Tuberculous Cattle Research Articles

In Tuberculous Cattle Transiting Early Peripartum Period Specific In Vitro PBMC Stimulation Induces an Increase of CD14 Expressing Cells, B B2 Cells, and CD25 Cells with Half IFNγ Production

In Tuberculous Cattle Transiting Early Peripartum Period Specific In Vitro PBMC Stimulation Induces an Increase of CD14 Expressing Cells, B B2 Cells, and CD25 Cells with Half IFNγ Production

Assessment of the frequency of Mycobacterium bovis shedding in the faeces of naturally and experimentally TB infected cattle.

AimsTo assess the prevalence of Mycobacterium bovis bacilli in faecal samples of tuberculous cattle, and to better understand the risk of environmental dissemination of bovine tuberculosis (TB) through the spreading of manure or slurry.Methods and ResultsFaecal samples were collected from 72 naturally infected cattle with visible lesions of TB that had reacted to the tuberculin skin test and 12 cattle experimentally infected with M. bovis. These were examined by microbial culture and PCR to assess the presence of M. bovis bacilli. There were no positive cultures from any naturally infected test reactor animal. A single M. bovis colony was cultured from a faecal sample from one of the experimentally infected animals. A single PCR positive result was obtained from the faecal sample of one naturally infected test reactor.ConclusionsThe prevalence of M. bovis in the faecal samples of TB‐infected cattle was extremely low.Significance and Impact of the StudyThe results suggest that the risk of spreading TB through the use of slurry or manure as an agricultural fertilizer is lower than that suggested in some historical literature. The results could inform a reconsideration of current risk assessments and guidelines on the disposal of manure and slurry from TB‐infected herds.

Cytokine Expression in Peripheral Blood Mononuclear Cell Cultures Obtained from Cattle with Different Stages of Natural Mycobacterium bovis Infection

In bovine tuberculosis (bTB), cellular, humoral, or both types of immune responses have been observed. The purpose of this study was to examine the immune status of tuberculous cows based on the differential cytokine gene expression associated with Th1 (IFN-γ, IL-2), or Th2 (IL-4, IL-10) responses. Twenty-three (23) cows belonging to a dairy herd located in a rural region of the State of Hidalgo, México, were selected for the study. Single Intradermal Comparative Cervical Tuberculin (SICCT) Test, Interferon-Gamma (IFN-γ) Release Assay (BOVIGAM), and Enzyme-Linked Immunosorbent Assay (ELISA) were used for detection of cattle infected by M. bovis. Thirteen cows were positive to all the tests (Group 1); ten cows were positive only to ELISA (Group 2), and the remaining Group (Group 3, control) included cows negative to all the tests. Peripheral blood mononuclear cells (PBMC) from animals were in vitro stimulated by bovin purified protein derivative (PPD), avian PPD, and Concanavalin A (Con A) mitogen for 72h. Changes in the levels of expression of mRNA of the respective cytokines was measured by Reverse Transcription-Polymerase Chain Reaction (RT-PCR) using β-actin gene as internal control. In group 1, PPD bovis and Con A-stimulated cells exhibited high production of IFN-γ, IL-2 and IL-4, but not IL-10. In contrast, PPD avium-stimulated cells displayed a low production of cytokine transcripts. In group 2, cells showed a significant production of IL-10 in response to bovine PPD (P< 0.001). In the control group, a high production of IFN-γ and IL-2 was observed only in Con A-stimulated cells. Post-mortem examinations in animals of group 1 showed slight and medium lesions in lymph nodes, whereas in group 2, the lesions were more extensive. Results indicate differences on gene expression levels of cytokines considered to determine balance in Th1/Th2 response among the evaluated groups. In addition, high levels of antibodies against M. bovis and high IL-10 expression in PBMC together are indicators of progressive bTB when both tuberculin test and IFN-γ assay are negative in tuberculous anergic cattle. Inclusion of serology and IL-10 cytokine expression in in the diagnosis checklist improves detection of infected cattle to help control bovine tuberculosis.

