Lesion development and immunohistochemical changes in granulomas from cattle experimentally infected with Mycobacterium bovis

Mycobacterium bovis, the causative agent of bovine tuberculosis, persists within granulomas. Formation of granulomas involves a complex array of immune activation and cellular migration. To examine temporal changes in granuloma development, we inoculated 32 cattle with M. bovis of deer origin. Tissues from 4 calves each were examined at 15, 28, 42, 60, 90, 180, 270, and 370 days after inoculation. Granulomas in the medial retropharyngeal lymph node were staged (I-IV) on the basis of cellular composition and the presence or absence of necrosis and peripheral fibrosis. Immunohistochemistry for inducible nitric oxide synthase (iNOS), CD68, CD4, CD8, and gamma/delta T cells was performed. Fifteen days after inoculation only stage I granulomas were seen, while between 28 and 60 days, there was a steady progression through granuloma stages such that by day 60, granulomas of all 4 stages were seen. Acid-fast bacilli were present in moderate-to-large numbers in stage I granulomas 15-60 days after inoculation. Stage IV granulomas contained large numbers of acid-fast bacteria. Abundant iNOS immunoreactivity was associated with granulomas from day 15 through day 60 but was minimal from day 90 to the termination of the experiment. The relative number of CD4+ and CD68+ cells remained constant throughout the study. In contrast, at time points >60 days, numbers of CD8+ and gamma/delta T cells diminished. Tuberculous granulomas are dynamic lesions that follow an orderly progression through disease stages. Diminished expression of iNOS and reduced numbers of CD8+ and gamma/delta T cells late in the progression of tuberculous granulomas may represent a failure of the host response to control infection.     

Distribution and activation of T-lymphocyte subsets in tuberculous bovine lymph-node granulomas

The immune response against mycobacterial infections is dependant upon a complex interaction between T lymphocytes and macrophages in the context of the granuloma. For this study, we performed the analysis of 18 stage I or II, and 13 stage III or IV granulomas found in lymph nodes from 8 experimentally and 2 naturally infected cattle. T-cell subpopulations (CD3(+), CD4(+), CD8(+), WC1(+), CD25(+)) were investigated by immunohistochemistry. In the majority of stage I/II lesions, CD8(+) and CD25(+) cells were predominantly found in the lymphocytic outer region of the granuloma, suggesting a possible role for activated CD8(+) cells in the initial attempt to restrain the granuloma growth. CD4(+) T cells appeared equally distributed in the lymphocytic mantle and in the internal areas of the granulomas. WC1(+) cells appeared interspersed among the macrophages. We speculated that this could indicate a role for these 2 subsets in the maintenance and the maturation of the granuloma. In stage III/IV lesions, all of the T-cell subsets investigated appeared interspersed among the mononuclear component of the granulomas. In general terms, there was a higher density of CD8(+) cells compared with CD4(+) cells. However, there was no sense of rimming effect for any of the investigated cell populations.     

Histological observations of bovine tuberculosis in lung and lymph node tissues from British deer

Deer are recognized as hosts of Mycobacterium bovis and assessing the role of wild cervids in perpetuating tuberculosis among cattle has motivated extensive research on several continents. In this paper, the histopathology of lymph node and lung tuberculous granulomas in M. bovis positive British deer is presented. The overall aim was to seek further insights into the potential for onward transmission from infected deer to other species, including cattle. Samples were obtained from an extensive survey of wild mammals in South-West England and from statutory tuberculosis surveillance. M. bovis culture-positive samples were characterised microscopically as to their stage of lesion advancement, number of acid-fast bacilli and granuloma encapsulation. Seventy percent of the deer developed granulomas containing far greater numbers of M. bovis bacilli than typically reported in cattle. Red and fallow deer had the largest number of poorly encapsulated granulomas often containing many hundreds of bacilli. The results are consistent with infected wild British deer being a potential source of environmental contamination and onward transmission to other species. However, further work on levels of bacillary shedding is required before this can be confirmed.     

