Biomedical subjects

D N Wedlock

Publications and source records attributed to D N Wedlock.

At least 19 recordsLinked to original sources

PubMed2006

Differences of gene expression in bovine alveolar macrophages infected with virulent and attenuated isogenic strains of Mycobacterium bovis.

Infection with Mycobacterium bovis is a significant human and animal health problem in many parts of the world. The first stage of pulmonary tuberculosis occurs after inhalation of the bacilli into an alveolus where they are ingested by resident macrophages. DNA microarray analysis was used to detect genes expressed in bovine lung alveolar macrophages infected with two isogenic strains of M. bovis, a virulent strain, ATCC35723 and an attenuated strain, WAg520 derived from ATCC35723. Chemokines, interleukin-8 and monocyte chemotactic protein 1, were more strongly expressed in ATCC35723-infected macrophages compared to WAg520-infected macrophages. Conversely, a group of genes, including fibrinogen-like protein 2 and legumain, were expressed at a higher level in macrophages infected with WAg520 compared to ATCC35723. Quantitative real-time PCR of a selected group of these differentially expressed genes confirmed enhanced levels of IL-8 mRNA in ATCC35723-infected macrophages compared to WAg520-infected macrophages. Microarray analysis of gene expression in macrophages infected with attenuated isogenic strains of M. bovis may identify key genes involved in early and protective immune responses to tuberculosis.

Animals↗

PubMed2005

Progress in the development of tuberculosis vaccines for cattle and wildlife.

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.

Animals↗

PubMed2005

Vaccination of cattle with Mycobacterium bovis culture filtrate proteins and CpG oligodeoxynucleotides induces protection against bovine tuberculosis.

Culture filtrate protein (CFP) vaccines have been shown to be effective in small animal models for protecting against tuberculosis while immunisation with these types of vaccines in cattle has been less successful. A study was conducted in cattle to evaluate the ability of selected adjuvants and immunomodulators to stimulate protective immune responses to tuberculosis in animals vaccinated with Mycobacterium bovis CFP. Seven groups of cattle (n=5) were vaccinated with M. bovis CFP formulated with either Emulsigen or Polygen adjuvant alone or in combination with a specific oligodeoxynucleotides (ODN), polyinosinic acid: polycytidylic acid (poly I:C) or poly I:C and recombinant granulocyte-macrophage colony stimulating factor. Two additional groups were vaccinated subcutaneously with BCG or non-vaccinated. In contrast to the strong interferon-gamma (IFN-gamma) responses induced by BCG, the CFP vaccines induced strong antibody responses but weak IFN-gamma responses. The addition of CpG ODN to CFP significantly enhanced cell-mediated responses and elevated antibody responses to mycobacterial antigens. Of the CFP vaccinated groups, the strongest IFN-gamma responses to CFP vaccines were measured in animals vaccinated with CFP/Emulsigen+CpG or CFP/Polygen+CpG. The animals in these two groups, together with those in the BCG and non-vaccinated groups were challenged intratracheally with virulent M. bovis at 13 weeks after the first vaccination and protection was assessed, by examination for presence of tuberculous lesions in the lungs and lymph nodes, 13 weeks later at postmortem. While BCG gave the best overall protection against tuberculosis, significant protection was also seen in animals vaccinated with CFP/Emulsigen+CpG. These results establish an important role for CpG ODN in stimulating protective Th1 responses to tuberculosis in cattle and indicate that a sub-unit protein vaccine can protect these animals against tuberculosis.

Adjuvants, Immunologic↗

PubMed2005

Oral vaccination of brushtail possums (Tichosurus vulpecula) with BCG: immune responses, persistence of BCG in lymphoid organs and excretion in faeces.