BoLA-DRB3 exon2 polymorphisms among tuberculous cattle: Nucleotide and functional variability and their association with bovine tuberculosis pathology

Bovine tuberculosis (bTB) is caused by Mycobacterium bovis and disseminated worldwide. In Argentina, the highest prevalence occurs in dairy areas. BoLA DRB3.2 is related to the adaptive immunity in mycobacterial infections. Genetic polymorphisms of this marker have been associated with resistance or susceptibility to bovine diseases. We evaluated the association between BoLA DRB3.2 polymorphisms and bTB pathology scores in dairy and beef cattle breeds of Argentina. Most bovines exhibited visible lesions compatible with tuberculosis and, furthermore, 150 (85.7%) were also positive by bacteriology. A pathology index showed a variable degree of disease, from 3 to 76 (median pathology score = 9 (IQR: 7–15)). Thirty-five BoLA DRB3.2 alleles were identified with an associated frequency from 16% to 0.3%, distributed 73% (n = 128) in heterozygosis and 27% (n = 47) in homozygosis, with 12 BoLA DRB3.2 alleles (*0101, *1101, *1501, *0201, *2707 *1001, *1002, *1201, *14011, *0501 *0902 and *0701) representing the 74.7% of the population variability. A functional analysis grouped them in 4 out of 5 clusters (A-D), suggesting a functional overlapping. Among the 90 identified genotypes, *1101/*1101, *1101/*1501 and *0101/*0101 were the most frequent (10%, 8.9% and 8.9%, respectively). No association was detected between the pathology scores and a specific DRB3.2 allele (p > .05). Animals infected with M. bovis spoligotype SB0153 showed a significantly higher pathology score than those affected by the spoligotype SB0145 (p = .018). Furthermore, the Aberdeen Angus breed exhibited highest pathological scores (p < .0001), which were associated with disseminated lesion, thus suggesting that the host component could be important to the disease progression.

Development and diagnostic validation of an ELISA based on an antigenic mixture for the detection of bovine tuberculosis

Programs for the eradication of bovine tuberculosis (bTB) focus on the tuberculin skin test (TST) and slaughter of reactor cattle. However, the disease remains an animal health concern in several countries and improving the efficiency of the TST has become a critical issue. The detection of Mycobacterium bovis antibodies in serum, within weeks after the TST, may be a rapid and inexpensive way to improve bTB control. This study reports the validation of an enzyme-linked immunosorbent assay (ELISA) to detect bovine tuberculosis as an ancillary test to TST in dairy farms in Argentina.The estimated validation parameters were within the established requirements of the World Organization for Animal Health (OIE). The test demonstrated high repeatability, with coefficients of variation <25%. High test reproducibility through interlaboratory testing was also found, with an estimated Pearson coefficient of 0.9648 (95% confidence intervals 0.9315–0.9820). The ELISA detected tuberculous cattle unidentified by the TST. Of 43 animals sent to slaughterhouses that were ELISA positive 15–17 days after a negative TST, 36 were confirmed as infected with M. bovis by histopathology and IS6110 PCR. According to ROC curve analysis of results of 145 cattle from M. bovis-free herds and the 36 M. bovis-infected cattle, at a corrected optical density cut-off point of 0.3853, specificity was 95.95% and the positive predictive value at this cut-off was 83.72%. The ELISA detection test validated in this study could be readily applied in dairy farms, to complement a prior TST and improve livestock health.

Tuberculin PPD Potency Assays in Naturally Infected Tuberculous Cattle as a Quality Control Measure in the Irish Bovine Tuberculosis Eradication Programme.

