Oral administration of Lactobacillus brevis KB290 to mice alleviates clinical symptoms following influenza virus infection.

UNLABELLED: Lactobacillus brevis KB290 (KB290), isolated from a traditional Japanese pickle 'Suguki', has been reported to have immunomodulatory effects. We investigated whether oral administration of KB290 has protective effects against influenza virus (IFV) infection in mice. After 14 days of administration of lyophilized KB290 suspended in phosphate-buffered saline by oral gavage, BALB/c mice were intranasally infected with 2 × MLD50 (50% mouse lethal dose) of IFV A/PR/8/34 (H1N1). Prophylactically administered KB290 significantly alleviated the loss of body weight and the deterioration in observational physical conditions induced by the infection. In addition, 7 days after infection, the levels of IFV-specific immunoglobulin (Ig)A in bronchoalveolar lavage fluid were significantly increased in mice fed KB290 compared with controls. Moreover, there was a significant elevation of serum interferon (IFN)-α in KB290 group mice, even at three and 7 days after infection, despite the administration of KB290 being stopped before IFV infection. Our results demonstrated that oral administration of KB290 before infection could alleviate IFV-induced clinical symptoms. Alleviation of clinical symptoms by KB290 consumption may have been induced by long-lasting enhancement of IFN-α production and the augmentation of IFV-specific IgA production. SIGNIFICANCE AND IMPACT OF THE STUDY: This study demonstrated that oral administration of Lactobacillus brevis KB290 (KB290), a probiotic strain derived from a Japanese traditional pickle, could protect against influenza virus (IFV) infection in mice. Our results demonstrated that continual intake of KB290 for 14 days prior to IFV infection alleviated clinical symptoms such as loss of body weight and deterioration in observational physical conditions induced by the infection. The beneficial effects of KB290 consumption may have been elicited by the long-lasting enhancement of interferon-α production and the augmentation of IFV-specific immunoglobulin A production.

Oral vaccination reduces the incidence of tuberculosis in free-living brushtail possums.

Bovine tuberculosis (Tb) caused by Mycobacterium bovis has proved refractory to eradication from domestic livestock in countries with wildlife disease reservoirs. Vaccination of wild hosts offers a way of controlling Tb in livestock without wildlife culling. This study was conducted in a Tb-endemic region of New Zealand, where the introduced Australian brushtail possum (Trichosurus vulpecula) is the main wildlife reservoir of Tb. Possums were trapped and vaccinated using a prototype oral-delivery system to deliver the Tb vaccine bacille Calmette-Guerin. Vaccinated and control possums were matched according to age, sex and location, re-trapped bimonthly and assessed for Tb status by palpation and lesion aspiration; the site was depopulated after 2 years and post-mortem examinations were conducted to further identify clinical Tb cases and subclinical infection. Significantly fewer culture-confirmed Tb cases were recorded in vaccinated possums (1/51) compared with control animals (12/71); the transition probability from susceptible to infected was significantly reduced in both males and females by vaccination. Vaccine efficacy was estimated at 95 per cent (87-100%) for females and 96 per cent (82-99%) for males. Hence, this trial demonstrates that orally delivered live bacterial vaccines can significantly protect wildlife against natural disease exposure, indicating that wildlife vaccination, along with existing control methods, could be used to eradicate Tb from domestic animals.

Animal-side serologic assay for rapid detection of Mycobacterium bovis infection in multiple species of free-ranging wildlife.

Numerous species of mammals are susceptible to Mycobacterium bovis, the causative agent of bovine tuberculosis (TB). Several wildlife hosts have emerged as reservoirs of M. bovis infection for domestic livestock in different countries. In the present study, blood samples were collected from Eurasian badgers (n=1532), white-tailed deer (n=463), brushtail possums (n=129), and wild boar (n=177) for evaluation of antibody responses to M. bovis infection by a lateral-flow rapid test (RT) and multiantigen print immunoassay (MAPIA). Magnitude of the antibody responses and antigen recognition patterns varied among the animals as determined by MAPIA; however, MPB83 was the most commonly recognized antigen for each host studied. Other seroreactive antigens included ESAT-6, CFP10, and MPB70. The agreement of the RT with culture results varied from 74% for possums to 81% for badgers to 90% for wild boar to 97% for white-tailed deer. Small numbers of wild boar and deer exposed to M. avium infection or paratuberculosis, respectively, did not cross-react in the RT, supporting the high specificity of the assay. In deer, whole blood samples reacted similarly to corresponding serum specimens (97% concordance), demonstrating the potential for field application. As previously demonstrated for badgers and deer, antibody responses to M. bovis infection in wild boar were positively associated with advanced disease. Together, these findings suggest that a rapid TB assay such as the RT may provide a useful screening tool for certain wildlife species that may be implicated in the maintenance and transmission of M. bovis infection to domestic livestock.

