Paratuberculosis control: a review with a focus on vaccination

Paratuberculosis poses a big challenge to Veterinary Medicine and in particular to ruminant production. Since the first description of the disease in 1895 in a cow from Oldenburg, Friesland, its etiological agent, Mycobacterium avium subsp. paratuberculosis (MAP), has been shown to cause the disease in the majority of wild and domestic ruminant species [1, 2]. This microbe is also present in many other hosts as well as the environment [3, 4]. Even though the most important mycobacterial infection in animals, bovine tuberculosis, has been successfully controlled in nearly all developed countries, the other important mycobacterial infection, paratuberculosis, remains an unsolved problem for the veterinary scientific community still incapable of reaching a consensus on the better way to deal with it. This is so despite large control efforts in different countries during the past three decades.

The mounting evidence showing that MAP is a factor in the pathogenesis of human inflammatory bowel disease (IBD) has increased the pressure to overcome this challenge. In spite of this, most of the undertakings are nevertheless based on the old principle that the only way to control an infectious disease is to eradicate its agent. This principle has worked well for some acute infections in times of survival struggle and profligate use of means but is increasingly difficult to apply because of demonstrated lack of efficacy and sustainability philosophy [5, 6]. We are no longer faced with a live or death dilemma due to infectious diseases, but we have to deal with a need to increase productivity for the sake of improved and prolonged use of scarce resources. From this perspective, it is necessary to simultaneously exploit the three classical main approaches to eradicate or reduce the impact of paratuberculosis in herds or flocks. These are: 1) to introduce management changes to decrease the transmission of MAP, 2) to apply test and cull practices to eliminate the sources of infection, 3) to vaccinate replacers in order to increase their resistance to infection. The advantages and drawbacks of these strategies will be briefly examined.

Pathogenic background

MAP distribution

If we take a general view of our knowledge on paratuberculosis, we should point out that MAP is not a classical infectious agent fully complying with Koch's postulates. Indeed, we know that many experimental infections fail to establish the infectious agent in the intestinal tissue and to cause the disease [30–33]. We also know that frequently the initial focal lesions do not progress to clinical stages. More recent evidence has revealed that it is not rare for herds with no clinical history of paratuberculosis and even with a history of negative fecal culture to occasionally show positive fecal culture results [34]. In addition, recent studies on paratuberculosis prevalence have revealed that as many as 60% of some national herds are actually infected [35]. Finally, Pickup and collaborators have shown that MAP is present in the environment at a previously unsuspected high frequency [4]. All this evidence indicates that MAP might be a necessary, but not a sufficient cause of paratuberculosis. Under these conditions, we should therefore ask ourselves: Is paratuberculosis eradication a realistic goal? Is it necessary? Is it profitable for the society in general? Answers to these questions are not readily available because we lack accurate information on the actual distribution of MAP and its potential impact on human health. Reviewing aspects of the pathogenesis and epidemiology may lay the grounds on which control alternative(s) to choose.

Forms of infection

Multiple forms of infection can be observed in MAP infected animals. The form present in an animal will not only depend on the progression of the infection or stage of the disease, but also on many other factors including an individual's genetic resistance or susceptibility to the pathogen, age at the time of infection, and previous exposure to other environmental mycobacteria. On Figure 1, we illustrate the balance between the infection and the animal's immune system and their corresponding forms of infection. According to different studies, about 46% of cattle, 51% of sheep, and 50% of goats in a MAP-contaminated environment do not show any signs of infection [36–38]. Since these animals live in a heavily contaminated environment, they must continuously be exposed to MAP, and, therefore, they either prevent the infection or very quickly clear up the establishment of local infection foci. Because it is not rare for such animals to carry MAP and plenty of experimental evidence has shown that administration of large amounts of MAP not always results in the development of a full blown infection, quite the opposite frequently produces very regressive lesions, the more likely explanation is that there is a balance between MAP and the host that in about half of the exposed individuals results in containing the infection (Figure 1). Beyond this balance point there are also different stages of infection. About 19% of cattle, 24% of sheep and 12% of goats carry an infection which is very focal and delimited. Around 17% and 9% of cattle and sheep, respectively, have multifocal forms. Of the animals presenting diffuse forms, approximately 19% of cattle, 16% of sheep and 38% of goats develop into diffuse forms which lead to animals showing clinical signs and to their death.

Vaccine types

Both live (non-attenuated and attenuated) and killed whole cell vaccines have been used against paratuberculosis. In a few cases, subunit vaccines consisting of sonicated bacteria, bacterial cell fractions or recombinant MAP antigens have been used but they have shown a much lower degree of protection [39, 40]. More recently, DNA vaccines, consisting of the inoculation of mammalian expression vectors containing MAP genes have also been used in mice, humans and sheep but not in cattle [41–47]. Most MAP vaccine formulations have been based on mycobacteria and a water-in-oil emulsion (olive, mineral, liquid, paraffin, etc). Some have also an irritant like pumice powder in order to increase and stimulate the local inflammatory response, and therefore enhance the immunogenicity of the vaccine. The goal of these vaccines is to establish a focus of inflammation where the antigens can permanently stimulate the host immune system. Under this principle, it would not be necessary to revaccinate animals because the slow liberation of antigens from the vaccination site keeps on stimulating the immune system, at least during the period before the age of initial clinical disease presentation.