AIMS: To determine immune responses, and the localisation and persistence of Mycobacterium bovis bacille Calmette-Guérin (BCG) in gut-associated lymphoid tissues (GALT) and other organs in possums vaccinated orally with lipid-formulated BCG vaccine. To determine the duration of excretion and longevity of survival of BCG in the faeces of vaccinated animals. METHODS: Possums (n=28) were vaccinated with lipid-formulated BCG (1 x 10(8) colony forming units (cfu) of formulated BCG) by the oral route. Control possums (n=17) were fed oral bait pellets containing formulation medium only. Possums were sacrificed at 3 days and at 1, 3, 6 and 8 weeks after vaccination or ingestion of bait. Proliferation responses to bovine purified protein derivative (PPD) were measured in lymphocytes from blood and mesenteric lymph nodes (MLN) and samples of lung, spleen, liver, MLN and Peyer's patches (PP) were cultured for the presence of BCG. The number of BCG organisms excreted in faeces and the duration of excretion were determined in eight vaccinated possums and eight control possums over a 3-week period. In a separate experiment, a further six possums were vaccinated with oral BCG vaccine (5-10 x 10(8) cfu BCG/possum) and their faeces collected over 48-72 h, for culture of BCG. The longevity of survival of BCG in these faeces was determined by storing faecal samples (n=12) under three different conditions: in an incubator (22.5 degrees C), and conditions which simulated the forest floor and open pasture. A proportion (1-2 g) of these faecal samples was collected after storage for 1, 3, 5, 8 or 20 weeks, and cultured for BCG. RESULTS: Possums vaccinated orally with BCG vaccine showed strong proliferation responses to bovine PPD in peripheral blood lymphocytes at 6-8 weeks post-vaccination (p.v.). Positive lymphocyte proliferation assay (LPA) responses to bovine PPD were first evident in MLN at 3 weeks p.v. BCG was cultured from MLN and PP in a proportion of animals at 3-8 weeks p.v. BCG was not cultured from sections of spleen, lung or liver at any time p.v. BCG was recovered in low to moderate numbers from the faeces of vaccinated possums for up to 7 days, and maximal numbers were cultured in faeces collected 48-72 h p.v. After storage for 1 week, BCG was cultured from all faecal samples placed in the incubator and from a proportion of faeces exposed to conditions similar to those on the forest floor and pasture. With the exception of one faecal sample stored under forest floor conditions which was culture-positive for BCG at 3 and 5 weeks, BCG was not cultured from any other faecal sample stored for more than 1 week. CONCLUSIONS: Ingestion of oral BCG vaccine by possums was associated with the development of strong cell-mediated immunity in both blood and MLN. Following oral vaccination with BCG, the organisms were localised and persisted in GALT but did not spread to the spleen, liver or lungs. BCG was shed in low to moderate numbers in the faeces for up to 7 days p.v. The viability of BCG excreted in faeces decreased rapidly, particularly when faeces were exposed to an open pasture environment. Oral vaccination of possums with formulated BCG is unlikely to result in undue contamination of the environment with BCG.

Adjuvants, Immunologic↗

PubMed2003

Vaccination with DNA vaccines encoding MPB70 or MPB83 or a MPB70 DNA prime-protein boost does not protect cattle against bovine tuberculosis.

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-gamma) 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-gamma 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.

Animals↗

PubMed2003

Revaccination of neonatal calves with Mycobacterium bovis BCG reduces the level of protection against bovine tuberculosis induced by a single vaccination.

Cattle may provide a suitable model for testing ways of improving tuberculosis vaccine efficacy in human infants. A vaccination and challenge study was undertaken in calves to determine the optimal time to vaccinate neonatal animals with Mycobacterium bovis bacillus Calmette-Guérin (BCG) for protection against tuberculosis and to determine whether revaccination with BCG was beneficial. Calves (10 per group) were vaccinated with BCG within 8 h of birth or at 6 weeks of age, when immune responses to antigens of environmental mycobacteria were detectable, or vaccinated at birth and revaccinated at 6 weeks. A control group was not vaccinated. BCG vaccination at birth induced strong antigen-specific gamma interferon (IFN-gamma) and interleukin-2 (IL-2) responses and antigen-specific activation in CD4(+), CD8(+), and WC1(+) gammadelta T-cell subsets from blood. The proportions of animals per group with macroscopic tuberculous lesions after challenge were 0/10 for BCG at birth, 1/9 for BCG at 6 weeks, 4/10 for the revaccinated group, and 10/10 for the nonvaccinated group. There was no significant difference in the levels of protection between groups vaccinated at birth or at 6 weeks, while animals vaccinated both at birth and at 6 weeks had significantly less protection than those vaccinated only at birth. The revaccinated calves that subsequently developed tuberculous lesions had significantly stronger IFN-gamma and IL-2 responses to bovine purified protein derivative after the BCG booster than those in the same group that did not develop lesions. The results indicated that BCG vaccination at birth induced a high level of immunity and that the sensitization of very young animals to antigens of environmental mycobacteria by 6 weeks of age did not affect the effectiveness of BCG. However, BCG revaccination of these young animals was contraindicated.