Efficacy of oral and parenteral routes of Mycobacterium bovis bacille Calmette-Guerin vaccination against experimental bovine tuberculosis in white-tailed deer (Odocoileus virginianus): a feasibility study.

We investigated the efficacy of oral and parenteral Mycobacterium bovis bacille Calmette-Guerin Danish strain 1331 (BCG) in its ability to protect white-tailed deer (Odocoileus virginianus) against disease caused by M. bovis infection. Twenty-two white-tailed deer were divided into four groups. One group (n=5) received 10(9) colony-forming units (cfu) BCG via a lipid-formulated oral bait; one group (n=5) received 10(9) cfu BCG in culture directly to the oropharynx, one group (n=6) was vaccinated with 10(6) cfu BCG subcutaneously, and one group served as a control and received culture media directly to the oropharynx (n=6). All animals were challenged 3 mo after vaccination. Five months postchallenge the animals were examined for lesions. Results indicate that both oral forms of BCG and parenterally administered BCG offered significant protection against M. bovis challenge as compared to controls. This study suggests that oral BCG vaccination may be a feasible means of controlling bovine tuberculosis in wild white-tailed deer populations.

The potential of oral vaccines for disease control in wildlife species.

Numerous infectious diseases caused by bacteria or viruses persist in developed and developing countries due to ongoing transmission among wildlife reservoir species. Such diseases become the target of control and management programmes in cases where they represent a threat to public health (for example rabies, sylvatic plague, Lyme disease), or livestock production (for example bovine tuberculosis, brucellosis, pseudorabies), or where they threaten the survival of endangered animal populations. In the majority of cases, lethal control operations are neither economically feasible nor publicly supported as a practical means for disease management. Prophylactic vaccination has emerged over the last 15 years as an alternative control strategy for wildlife diseases, mainly driven by the success of widescale oral rabies vaccination programmes for meso-carnivores in North America and Northern Europe. Different methods have been trialled for the effective delivery of wildlife vaccines in the field, however oral vaccination remains the most widely used approach. Successful implementation of an oral wildlife vaccine is dependent on a combination of three components: an efficacious immunogen, a suitable delivery vehicle, and a species-specific bait. This review outlines the major wildlife disease problems for which oral vaccination is currently under consideration as a disease management tool, and also focuses on the technological challenges that face wildlife vaccine development. The major conclusion is that attenuated or recombinant live microbes represent the most widely-used vaccines that can be delivered by the oral route; this in turn places major emphasis on effective delivery systems (to maintain vaccine viability), and on selective baiting systems, as the keys to wildlife vaccine success. Oral vaccination is a valuable adjunct or alternative strategy to culling for the control of diseases which persist in wildlife reservoirs.

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.

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.

Advances in understanding disease epidemiology and implications for control and eradication of tuberculosis in livestock: the experience from New Zealand.

A deteriorating tuberculosis problem in cattle and deer in New Zealand has been halted and then reversed over the last decade. Mycobacterium bovis infection in both wild and domestic animal populations has been controlled. This has been achieved by applying a multi-faceted science-based programme. Key features of this have been a comprehensive understanding of the epidemiology of tuberculosis in animals, confidence in sampling wild animal populations, effective application of diagnostic tests in cattle and deer, and the ability to map M. bovis genotypes.

Progress in the development of vaccines and diagnostic reagents to control tuberculosis in cattle.