Implications and challenges of tuberculosis in wildlife ungulates in Portugal: a molecular epidemiology perspective

Mycobacterium bovis and, more rarely, Mycobacterium caprae, may cause zoonotic bovine tuberculosis (bTB) in an extensive range of animal species. In Portugal, during 2009, a remarkable raise of bTB incidence was registered in cattle along with an increase of new cases in wildlife. In this work, we reassess and update the molecular epidemiology of bTB in wild ungulates by including 83 new M. bovis and M. caprae isolates from wild boar and red deer obtained during 2008-2009. Spoligotyping identified 27 patterns in wild ungulates, including 11 patterns exclusive from deer and five from wild boar. The genetic relatedness of wildlife and livestock isolates is confirmed. However, the relative prevalence of the predominant genotypes is different between the two groups. Contrasting with the disease in livestock, which is widespread in the territory, the isolation of bTB in wildlife is, apparently, geographically localized and genotypic similarities of strains are observed at the Iberian level.     

MHC class II-restricted, CD4(+) T-cell proliferative responses of peripheral blood mononuclear cells from Mycobacterium bovis-infected white-tailed deer

White-tailed deer are significant wildlife reservoirs of Mycobacterium bovis for cattle, predators, and, potentially, humans. Infection of cattle with M. bovis stimulates an antigen-specific T-cell response, with both CD4(+) and CD8(+) cells implicated in protective immunity. Few studies, however, have examined lymphocyte subset responses to experimental M. bovis infection of white-tailed deer. In this study, a flow cytometric proliferation assay was used to determine the relative contribution of individual peripheral blood mononuclear cell subsets of M. bovis-infected white-tailed deer in the recall response to M. bovis antigen. Naive deer were challenged with M. bovis by cohabitation with infected deer. These M. bovis-challenged deer developed significant in vivo (delayed-type hypersensitivity) and in vitro (proliferative) responses to M. bovis purified protein derivative (PPD). At necropsy, typical tuberculous lesions containing M. bovis were detected within lungs and lung-associated lymph nodes of infected deer. The predominant subset of lymphocytes that proliferated in response to in vitro stimulation with PPD was the CD4(+) subset. Minimal proliferative responses were detected from CD8(+), gamma delta TCR(+), and B-cells. Addition of monoclonal antibodies specific for MHC II antigens, but not MHC I or CD1 antigens, abrogated the proliferative response. Together, these findings indicate that while CD4(+) cells from infected deer proliferate in the recall response to M. bovis antigens, this response is not sufficient to clear M. bovis and immunologic intervention may require stimulation of alternate subsets of lymphocytes.     

Evaluation of Spoligotyping and MIRU-VNTR for Mycobacterium bovis in Xinjiang, China

Mycobacterium bovis, the causative agent of bovine tuberculosis (bTB) in cattle, is also a pathogen for human and other mammals. In this study, 406 cows were screened for bTB by both single intradermal comparative cervical tuberculin (SICCT) test and IFN-γ assay. 135 M. bovis were isolated from 31 SICCT and IFN-γ double-positive cows in Xinjiang Uygur Autonomous Region of China. Spoligotyping and MIRU-VNTR were evaluated for genotyping, and 4 and 7 genotypes were identified, respectively. A new combination of nine MIRU-VNTR loci was most discriminative for M. bovis clones from Xinjiang. Interestingly, two new spoligotypes (SB1903 and SB1904) and special repeat numbers of three loci (ETR-D, QUB 1895 and QUB 3336) were discovered in this study. These results indicated a specific epidemic conservation in Xinjiang, China. M. bovis strains with the unique genotypes were isolated from the herds maintaining parent cows imported from the bTB-free countries, suggesting a possible transmission from the local breed of Xinjiang brown cattle.     

