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.     

Bovine TB and the development of new vaccines

Bovine tuberculosis (bTB) is caused by Mycobacterium bovis. The incidence of bTB is increasing in cattle herds of developed countries that have a wild life reservoir of M. bovis, such as the UK, New Zealand and the USA. The increase in the incidence of bTB is thought to be due, at least in part, to a wildlife reservoir of M. bovis. M. bovis is also capable of infecting humans and on a worldwide basis, M. bovis is thought to account for up to 10% of cases of human TB [Cosivi O, Grange JM, Daborn CJ et al. Zoonotic tuberculosis due to Mycobacterium bovis in developing countries. Emerg Infect Dis 1998;4(1):59–70]. Thus, the increased incidence of bTB, besides being a major economic problem, poses an increased risk to human health. In the UK, the incidence of bTB continues to rise despite the use of the tuberculin test and slaughter control policy, highlighting the need for improved control strategies. Vaccination of cattle, in combination with more specific and sensitive diagnostic tests, is suggested as the most effective strategy for bovine TB control. The only vaccine currently available for human and bovine TB is the live attenuated Bacille Calmette Guerin (BCG). BCG is thought to confer protection through the induction of Th1 responses against mycobacteria. However, protection against TB conferred by BCG is variable and to this date the reasons for the successes and failures of BCG are not clear. Therefore, there is a need to develop vaccines that confer greater and more consistent protection against bTB than that afforded by BCG. Given that BCG is currently the only licensed vaccine against human TB, it is likely that any new vaccine or vaccination strategy will be based around BCG. In this review we discuss immune responses elicited by mycobacteria in cattle and the novel approaches emerging for the control of bovine TB based on our increasing knowledge of protective immune responses.     

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.     

BCG vaccination against tuberculosis in European badgers (Meles meles): a review

Tuberculosis (TB) is a significant animal health problem in many parts of the world, and reservoirs of infection in wild animals complicate disease control efforts in farmed livestock, particularly cattle. Badgers (Meles meles) are a significant wildlife reservoir of Mycobacterium bovis infection for cattle in the United Kingdom (UK) and Republic of Ireland (ROI). Vaccination of badgers using an M. bovis strain bacille Calmette-Guérin (BCG) vaccine could potentially be an option in the national TB eradication strategy. Wildlife vaccination has been used successfully for other diseases in wildlife species, and may have a role to play in reducing M. bovis transmission at the wildlife-livestock interface. Research to date has provided evidence that BCG is protective in badgers, and a parenteral badger BCG vaccine has been licensed in the UK. Further research is required to develop effective strategies for vaccine deployment and to determine the effect of badger vaccination on cattle TB incidence.     

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.     

Route of BCG administration in possums affects protection against bovine tuberculosis

The Australian brushtail possum (Trichosurus vulpecula) is the major wildlife reservoir of Mycobacterium bovis in New Zealand. Control of bovine tuberculosis in farmed animals requires measures to reduce the transmission of M. bovis from wildlife. Possums were vaccinated with BCG intranasally by aerosol spray, orally or subcutaneously to compare the efficacy of these three routes on protection against challenge with virulent M. bovis. Possums vaccinated with BCG by the intranasal or subcutaneous routes had a marked reduction in severity of disease compared to possums which had been unvaccinated or orally vaccinated. The severity of the disease was assessed by changes in body weight and pathology. BCG vaccination by all three routes resulted in reduced dissemination of M. bovis to the spleen and liver following challenge. Intranasal and oral BCG vaccination induced lower mean peripheral blood lymphocyte blastogenic responses to bovine PPD than subcutaneous vaccination, indicating that these responses did not correlate well with protection from the disease. Given a suitable delivery system, aerosol vaccination of possums, used in conjunction with other control measures, may be a suitable method of reducing the spread of M. bovis from wildlife to domestic animals.     

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.     

Immunological approaches to the control of tuberculosis in wildlife reservoirs

Attempts to eradicate tuberculosis from cattle and farmed deer in some countries have been frustrated by the existence of wildlife reservoirs of Mycobacterium bovis infection. Possum control programmes in New Zealand using poisons have shown clearly that the brushtail possum is an important source of infection for cattle and farmed deer, and the sum of evidence strongly suggests that badgers serve as a source of infection for cattle in the UK. Bovine tuberculosis can only be eradicated from these countries by controlling M. bovis infection in both wildlife and domestic animals. The most promising options for control of M. bovis infection in wildlife in the longer term include the development of a tuberculosis vaccine for wildlife and a strategy for biological control of possums. The aim of this review is to address the problems and approaches involved in the control of wildlife tuberculosis from an immunological perspective.     