Animals↗

PubMed2002

New generation vaccines and delivery systems for control of bovine tuberculosis in cattle and wildlife.

Advances in the understanding of protective immune responses to tuberculosis are providing opportunities for the rational development of improved vaccines for bovine tuberculosis. Protection requires activation of macrophages through stimulation of a Th 1 type immune response. Ideally, a vaccine for cattle should induce protection without causing animals to react in a tuberculin test when exposed to Mycobacterium bovis. A number of new tuberculosis vaccines including attenuated M. bovis strains, killed mycobacteria, protein and DNA vaccines have been developed and many of these are being assessed in cattle. The requirements for a tuberculosis vaccine for wildlife differ from those for cattle. The major goal of a wildlife vaccine is to prevent the transmission of M. bovis to cattle and other wildlife. Although there are a number of technical problems associated with the development of a vaccine delivery system for wildlife, attenuated M. bovis vaccines administered via oral baits or aerosol spray to possums have already been shown to reduce the severity of a subsequent M. bovis infection.

Animals↗

PubMed2002

Effect of different adjuvants on the immune responses of cattle vaccinated with Mycobacterium tuberculosis culture filtrate proteins.

The development of improved vaccines for bovine tuberculosis is urgently required as a cost effective solution for control and eventual eradication of tuberculosis in domestic animals. Studies in small animal models of tuberculosis have shown that vaccination with culture filtrate proteins (CFP), prepared from Mycobacterium tuberculosis or M. bovis, can induce cellular immune responses and confer a level of protection against aerogenic challenge with virulent mycobacteria. As a first step in the development of a mycobacterial CFP vaccine for protection of cattle against bovine tuberculosis, the immune responses of cattle vaccinated with short-term culture filtrate proteins (ST-CFP) from M. tuberculosis and formulated with different adjuvants were compared with those vaccinated with bacille Calmette-Guerin (BCG). The adjuvants included dimethyldioctyldecyl ammonium bromide (DDA), diethylaminoethyl (DEAE)-dextran, and ST-CFP adsorbed onto polystyrene beads. Vaccination with ST-CFP/DEAE-dextran induced high levels of interleukin-2 (IL-2) but low levels of interferon-gamma (IFN-gamma) from whole-blood cultures stimulated with M. tuberculosis ST-CFP in comparison with the strong IFN-gamma and IL-2 responses induced after vaccination with BCG. ST-CFP/DEAE-dextran also induced a strong antigen-specific immunoglobulin antibody response with both immunoglobulin G1 (IgG1) and IgG2 isotypes. Vaccination with ST-CFP/beads induced a weak IgG1-biased antibody response but no IFN-gamma or IL-2 response. DDA did not induce significant immune responses in animals vaccinated with ST-CFP. In comparison to the moderate delayed-type hypersensitivity (DTH) responses induced by vaccination with subcutaneous BCG, none of the ST-CFP vaccines induced a significant DTH response to either M. tuberculosis ST-CFP or bovine purified protein derivative (PPD). While the ST-CFP vaccines used in this study have not induced strong antigen-specific cellular immune responses in cattle comparable to those induced by BCG, they are immunogenic in cattle and it may be possible to overcome this problem by using adjuvants that more effectively promote IFN-gamma responses in this species.

Adjuvants, Immunologic↗

PubMed2001

In vitro control of Mycobacterium bovis by macrophages.

Mycobacterium bovis is frequently seen inside macrophages in vivo. The outcome of M. bovis infection depends on T cell interactions with macrophages, however mycobacteria are thought to be relatively resistant to macrophage killing. Little is known about the immunological mechanisms which control intracellular growth of M. bovis, and in the absence of T cell help the organism is capable of intracellular survival and replication. We have investigated the role of macrophages in controlling growth of virulent M. bovis or M. bovis BCG in vitro. At a multiplicity of infection of 5:1, macrophages from a range of animal species including cattle, deer, possums, ferrets and mice restricted growth of BCG while M. bovis grew progressively. Inter-species variation in controlling growth of M. bovis by alveolar macrophages was observed. Pre-treatment of macrophages with interferon-gamma and lipopolysaccharide inhibited intracellular growth of M. bovis. Addition of freshly recruited macrophages further inhibited M. bovis, and intracellular growth was arrested by activated fresh macrophages. Our observations suggest that naïve macrophages can prevent BCG growth, while T cell activation in conjunction with freshly recruited macrophages is required for preventing growth of M. bovis.