Recent advances in the management of bovine tuberculosis in free-ranging wildlife

Established foci of Mycobacterium bovis (the causative agent of bovine tuberculosis [bTB]) in free-ranging wildlife are currently under various stages of management on three continents (Africa, Europe and North America) and in New Zealand. Other, as yet undiagnosed, foci seem likely to exist elsewhere. The complex roles that these wildlife foci play in the ecology of bTB remain among the greatest challenges facing bTB control globally. Conceptually, management of bTB in free-ranging wildlife can be thought of as progressing from the discovery of an outbreak through frequently overlapping stages of epidemiological characterization, initial control, simulation and forecasting, focused control, and verification of eradication. Surveillance in its various forms remains a critical component of assessment throughout. Since the Fourth International M. bovis Conference in 2005, research on management of bTB in free-ranging wildlife has encompassed such areas as the human dimensions of wildlife management, mitigation of bTB risks from wildlife on cattle farms, vaccine biology, and epidemiology, with a major contribution from simulation modeling. In order to advance the actual field management of bTB, however, research must be sufficiently grounded to aid development of practical, affordable and politically defensible management interventions which stand a reasonable chance of being implemented. The current management of two wildlife reservoirs of bTB, brushtail possums (Trichosurus vulpecula) in New Zealand, and white-tailed deer (Odocoileus virginianus) in Michigan, USA, serve as contrasting examples of different wildlife management strategies aimed at achieving a common goal. In New Zealand, the importance of agricultural export markets and the status of the possum as a non-native pest have facilitated direct, aggressive management of the disease reservoir, resulting in considerable progress towards bTB freedom since 1994. In Michigan, the relative importance of the hunting economy and of whitetails as a game animal have made such aggressive culling politically untenable. This has forced reliance upon publicly supported, and implemented, management tools, and so provided impetus to better understand social support for wildlife management policy, its limitations, and ways to employ it in disease control policy development.     

Progress in the control of bovine tuberculosis in Spanish wildlife

Despite the compulsory test and slaughter campaigns in cattle, bovine tuberculosis (bTB) is still present in Spain, and the role of wildlife reservoirs is increasingly recognized. We provide an update on recent progress made in bTB control in Spanish wildlife, including aspects of epidemiology, surveillance, host-pathogen interaction and wildlife vaccination. At the high densities and in the particular circumstances of Mediterranean environments, wild ungulates, mainly Eurasian wild boar and red deer, are able to maintain Mycobacterium bovis circulation even in absence of domestic livestock. Infection is widespread among wild ungulates in the south of the country, local infection prevalence being as high as 52% in wild boar and 27% in red deer. Risk factors identified include host genetic susceptibility, abundance, spatial aggregation at feeders and waterholes, scavenging, and social behaviour. An increasing trend of bTB compatible lesions was reported among wild boar and red deer inspected between 1992 and 2004 in Southwestern Spain. Sporadic cases of badger TB have been detected, further complicating the picture. Gene expression profiles were characterized in European wild boar and Iberian red deer naturally infected with M. bovis. The comparative analysis of gene expression profiles in wildlife hosts in response to infection advanced our understanding of the molecular mechanisms of infection and pathogenesis, revealed common and distinctive host responses to infection and identified candidate genes associated with resistance to bTB and for the characterization of host response to infection and vaccination. Ongoing research is producing valuable knowledge on vaccine delivery, safety and efficacy issues. Baits for the oral delivery of BCG vaccine preparations to wild boar piglets were developed and evaluated. The use of selective feeders during the summer was found to be a potentially reliable bait-deployment strategy. Safety experiments yielded no isolation of M. bovis BCG from faeces, internal organs at necropsy and the environment, even after oral delivery of very high doses. Finally, preliminary vaccination and challenge experiments suggested that a single oral BCG vaccination may protect wild boar from infection by a virulent M. bovis field strain.     

