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Context The Australian brushtail possums (Trichosurus vulpecula) introduction to New Zealand has exacted a heavy toll on native biodiversity and presented the country with its greatest wildlife reservoir host for bovine tuberculosis (TB).... more
Context The Australian brushtail possums (Trichosurus vulpecula) introduction to New Zealand has exacted a heavy toll on native biodiversity and presented the country with its greatest wildlife reservoir host for bovine tuberculosis (TB). Management efforts to control both possums and TB have been ongoing for decades, and the biology of possums has been studied extensively in Australia and New Zealand over the past 50 years; however, we still do not have a clear understanding of its home-range dynamics. Aims To investigate determinants of home range size by using a uniquely large dataset in the Orongorongo Valley, a highly monitored research area in New Zealand and compare our findings with those of other studies. Methods Possum density was estimated, for subpopulations on four 13-ha cage-trap grids, by the spatially explicit capture–mark–recapture analysis of trapping data from 10 consecutive months. Home ranges were estimated from trap locations using a 100% minimum convex polygon...
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Protection of cattle against bovine tuberculosis by vaccination could be an important control strategy in countries where there is persistence of Mycobacterium bovis infection in wildlife and in developing countries where it is not... more
Protection of cattle against bovine tuberculosis by vaccination could be an important control strategy in countries where there is persistence of Mycobacterium bovis infection in wildlife and in developing countries where it is not economical to implement a 'test and slaughter' control programme. Early field trials with Bacille Calmette Guerin (BCG) M. bovis vaccine in cattle produced disappointing results, with induction of tuberculin skin-test reactivity following vaccination and low levels of protection. However, recent studies using a low dose of BCG vaccine in cattle have produced more encouraging results and field trials should now be carried out in developing countries to determine whether this low dose BCG vaccination strategy will reduce the spread of infection. The options for new candidate tuberculosis vaccines have increased markedly in the last decade with the advent of new attenuated strains of M. bovis, and sub-unit protein and recombinant DNA vaccines. Some of these new types of vaccines have recently been tested in cattle. New attenuated M. bovis vaccines induced greater protection than BCG vaccine in cattle which had been sensitized to environmental mycobacteria prior to vaccination. In contrast, it has proved difficult to stimulate appropriate immune responses in cattle necessary for protection with sub-unit protein and recombinant DNA vaccines and better immunological adjuvants are required for these types of vaccines. Progress in the development of new tuberculosis vaccines has been very rapid in the past decade and the prospects for vaccination to control and eradicate bovine tuberculosis are encouraging.
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Oral-delivery Mycobacterium bovis bacillus Calmette-Guérin (BCG) vaccine in a lipid matrix has been shown to confer protection against M. bovis infection and reduce the severity of tuberculosis (TB) when fed to brushtail possums... more
Oral-delivery Mycobacterium bovis bacillus Calmette-Guérin (BCG) vaccine in a lipid matrix has been shown to confer protection against M. bovis infection and reduce the severity of tuberculosis (TB) when fed to brushtail possums (Trichosurus vulpecula), the major wildlife vector of bovine TB in New Zealand. Here we demonstrate the feasibility of aerial delivery of this live vaccine in bait form to an M. bovis-infected wild possum population, and subsequently assess vaccine uptake and field efficacy. Pre-trial studies indicated a resident possum population at very low density (<0.6 possums/ha) at the field site, with a 5.1% prevalence of macroscopic TB lesions. Pilot studies indicated that flavoured lipid matrix baits in weather-proof sachets could be successfully sown aerially via helicopter and were palatable to, and likely to be consumed by, a majority of wild possums under free-choice conditions. Subsequently, sachet-held lipid baits containing live BCG vaccine were sown at 3 ...
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Bovine tuberculosis (TB) continues to be an intractable problem in many countries, particularly where "test and slaughter" policies cannot be implemented or where wildlife reservoirs of infection serve as a recurrent source of... more
Bovine tuberculosis (TB) continues to be an intractable problem in many countries, particularly where "test and slaughter" policies cannot be implemented or where wildlife reservoirs of infection serve as a recurrent source of infection for domestic livestock. Alternative control measures are urgently required and vaccination is a promising option. Although the bacille Calmette-Guérin (BCG) vaccine has been used in humans for nearly a century, its use in animals has been limited, principally as protection against TB has been incomplete and vaccination may result in animals reacting in the tuberculin skin test. Valuable insights have been gained over the past 25 years to optimise protection induced by BCG vaccine in animals and in the development of tests to differentiate infected from vaccinated animals (DIVA). This review examines factors affecting the efficacy of BCG vaccine in cattle, recent field trials, use of DIVA tests and the effectiveness of BCG vaccine in other d...