Analysis of Variance↗

PubMed2001

Vaccination of animals against Mycobacterium bovis.

Vaccination could potentially be used as a practical means of controlling bovine tuberculosis in countries in which a wildlife reservoir of the disease is present, and also in those countries which cannot afford conventional control strategies. An understanding of the processes involved in the protective immune response to tuberculosis is desirable for the rational development and testing of new vaccines for tuberculosis. The authors review current knowledge regarding the processes involved in protective immune responses to tuberculosis, much of which has been derived from studies in mice. This knowledge is discussed in relation to the problem of using vaccination to induce protective immunity in cattle, deer and wildlife. Challenge models have now been developed to test candidate vaccines in many domestic animals and wildlife species and these models are being used to evaluate tuberculosis vaccines. Most studies of the efficacy of tuberculosis vaccines in target animals have focused on the use of bacillus Calmette-Guérin (BCG), an attenuated strain of Mycobacterium bovis. Recent advances in immunology and the molecular biology of mycobacteria have greatly increased the options for candidate vaccines and future studies will test new types of vaccines including new attenuated strains of M. bovis, sub-unit protein vaccines and recombinant deoxyribonucleic acid vaccines. Several of these vaccines have shown promising results when tested in small animal models. Although progress has been made in the development of vaccine delivery systems for animals, the technical problems associated with vaccination of wildlife remain a challenge.

Animals↗

PubMed2000

Marsupial cytokines. Structure, function and evolution.

The cytokines are an important group of molecules involved in coordinating the many and varied components of the immune system. These molecules have been extensively studied in model eutherian mammals such as mice but comparatively little is known about the cytokine network of marsupials. Such information will be invaluable in elucidating fundamental aspects of the marsupial immune system and will also highlight parallels and differences between the immune systems of marsupials and eutherians. Given the importance of these goals, our groups have recently begun to tackle this lack of knowledge of the marsupial cytokine system and have met with considerable success in the face of the rapid rate of change of these proteins. This has led to the isolation of the full-length sequences encoding marsupial orthologues of tumour necrosis factor (TNF), lymphotoxins alpha and beta (LT-alpha and beta), interleukin-1 beta (IL-1beta), and interleukin-10 (IL-10). Here we review what has been learnt about structural, functional and evolutionary aspects of these marsupial cytokines as well as briefly describing more recent work in progress and future directions in this field.

Animals↗

PubMed2000

Effects of yeast expressed recombinant interleukin-2 and interferon-gamma on physiological changes in bovine mammary glands and on bactericidal activity of neutrophils.

The physiological effects of intramammary infusions of recombinant bovine cytokines in six lactating dairy cows on the quality and yield of milk and the bactericidal activity of milk neutrophils were investigated. Recombinant bovine interleukin-2 (rboIL-2) and interferon-gamma (rboIFN-gamma) were produced in the yeast Pichia pastoris. Two animals were given rboIL-2 (2 x 10(5) units) in two quarters, two animals were given rboIFN-gamma (6.5 x 10(5) units) in two quarters, and the other two cows received a dose of rboIL-2 in one quarter and rboIFN-gamma in a second quarter. In addition, each animal was given phosphate-buffered saline (PBS) in the other two quarters as a control. Somatic cell counts and conductivity of the fore milk were monitored before and after infusion. Neutrophils were isolated from quarter milk samples 36 h after infusion of cytokine or PBS and their bactericidal activities against Staphylococcus aureus were measured in vitro with a colorimetric assay. Quarters infused with rboIL-2 or rboIFN-gamma showed significant but transitory increases in both milk somatic cell counts and conductivity when compared with preinfusion values and with control quarters. There were minimal effects on daily milk yield. Neutrophils isolated from milk from quarters infused with rboIL-2 showed enhanced bactericidal activity against Staph. aureus. The bacterial killing from rboIL-2 treated quarters was significantly greater, with a mean of 63.5% compared with a mean of 5.4% for neutrophils taken from uninfected quarters to which PBS had been administered. The bactericidal activities for quarters treated with rboIFN-gamma and infected quarters treated with PBS were 15.0 and 30.0% respectively. The results indicate that intramammary infusions of rboIL-2 and rboIFN-gamma to lactating cows are well tolerated, and that rboIL-2 can activate milk neutrophils and augment their bactericidal activity.