The Irish Bovine Tuberculosis (bTB) eradication programme operates under national legislation and fulfills OIE and EU trade requirements. Tuberculin purified protein derivative (PPD), a preparation obtained from the heat-treated products of growth and lysis of Mycobacterium bovis or Mycobacterium avium (as appropriate), is critical to the diagnosis of tuberculosis (TB). Standardization of Tuberculin PPD potency, the relative activity in sensitized animals compared to a reference standard, is essential to underpin the reliability of certification for international trade and to ensure that disease eradication programmes are effective and efficient. A Bovine International Standard Tuberculin PPD (BIS) was established by the WHO in 1986 and is used to determine comparative potencies of Tuberculin PPDs. Ideally, Tuberculin PPD potency should be evaluated in the species in which the tuberculin will be used but due to practical difficulties in performing potency assays in cattle, for routine PPD production, they are usually assayed in guinea pigs. Low potency tuberculin PPD is less efficient and thus inferior for bTB diagnosis. Difficulties experienced in the Irish bTB eradication programme have included the supply of sub-standard potency, and thus inferior, bovine (M. bovis) Tuberculin PPD in the late 1970s. The purpose of this paper is to outline the critical role of Tuberculin PPD assays carried out on naturally infected tuberculous cattle, as required by the OIE and under EU legislation in the quality control for the Irish Bovine Eradication Programme. Such assays ensure that the Tuberculin PPD used meets the diagnostic sensitivity and specificity requirements to underpin a successful national eradication programme.

Tuberculosis Caused by Mycobacterium bovis in a Sheep Flock Colocated with a Tuberculous Dairy Cattle Herd in Central Ethiopia.

Mycobacterium bovis (M. bovis) has an exceptionally wide host range including sheep. Information on tuberculosis (TB) in sheep is scarce, and there appears to be conflicting opinions about the relative susceptibility of sheep to infection. In Ethiopia, there was no single previous report on tuberculosis in sheep, though mixed farming of cattle and sheep is a common practice. In this study, following the observation of TB-like lesions on sheep died from sheep flock kept in contact with cattle herd, further investigation was conducted on the flock to assess the magnitude of the infection and identify and characterize the causative M. bovis strain. An outbreak investigation was carried out on 26 eligible sheep out of 33 sheep found on the farm. Comparative intradermal tuberculin (CIDT) test, postmortem examination, Mycobacterium culturing, and spoligotyping were the techniques used in the study. The prevalence of TB in the tested sheep was 15% (4/26). All the sheep that were positive to CIDT had gross lesions suggestive of TB. Three of the positive sheep had extensive and multiple lesions. M. bovis was isolated from all four sheep and the strain was identified as spoligotype SBO134. The in-contact dairy cows were screened for TB and 98% (45/46) of the cows tested positive to CIDT. Furthermore, the same strain, SB0134, was also isolated from the two in-contact cows. The isolation of a matching genotype (SB0134) of M. bovis from both species sharing a known epidemiologic link strongly suggests that the sheep flock might have acquired the pathogen from the dairy cows. This warrants strict physical separation of the sheep flock from the cattle herd to prevent such interspecies transmission of M. bovis.

Hematological and serum biochemical alterations in White Fulani cattle with tuberculous lesions

This study assessed hematological and serum biochemical alterations associated with the occurrence of tuberculous lesions in slaughtered White Fulani cattle at Nsukka abattoir, Enugu State, Nigeria. Diagnosis was confirmed by the immunochromatographic technique and histopathology. Out of 567 cattle examined, ten (1.76%) had tuberculous lesions. The tuberculous cattle had normocytic normochromic anemia, leukocytosis, lymphocytosis, eosinophilia and an increased erythrocyte sedimentation rate. They also had significantly lower (p &lt; 0.05) serum alanine aminotransferase, albumin and urea, and significantly higher (p &lt; 0.05) serum globulin than the apparently healthy control animals. We concluded that the occurrence of tubercles in cattle was associated with alterations in hematological and serum biochemical parameters, which may be relevant to the establishment of an antemortem diagnosis of tuberculosis.