Farm and slaughter survey of bovine tuberculosis in captive deer in Switzerland

In 1998, a survey was conducted by postal questionnaire to gather basic knowledge about the management, health and productivity of captive deer in Switzerland. In addition, lymph nodes were collected from slaughtered deer from 124 of the 262 holdings surveyed, and tested for Mycobacterium bovis and Mycobacterium tuberculosis. The total farmed deer population was 8389 animals kept on 485 holdings; 87 per cent were fallow deer, 8 per cent red deer, 4 per cent sika deer, and there were small numbers of other species. The median herd sizes were 12 for fallow deer and eight for red deer. Few owners had handling facilities or crushes. In none of the lymph nodes examined were lesions typical of bovine tuberculosis observed, and neither M bovis nor M tuberculosis was cultivated from any of the samples.     

Lesion development in white-tailed deer (Odocoileus virginianus) experimentally infected with Mycobacterium bovis

The recent discovery of tuberculosis in free-living white-tailed deer in northeastern Michigan underscores the need for increased understanding of the pathogenesis of tuberculosis in wildlife species. To investigate lesion development in white-tailed deer, 32 deer were experimentally infected by intratonsilar instillation of 300 colony-forming units of Mycobacterium bovis. Three deer each were euthanatized and examined at days 15, 28, 42, and 56 after inoculation, and five deer each were euthanatized and examined at days 89, 180, 262, and 328 after inoculation. Microscopic lesions first were seen in the medial retropharyngeal lymph node and lung 28 and 42 days after inoculation, respectively. Lung lesions were present in 12 (38%) of 32 deer, involving 23 lung lobes. Left caudal and right middle and caudal lobes were involved in 17 (74%) of the 23 affected lung lobes. Lesions in the medial retropharyngeal lymph node first appeared as granulomas composed of aggregates of macrophages and Langhans-type giant cells. Some early granulomas contained centrally located neutrophils. As granulomas developed, neutrophils were replaced with a central zone of caseous necrosis that first showed signs of mineralization 42 days after inoculation. Granulomas increased in size as the zone of caseous necrosis expanded. Peripheral fibrosis, first seen at 56 days after inoculation, progressed to only a thin fibrous capsule by 328 days after inoculation. By the termination of the study, the central necrotic core of the granuloma contained abundant liquefied necrotic material and grossly resembled an abscess. Although tuberculous lesions in white-tailed deer follow a developmental pattern similar to that in cattle, fibrosis is less pronounced and the advanced lesions may liquefy, a change seldom reported in cattle. An understanding of lesion development will aid in the identification of the spectrum of disease that may be seen in this important wildlife reservoir of tuberculosis.     

Blood culture and stimulation conditions for the diagnosis of tuberculosis in cervids by the Cervigam assay

Mitogen- and antigen-induced interferon-gamma (IFN-gamma) responses of peripheral blood leucocytes from cervids were evaluated by a commercial whole-blood assay. The assay was applied to Mycobacterium bovis-infected white-tailed deer and reindeer, M bovis BCG-vaccinated white-tailed deer and elk, and unvaccinated, uninfected white-tailed deer, fallow deer, elk and reindeer. The responses of the M bovis-infected white-tailed deer to pokeweed mitogen (PWM) varied with time and between individuals. The responses of the M bovis-infected reindeer to PWM and M bovis purified protein derivative (PPD) were positively associated. Samples from tuberculosis-free captive herds in various parts of the USA were also evaluated. Four per cent of fallow deer, 20 per cent of elk, 44 per cent of white-tailed deer, and 91 per cent of reindeer had responses to PWM exceeding 0.25 Delta optical density, that is, PWM stimulation minus no stimulation. The specificity of the responses to M bovis PPD and a Mycobacterium tuberculosis complex-specific antigen rESAT-6:CFP-10, excluding animals not responding to PWM, ranged from 78 per cent to 100 per cent and was dependent upon the species and the positive response cut-off value. The results show that the commercial assay is valid for the detection of TB in reindeer; however, further development of the assay will be required before it is used in surveillance programmes for white-tailed deer, fallow deer, and elk.     