Animals↗

PubMed2000

Vaccination of cattle with Mycobacterium bovis culture filtrate proteins and interleukin-2 for protection against bovine tuberculosis.

In this study vaccines prepared from culture filtrate proteins (CFP) of Mycobacterium bovis and interleukin-2 (IL-2) were tested in cattle for their capacity to stimulate immune responses and to protect against an intratracheal challenge with virulent M. bovis. Nine groups of cattle were vaccinated with combinations of different doses of CFP and bovine IL-2 mixed with a monophosphoryl lipid A (MPL) adjuvant. An additional group was vaccinated with M. bovis BCG. Immune responses in b1P-IL-2-vaccinated animals differed from those seen in BCG-vaccinated animals by inducing high antigen-specific antibody responses and low levels of gamma interferon and IL-2 released from purified protein derivative-stimulated whole-blood cultures. In a concurrent experiment, additional animals were added to the high-dose CFP-IL-2, MPL control, and BCG groups and these expanded groups of animals were challenged intratracheally with virulent M. bovis. Although the lung lesion scores were significantly lower for both the CFP-IL-2-and BCG-vaccinated groups compared to the MPL control group, the overall level of protection was greatest for the BCG-vaccinated animals. There were more animals with extrathoracic spread of disease in the CFP-IL-2 group than in the other groups. While vaccination of cattle with M. bovis CFP gave an encouraging reduction in tuberculous lesions and did not induce a delayed-type hypersensitivity response to PPD, future CFP vaccines must prevent any extrathoracic spread of disease.

Animals↗

PubMed1999

Molecular cloning and physiological effects of brushtail possum interleukin-1beta.

Interleukin-1beta (IL-1beta) was isolated from LPS-stimulated brushtail possum alveolar macrophages using PCR primers based on conserved regions of mammalian IL-1beta. The complete cDNA was cloned by 5' and 3' rapid amplification of cDNA ends (RACE). The predicted protein of 269 amino acids shared 4346% identity with several mammalian IL-1beta proteins. Constructs were made to express the mature IL-1beta in Escherichia coli and two recombinant IL-1beta proteins, rpIL-1beta1 and rpIL-1beta2, which differed in length by four amino acids at the N-terminus, were produced. Both proteins induced a weak proliferative response in a possum thymocyte assay. Possums injected intravenously with 100 microg of rpIL-1beta1 or rpIL-1beta2 showed profound changes in body temperature and numbers of circulating leukocytes. A sharp decrease in temperature occurred within 2 h of administration followed by an elevation of temperature peaking at 24 h. The smaller rpIL-1beta1 protein had a greater effect on temperature than rpIL-1beta2. Both rpIL-1beta proteins caused a marked decrease in number of neutrophils and lymphocytes at 2-6 h after injection. At 24 h after injection, neutrophil and lymphocyte numbers were elevated 6.0-fold and 2.6-fold, respectively in the possums injected with rpIL-1beta1 and 3.9-fold and 1.5-fold, respectively in the possums injected with rpIL-1beta2. Fibrinogen levels were elevated at 24 and 72 h after injection with both proteins. In comparison, neither recombinant bovine IL-1beta (rbIL-1beta) nor PBS had significant effects on body temperature or blood haematology. The studies have shown that the two recombinant forms of IL-1beta were biologically active in possums and that the IL-1beta with four fewer amino acids at the N-terminus was the more active.

Amino Acid Sequence↗

PubMed1999

Physiological effects and adjuvanticity of recombinant brushtail possum TNF-alpha.

