P.D. van Helden, E.G. Hoal/ Comparative ology, Microbiology and Infectious Diseases 36 (2013)287-294 It is often said that Mycobacterium bovis BCG, the current we also know that many species appear to be quite suscep- vaccine against tuberculosis, is arguably the most fre tible since TB is commonly reported in them, e.g. badgers, quently given human vaccine in the world today. It has been possums, cattle, buffalo, lions, lechwe, deer[13-16].On in use since the 1920s, despite ongoing argument about the other hand tb (or mycobacterial disease) has never ol its efficacy and complications in its usage [1, 2. Owing to rarely been reported in many other species, such as dogs this, some low incidence countries such as the USA, do not or horses. In some susceptible species, vaccination stud routinely vaccinate with BCG 3 One of the factors in this ies have been done, with some studies reporting various controversy is that there are now many different deriva levels of protection, using BCG or new candidate vaccines tives of BCG [ 4, which may not be comparable in their [17-26 However, it is not clear whether some animal effect, making any analysis of outcome extremely complex, strains or species or a proportion of any given species is and inter-trial or meta-analyses impossible owing to lack resistant to TB, or whether a proportion can acquire immu of power [1, 2,5]. Furthermore, BCG can be administered in nity under the appropriate conditions. different ways and at different ages 3, complicating an comparative analysis. Existing meta-analysis of BCG vacci- 3. Animal models nation suggests efficacy ranging from negative to positive effects, or protection preferentially against TB meningitis Various options exist, but the mouse model is still the and military TB, although it has been concluded that overall most frequently used Mice are relatively cheap and can be he risk of TB in humans are reduced by 50%[1, 2). Despite housed reasonably easily under containment for pathogen problems of analysis it cannot be claimed that BCG vaccina- challenge. Many well-characterized strains of mice are tion against TB is highly effective It is probably safe to say available, and there is a variety of reagents and kits avail- that BCG performs best at limiting disseminated disease able for mouse research. Few if any specific reagents or and mortality in children 1, 6]. It cannot reliably prevent lines exist for other animal models. However, there are infection or protect against pulmonary tB disease in adults some major differences between mice, other ani Thus, BCG as a vaccine is not sterilizing and there is a clear humans one notable difference is that mice do not form reason for development of new vaccine candidates (like he classic granuloma seen in most other vertebrates with wise, BCG is not a sterilizing vaccine in animals). Any nev TB. For this and many other reasons, this model may there candidate will have to be demonstrably better than the cur- fore have major deficiencies. Noting this, some researchers rent live BCG, in terms of efficacy and safety, the latter have invested in non-human primates as a model sys being a problem with BCG use, particularly in HIV posi- tem [27-29, but this is an expensive and time consuming route, albeit perhaps self-evidently better than others. Ide does not stop infection, latent TB or reactivation, or guar ally, potential vaccines for a particular species should be antee sterilizing immunity and we may perhaps find better tested in either the same animal, or at least a closely solutions using a post-exposure or therapeutic vaccine[8]. related species such as bovids for cattle or buffalo vaccines Perhaps the most difficult problem with TB or my Each existing model has its own problems and erial immunity is that the organism hides very successfully cies, the details of which are beyond the scope of this inside the macrophage or granuloma. For this reason, it paper to discuss in detail. However, a common limita- Is comm hought that T-cell immunity is critical and tion is that many lab-based models are not exposed to hat B-cells are of lesser importance 9-12]. Developing a the environmental stimuli experienced by free-living crea- vaccine to deliver immunity based on this premise is of tures, which include organisms such as other bacteria, fungi course complex, unlike the well-known successful humoral and parasites, which alone may render their hosts defi- response vaccines against viral diseases. cient as models. In addition. most animal models are inbred strains. If we postulate that genetics is a major determinant 2. Prior or acquired immunity? of resistance or susceptibility [30-33]. this alone makes such models tricky to understand since the strain may Since the overwhelming majority of infected humans be inherently resistant or susceptible. Even if not inbred never develop symptomatic (TB)disease, one may con- certain animal species may be innately resistant, where clude that they have inherent or innate immunity which vaccination is unnecessary, or innately susceptible, being an cope with theinfection, rendering them resistant. How- perhaps intractable to vaccination. This variation may of ever, it is also clear that at least in some communities, BCG course also be found within an out-bred population. due to genetic variation. Recognizing the limitations in labo- sonably conclude that vaccination can confer some degree ratory based animal models and the importance of dealing of acquired immunity. One may thus postulate that in the with research close to the real problem, a number of groups absence of immunosuppression, most humans are inhe have investigated larger animals such as deer or cattle for ently resistant via innate immunity: some may become trial vaccination. These animals are arguably a good choice resistant with the correct stimulus, but that some ma ince they are potentially susceptible, are at risk for disease, not be able to acquire immunity to TB In the absence of are economically important and impact on ecosystems. a sound body of research and data concerning diffe eren In addition, they present with large genetic and environ species of animals, we cannot make statements with the mental heterogeneity, which better reflects what we may same level of confidence for these species. However, w expect in trials under real life conditions. know that different mouse strains show varying levels of For this reason, apart from laboratory-based animal resistance or susceptibility to mycobacterial infection and models, a number of attempts to vaccinate free-living288 P.D. van Helden, E.G. Hoal / Comparative Immunology, Microbiology and