune response (AMIR) to ovalbumin (OVA) or hen-egg white lysozyme (HEWL) and cell-mediated immune response (CMIR) such as delayed-type hypersensitivity (DTH) to mycobacteria have been proposed as quantitative traits for selective breeding to improve animal health. However, DTH to mycobacteria may confound diagnosis of tuberculosis in cattle. Candida albicans, a yeast also known to induce DTH, was tested as an alternative for DTH induction and testing since it is not a target of regulatory diagnostic tests. Other objectives were to determine if both AMIR and CMIR in cattle receiving Quil A as adjuvant were equivalent to corresponding responses induced by Freund's complete adjuvant (FCA). Forty lactating Holstein cows were randomly assigned to two treatment groups, which received ovalbumin (OVA) and C. albicans adjuvanted with FCA and Freund's incomplete adjuvant (FIA) on days 0 and 14, respectively, or Quil A on days 0 and 14. The FCA was used as adjuvant and as a source of Mycobacterium tuberculosis-induced DTH. Testing for DTH was performed on day 21 with killed C. albicans whole cell (CaWC), a purified extract from C. albicans (candin) and M. phlei purified protein (phlein). Both primary and secondary antibody responses to OVA were statistically significant and similar in both FCA and Quil A treatment groups. No significant differences were detected in immunoglobulin G (IgG) isotypic-mediated responses to OVA or candin between groups. C. albicans adjuvanted with Quil A induced DTH reaction similar to those induced by C. albicans and mycobacteria in FCA.

Appraisal of interpretation criteria for the comparative intradermal tuberculin test for diagnosis of tuberculosis in cattle in central Ethiopia.

Accurate detection and removal of infected cattle, using immunodiagnostic tests such as the comparative intradermal tuberculin (CIDT) test, are the basis of control strategies for bovine tuberculosis (TB). According to the Office des Internationale Epizooties recommendation, the cutoff point for positivity of the CIDT test, calculated as the difference between skin thicknesses after bovine tuberculin (B) and avian tuberculin (A) injections (B - A), is >4 mm. This cutoff point is used worldwide, although it is likely that local conditions influence test performance. Thus, this study was formulated to determine CIDT test cutoff points applicable to cattle in central Ethiopia. Receiver operating characteristic analysis was performed for the CIDT test, using data from 186 Bos indicus (zebu) and Bos taurus (Holstein) cattle. Detailed postmortem examination for the presence of TB lesions was used to define disease status. At a cutoff of >2 mm, CIDT test sensitivity was 69% (95% confidence interval [95% CI], 58.5 to 79%), while it was 59% (95% CI, 49 to 69%) at a cutoff of >4 mm. In contrast, specificities of the CIDT test at these two cutoff values were identical, at 97% (95% CI, 89 to 100%). Thus, the maximum sensitivity of the CIDT test can be realized using a >2-mm cutoff without affecting specificity. The apparent prevalence was significantly (chi(2) = 13.56; P < 0.001) higher at a cutoff of >2 mm (16.0%; n = 5,424) than at a >4-mm cutoff (13.5%; n = 5,424). Nonetheless, no significant difference (chi(2) = 2.15; P = 0.14) in true prevalence was observed at a cutoff of >2 mm (19.6%) and at a cutoff of >4 mm (18.5%). Thus, our study demonstrates the importance of defining local, relevant cutoff values to maximize test sensitivity, and we suggest the application of the >2-mm cutoff for testing of cattle in central Ethiopia.

Comparison of histologic techniques for the diagnosis of bovine tuberculosis in the framework of eradication programs

Rapid diagnosis of tuberculosis in cattle reacting positive in antemortem assays is crucial in countries where eradication programs are operated to confirm the presence of the infection in tuberculosis-free herds. This study evaluated the accuracy of histopathologic examination by hematoxylin and eosin and Ziehl-Neelsen (ZN) staining applied in this framework, when suspected lesions are caused by low infectious doses and are detected in early stages of the disease. For this purpose, histologic methods were compared with mycobacterial culture as reference test on suspected lymph node samples from 173 cattle reacting positive in antemortem tests. Histopathology demonstrated high sensitivity (93.4%) and specificity (92.3%), while ZN sensitivity and specificity were respectively 33.9% and 100%. There was good agreement between histopathology and bacterial culture, suggesting that histopathologic examination is a reliable tool for rapid diagnosis in countries where active tuberculosis eradication programs allow the prompt identification and elimination of reactor cattle. Histopathology permits identification of typical mycobacterial lesions and its differentiation from other causes.