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Mycobacterium avium subspecies paratuberculosis (MAP) causes Johne's disease in ruminants, which is characterized by chronic progressive granulomatous enteritis. The infection leads to wasting and weight loss in the animals and... more
Mycobacterium avium subspecies paratuberculosis (MAP) causes Johne's disease in ruminants, which is characterized by chronic progressive granulomatous enteritis. The infection leads to wasting and weight loss in the animals and eventually death, causing considerable production losses to the agricultural industry worldwide. Currently available ELISA- and PCR-based diagnostic tests have limited sensitivity and specificity during early MAP infection in cattle, suggesting that there is an urgent demand for alternative diagnostic tests. Circulating microRNA (miRNA) have recently gained attention as potential biomarkers for several diseases in humans. However, knowledge and use of miRNA as biomarkers in diseases of ruminants, including Johne's disease, are very limited. Here we used NanoString nCounter technology (NanoString, Seattle, WA), a digital platform for amplification-free and hybridization-based quantitative measurement of miRNA in the sera of noninfected and naturally MA...
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BCG vaccination sensitises cattle to bovine tuberculin which compromises the use of the current bovine TB surveillance tests. Although the performance of a blood test (that utilises antigens expressed by Mycobacterium bovis but not by... more
BCG vaccination sensitises cattle to bovine tuberculin which compromises the use of the current bovine TB surveillance tests. Although the performance of a blood test (that utilises antigens expressed by Mycobacterium bovis but not by BCG) capable of discriminating infected from vaccinated animals (DIVA interferon-gamma test, DIT) has been evaluated in naturally infected TB field reactors, there is a need to perform similar analysis in a BCG vaccinated/M. bovis infected population. Furthermore, we explored different scenarios under which a DIT may be implemented alongside BCG vaccination: (i) serial testing to resolve potential false positive skin test results; or (ii) standalone test to replace the single intradermal comparative cervical tuberculin skin test (SICCT). Our results demonstrated significantly better relative test sensitivity when the DIT was evaluated in a serial test scenario. Direct comparison of pre and post skin test blood samples revealed that the SICCT test induc...
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The gamma interferon (IFN-γ) test has been used for many years as an ancillary test in the detection of bovine tuberculosis. We investigated the effect of skin testing and the length of time between blood collection and processing on the... more
The gamma interferon (IFN-γ) test has been used for many years as an ancillary test in the detection of bovine tuberculosis. We investigated the effect of skin testing and the length of time between blood collection and processing on the performance of the IFN-γ test. A series of blood samples were taken from groups of experimentally infected cattle ( n = 10), naturally infected ( n = 11), and uninfected animals ( n = 12) that were examined with a caudal fold skin test. Blood was taken on the day of tuberculin injection, 3 d later when the skin tests were read, and 11-19 d post-tuberculin injection, and was processed for the IFN-γ test at 8, 30, and 36 h postcollection. There were significant decreases in the IFN-γ responses with increasing time between blood collection and sample processing. Significantly greater responses were observed in both the purified protein derivative (PPD) and early secretory antigenic target protein 6/culture filtrate protein 10 IFN-γ tests for samples pr...
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Tuberculosis (TB) is a disease caused by Mycobacterium tuberculosis or Mycobacterium bovis and still remains one of the world's biggest global health burdens. Recently, engineered polyhydroxyalkanoate (PHA) biobeads that were produced... more
Tuberculosis (TB) is a disease caused by Mycobacterium tuberculosis or Mycobacterium bovis and still remains one of the world's biggest global health burdens. Recently, engineered polyhydroxyalkanoate (PHA) biobeads that were produced in both Escherichia coli and Lactococcus lactis and displayed mycobacterial antigens were found to induce significant cell-mediated immune responses in mice. We observed that such PHA beads contained host cell proteins as impurities, which we hypothesized to have the potential to induce immunity. In this study, we aimed to develop PHA beads produced in mycobacteria (mycobacterial PHA biobeads [MBB]) and test their potential as a TB vaccine in a mouse model. As a model organism, nonpathogenic Mycobacterium smegmatis was engineered to produce MBB or MBB with immobilized mycobacterial antigens Ag85A and ESAT-6 on their surface (A:E-MBB). Three key enzymes involved in the poly(3-hydroxybutyric acid) pathway, namely, β-ketothiolase (PhaA), acetoacetyl-c...