Infectious Diseases 36 (2013) 287–294 Itis often said that Mycobacterium bovis BCG,the current vaccine against tuberculosis, is arguably the most fre￾quently givenhumanvaccine inthe worldtoday.Ithas been in use since the 1920s, despite ongoing argument about its efficacy and complications in its usage [1,2]. Owing to this, some low incidence countries such as the USA, do not routinely vaccinate with BCG [3]. One of the factors in this controversy is that there are now many different deriva￾tives of BCG [4], which may not be comparable in their effect, making any analysis of outcome extremely complex, and inter-trial or meta-analyses impossible owing to lack of power [1,2,5]. Furthermore, BCG can be administered in different ways and at different ages [3], complicating any comparative analysis. Existing meta-analysis of BCG vacci￾nation suggests efficacy ranging from negative to positive effects, or protection preferentially against TB meningitis and military TB, although it has been concluded that overall the risk of TB in humans are reduced by 50% [1,2]. Despite problems of analysis it cannot be claimed that BCG vaccina￾tion against TB is highly effective. It is probably safe to say that BCG performs best at limiting disseminated disease and mortality in children [1,6]. It cannot reliably prevent infection or protect against pulmonary TB disease in adults. Thus, BCG as a vaccine is not sterilizing and there is a clear reason for development of new vaccine candidates (like￾wise, BCG is not a sterilizing vaccine in animals). Any new candidate will have to be demonstrably better than the cur￾rent live BCG, in terms of efficacy and safety, the latter being a problem with BCG use, particularly in HIV posi￾tive neonates [7]. BCG is also a pre-exposure vaccine which does not stop infection, latent TB or reactivation, or guar￾antee sterilizing immunity and we may perhaps find better solutions using a post-exposure or therapeutic vaccine [8]. Perhaps the most difficult problem with TB or mycobac￾terial immunity is thatthe organismhides very successfully inside the macrophage or granuloma. For this reason, it is commonly thought that T-cell immunity is critical and that B-cells are of lesser importance [9–12]. Developing a vaccine to deliver immunity based on this premise is of course complex, unlike the well-knownsuccessfulhumoral response vaccines against viral diseases. 2. Prior or acquired immunity? Since the overwhelming majority of infected humans never develop symptomatic (TB) disease, one may con￾clude that they have inherent or innate immunity which cancope withthe infection, rendering themresistant.How￾ever, it is also clear that at least in some communities, BCG vaccination provides some protection, thus one may rea￾sonably conclude that vaccination can confer some degree of acquired immunity. One may thus postulate that in the absence of immunosuppression, most humans are inher￾ently resistant via innate immunity; some may become resistant with the correct stimulus, but that some may not be able to acquire immunity to TB. In the absence of a sound body of research and data concerning different species of animals, we cannot make statements with the same level of confidence for these species. However, we know that different mouse strains show varying levels of resistance or susceptibility to mycobacterial infection and we also know that many species appear to be quite suscep￾tible since TB is commonly reported in them, e.g. badgers, possums, cattle, buffalo, lions, lechwe, deer [13–16]. On the other hand, TB (or mycobacterial disease) has never or rarely been reported in many other species, such as dogs or horses. In some susceptible species, vaccination stud￾ies have been done, with some studies reporting various levels of protection, using BCG or new candidate vaccines [17–26]. However, it is not clear whether some animal strains or species or a proportion of any given species is resistant to TB, or whether a proportion can acquire immu￾nity under the appropriate conditions. 3. Animal models Various options exist, but the mouse model is still the most frequently used. Mice are relatively cheap and can be housed reasonably easily under containment for pathogen challenge. Many well-characterized strains of mice are available, and there is a variety of reagents and kits avail￾able for mouse research. Few if any specific reagents or lines exist for other animal models. However, there are some major differences between mice, other animals and humans. One notable difference is that mice do not form the classic granuloma seen in most other vertebrates with TB. For this and many other reasons, this model may there￾fore have major deficiencies. Noting this, some researchers have invested in non-human primates as a model sys￾tem [27–29], but this is an expensive and time consuming route, albeit perhaps self-evidently better than others. Ide￾ally, potential vaccines for a particular species should be tested in either the same animal, or at least a closely related species such as bovids for cattle or buffalo vaccines. Each existing model has its own problems and deficien￾cies, the details of which are beyond the scope of this paper to discuss in detail. However, a common limita￾tion is that many lab-based models are not exposed to the environmental stimuli experienced by free-living crea￾tures, whichinclude organisms suchas other bacteria,fungi and parasites, which alone may render their hosts defi- cient as models. In addition, most animal models are inbred strains. If we postulate that genetics is a major determinant of resistance or susceptibility [30–33], this alone makes such models tricky to understand, since the strain may be inherently resistant or susceptible. Even if not inbred, certain animal species may be innately resistant, where vaccination is unnecessary, or innately susceptible, being perhaps intractable to vaccination. This variation may of course also be found within an out-bred population, due to genetic variation. Recognizing the limitations in labo￾ratory based animal models and the importance of dealing with research close to the real problem, a number of groups have investigated larger animals such as deer or cattle for trial vaccination. These animals are arguably a good choice, since they are potentially susceptible, are at risk for disease, are economically important and impact on ecosystems. In addition, they present with large genetic and environ￾mental heterogeneity, which better reflects what we may expect in trials under real life conditions. For this reason, apart from laboratory-based animal models, a number of attempts to vaccinate free-living