Study of peripheral blood cell populations involved in the immune response of goats naturally infected with Mycobacterium caprae

Tuberculosis in goats caused by Mycobacterium bovis and Mycobacterium caprae has noteworthy sanitary and economic implications. Current diagnostic assays are based on cellular immunity and although they have demonstrated a high sensitivity, some animals remain undetected. In the present study, flow cytometry has been used to determine changes in CD4+, CD8+ and CD25+ T cell populations in peripheral blood from naturally infected goats. Proportion of lymphocytes producing PPD-specific interferon-gamma (IFN-γ) was calculated and an ELISA for detection of PPD-specific IFN-γ was performed to measure the cytokine in plasma. The infected goats showed percentages of CD4+ T cells between 27.31% and 47.23% and there were not significant differences (p=0.113) with the non-infected control goats although the mean percentage was lower in this group. Regarding CD8+ T cells, a higher percentage was observed in healthy goats compared to controls (p=0.081). The mean percentage of lymphocytes expressing CD25 without antigen stimulation (30.65±3.91) was higher in lesion and/or culture-positive animals than in the controls (21.84±1.21; p=0.053). The percentage of CD4+/IFN+ T cell population stimulated with bovine PPD was a reliable marker of infection, since the mean percentage in the infected goats was significantly higher than in the controls (p<0.05). Tuberculosis in goats caused by M. caprae induced changes in cellular populations similar to those described for M. bovis in cattle.     

Bovine tuberculosis infection in wild mammals in the South-West region of England: a survey of prevalence and a semi-quantitative assessment of the relative risks to cattle

In the United Kingdom, badgers are implicated in the transmission of Mycobacterium bovis to cattle, but little information is available on the potential role of other wild mammals. This paper presents the results of the largest systematic UK survey of M. bovis infection in other wild mammals. Mammal carcasses (4715) from throughout the South-West region of England were subjected to a systematic post mortem examination, microbiological culture of tissues and spoligotyping of isolates. Infection was confirmed in fox, stoat, polecat, common shrew, yellow-necked mouse, wood mouse, field vole, grey squirrel, roe deer, red deer, fallow deer and muntjac. Prevalence in deer may have been underestimated because the majority were incomplete carcasses, which reduced the likelihood of detecting infection. Infected cases were found in Wiltshire, Somerset, Devon and Cornwall, Gloucestershire and Herefordshire. Lesions were found in a high proportion of spoligotype-positive fallow, red and roe deer, and a single fox, stoat and muntjac. M. bovis spoligotypes occurred in a similar frequency of occurrence to that in cattle and badgers. Data on prevalence, pathology, abundance and ecology of wild mammals was integrated in a semi-quantitative risk assessment of the likelihood of transmission to cattle relative to badgers. Although most species presented a relatively low risk, higher values and uncertainty associated with muntjac, roe, red and in particular fallow deer, suggest they require further investigation. The results suggest that deer should be considered as potential, although probably localised, sources of infection for cattle.     

An outbreak of Mycobacterium bovis infection in fallow deer (Dama dama)

In an outbreak of Mycobacterium bovis infection in fallow deer in South Australia, 3 herds related by recent movement of deer were infected. From these 3 infected herds, 47 of 51 animals were tuberculosis at necropsy. A range of lesions was seen most of which differed from classical bovine tuberculosis in that pus was a white liquid, fibrous encapsulation was not marked and calcification was rare. Histopathology was of classical tuberculosis. M. bovis was cultured from lesions and M. avium-intracellulare was cultured from one deer with no visible lesions. The source of M. bovis infection has not been determined.     

Phagolysosome maturation of macrophages was reduced by PE_PGRS 62 protein expressing in Mycobacterium smegmatis and induced in IFN-γ priming