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.     

A review of M. bovis BCG protection against TB in cattle and other animals species

Bovine tuberculosis (TB) causes severe economic losses in livestock due to low production, animal deaths and condemnation of carcasses. It is also an important constraint in international trade of animals and animal products. A scientific committee in Great Britain in 1997 concluded that the development of a cattle vaccine would be the best option for long-term control of TB. However, vaccination of cattle currently is not accepted because the vaccine interferes with the skin reaction to the tuberculin test in the field. Efficacy of M. bovis BCG in protecting bovine and other animal species against tuberculous infection has received much study. Vaccination of cattle prevents the spread of the disease in populations by reducing the number and size of the lesions, and the load of bacteria (rather than by preventing infection). We review the literature about the efficacy of BCG in protecting cattle and other animal species against infection with field strains of M. bovis and discusses its potential use in programs of TB control in high-prevalence populations.     

Improved immunogenicity of DNA vaccination with mycobacterial HSP65 against bovine tuberculosis by protein boosting

A scientific review for the government of the United Kingdom has recommended that the development of a cattle vaccine against bovine tuberculosis holds the best prospects to control this disease in the national herd. As BCG vaccination of cattle results in variable degrees of protection, novel vaccine strategies that could replace or supplement BCG are required. In this study, the mycobacterial antigen HSP65 was used to determine whether priming cattle with a plasmid DNA vaccine and subsequently boosting with the recombinant protein in adjuvant (heterologous prime-boost approach) would result in improved and more homogenous immune responses over immunising with plasmid DNA or protein in adjuvant alone. The results demonstrated that strong, and compared to protein or DNA vaccination protocols alone, more homogenous, cellular immune responses were induced in cattle vaccinated with the prime-boost regimen. In addition, DNA prime-protein boost vaccination as well as protein vaccination resulted in stronger humoral immune responses with a balanced IgG profile compared to DNA vaccination alone. Importantly, none of the vaccination protocols sensitised cattle to the intradermal tuberculin test suggesting that TB subunit vaccines can be designed to allow the continued use of the tuberculin test to discriminate between vaccinated cattle and those infected with Mycobacterium bovis.     

Protection against bovine tuberculosis induced by oral vaccination of cattle with Mycobacterium bovis BCG is not enhanced by co-administration of mycobacterial protein vaccines

Mycobacterium bovis bacille Calmette-Guérin (BCG) delivered to calves by the oral route in a formulated lipid matrix has been previously shown to induce protection against bovine tuberculosis. A study was conducted in cattle to determine if a combination of a low dose of oral BCG and a protein vaccine could induce protective immunity to tuberculosis while not sensitising animals to tuberculin. Groups of calves (10 per group) were vaccinated by administering 2 × 10(7)colony forming units (CFU) of BCG orally or a combination of 2 × 10(7)CFU oral BCG and a protein vaccine comprised of M. bovis culture filtrate proteins (CFP) formulated with the adjuvants Chitin and Gel 01 and delivered by the intranasal route, or CFP formulated with Emulsigen and the TLR2 agonist Pam(3)CSK(4) and administered by the subcutaneous (s.c.) route. Two further groups were vaccinated with the CFP/Chitin/Gel 01 or CFP/Emulsigen/Pam(3)CSK(4) vaccines alone. Positive control groups were given 10(8)CFU oral BCG or 10(6)CFU s.c. BCG while a negative control group was non-vaccinated. All animals were challenged with M. bovis 15 weeks after vaccination and euthanized and necropsied at 16 weeks following challenge. Groups of cattle vaccinated with s.c. BCG, 10(8)CFU or 2 × 10(7)CFU oral BCG showed significant reductions in seven, three and four pathological or microbiological disease parameters, respectively, compared to the results for the non-vaccinated group. There was no evidence of protection in calves vaccinated with the combination of oral BCG and CFP/Emulsigen/Pam(3)CSK(4) or oral BCG and CFP/Chitin/Gel 01 or vaccinated with the protein vaccines alone. Positive responses in the comparative cervical skin test at 12 weeks after vaccination were only observed in animals vaccinated with s.c. BCG, 10(8)CFU oral BCG or a combination of 2 × 10(7)CFU oral BCG and CFP/Chitin/Gel 01. In conclusion, co-administration of a protein vaccine, administered by either systemic or mucosal routes with oral BCG did not enhance the protection conferred by administration of oral BCG alone.     