Research Interests: Biology, Medicine, Multidisciplinary, Tuberculosis, Mycobacterium tuberculosis, and 12 moreEscherichia coli, Mice, Biopolymers, Female, Animals, Applied Environmental Microbiology, Vaccination, Polyhydroxyalkanoates, Polyesters, Mycobacterium bovis, Tuberculosis vaccines, and Mycobacterium smegmatis
A long-term study was undertaken to monitor immune responses, faecal cultures and clinical disease in sheep experimentally infected with Mycobacterium avium subspecies paratuberculosis (Map) strain Telford. New Zealand Merino lambs (N=56)... more
A long-term study was undertaken to monitor immune responses, faecal cultures and clinical disease in sheep experimentally infected with Mycobacterium avium subspecies paratuberculosis (Map) strain Telford. New Zealand Merino lambs (N=56) were challenged with three oral doses of Map suspension. The lambs were weighed and faecal and blood samples obtained at different time-points. At 63 weeks post-challenge, surviving sheep were euthanised and samples of liver, ileo-caecal valve and mesenteric lymph node were collected for histopathology and Map culture. High IFN-γ and antibody responses were evident as early as 8 weeks post-C1 which persisted until the end of the trial. Approximately 92% of the sheep shed Map in faeces at 36 weeks post-challenge, with the prevalence decreasing to around 40% at the end of the trial. Thirteen sheep progressively lost weight and were euthanised between weeks 32 and 58 post-challenge. Nearly 58% of surviving sheep exhibited histo-pathological lesions in...
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Methane is produced in the rumen of ruminant livestock by methanogens and is a major contributor to agricultural greenhouse gases. Vaccination against ruminal methanogens could reduce methane emissions by inducing antibodies in saliva... more
Methane is produced in the rumen of ruminant livestock by methanogens and is a major contributor to agricultural greenhouse gases. Vaccination against ruminal methanogens could reduce methane emissions by inducing antibodies in saliva which enter the rumen and impair ability of methanogens to produce methane. Presently, it is not known if vaccination can induce sufficient amounts of antibody in the saliva to target methanogen populations in the rumen and little is known about how long antibody in the rumen remains active. In the current study, sheep were vaccinated twice at a 3-week interval with a model methanogen antigen, recombinant glycosyl transferase protein (rGT2) formulated with one of four adjuvants: saponin, Montanide ISA61, a chitosan thermogel, or a lipid nanoparticle/cationic liposome adjuvant (n = 6/formulation). A control group of sheep (n = 6) was not vaccinated. The highest antigen-specific IgA and IgG responses in both saliva and serum were observed with Montanide ...
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The tuberculin skin test is the primary screening test for the diagnosis of bovine tuberculosis (TB) and use of this test has been very valuable in control of this disease in many countries. However, the test lacks specificity when cattle... more
The tuberculin skin test is the primary screening test for the diagnosis of bovine tuberculosis (TB) and use of this test has been very valuable in control of this disease in many countries. However, the test lacks specificity when cattle have been exposed to environmental mycobacteria or vaccinated with Mycobacterium bovis bacille Calmette-Guérin (BCG). Recent studies have shown that use of three or four recombinant mycobacterial proteins, including ESAT6, CFP10, Rv3615c and Rv3020c or a peptide cocktail derived from these proteins, in the skin test greatly enhanced test specificity with minimal loss in test sensitivity. These proteins are present in members of the pathogenic Mycobacterium tuberculosis complex, but absent or not expressed by the majority of environmental mycobacteria or by the BCG vaccine strain. To produce a low cost skin test reagent these proteins were displayed at high density on polyester beads by translational fusion to a polyhydroxyalkanoate (PHA) synthase w...
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Bovine tuberculosis costs New Zealand more than $80 million per year, mostly because extensive areas of the country are occupied by brushtail possums infected with Mycobacterium bovis. AgResearch has a major programme to produce new live... more
Bovine tuberculosis costs New Zealand more than $80 million per year, mostly because extensive areas of the country are occupied by brushtail possums infected with Mycobacterium bovis. AgResearch has a major programme to produce new live tuberculosis vaccines that can be delivered to possums. Primary work involved development of molecular biological methods to enable genetic manipulation of M. bovis, including the production of random and specific mutants. Many avirulent mutants of M. bovis have been produced and their vaccine efficacy has been compared to BCG in guinea pigs. Selected mutants that perform at least as well as BCG are retested in guinea pigs using an extended vaccination protocol in which animals are pre-sensitized to environmental mycobacteria to mimic natural exposure. Ten candidate vaccines that have induced good protection in guinea pigs have been subsequently tested as vaccines in possums. While the protective efficacy of an M. bovis mutant inoculated into guinea pigs reliably indicated that some protection would be induced in possums, the most protective mutant in guinea pigs was different from that in possums. This illustrates the importance of testing in the target species as part of new vaccine development. An important outcome of this work was the identification of an operon in M. bovis whose inactivation produced an avirulent M. bovis vaccine candidate that was better than BCG in protecting possums from experimental tuberculosis. Allelic exchange methods are now being used to produce vaccine strains with multiple specific mutations to improve safety and immunological characteristics.