The sharp rise of bovine tuberculosis (TB) in Great Britain and the continuing problem of wild life reservoirs in countries such as New Zealand and Great Britain have resulted in increased research efforts into the disease. Two of the goals of this research are to develop (1) cattle vaccines against TB and (2) associated diagnostic reagents that can differentiate between vaccinated and infected animals (differential diagnosis). This review summarises recent progress and describes efforts to increase the protective efficacy of the only potential TB vaccine currently available, Mycobacterium bovis BCG, and to develop specific reagents for differential diagnosis. Vaccination strategies based on DNA or protein subunit vaccination, vaccination with live viral vectors as well as heterologous prime-boost scenarios are discussed. In addition, we outline results from studies aimed at developing diagnostic reagents to allow the distinction of vaccinated from infected animals, for example antigens that are not expressed by vaccines like Mycobacterium bovis Bacille-Calmette-Guérin, but recognised strongly in Mycobacterium bovis infected cattle.

The effectiveness of parallel gamma-interferon testing in New Zealand's bovine tuberculosis eradication programme.

In bovine tuberculosis (bTB) eradication programmes, especially where prevalence is low, sensitivity of testing in infected herds must be maximised to reduce the possibility of recrudescence of prior infection and the risk to other herds via animal movement. The gamma-interferon (γ-IFN) assay applied in parallel with intradermal tuberculin testing has been shown to increase test sensitivity. The aim of this work was to substantiate this effect in the field. A retrospective observational study was conducted on 239 New Zealand cattle breeding and dairy herds with bTB infection between 1 July 2011 and 1 September 2015 to evaluate the outcomes of new policy introduced in 2011. The investigation defined the number and proportion of reactors (animals testing positive and slaughtered) found with lesions of bTB in intradermal caudal fold testing (CFT) and parallel γ-IFN testing, at the breakdown test or first whole herd test after breakdown, WHT(1), and at the final or projected final whole herd test, WHT(F). Parallel γ-IFN testing was used in 26.8% of the 239 herds at WHT(1), and 430 animals in 49 herds were deemed reactors. One hundred and sixty (37.2%) of these reactors from 32 herds were found to have bTB lesions, despite having been negative to caudal fold testing. These 160 infected animals accounted for 29.6% of all infection found at WHT(1). At WHT(F), parallel γ-IFN testing was conducted on 93 herds and detected a total of 122 reactors in 49 herds, in addition to those found by CFT. Twenty-one of these reactors, from 13 herds, had bTB lesions at slaughter, accounting for 67.7% of all reactors found with bTB at WHT(F). Eleven of these 13 herds would have had their movement restrictions revoked based on a negative herd CFT alone, and could potentially have caused outward transmission of bTB to other herds, as well as experiencing recrudescent breakdowns. We conclude that γ-IFN testing in infected herds, in parallel with intradermal tuberculin testing, is a valuable tool in a bTB eradication programme, as it enables higher test sensitivity at both herd and animal level. The use of the γ-IFN test over a risk cohort early in a breakdown assists in removal of early infection and some cases of anergy to intradermal tuberculin testing. Parallel γ-IFN with compulsory slaughter of reactors should be considered in breeding and dairy herds in conjunction with tuberculin testing before movement control is revoked, and will assist in achieving TB freedom on a herd level and nationally.

The efficacy of live tuberculosis vaccines after presensitization with Mycobacterium avium.

The variable efficacy of BCG in humans has been extensively documented but its cause is still not well understood. One possible reason for this variation is the effect of presensitization with environmental mycobacteria. To investigate in guinea pigs the effects of presensitization with well characterized Mycobacterium avium strains on the vaccine efficacy of BCG and of two recently developed, avirulent strains of Mycobacterium bovis. Two strains of M. avium containing the DNA insertion element IS901 (M. avium+) and two strains not containing this element (M. avium-) were inoculated subcutaneously or by oral administration into guinea pigs to assess their virulence in these animals and their ability to induce delayed type hypersensitivity to tuberculins. Subsequently, groups of guinea pigs presensitized with orally administered M. avium+ and M. avium- and a control group were vaccinated with BCG, or one of two newly attenuated strains of M. bovis. All groups were then challenged by the aerosol route with virulent M. bovis. Vaccine efficacy was assessed 5 weeks later by the presence of macroscopic lesions and bacterial counts of spleen and lung. No macroscopic lesions were observed in any of the guinea pigs inoculated with strains of M. avium+ or M. avium- and all animals gave delayed-type hypersensitivity skin-test reactions to avian PPD. In the vaccine experiment, presensitization with orally administered M. avium+ alone produced a low level of protection against subsequent challenge with virulent M. bovis. In the absence of presensitization with M. avium or after presensitization with an M. avium- strain, BCG and two attenuated strains of M. bovis produced significant levels of protection. No additional protection was observed in lungs of guinea pigs presensitized with M. avium+ and subsequently vaccinated with BCG. In contrast, both newly attenuated strains of M. bovis induced significant protection in lungs after such presensitization. Presensitization of guinea pigs by the oral administration of M. avium+ provides a model for testing vaccines under conditions where the efficacy of BCG has been compromised by prior sensitization with environmental mycobacteria.