Vaccines designed to protect against Mycobacterium tuberculosis infection may aid the identification of novel vaccine constructs and diagnostic antigens for bovine tuberculosis

Vaccination has been identified as a promising control strategy for tuberculosis in both humans and cattle. Recent heterologous prime-boost approaches combining BCG vaccination with subunit boosts have shown considerable promise in both fields. However, the identification of further protective antigens is still a research priority. In this paper we have established the response hierarchy in Mycobacterium bovis infected or BCG vaccinated cattle of 6 Mycobacterium tuberculosis-derived proteins that were protective against M. tuberculosis in the guinea pig aerosol challenge model. Two of these proteins, Rv1806 and Rv3812, were recognised most frequently in cattle and therefore constitute potential subunit vaccine candidates that merit further evaluation in cattle. Their epitopes were mapped in infected cattle and were shown to be located primarily in the non-conserved regions of these PE/PE-PGRS protein family members. The aim of this study was to ascertain the presence of any correlation between the immunogenicity of defined antigens in different animal species. A weak association between guinea pig immunogenicity (as measured by protection) and antigenicity in M. bovis infected or BCG vaccinated cattle was found.     

牛结核病新型疫苗的研究现状

在过去的20年里,人们应用分子生物学技术来研制更为有效的结核病疫苗,新型候选疫苗大量涌现。这些新型疫苗主要包括减毒活疫苗、重组活疫苗、亚单位疫苗和核酸疫苗。对防制牛结核病的各种新型疫苗也进行了相应的研究,并取得了令人振奋的进展。各种新型疫苗各有优缺点。目前看来,重组卡介苗和DNA疫苗被认为是最有前途的候选疫苗。但是,所有候选疫苗共同的也是致命的缺点是免疫保护力低。因此,牛结核病疫苗研制的主要努力方向还是在研究分支杆菌免疫机制和免疫失败原因的基础上,进一步增强现有候选疫苗的免疫效力或研制更为有效的新型疫苗。     

Progress in the control of bovine tuberculosis in Spanish wildlife

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

Trial design to estimate the effect of vaccination on tuberculosis incidence in badgers

The principal wildlife reservoir of Mycobacterium bovis in Ireland is the European badger. Studies in the Republic of Ireland (RoI) have shown that badgers culled in association with cattle herd tuberculosis breakdowns (focal culling) have a higher prevalence of infection than the badger population at large. This observation is one rationale for the medium term national strategy of focal badger culling. A vaccination strategy for the control of bovine tuberculosis (bTB) in badgers is a preferred long-term option. The Bacillus Calmette-Guérin (BCG) vaccine has been shown to decrease disease severity in captive badgers under controlled conditions. As the vaccine has been tested in a controlled environment with precise information on infection pressure, it cannot be assumed a priori that the effects of vaccination are similar in the wild, where other environmental and/or ecological factors prevail. For this reason we have designed a vaccine field trial to assess the impact of vaccination on the incidence of TB infection in a wild badger population. The selected study area for the vaccine trial (approximately 755 square kilometers) is divided into three zones each of which has similar characteristics in terms of size, number of main badger setts, cattle herds, cattle and land classification type. Three vaccination levels (100%, 50% and 0%) will be allocated to the three zones in a way that a gradient of vaccination coverage North to South is achieved. The middle zone (zone B) will be vaccinated at a 50% coverage but zone A and C will be randomly allocated with 100% or 0% vaccination coverage. Vaccination within zone B will be done randomly at individual badger level. The objective of this paper is to describe the design of a field tuberculosis vaccination trial for badgers, the epidemiological methods that were used to design the trial and the subsequent data analysis. The analysis will enable us to quantify the magnitude of the observed vaccination effect on M. bovis transmission in badgers under field conditions and to improve our knowledge of the biological effects of vaccination on susceptibility and infectiousness.     