Research Interests: Genetics, Microbiology, Chemistry, Veterinary Microbiology, Biology, and 15 moreMedicine, Veterinary, Tuberculosis, Mutation, Animals, New Zealand, Bovine Tuberculosis, Vaccination, Cattle, Guinea Pig, Veterinary Sciences, Mycobacterium bovis, Mycobacterium avium, Tuberculosis vaccines, and Guinea pigs
Methane emissions from livestock are a significant contributor to greenhouse gas emissions and have become a focus of research activities, especially in countries where agriculture is a major economic sector. Understanding the complexity... more
Methane emissions from livestock are a significant contributor to greenhouse gas emissions and have become a focus of research activities, especially in countries where agriculture is a major economic sector. Understanding the complexity of the rumen microbiota, including methane-producing Archaea, is in its infancy. There are currently no robust, reproducible and economically viable methods for reducing methane emissions from ruminants grazing on pasture and novel innovative strategies to diminish methane output from livestock are required. In this review, current approaches towards mitigation of methane in pastoral farming are summarised. Research strategies based on vaccination, enzyme inhibitors, phage, homoacetogens, defaunation, feed supplements, and animal selection are reviewed. Many approaches are currently being investigated, and it is likely that more than one strategy will be required to enable pastoral farming to lower its emissions of methane significantly. Different strategies may be suitable for different farming practices and systems.
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To develop an understanding of the immune responses of ruminants to methanogens, and to provide proof of a concept that harnessing the immune system of ruminants is a potentially viable approach to mitigate greenhouse gas emissions from... more
To develop an understanding of the immune responses of ruminants to methanogens, and to provide proof of a concept that harnessing the immune system of ruminants is a potentially viable approach to mitigate greenhouse gas emissions from agriculture. Four subcellular fractions, namely cytoplasmic, two cell-wall preparations, and cell wall-derived proteins were prepared from Methanobrevibacter ruminantium M1. Twenty sheep (10 months of age) were vaccinated with these fractions or with whole cells (n=4 per group). Sheep were re-vaccinated once after 3 weeks, and antibody responses to M. ruminantium M1 antigens in sera and saliva measured using ELISA at 2 weeks after the second vaccination. Antigens recognised by the antisera were visualised using Western blotting. The antisera were tested in vitro for their impact on M. ruminantium M1, measuring the effect on cell growth, methane production, and ability to induce agglutination. Basal levels (pre-vaccination) of antibodies against M. ruminantium M1 antigens were low. Vaccination with the antigenic fractions induced strong antibody responses in serum. Both IgG and IgA responses to methanogen antigens were detected in saliva following vaccination. Western blot analysis of the antisera indicated reactivity of antibodies, and a wide range of proteins was present in the different methanogen fractions. Antisera against the various fractions agglutinated methanogens in an in-vitro assay. In addition, these antisera decreased the growth of a pure culture of a methanogen and production of methane in vitro. Antigens from methanogens are immunogenic in ruminants, and antisera from sheep vaccinated with fractions of methanogens have a significant impact on these organisms, inducing cell agglutination, and decreasing growth of methanogens and production of methane. Only antisera to selected methanogen fractions were able to achieve these effects. The results demonstrate the feasibility of a vaccination strategy to mitigate emission of methane.
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The whole blood interferon-γ (IFN-γ) test has proven to be a practical ancillary test for re-testing cattle for bovine tuberculosis 8–28 days following tuberculin skin testing. An improvement in the specificity of the IFN-γ test could... more
The whole blood interferon-γ (IFN-γ) test has proven to be a practical ancillary test for re-testing cattle for bovine tuberculosis 8–28 days following tuberculin skin testing. An improvement in the specificity of the IFN-γ test could further reduce culling of false positive animals. The primary aim of this study was to evaluate a single mycobacterial antigen, ESAT-6 in the IFN-γ test for use in skin test-positive cattle. These skin test-positive cattle comprised 51 Mycobacterium bovis-infected animals from tuberculosis-infected herds and 85 non-infected animals from tuberculosis-free herds. The test based on ESAT-6 had a higher specificity than the test based on purified protein derivative (PPD) tuberculin, but this was offset by a small decrease in sensitivity. Use of a lower cut-off in the ESAT-6-based test improved the sensitivity, while still maintaining a very high specificity. A secondary aim in the study was to assess the ESAT-6 and PPD-based tests for detecting bovine tuberculosis in skin test-negative animals from a persistently infected herd. The PPD-based test detected the majority of the lesioned or M. bovis-culture positive animals, while the ESAT-6-based test detected a smaller proportion. The false negatives in the IFN-γ test from both the skin test-negative and positive groups were predominantly M. bovis-culture positive animals with no visible lesions. The current study has shown that a defined specific antigen such as ESAT-6 can markedly improve the specificity of the IFN-γ test for re-testing skin test-positive animals. An ESAT-6-based IFN-γ test could be particularly useful to reduce the false positive rate, yet still maintain an acceptable level of sensitivity.