Spectrum of antibody profiles in tuberculous elephants, cervids, and cattle

Using multi-antigen print immunoassay and DPP® VetTB Assay approved in the United States for testing captive cervids and elephants, we analyzed antibody recognition of MPB83 and CFP10/ESAT-6 antigens in Asian elephants (Elephas maximus) infected with Mycobacterium tuberculosis and in white-tailed deer (Odocoileus virginianus), fallow deer (Dama dama), elk (Cervus elaphus), and cattle (Bos taurus) infected with Mycobacterium bovis. Serum IgG reactivity to MPB83 was found in the vast majority of tuberculous cattle and cervid species among which white-tailed deer and elk also showed significant CFP10/ESAT-6 recognition rates with added serodiagnostic value. In contrast, the infected elephants developed antibody responses mainly to CFP10/ESAT-6 with MPB83 reactivity being relatively low. The findings demonstrate distinct patterns of predominant antigen recognition by different animal hosts in tuberculosis.

Potential for rapid antibody detection to identify tuberculous cattle with non-reactive tuberculin skin test results

BackgroundBovine tuberculosis (TB) control programs generally rely on the tuberculin skin test (TST) for ante-mortem detection of Mycobacterium bovis-infected cattle.ResultsPresent findings demonstrate that a rapid antibody test based on Dual-Path Platform (DPP®) technology, when applied 1-3 weeks after TST, detected 9 of 11 and 34 of 52 TST non-reactive yet M. bovis-infected cattle from the US and GB, respectively. The specificity of the assay ranged from 98.9% (n = 92, US) to 96.0% (n = 50, GB) with samples from TB-free herds. Multi-antigen print immunoassay (MAPIA) revealed the presence of antibodies to multiple antigens of M. bovis in sera from TST non-reactors diagnosed with TB.ConclusionsThus, use of serologic assays in series with TST can identify a significant number of TST non-reactive tuberculous cattle for more efficient removal from TB-affected herds.

Cross-Sectional Study and Comparison of Different Diagnostic Methods of Bovine Tuberculosis in Gondar Elfora Abattoir, Ethiopia

Background: The prevalence of bovine tuberculosis is high in developing countries due to lack of awareness, good diagnostic methods and prevention strategies. Therefore, this study aims to estimate the prevalence of bovine tuberculosis and compare the efficacy of different procedures of diagnosis. Methods: A cross sectional study was conducted at Gondar Elfora abattoir from December, 2005 to June, 2006. To this effect, comparison has been made between the detailed postmortem examination and routine meat inspection procedures with gold standard culture result. Result: Out of 402 animals examined at slaughter, 15.9% were diagnosed with gross tuberculous lesions by detailed laboratory examination. Routine abattoir inspection detected only 2.9% of the tuberculous cattle. From 64 cattle considered tuberculous, 10 show growth in Lowenstein-Jensen. The average number of lesions per infected cattle was 1.6% and 55.5% of cattle with tuberculous lesions possessed single lesion. All the traits (including sex, age and body condition score) measured in relation to tuberculous lesions did not show a statistically significant difference among the categories. The sensitivity of routine meat inspection was 18.8% with detailed postmortem examination and 30% with culture in comparison with 83.3% specificity. There was a poor agreement (k=0.18) between routine meat inspection and detailed postmortem examination procedures. Similarly, a poor agreement (k=0.12) was obtained between routine meat inspection procedure and culture result. Conclusion: Relatively higher prevalence was recorded, and there is a need to improve the sensitivity of routine abattoir inspection procedures to diagnose tuberculous.

Serological Follow-up of Tuberculosis in a Wild Boar Population in Contact with Infected Cattle.