Repaso de las pruebas existentes para el diagnóstico de la tuberculosis en especies no bovinas

Bovine tuberculosis is an important disease that has impacts on regional and international trade. The disease can affect both social and economic stability and have a deleterious affect on species diversity. The intradermal tuberculin test has been in use for almost a century and, despite the technological advances of the last two decades, is still the only prescribed test for the diagnosis of tuberculosis in cattle. Many other species of animal, including humans, can be infected with Mycobacterium bovis. This paper reviews the various tests that have been used by researchers for detecting infection with M. bovis in a variety of animal species, and attempts to prioritise or comment on the importance of having appropriately validated diagnostics for the different species. The difficulties of test validation using small numbers of animals, especially when tuberculosis occurs in only a few instances or the species of animal affected is rare and/or valuable, are discussed.

The use of Ag85 complex as antigen in ELISA for the diagnosis of bovine tuberculosis in dairy cows in Brazil.

Several enzyme-linked immunosorbent assay (ELISA) methods have been investigated to evaluate their performance in the diagnosis of tuberculosis in cattle. Increased production of antibodies to the proteins of antigen 85 complex (Ag85) after experimental and natural infection in cattle shows that they are strongly immunogenic in vivo. The purpose of this study was to evaluate the use of Ag85 as an antigen in ELISA for the diagnosis of bovine tuberculosis in dairy cows in Brazil. The test groups consisted of 46 serum samples from intradermal tuberculin test (ITT)-positive animals (Group A) and 46 samples from ITT-negative animals (Group B). Group C comprised 12 samples from a tuberculosis-free herd and was the control group of the study. Samples were tested in an ELISA using Ag85 as antigen. Differences between the mean ODs of groups A and B and A and C were significant (P < 0.01), but no significant difference (P > 0.05) was observed between groups B and C. The sensitivity of the ELISA using Ag85 was 91.3% (42/46) and its specificity was 94.8% (55/58). These results were not significantly different (P > 0.05) from those observed in a previous study of an ELISA using purified protein derivative (PPD). We concluded that, although Ag85 can be used as antigen for ELISA tests in the diagnosis of bovine tuberculosis with good sensitivity and specificity rates, no significant advantages were observed in relation to the ELISA using PPD that could justify the purification and utilization of Ag85 as a single antigen in routine methods of diagnosis.

An abattoir monitoring system for diagnosis of tuberculosis in cattle in Baja California, Mexico

Objective To implement a disease monitoring system in federal, municipal, and private abattoirs in Baja California, Mexico and to estimate annual prevalence of tuberculosis (TB) in beef and dairy cattle slaughtered and inspected in 1995 and 1996. Design Epidemiologic survey. Animals About 200,000 cattle (95% beef, 5% dairy). Procedures Lymph node and tissue specimens with lesions suggestive of TB were fixed in neutral-buffered 10% formalin and embedded in paraffin. Sections were stained with H&amp;E and Ziehl-Neelsen and examined for typical tuberculous lesions and acid-fast bacilli. Annual prevalence and 95% confidence intervals were estimated. Results Prevalence of TB in all slaughtered cattle was 0.12 and 0.46% in 1995 and 1996, respectively (beef cattle, 0.02 and 0.05%, respectively; dairy cattle, 2.0 and 8.3%, respectively). Tuberculosis cases/1,000 slaughtered cattle were linearly associated with monthly volumes of tissue submissions. Clinical Implications It is critical to quantify the monitoring activity at abattoirs to better estimate the prevalence of TB in slaughtered cattle. Annual prevalence of TB was significantly greater in dairy cattle than in beef cattle. Veterinarians and cattle producers in this region are encouraged to develop and work on herd plans aimed at controlling and eradicating TB. (J Am Vet Med Assoc 1997;211:709–711)

Evaluation of a five-antigen ELISA for diagnosis of tuberculosis in cattle and Cervidae

Objective To determine the validity of a 5-antigen ELISA for detection of tuberculosis in cattle and Cervidae. Design Cross-sectional observational study. Sample Population Serum samples collected from 5,304 cattle in 23 herds and 1,441 Cervidae in 12 herds. Procedure Discriminant analysis was used to determine the linear combination of antigens that accurately predicted the true Mycobacterium bovis infection status of the most animals. The resulting classification functions then were used to calculate the percentage of animals that were correctly classified (ie, sensitivity and specificity). The kappa statistic was calculated to evaluate different combinations of test results, Results Of the 23 cattle herds, 4 dairy and 2 beef herds were considered infected. Of the 12 Cervidae herds, 5 were considered infected. For cattle, the specificity and sensitivity of ELISA, using the discriminant function, were 56.4 and 65.6%, respectively. For Cervidae, the specificity and sensitivity of ELISA, using the discriminant function, were 78.6 and 70.0%, respectively. Clinical Implications Results suggest that the 5- antigen ELISA would not be a good test for tuberculosis, especially in cattle, if used alone. However, when results of the ELISA and tuberculin test were interpreted in parallel, sensitivity of the combination was greater than sensitivity of either test alone. Similarly, when results of the 2 tests were interpreted in series, specificity of the combination was greater than specificity of either test alone. (J Am Vet Med Assoc 1996; 209:962-966)