Mycobacterium bovis parasitizes host macrophages and has developed strategies to survive within macrophages. Research on mycobacteria-specific PE_PGRS genes indicates that they code for cell surface proteins that may influence virulence. To further elucidate the molecular pathogenesis of tuberculosis and host response to M. bovis, we explored the mechanisms by which PE_PGRS62 protein increase persistence of mycobacterium within host macrophages. We found that the M. smegmatis strain expressing M. bovis PE_PGRS 62 protein reduced phagolysosome maturation in human macrophages, and significantly decreased the mRNA expression of IL-1β in a dose- and time-dependent. We identified that IFN-γ priming of macrophages immediately prior to infection with PE_PGRS62 expressing M. smegmantis, enhanced the maturation of phagolysosomes and induced IL-1β production both that the protein and mRNA levels and further activated the NF-κB pathway. Overall, we demonstrated that PE_PGRS62 protein altered the immune environment of the host cells, which suggested that the pathogenic PE_PGRS62 protein altering the immune mechanism might be involved in the pathogenesis of mycobacterial disease and hence influenced host cell responses to M. bovis infection.     

硒对树突状细胞和巨噬细胞功能的调控作用

旨在探讨硒对树突状细胞（dendritic cells,DCs）和巨噬细胞功能的影响,试验将硒分别与髓源性树突状细胞（bone marrow derived dendritic cells,BMDCs）和腹腔巨噬细胞共同培养后,用流式细胞术检测未成熟BMDCs和巨噬细胞的吞噬活性以及BMDCs上的MHCⅡ、CD86、CD80和CD40的表达量,测定经硒处理后的成熟BMDCs对刺激同种异体淋巴细胞增殖和抗原递呈能力,并用ELISA检测BMDCs和巨噬细胞上清液中细胞因子（IL-12、IL-1β、IFN-γ、IL-6、IL-10、TNF-α、NO）水平的变化。结果显示,当硒的质量浓度在0.18~0.09 mg·L^-1时,BMDCs和巨噬细胞的吞噬活性显著增强（P<0.05）,并且BMDCs上的MHCⅡ、CD86和CD80的表达量显著升高（P<0.05）,对刺激同种异体淋巴细胞的增殖和抗原递呈能力也显著增强（P<0.05）。此外,在BMDCs的上清中,IFN-γ、IL-12和IL-10的含量显著升高（P<0.05）;在巨噬细胞的上清中,IFN-γ、TNF-α和NO的含量显著升高（P<0.05）。结果表明,一定质量浓度的硒可以增强树突状细胞和腹腔巨噬细胞的功能,值得进一步探究硒对免疫功能的影响。     

Bovine tuberculosis in Europe from the perspective of an officially tuberculosis free country: trade, surveillance and diagnostics

Switzerland has been officially free of bovine tuberculosis (OTF) since 1960. Since 1980 the control of bovine tuberculosis (bTB) has been reduced to passive abattoir surveillance. Isolated cases of bTB, partly due to reactivation of human Mycobacterium bovis infections with subsequent transmission to cattle, have been noticed in the last years. In Europe, the overall prevalence of bTB is slightly increasing. Both OTF and non-OTF countries report increases in the proportion of bTB positive cattle herds. Current bTB eradication and control programs in Europe are facing a range of challenges. Whole herd depopulation is becoming a less attractive option for economic reasons and due to animal welfare concerns. Live animal trade is increasing both at national and international levels. Regarding these tendencies and taking into account the chronicity of bTB infection, pre-movement testing is becoming increasingly important as a central tool for eradication and for protection against re-introduction of bTB. Pre-movement testing, however specifically focuses on the infection status in individuals, requiring a high level of diagnostic accuracy to correctly diagnose infected animals. Current screening tests for bTB, however, have been designed to meet demands as herd tests. This illustrates that the modification of existing and/or the development of new diagnostics for bTB might be needed. The tuberculin skin test (TST), the primary screening test for bTB may in certain situations have low sensitivity. The interferon gamma (IFN-γ) assay is accepted to be more sensitive compared to TST. Reduced specificity, however, especially in areas of low bTB prevalence raises concerns. New antigen combinations including Rv3615c, OmpATb and others have been shown to complement ESAT-6 and CFP-10 in the whole blood IFN-γ assay and resulted in improved sensitivity (compared to ESAT-6 and CFP-10) and specificity (compared to tuberculins). Lesion detection after slaughter represents a cost-effective procedure for passive surveillance of bTB, especially in areas of low prevalence or in regions free of bTB; however, its sensitivity is very low. This illustrates that trade is linked with a certain risk to re-introduce bTB in OTF regions or countries and that there may be delays in detecting a re-introduction of bTB. In conclusion, regarding the fact that some parameters linked with bTB programs are changing, the development of improved diagnostic tests with a high reliability for use as individual animal tests will be important for future eradication of bTB, in line with international commitment to high standard animal health programs.     