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Setting: Bovine tuberculosis is a problem in a number of countries and protection of cattle by vaccination could be an important control strategy.Objectives: To determine the ability of DNA vaccines, which express the mycobacterial... more
Setting: Bovine tuberculosis is a problem in a number of countries and protection of cattle by vaccination could be an important control strategy.Objectives: To determine the ability of DNA vaccines, which express the mycobacterial antigens MPB83 and MPB70 and a DNA prime-protein boost strategy to stimulate immune responses in cattle and protect against bovine tuberculosis.Design: Groups of cattle (n=10) were vaccinated with MPB83 DNA, MPB70 DNA, or MPB70 DNA followed by MPB70 protein or injected with BCG or control plasmid DNA. Animals were challenged intratracheally with virulent Mycobacterium bovis at 13 weeks and protection assessed 17 weeks later at postmortem.Results: In contrast to the strong cellular immune responses induced by BCG, the DNA vaccines induced minimal interferon-gamma (IFN-γ) and interleukin-2 (IL-2) responses. Cattle primed with MPB70 DNA and boosted with MPB70 protein induced a strong antibody response and a weak IFN-γ response. BCG gave significant reduction in four pathological parameters of disease while the DNA vaccines and MPB70 DNA/protein did not protect animals against challenge with M. bovis. Moreover, cattle vaccinated with MPB70 DNA/protein had a significantly higher proportion of animals with severe lung lesions (>100 lesions) than the MPB70 DNA alone or the control group. Increased bovine PPD-specific IL-4 mRNA expression in cattle, post-challenge, correlated with the presence of tuberculous lung lesions.Conclusion: Vaccination of calves with MPB70 or MPB83 DNA vaccines or with a more immunogenic MPB70 DNA prime-protein boost strategy did not induce protection against bovine tuberculosis.
Research Interests: Immune response, Membrane Proteins, Tuberculosis, Animals, Bovine Tuberculosis, and 13 moreDNA vaccines, Cattle, T lymphocytes, mRna expression levels, Control Strategy, DNA vaccine, Pulmonary Tuberculosis, Mycobacterium bovis, Plasmid DNA, Control Group, Interleukin, Interferon gamma, and BCG vaccine
Vaccination against bovine tuberculosis is likely to become an important disease control strategy in developing countries, which cannot afford a test and slaughter control programme, or in countries which have a wildlife reservoir of... more
Vaccination against bovine tuberculosis is likely to become an important disease control strategy in developing countries, which cannot afford a test and slaughter control programme, or in countries which have a wildlife reservoir of Mycobacterium bovis infection. In the past decade, considerable progress has been made in the development and evaluation of tuberculosis vaccines for cattle and for a range of wildlife maintenance hosts including possums, badgers, deer and African buffaloes. Experimental challenge systems have been established for the different target species and the resulting disease process has mimicked that seen in the field. In cattle, neonatal vaccination with BCG appeared to be more effective than vaccination of 6-month-old calves and in most situations no other vaccine has been shown to be better than BCG. However, prime-boost strategies involving combinations of BCG with a protein or DNA vaccine, to improve on BCG vaccination alone, have produced very encouraging results. Differential diagnostic tests have been developed using mycobacterial antigens that are only present in virulent M. bovis to differentiate between BCG-vaccinated and M. bovis-infected cattle. BCG vaccine has been shown to reduce the spread of tuberculous lesions in a range of wildlife species and a prototype oral bait delivery system has been developed. Prospects for the development of improved vaccines against bovine tuberculosis are promising and vaccination approaches could become very valuable in the control and eradication of bovine tuberculosis.
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Bovine tuberculosis remains a major economic and animal welfare concern worldwide. As part of control strategies, cattle vaccination is being considered. This approach, used alongside conventional control policies, also requires the... more
Bovine tuberculosis remains a major economic and animal welfare concern worldwide. As part of control strategies, cattle vaccination is being considered. This approach, used alongside conventional control policies, also requires the development of vaccine compatible diagnostic assays to distinguish infected from vaccinated animals (DIVA). In this review we discuss recent advances in DIVA development based on the detection of host cellular immune responses by blood testing or skin testing approaches.