The safety and immunogenicity of Bacillus Calmette-Guérin (BCG) vaccine in European badgers (Meles meles)

European badgers (Meles meles) are a wildlife reservoir for Mycobacterium bovis (M. bovis) in Great Britain (GB) and the Republic of Ireland and therefore constitute a potential source of infection for cattle. Reduction of badger densities in the Republic of Ireland has resulted in an associated reduction in the risk of a herd break-down with bovine tuberculosis and a study to determine whether this is also the case in GB has been running since 1997. If badgers are a significant source of M. bovis infection for cattle, vaccinating badgers with Bacillus Calmette-Guérin (BCG) might prove to be a long term, cost-effective strategy for controlling bovine tuberculosis whilst preserving badger populations. As a first step towards BCG vaccination of wild badgers, it was necessary to demonstrate safety of the vaccine in captive badgers. Therefore, captive badgers were vaccinated with a commercial source of BCG that is already licensed for administration to humans in GB-BCG Danish SSI. Using a protocol prescribed by the Veterinary Medicines Directorate (VMD) of GB, badgers were vaccinated with two consecutive doses of BCG via either the subcutaneous (s.c.) or intra-muscular (i.m.) routes. The first dose was high, ranging from 16 to 22 x 10(7) colony-forming units (CFU), and was followed 15 weeks later by a lower dose in the range of 4-7 x 10(5)CFU. Local reaction at the site of injection and general responses (body temperature, haematology and blood serum chemistry), behaviour and excretion of BCG were monitored for 28 weeks from the time of the first vaccination. The only side-effect observed was the occurrence of localised swelling at the site of BCG injection that disappeared 48 days after i.m. vaccination but persisted longer in the group vaccinated by the s.c. route. Immunological responses were measured at regular intervals. Strong cellular responses were observed 13 days after the first vaccination, which persisted for 76 days. The lower dose induced a weaker and shorter-lived response.     

A DNA vaccine encoding MPB83 from Mycobacterium bovis reduces M. bovis dissemination to the kidneys of mice and is expressed in primary cell cultures of the European badger (Meles meles)

Nucleic acid (DNA) vaccination against tuberculosis in the European badger (Meles meles) is one approach to addressing the escalating problem of bovine tuberculosis in Great Britain. The aim of vaccination is to reduce the burden of tuberculosis within the badger population and the shedding of Mycobacterium bovis to levels that would break the transmission of infection to cattle. To this end, the vaccine would be required to limit the amount of disseminated tuberculosis in the badger, especially dissemination to the kidney from where M. bovis can be shed in the urine. A promising candidate DNA vaccine encoding a 26 kDa major antigen (MPB83) of M. bovis was evaluated in a mouse model of disseminated M. bovis infection. Using the DNA vaccine, protection against infection of the kidney was found to be greater than that achieved with the current live vaccine, Bacille Calmette-Guerin (BCG). Kidney tissue and skeletal muscle from the badger was used to derive primary cell cultures in which to examine the expression of MPB83 following transfection with the DNA vaccine. Kidney cortex gave rise to a monotypic culture of epithelial cells whilst the muscle gave rise to a mixed culture of fibroblasts and myoblasts. During culture the myoblasts differentiated into multinucleated myotubes, verified by immunofluorescent detection of mammalian desmin. Successful expression of MPB83 by transfected epithelial and myotube cells was confirmed by immunofluorescence using a monoclonal antibody specific to the protein. These observations fulfil the early requirements for the development of a DNA vaccine for badger tuberculosis.     

Development of cattle TB vaccines in the UK

In 1996, an independent scientific committee chaired by Professor John Krebs, tasked to review the problem of bovine tuberculosis (TB) in GB, concluded that vaccination of cattle offered the best long-term solution for controlling the disease in the National Herd. This view has been re-affirmed recently in the House of Commons Environment, Food and Rural Affairs Committee's report on Bovine TB (2004) and by the findings of the Independent Scientific Group Vaccine Scoping Sub-committee. Significant progress in developing TB vaccines for cattle has been made over the last 5 years. Specifically: (i) DNA or protein subunit vaccines used in combination with BCG have been shown to give superior protection against experimental challenge in cattle than BCG (heterologous prime-boost); (ii) prototype reagents that allow discrimination between vaccinated and infected animals have been developed; and (iii) and correlates of disease severity have been identified that can predict the success or failure of vaccination. These significant advances are detailed in this review with a summary of future directions that TB vaccine development for cattle is likely to take.