An Assessment of the Single Intradermal Comparative Test in the Diagnosis of Bovine Tuberculosis in Cattle Previously Vaccinated Against Johne's Disease

An Assessment of the Single Intradermal Comparative Test in the Diagnosis of Bovine Tuberculosis in Cattle Previously Vaccinated Against Johne's Disease

Societies and Academies

Societies and Academies

THE COMPLEMENT FIXATION TEST IN THE DIAGNOSIS OF TUBERCULOSIS

The great desirability of reaching an early diagnosis in tuberculous infection has led to much work being done in laboratories in the search for some test which would make such a diagnosis possible, with the result that the various tuberculin reactions were discovered, as well as other methods, as the serum skin reaction of Bronfenbrenner.1All of these diagnostic methods, however, have proved more or less unsatisfactory in thes differentiation of active and inactive tuberculosis and a criterion of cure. As early as 1910, Ruppel and Rickmann2made use of complement fixation in the diagnosis of tuberculosis in cattle, using old tuberculin as an antigen, and obtained positive results only in tuberculous animals, although no fixation occurred in some known to be tuberculous. Stimulated by this work, numerous investigators, notably Caulfield and Beatty,3Calmette and Massol,4and Hammer,5demonstrated that complement fixation could be obtained

The Diagnosis of Tuberculosis in Cattle by Inoculation of Guinea-Pigs with Anti-Formined Fæeces

The Diagnosis of Tuberculosis in Cattle by Inoculation of Guinea-Pigs with Anti-Formined Fæeces

The complement test in the diagnosis of tuberculosis in cattle

The complement test in the diagnosis of tuberculosis in cattle

FURTHER STUDIES ON THE OPHTHALMO-TUBERCULIN REACTION IN CATTLE

Our conclusions differ in a few points from those of our first work on this subject. We are able to sum up our observations in this series of experiments as follows: 1. The ophthalmo-tuberculin test is of limited value in the diagnosis of tuberculosis in cattle. In some cases the reaction is very slight (hyperaemia). In others more pronounced congestion with profuse exudates are noted. Accuracy of observation is important. We are inclined to rely primarily on the results of the first instillation of tuberculin. Second instillations in a few instances elicit reaction in non-tubercular animals. 2. In the majority of animals tested the reaction increased in its intensity with each subsequent instillation of tuberculin. This fact indicates the development of a local hypersusceptibility or anaphylaxis associated with a partial immunity; von Pirquet calls this condition "allergie" (9). 3. It is possible in some cases to create a condition of "allergie" in healthy cattle, when spaced instillations of tuberculin are made. It is evident, therefore, that the result of the first instillation of tuberculin should be made the only basis of diagnosis. Rosenau and Anderson (II) have recently called attention to this point in regard to the human subject. 4. When repeated instillations of tuberculin are made on the conjunctiva at short intervals (twenty-four hours, etc.) a local immunity results (No. 3D et al.). If the instillations are separated two weeks or more anaphylaxis results. 5. We, therefore, hold that if tuberculin (0.1 cubic centimeter) is carefully instilled into the conjunctival sac and if careful comparison of the instilled eye with the opposite eye shows that a reaction of varying intensity results in from ten to twelve hours after the first instillation, a tubercular lesion is present. 6. In our first report (7) we were inclined to believe that subcutaneous tubercular injection given previous to the ocular test would slightly inhibit it. We have since become convinced that this is true only to a limited extent, and that in some cases the ophthalmo-reaction is exaggerated by a subcutaneous injection of tuberculin. 7. The primary ophthalmo-tuberculin reaction is in direct proportion to the extent of the tubercular processes in the body. The more extensive the tubercular processes, the more anaphylactic the animal is. This is in direct variance with the condition in the usual subcutaneous tuberculin test. (See Necropsy Report.) 8. We are inclined to believe that the ophthalmo-tuberculin test will reveal tuberculosis at as early a state as the usual subcutaneous test. 9. The ophthalmo-reaction is of no value in determining whether vaccinated cattle are actively tubercular or not, or in demonstrating any hypersusceptibility in the offspring of tubercular cattle. 10. The cutaneous test from our brief series of experiments does not seem to be as accurate as the ophthalmic test. This conclusion has been reached by several investigators.