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Bovine tuberculosis remains a major economic and animal welfare concern worldwide. Cattle vaccination is being considered as part of control strategies. This approach, used alongside conventional control policies, also requires the... more
Bovine tuberculosis remains a major economic and animal welfare concern worldwide. Cattle vaccination is being considered as part of control strategies. This approach, used alongside conventional control policies, also requires the development of vaccine-compatible diagnostic assays to distinguish vaccinated from infected animals (DIVA). We discuss progress made on optimizing the only potentially available vaccine, bacille Calmette Guérin (BCG), and on strategies to improve BCG efficacy. We also describe recent advances in DIVA development based on the detection of host cellular immune responses by blood-testing or skin-testing approaches. Finally, to accelerate vaccine development, definition of host biomarkers that provide meaningful stage-gating criteria to select vaccine candidates for further testing is highly desirable. Some progress has also been made in this area of research, and we summarize studies that defined either markers predicting vaccine success or markers that corr...
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Research Interests: Immunology, Immune response, Australia, Gene expression, Sequence Analysis, and 17 morePhylogeny, Cell line, Humans, Escherichia coli, Animals, Polymerase Chain Reaction, Immunology & Cell Biology, Molecular cloning, Cytotoxic Activity, Tumor necrosis factor-alpha, Amino Acid Profile, Amino Acid Sequence, Base Sequence, Opossums, Fusion Protein, Biochemistry and cell biology, and Alveolar Macrophage
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In order to limit the number and impact of exotic pest invasions, leading-edge technologies must be embraced and embedded within integrated national and international biosecurity systems. Outlined here are recent advances in the detection... more
In order to limit the number and impact of exotic pest invasions, leading-edge technologies must be embraced and embedded within integrated national and international biosecurity systems. Outlined here are recent advances in the detection of exotic pests, and prospects for the early recognition of disease. Applications of new tools are described, using our understanding of the genomes of pathogens and vectors. In addition, the role of mathematical and simulation models to aid both biosecurity planning, and decision making in the face of an epidemic, are discussed, and recent attempts to unify epidemiology and evolutionary dynamics are outlined. Given the importance of emerging diseases and zoonoses, the need to align human and veterinary surveillance within fully integrated systems is underlined.
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Alveolar macrophages (AM) form the first line of defence against most respiratory pathogens and, unlike tissue macrophages, are constantly exposed to a wide variety of antigenic stimuli. In this study we investigated the in vitro effects... more
Alveolar macrophages (AM) form the first line of defence against most respiratory pathogens and, unlike tissue macrophages, are constantly exposed to a wide variety of antigenic stimuli. In this study we investigated the in vitro effects of IFN-gamma and LPS on growth of virulent Mycobacterium bovis and M. bovis bacille Calmette-Guerin (BCG) in bovine AM. Bovine AM were purified from bronchial lavage fluid and cultured in serum-free medium. Pretreatment of bovine AM with IFN-gamma resulted in growth inhibition of M. bovis BCG but only partially inhibited growth of virulent M.bovis. Enhanced inhibition of virulent M.bovis by bovine AM required sequential stimulation with IFN-gamma and LPS and was associated with increased induction of nitric oxide (NO) and IL-12 mRNa. Growth inhibition of M. bovis was not affected by treatment of macrophages with the L-arginine analogue, NG-monomethyl-L-arginine although this treatment decreased NO production. These results suggest that a second activation signal in the form of TNF-alpha or LPS may be required to induce bacteriostasis of virulent M. bovis by bovine AM in vivo. The ability of bovine AM to respond to activation stimuli in vitro suggests that these cells may play an important role in preventing establishment of intracellular bacterial infections in the lung.
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Animal models indicate that exposure to environmental strains of mycobacteria can modulate immune responses and influence the effectiveness of live mycobacterial vaccines. Here, we describe that between the two recently reported... more
Animal models indicate that exposure to environmental strains of mycobacteria can modulate immune responses and influence the effectiveness of live mycobacterial vaccines. Here, we describe that between the two recently reported Mycobacterium avium isolates, strain WAg 206 (but not strain WAg 207) interferes with human monocyte-derived dendritic cell (MDDC) activation. WAg 206, unlike WAg 207, did not elicit inflammatory cytokine production (TNFalpha, IL-1beta, IL-12) or costimulatory molecule expression (HLA-DR, CD83, CD80, CD86) by human MDDCs in vitro. These data highlight the potential for environmental mycobacteria to modulate immune responses in humans, and suggest a mechanism by which earlier exposure to such microbes may compromise the efficacy of live mycobacterial vaccines, as has been observed in some human BCG vaccine trials.