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.

Improving protective efficacy of BCG vaccination for wildlife against bovine tuberculosis.

Possums are a wildlife vector of bovine tuberculosis in New Zealand. Vaccination of possums with BCG is being considered as a measure to control the spread of bovine tuberculosis to cattle and deer. Delivery via oral bait is feasible but BCG is degraded in the stomach. The aim was to determine whether ranitidine (Zantac) would reduce gastric acidity and enhance the efficacy of intragastrically administered BCG. A dose of 75 mg reduced gastric acidity for at least 4 h. Thus, possums were vaccinated intragastrically with BCG after receiving 75 mg ranitidine or ranitidine or BCG alone, as controls, before challenge with virulent Mycobacterium bovis. Proliferative responses of blood lymphocytes to M. bovis antigens after vaccination were significantly higher in possums given ranitidine/BCG compared to controls and seven weeks after challenge they had significantly lower lung weights and spleen bacterial counts than ranitidine alone controls. Vaccination with BCG alone only gave a reduction in loss in body weight. Agents that reduce gastric acidity may be useful in formulating BCG for oral bait delivery to wildlife for vaccination against bovine tuberculosis.

Epidemiology and control of Mycobacterium bovis infection in brushtail possums (Trichosurus vulpecula), the primary wildlife host of bovine tuberculosis in New Zealand.

The introduced Australian brushtail possum (Trichosurus vulpecula) is a maintenance host for bovine tuberculosis (TB) in New Zealand and plays a central role in the TB problem in this country. The TB-possum problem emerged in the late 1960s, and intensive lethal control of possums is now used to reduce densities to low levels over 8 million ha of the country. This review summarises what is currently known about the pathogenesis and epidemiology of TB in possums, and how the disease responds to possum control. TB in possums is a highly lethal disease, with most possums likely to die within 6 months of becoming infected. The mechanisms of transmission between possums remain unclear, but appear to require some form of close contact or proximity. At large geographic scales, TB prevalence in possum populations is usually low (1-5%), but local prevalence can sometimes reach 60%. Intensive, systematic and uniform population control has been highly effective in breaking the TB cycle in possum populations, and where that control has been sustained for many years the prevalence of TB is now zero or near zero. Although some uncertainties remain, local eradication of TB from possums appears to be straightforward, given that TB managers now have the ability to reduce possum numbers to near zero levels and to maintain them at those levels for extended periods where required. We conclude that, although far from complete, the current understanding of TB-possum epidemiology, and the current management strategies and tactics, are sufficient to achieve local, regional, and even national disease eradication from possums in New Zealand.

Epidemiology, diagnostics, and management of tuberculosis in domestic cattle and deer in New Zealand in the face of a wildlife reservoir.

The control of tuberculosis (TB) in cattle and farmed deer in New Zealand has been greatly influenced by the existence of a wildlife reservoir of Mycobacterium bovis infection, principally the Australian brushtail possum (Trichosurus vulpecula). The reduction in possum numbers in areas with endemic M. bovis infection through vigorous vector control operations has been a major contributor to the marked reduction in the number of infected cattle and farmed deer herds in the past two decades. Management of TB in cattle and farmed deer in New Zealand has involved a combination of vector control, regionalisation of diagnostic testing of cattle and deer herds, abattoir surveillance and movement control from vector risk areas. Accurate diagnosis of infected cattle and deer has been a crucial component in the control programme. As the control programme has evolved, test requirements have changed and new tests have been introduced or test interpretations modified. Subspecific strain typing of M. bovis isolates has proved to be a valuable component in the epidemiological investigation of herd breakdowns to identify whether the source of infection was domestic livestock or wildlife. New initiatives will include the use of improved models for analysing diagnostic test data and characterising disease outbreaks leading to faster elimination of infection from herds. The introduction of the National Animal Identification Tracing programme will allow better risk profiling of individual herds and more reliable tracing of animal movements. TB in cattle and farmed deer in New Zealand can only be controlled by eliminating the disease in both domestic livestock and the wildlife reservoir.