Characterization of selected genes upregulated in non-tuberculous European wild boar as possible correlates of resistance to Mycobacterium bovis infection

Bovine tuberculosis (bTB), caused by Mycobacterium bovis (Mycobacterium tuberculosis complex), is a zoonotic disease that affects cattle and wildlife worldwide. These animal hosts can serve as reservoirs of infection, thus increasing the risk of human exposure and infection. In this study we quantified by RNA macroarray fluorescent hybridization and real-time RT-PCR the mRNA levels of genes differentially expressed in oropharyngeal tonsils and mandibular lymph nodes of three and seven individual non-tuberculous and tuberculous wild boars naturally exposed to M. bovis, respectively. These results demonstrated upregulation of two genes, complement component 3 (C3) and methylmalonyl-CoA mutase (MUT), in the non-tuberculous wild boars. These upregulated genes may contribute to resistance of wild boars to bTB by modifying the innate immunity, which limits the ability of the mycobacterium to infect and persist within macrophages. The C3 and MUT genes, therefore, are likely to be good candidates to study as markers of bTB resistance using functional genomics in animal model systems. Identification of genes upregulated in wild animals resistant to bTB contributes to our understanding of the mechanisms of protective immunity and resistance to mycobacterial organisms.     

Larval toxocarosis in sheep: the immunohistochemical characterization of lesions in some affected organs

Morphological and immunohistochemical responses of lambs following oral infection with 10,000 infective Toxocara canis (T. canis) eggs were studied up to 28 days post-infection (pi). The small intestine, liver, lungs and brain of both infected and control lambs were examined using the routine histological methods for paraffin sections and immunohistochemical techniques for frozen tissue sections. Eosinophil-rich hepatic granulomas and diffuse T. canis-induced pulmonary inflammation were the most prominent pathological features. CD2+, CD4+, CD8+, IgM bearing cells, and macrophages were demonstrated in the liver granulomas. The observed histomorphologic changes were similar to other paratenic hosts. It was concluded that larval toxocarosis followed the classical migratory path in the infected lambs.     

Pathology of naturally occurring bovine tuberculosis in England and Wales

The aim of this study was to obtain a contemporary data set of pathology in tuberculin reactor and in-contact cattle in England and Wales. Four hundred animals (200 reactors and 200 in-contacts) from 242 farms located in 14 counties in Western England and Wales were examined. The mean number of lymph nodes (LNs) with tuberculosis (TB)-like lesions per TB-confirmed animal was 1.7 in reactors and 1.5 in in-contact animals. Tuberculous lesions in both reactor and in-contact animals were most commonly observed in the LNs of the thorax, followed by the head and abdomen, particularly the mediastinal, retropharyngeal and tracheobronchial LNs. Twenty-five reactors had macroscopic lesions in the palatine tonsils. Among TB-confirmed cattle, 27% of reactors and 9% of in-contact animals had gross TB-like lesions in the lungs, particularly in the caudal lobes. Gross lesions that were not TB-confirmed were parasitic granulomas (45%), bacterial or mycotic club-forming pyogranulomas (27%) and bacterial abscesses (23%). Diagnostic sensitivity was maximised when bacteriology and histopathology were used concurrently. Stage IV granulomas, alone or in combination with other stages, constituted 63% of lesions, while 16% of lesions were stage I/II granulomas. Caseous necrosis and calcification were common features of the granulomas encountered in natural Mycobacterium bovis infections, even with pathology limited to a small number of sites. Granulomas often covered large areas of histological sections and typically contained only small numbers of acid fast bacilli.