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A population of wild brushtail possums (Trichosurus vulpecula) in which bovine tuberculosis was endemic was vaccinated with live bacille Calmette-Guérin (BCG) to determine the efficacy of vaccination. The population on the 56 hectare site... more
A population of wild brushtail possums (Trichosurus vulpecula) in which bovine tuberculosis was endemic was vaccinated with live bacille Calmette-Guérin (BCG) to determine the efficacy of vaccination. The population on the 56 hectare site was monitored bimonthly over 2 years using a capture-release regime. During the study tuberculosis was diagnosed by clinical and post mortem examination. Possums were vaccinated with BCG by both intranasal aerosol and conjunctival instillation. Possums were revaccinated on average every 5 months. Over the 2 years, 300 possums were recruited to the study with 149 being allocated to the vaccination group. There were significantly fewer cases of tuberculosis in the vaccinated (4 cases) than in the unvaccinated group (13 cases; P=0.023). The vaccine efficacy was 69%. An attempt was made to increase the incidence of disease by releasing onto the site possums that had been experimentally infected with a strain of M. bovis unknown in the area. However, this did not result in any additional cases. BCG vaccine was shown to have a level of efficacy which could be of assistance in controlling tuberculosis in wild possum populations. The future use of vaccination for the control of tuberculosis in wild possum populations is discussed.
Research Interests: Microbiology, Zoology, Sheep, Female, Animals, and 6 moreMale, Feces, SHEEP DISEASES, For, Veterinary Sciences, and Eosinophilia
A study was undertaken to indicate the importance of different causes of death in goats and to investigate the management factors which influence these problems. Over a 15 month period, 324 dead goats were received from 67 farms in the... more
A study was undertaken to indicate the importance of different causes of death in goats and to investigate the management factors which influence these problems. Over a 15 month period, 324 dead goats were received from 67 farms in the Horowhenua, Wairarapa, Wanganui and Wellington regions. Although a wide range of diseases was encountered in the study, the major causes of mortality could be divided into 4 groups: problems directly related to management, microbial diseases, nematode parasitism, and trace element related deficiencies and toxicities. The highest proportion of deaths related directly to management problems and included deaths from hypothermia, mismothering, premature birth, ruminal acidosis, pregnancy toxaemia, trauma, and plant and chemical toxicities. In larger flocks, microbial diseases including Pasteurella pneumonia and yersiniosis were major problems. Deaths from nematode parasitism were predominantly observed in goats 12 months of age and older. White muscle disease (selenium/vitamin E deficiency) was the major trace element deficiency causing death in goats. The influence of factors including age of goat, flock size and management practices on the major causes of death are discussed.
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Groups of six male goats were inoculated intratracheally and intranasally with either caprine herpesvirus followed 6 days later by Pasteurella haemolytica, canine herpesvirus alone or P. haemolytica alone. Pneumonic lesions were observed... more
Groups of six male goats were inoculated intratracheally and intranasally with either caprine herpesvirus followed 6 days later by Pasteurella haemolytica, canine herpesvirus alone or P. haemolytica alone. Pneumonic lesions were observed in five of the six goats inoculated with caprine herpesvirus followed by P. haemolytica and in three of the six goats inoculated with P. haemolytica alone, but were not observed in goats inoculated with caprine herpesvirus alone or in non-infected controls. Pasteurella haemolytica was isolated from seven of eight lungs with pneumonia, but only from one of sixteen lungs without pneumonia. The lesions ranged from fatal acute exudative necrotising pneumonia to predominantly proliferative pneumonia. Half of the caprine herpesvirus-inoculated goats developed a clinical catarrhal rhinitis five days post-inoculation and the only virus-specific histopathological lesion was a mild tracheitis. Canine herpesvirus was recovered from the nasal swabs of all caprine herpesvirus- inoculated goats developed a clinical catarrhal rhinitis five days post-inoculation and the only virus-specific histopathological lesion was a mild tracheitis. Canine herpesvirus was recovered from the nasal swabs of all canine herpesvirus-inoculated goats and from the lungs of three goats inoculated with caprine herpesvirus alone. The experimental inoculations demonstrated that P. haemolytica alone can produce pneumonia in goats. In addition, the study showed that caprine herpesvirus readily proliferates in the upper respiratory tract and lungs of goats but the role of caprine herpesvirus in the aetiology of pneumonia remains uncertain.