The present paper describes the physiological properties of recombinant possum TNF-alpha and an adjuvant effect on antibody responses to the model protein antigen, keyhole limpet haemocyanin (KLH). For these studies recombinant possum TNF-alpha was produced in the yeast Pichia pastoris. The recombinant cytokine was secreted into the culture medium and purified by gel filtration. Possum TNF-alpha produced in this expression system was N-glycosylated and bioactive in two different assays. In a murine fibroblast L929 cytotoxicity assay, the possum TNF-alpha had lower specific activity compared to human TNF-alpha, while in a possum-specific assay, possum TNF-alpha enhanced the proliferation of PHA-stimulated possum thymocytes and was more active than human TNF-alpha. The physiological effect of the recombinant possum TNF-alpha was investigated in groups of possums administered doses of 6, 30 or 150 micrograms of cytokine. For each dose, TNF-alpha caused profound effects on the numbers of circulating leucocytes characterized by a three-to-four-fold increase in neutrophil numbers at 6-24 h after injection and an initial sharp decrease in lymphocyte numbers. The efficacy of TNF-alpha as an immunological adjuvant was determined in possums administered KLH (125 micrograms) in an aqueous or Al(OH)3-based formulation with or without added recombinant TNF-alpha (150 micrograms). Serum antibody responses to KLH were monitored by ELISA. The TNF-alpha stimulated two-fold and four-fold increases in antibody levels in aqueous and Al(OH)3-based vaccine formulations, respectively. The strongest antibody responses were observed in the group of possums that received KLH formulated in Al(OH)3 with addition of TNF-alpha.

Adjuvants, Immunologic↗

PubMed1999

Immune responses induced in cattle by virulent and attenuated Mycobacterium bovis strains: correlation of delayed-type hypersensitivity with ability of strains to grow in macrophages.

Comparison of immune responses induced in cattle by virulent and attenuated strains of Mycobacterium bovis will assist in identifying responses associated with resistance or susceptibility to disease. Four strains of M. bovis, one which is virulent in guinea pigs (WAg201) and three which are attenuated in guinea pigs (an isoniazid-resistant strain [WAg405], ATCC 35721, and BCG) were compared for their abilities to induce immune responses in cattle and to grow in bovine lung alveolar macrophage cultures. Extensive macroscopic lesions were found only in cattle inoculated with the virulent M. bovis strain. Strong antibody responses to M. bovis culture filtrate, as well as persistently high levels of gamma interferon and interleukin-2 released from purified protein derivative (PPD)-stimulated peripheral blood lymphocyte cultures, were observed in the cattle inoculated with the virulent strain compared to those inoculated with the attenuated strains. All cattle inoculated with the virulent strain or two of the attenuated strains (WAg405 and ATCC 35721) elicited strong delayed-type hypersensitivity responses to PPD in skin tests, while animals inoculated with BCG induced only a weak response. The three strains which produced strong skin test responses proliferated well in bovine alveolar macrophages and induced high levels of proinflammatory cytokine mRNAs compared to BCG. Our study showed that skin test responsiveness to PPD correlated with the ability of the strains to grow in alveolar macrophages rather than to their pathogenicity in cattle.

Animals↗

PubMed1998

Nucleotide sequence of a marsupial interleukin-10 cDNA from the Australian brushtail possum (Trichosurus vulpecula).

A cDNA encoding a marsupial interleukin-10 (IL-10), was isolated from Australian brushtail possum alveolar macrophages. The cDNA of 1604 bp had an open reading frame of 522 bp coding for a protein of 174 amino acids. Its deduced amino acid sequence had an identity of 60% with cat, 58% with pig, 56% with human and cow, 52% with mouse and 53% with rat IL-10. The expression of IL-10 was up-regulated in both LPS-stimulated and Mycobacterium bovis-infected possum alveolar macrophages.

Amino Acid Sequence↗

PubMed1997

Antigen-induced interferon-gamma and interleukin-2 responses of cattle inoculated with Mycobacterium bovis.

Bovine purified protein derivative (PPD)-induced interferon-gamma (IFN-gamma) and interleukin-2 (IL-2) mRNA expression was measured in peripheral blood lymphocyte cultures of cattle inoculated with Mycobacterium bovis and compared to cytokine protein levels as measured by IFN-gamma enzyme-linked immunosorbent assay and IL-2 bioassay. For individual animals, positive correlations were observed between mRNA and protein levels of bovine PPD-induced IFN-gamma and IL-2, although the correlations were stronger for IFN-gamma. Expression of these two cytokines also correlated with responses from a comparative intradermal test and a M. bovis antibody test. At 7 and 20 weeks after inoculation, bovine PPD-induced IFN-gamma and IL-2 mRNA expression was detected in all animals with tuberculous lesions and in a proportion of the M. bovis-inoculated animals with no lesions. Correlation of antigen-induced IFN-gamma and IL-2 with other immune parameters suggests that these two cytokines play an important role in the immune response to bovine tuberculosis.

Animals↗