Vaccination of cattle against Mycobacterium bovis.

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.

Towards more accurate diagnosis of bovine tuberculosis using defined antigens.

Diagnostic accuracy is of paramount importance in test-and-slaughter programmes for the eradication of bovine tuberculosis (TB). Currently applied methods, such as in vivo skin testing and in vitro interferon-gamma (IFN- gamma) testing, utilize purified protein derivatives (PPDs), which are poorly-defined mixtures containing many individual antigenic components. It is known that false-positive responses to these reagents can occur in cattle which are not infected with TB, largely because of that antigenic complexity. This paper reviews recent approaches to the characterization of more precisely defined diagnostic tools which can be used to develop tests with greater specificity. For example, the low mass secreted protein ESAT-6 has been shown to be capable of differentiating TB-infected cattle from those which develop responsiveness to PPD through contact with environmental mycobacteria or vaccination with BCG. The information which has accumulated in recent years has shown that the increased specificity is associated with some decrease in test sensitivity, but the overall advantages of being able to make precise diagnostic decisions will have significant advantages in many situations.

Use of the bovine model of tuberculosis for the development of improved vaccines and diagnostics.

Over the past few years there has been a resurgence in research into bovine tuberculosis due to the sharp rise of the disease in countries such as Great Britain and to the continuing problem of wild-life reservoirs in countries such as New Zealand. One of the goals of this research is to develop cattle vaccines against TB. The initial testing of candidate vaccines is carried out in laboratory animals, initially mice and subsequently guinea pigs. A unique feature of the cattle vaccination programme is that candidate vaccines which show promise in laboratory models can then be tested in the natural host species, cattle, before progressing to clinical trials. This is a major advantage over the strategy for developing a vaccine for human tuberculosis where, of course, it is impossible to test a candidate vaccine by experimentally challenging the host species with the pathogen. The most commonly used model for testing vaccine candidates in cattle consists of an intra-tracheal challenge of between 10(3) and 10(4) colony forming units of Mycobacterium bovis. The pathology observed following challenge is similar to human tuberculosis giving rise to a marked granulomatous reaction and a predominantly cellular immune response. Using this model we have been able to make a number of significant advances towards a bovine TB vaccine. First we have developed antigen cocktails that, when used in a whole blood gamma interferon assay, can differentiate between M. bovis infected and BCG vaccinated animals. Next we have developed immune correlates of pathology, which allow us to assess whether the vaccine is protecting animals against challenge before post mortem examination. Finally we have been able to use the model to develop a vaccine that improves the efficacy of BCG against M. bovis challenge.

Increased protection against bovine tuberculosis in the brushtail possum (Trichosurus vulpecula) when BCG is administered with killed Mycobacterium vaccae.

SETTING: The Australian brushtail possum is the major wildlife reservoir for Mycobacterium bovis infection in New Zealand. Development of an effective tuberculosis vaccine for possums will reduce the spread of infection to cattle and farmed deer. OBJECTIVES: To determine whether killed M. vaccae can improve the efficacy of vaccination with M. bovis bacillus Calmette Guerin (BCG) against bovine tuberculosis in the possum. DESIGN: Groups of possums (n=6-8) were vaccinated via intranasal and intraconjunctival routes with BCG alone or BCG in combination with heat-killed M. vaccae. Controls were non-vaccinated or vaccinated with heat-killed M. vaccae alone. After challenge with virulent M. bovis, protection was assessed by a reduction in loss of body weight and bacterial counts in lungs and spleens. Blood lymphocyte proliferative responses to M. bovis purified protein derivative were monitored throughout. RESULTS: The earliest lymphocyte responses following vaccination were from animals inoculated with BCG plus 100 microg heat-killed M. vaccae. Loss of body weight was significantly reduced in all BCG-vaccinated groups compared control groups. Spleen bacterial counts were significantly lower in animals vaccinated with M. vaccae plus BCG compared to the non-vaccinated group. Furthermore, vaccination with 100 microg M. vaccae plus BCG significantly reduced spleen bacterial counts compared to vaccination with BCG alone. CONCLUSION: The possum infection model is one of the first to show that novel vaccine strategies may offer better protection against tuberculosis than BCG alone.

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.