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Cross-protection experiments were undertaken to investigate reasons for contagious ecthyma (CE) virus-vaccination failures. Vaccination with sheep-passaged or with cell culture-passaged virus did not protect lambs against development of... more
Cross-protection experiments were undertaken to investigate reasons for contagious ecthyma (CE) virus-vaccination failures. Vaccination with sheep-passaged or with cell culture-passaged virus did not protect lambs against development of lesions after challenge inoculation with sheep-passaged virus. However, lesions which developed after challenge exposure with sheep-passaged CE virus healed significantly faster than did those induced by the initial vaccination with sheep-passaged virus (P less than 0.001). A significant decrease in the healing time was not observed for lambs initially vaccinated with cell culture-passaged CE virus after challenge exposure with sheep-passaged virus (P greater than 0.05). Protection was evident when lambs were challenge inoculated with the less virulent cell culture-passaged virus and cross-protection between the ST and CSL isolates was detected. This study indicated that complete protection even against homologous strain challenge was not achieved. Antigenic differences between vaccinal and field strains appeared an unlikely cause of vaccination failures. It also is evident that cell culture-propagated CE virus preparations are less effective for vaccination purposes than are those propagated in sheep.
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Research Interests:
Results of cross-neutralization tests of 4 isolates of contagious ecthyma (CE) virus and their antisera indicated that the isolates were neutralized to various degrees by the CE virus antisera. Cross-reactions among isolates were... more
Results of cross-neutralization tests of 4 isolates of contagious ecthyma (CE) virus and their antisera indicated that the isolates were neutralized to various degrees by the CE virus antisera. Cross-reactions among isolates were unilateral, but not bilateral, and therefore grouping of CE virus isolates by neutralization tests was not possible to obtain. Structural analysis of polypeptides from 11 isolates of CE virus by sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicated that the profiles obtained were similar to one another, except for differences observed in the molecular weight region of 37,000 to 44,000. On the basis of these differences, the isolates could be classified into 4 groups. The polypeptides which varied among the different isolates were shown to be in the surface component of the virion and appeared to be components of the surface tubules. Unilateral cross-reactions detected in cross-neutralization tests were found to correlate with the 37,000 to 44,000 dalton polypeptide grouping of the isolates.
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An effective oral bacille Calmette-Guérin (BCG) vaccine would have advantages for use in humans and as an oral bait vaccine for protecting wild-life against bovine tuberculosis. To compare the level of protection against tuberculosis in... more
An effective oral bacille Calmette-Guérin (BCG) vaccine would have advantages for use in humans and as an oral bait vaccine for protecting wild-life against bovine tuberculosis. To compare the level of protection against tuberculosis in intraduodenally BCG-vaccinated possums with those vaccinated intragastrically in order to determine whether degradation of BCG in the stomach lowers vaccine efficacy. Three groups of five possums were vaccinated with BCG by the intraduodenal, intragastric or subcutaneous routes, with a fourth group serving as unvaccinated controls. The animals were later challenged intratracheally with a low dose of virulent Mycobacterium bovis. Possums vaccinated intraduodenally with BCG had significantly greater lymphocyte blastogenic responses to bovine purified protein derivative (PPD) and lower lung bacterial counts in comparison with intragastrically vaccinated animals. In comparison with unvaccinated animals, all of the BCG-vaccinated groups had significant protection against M. bovis infection as assessed by changes in body weight, lung weight and reduction in numbers of mycobacteria and granulomas in the spleen. The enhanced immune responses and protection against bovine tuberculosis observed in the intraduodenally BCG-vaccinated possums indicated that if BCG vaccine is protected from degradation in the stomach its efficacy should improve.
Research Interests: Tuberculosis, Humans, Child, Animals, Cattle, and 4 moreOpossums, Mycobacterium bovis, Duodenum, and BCG vaccine
Research Interests:
The Mycobacterium tuberculosis complex includes Mycobacterium bovis, which causes tuberculosis in most mammals, including humans. In previous work, it was shown that M. bovis ATCC 35721 has a mutation in its principal sigma factor gene,... more
The Mycobacterium tuberculosis complex includes Mycobacterium bovis, which causes tuberculosis in most mammals, including humans. In previous work, it was shown that M. bovis ATCC 35721 has a mutation in its principal sigma factor gene, sigA, causing a single amino acid change affecting binding of SigA with the accessory transcription factor WhiB3. ATCC 35721 is avirulent when inoculated subcutaneously into guinea pigs but can be restored to virulence by integration of wild-type sigA to produce M. bovis WAg320. Subsequently, it was surprising to discover that WAg320 was not virulent when inoculated intratracheally into the Australian brushtail possum (Trichosurus vulpecula), a marsupial that is normally very susceptible to infection with M. bovis. In this study, an in vivo complementation approach was used with ATCC 35721 to produce M. bovis WAg322, which was virulent in possums, and to identify the virulence-restoring gene, phoT. There are two point deletions in the phoT gene of AT...