There is an increasing concern in several European countries over the role that tuberculosis (TB)-infected wild boar may play in the progress of bovine TB eradication campaigns. In 2004, as a consequence of the detection of a TB focus in wild boar from a National Game Reserve (NGR) located in southern Catalonia, a surveillance programme based on post-mortem inspection for detection of macroscopic TB-like lesions (TBLL) was initiated in the affected area. The source of infection for wild boar was linked to a tuberculous cattle herd located in the same area. Besides, the results of the surveillance programme in wild boar were used for the validation of an indirect enzyme-linked immunosorbent assay (ELISA) specific for Mycobacterium tuberculosis complex (MTBC) IgG antibodies. Using this ELISA, a seven-year serological study of MTBC in wild boar from the NGR was conducted in 173 animals (93 adults, 44 juveniles-yearlings and 36 piglets) culled between 2004 and 2010. ELISA results and presence of TBLL showed excellent agreement for adult and juvenile wild boar (Kappa index=0.85; 95% CI: 0.76-0.95). Of the thirty-eight adults, yearlings and juveniles classified as positives by the ELISA, 34 (89%) showed TBLL at necropsy. In contrast, none of the ELISA-positive wild boar piglets (n=20) showed TBLL, suggesting the detection of early antibody responses to the infection. Overall, this study contributes to the knowledge of wild boar humoral responses to MTBC. The results also highlight the usefulness of this serological test for wild boar TB surveillance.

TALE nickase-mediated SP110 knockin endows cattle with increased resistance to tuberculosis.

Transcription activator-like effector nuclease (TALEN)-mediated genome modification has been applied successfully to create transgenic animals in various species, such as mouse, pig, and even monkey. However, transgenic cattle with gene knockin have yet to be created using TALENs. Here, we report site-specific knockin of the transcription activator-like effector (TALE) nickase-mediated SP110 nuclear body protein gene (SP110) via homologous recombination to produce tuberculosis-resistant cattle. In vitro and in vivo challenge and transmission experiments proved that the transgenic cattle are able to control the growth and multiplication of Mycobacterium bovis, turn on the apoptotic pathway of cell death instead of necrosis after infection, and efficiently resist the low dose of M. bovis transmitted from tuberculous cattle in nature. In this study, we developed TALE nickases to modify the genome of Holstein-Friesian cattle, thereby engineering a heritable genome modification that facilitates resistance to tuberculosis.

An outbreak of tuberculosis in Lleyn sheep in the UK associated with clinical signs

This case report describes an outbreak of Mycobacterium bovis infection a Lleyn sheep flock associated with clinical signs of illthrift. There was no known direct contact with tuberculous cattle although...

An outbreak of tuberculosis in Lleyn sheep in the UK associated with clinical signs

This case report describes an outbreak of Mycobacterium bovis infection a Lleyn sheep flock associated with clinical signs of illthrift. There was no known direct contact with tuberculous cattle although...

Estimation of the Relative Sensitivity of the Comparative Tuberculin Skin Test in Tuberculous Cattle Herds Subjected to Depopulation

Bovine tuberculosis (bTB) is one of the most serious economic animal health problems affecting the cattle industry in Great Britain (GB), with incidence in cattle herds increasing since the mid-1980s. The single intradermal comparative cervical tuberculin (SICCT) test is the primary screening test in the bTB surveillance and control programme in GB and Ireland. The sensitivity (ability to detect infected cattle) of this test is central to the efficacy of the current testing regime, but most previous studies that have estimated test sensitivity (relative to the number of slaughtered cattle with visible lesions [VL] and/or positive culture results) lacked post-mortem data for SICCT test-negative cattle. The slaughter of entire herds (“whole herd slaughters” or “depopulations”) that are infected by bTB are occasionally conducted in GB as a last-resort control measure to resolve intractable bTB herd breakdowns. These provide additional post-mortem data for SICCT test-negative cattle, allowing a rare opportunity to calculate the animal-level sensitivity of the test relative to the total number of SICCT test-positive and negative VL animals identified post-mortem (rSe). In this study, data were analysed from 16 whole herd slaughters (748 SICCT test-positive and 1031 SICCT test-negative cattle) conducted in GB between 1988 and 2010, using a Bayesian hierarchical model. The overall rSe estimate of the SICCT test at the severe interpretation was 85% (95% credible interval [CI]: 78–91%), and at standard interpretation was 81% (95% CI: 70–89%). These estimates are more robust than those previously reported in GB due to inclusion of post-mortem data from SICCT test-negative cattle.

Trichoderma pseudokoningii Rifai isolation from Egyptian immunocompromised cattle with Mycobacterium bovis infection

AbstractRecently, Trichoderma species have emerged as potent fungal pathogens in immunocompromised humans. We report the first three cases of Trichoderma pseudokoningii Rifai pulmonary infection in the Egyptian tuberculous dairy cattle with Mycobacterium bovis, from the heart of a generalized bovine TB in a cow over 5 years old, a mediastinal lymph node of pulmonary TB in a cow of 3 years old, and a lung of mixed pulmonary and digestive BTB in a cow of 4 years old. We have also developed a pathogenisity test technique for Trichoderma pseudokoningii Rifai infection in 3 G. pigs by intraperitoneal injection of 2 G. pigs with mixed infection of Mycobacterium bovis and Trichoderma pseudokoningii Rifai; death of both animals 14 days, thereafter, and by injection of 1 G. pig with single infection of Trichoderma pseudokoningii Rifai; death of animal 21 days, thereafter. We did not report any animal case along review of literature.

Mycobacterium tuberculosis infection in grazing cattle in central Ethiopia

A preliminary study to characterise mycobacteria infecting tuberculous cattle from two different management systems in central Ethiopia was carried out. Approximately 27% of isolates from grazing cattle were Mycobacterium tuberculosis, while cattle in a more intensive-production system were exclusively infected with M. bovis. The practice of local farmers discharging chewed tobacco directly into the mouths of pastured cattle was identified as a potential route of human-to-cattle transmission of M. tuberculosis.

Detection of Mycobacterium bovis DNA in nasal swabs from tuberculous cattle by a multiplex PCR

Detection of tuberculosis in cattle relies on the intradermal tuberculin test (ITT), but a definitive diagnosis requires identification of the pathogen after the animal is slaughtered. DNA in nasal swabs from 50 cows was analyzed by m-PCR, targeting for the RvD1-Rv2031c and IS6110 sequences. M. bovis was identified in two of 34 tuberculous cows (5.9%). The use of mPCR of nasal swabs as an in vivo diagnostic tool for bovine tuberculosis is suggested.

Repeat tuberculin skin testing leads to desensitisation in naturally infected tuberculous cattle which is associated with elevated interleukin-10 and decreased interleukin-1 beta responses.

The principal surveillance tool used to control bovine tuberculosis in cattle is the removal of animals that provide a positive response to the tuberculin skin-test. In this study we performed a longitudinal investigation of the immunological and diagnostic consequences of repeated short-interval skin-tests in cattle naturally infected with Mycobacterium bovis. Tuberculin skin-test positive cattle were subjected to up to four further intradermal comparative cervical skin-tests at approximately 60-day intervals. A significant progressive reduction in the strength of the skin-test was observed after successive tests. In contrast, the magnitude of interferon-γ (IFN-γ) responses was not influenced by repeat skin-testing either transiently around the time of each skin-test or longitudinally following repeated tests. A significant boost in blood interleukin-10 (IL-10) production was observed within 3 days following each skin-test although the magnitude of this boosted response returned to lower levels by day 10 post-test. The application of a novel multiplex assay to simultaneously measure seven cytokines and chemokines also identified that skin-testing resulted in a significant and progressive reduction in antigen specific interleukin-1β (IL-1β) whilst confirming stable IFN-γ and elevated IL-10 responses in the blood. Therefore, we have demonstrated that in cattle naturally infected with M. bovis, repeat short-interval skin-testing can lead to a progressive reduction in skin-test responsiveness which has potential negative consequences for the detection of infected animals with marginal or inconclusive skin-test responses. The desensitising effect is associated with decreased IL-1β and elevated IL-10 responses, but importantly, does not influence antigen specific IFN-γ responses.