Vaccination age

Paratuberculosis vaccines are recommended for exclusive use in very young animals on the grounds that this is necessary to prevent infection and to decrease interfering responses with the diagnosis of tuberculosis. Actually, the experience on animals older than 1 month is rather scarce, however recent studies on the pathogenesis as well as some field data suggest that vaccination of adult or subadult animals might have some management (no need for separate handling, vaccination of only replacers) and therapeutic (stronger humoral and cellular responses) advantages that need to be taken into account [48, 49]. More recent evidence form Australian sheep vaccination trials indicate that there might be an age threshold for vaccine efficacy that can be drawn at around 8 months of age [50].

Half of the studies are field trials where animals were naturally exposed to MAP. In these studies, results were assessed either by comparison between initial prevalence before vaccination, and final prevalence some time post-vaccination, or by following up a matched group within the same herd or flock. The later type of studies, when the control group is housed with vaccinated animals, frequently underestimates the positive effects of vaccination, because as herd immunity increases, bacterial shedding into the environment is reduced and thus the probability of a natural infection in the control group is also reduced. In three experiments the assessment was done using control unvaccinated herds, and one consisted of a questionnaire on clinical incidence in farms before and after using vaccination.

The vast majority of studies on all species showed positive reductions in all examined variables (Figure 2), that in cattle resulted in average reductions of 96.0%, 72.6% and 57.5% for production, epidemiological or pathogenetic effects, respectively. In sheep these reductions were of 67.5%, 76.4% and 89.7% and in goats of 45.1%, 79.3% and 94.8%, clearly demonstrating that MAP vaccination works well in all three species. The widest spread in reduction percentages, including several negative reduction rates, was observed with the epidemiological effects variable, which represents culture data. These differences are probably due to inherent aspects of each variable, since frequently the same study that gave negative reduction rates with the epidemiological variable, showed much better reduction results with the other variables, specially for the production effects variable. Most studies reported culture data as positive or negative result and did not include data on quantification of bacterial load in the sample. Thus, vaccinated animals with clinical signs reduction were still infected and excreted bacteria. This would imply that even though the amount of bacterial shedding might have been reduced, the proportion of shedding animals might have not. As a consequence, this would be in agreement with the widely accepted concept that, in general, current MAP vaccines can contain the infection and dramatically decrease clinical signs in a herd, but do not completely clear the infection.

Except for a few cases, vaccination in cattle was applied at early ages, in the first weeks of life, while in sheep more studies included adult sheep. The largest sample size studies, up to 150,000 animals, were done in cattle and preferentially recorded production effects in terms of paratuberculosis culling rates, since measurement of the other variables is much more time-consuming and costly. This is also reflected in the median sample size for the studies that looked at the production variable, 876, 700, and 4975 animals for cattle, sheep and goat studies, respectively, while studies analyzing the epidemiological or pathogenetic variables had median sizes around 100 or less.

The range of study length was between a few months and 16 years covering a period of 74 years. A large increase in sheep studies in the last decade coincided with the availability of the successful small ruminant commercial vaccine Gudair™ and its extended application in Australia and New Zealand. In the majority of the studies (68 experiments) killed vaccines were used. Most experiments used MAP strain 316F from Weybridge, nine used Strain 18 (now known to be M. avium subsp. avium rather than MAP [56]), and the rest used local isolates or subunit vaccines consisting of recombinant proteins. Not surprisingly, 316F is the most frequently used strain in sheep studies, since the above mentioned commercial vaccine for sheep and goats is based on this strain. Bacterial content varied widely, from 1000 CFU to 3 × 10⁹ CFU, and from 2.5 mg to 100 mg. The large majority of studies used some type of oily adjuvant (mineral oil, olive oil, liquid paraffin etc.) and in very few cases alum. In one study [57], AquaVax experimental vaccine was used, which contains no adjuvant but saline instead. More recent studies have started using other newer adjuvants such as MPLA, RIBI, cytokines, DDA, QS21, and lipid formulations, some of them with good results.

A wide variation in the efficacy of vaccines was observed, especially in cattle and sheep, where negative reductions are described in some studies. However, the overall results are pretty homogeneous, with very small error ranges due to the large numbers of observations included. In general, vaccine strain or administration route differences do not seem to substantially alter the outcome of vaccination, however, type of antigen formulation or adjuvant appears to have been important in a few experimental studies where different formulations were compared side by side [58].

The goal of this review was to evaluate vaccination as a whole, summarizing the results into a single table for each type of measure used to determine vaccine success. This analysis has revealed that, in average, vaccination has a positive effect. However, since the efficacy figures are rather poor in comparison to vaccines for other microorganisms, it is relevant to at least try to discuss possible reasons for some of the low success rates. In order to simplify, one possible approach is to find an explanation for studies where vaccination performed below the average. Production effect studies where the measurement was total mortality may be considered flawed because in most cases mortality did not differentiate between paratuberculosis and other pathologies, possibly diluting the "vaccine effect". This is evident in the case of sheep and goat trials were adults were considered. Since the more sensitive part of the population might have already died of paratuberculosis before vaccination the remaining animals could be considered more resistant to paratuberculosis, and therefore, less likely to show any effect of vaccine protection. Because young animal studies showed larger effects, this becomes a very likely explanation for low reduction rates. An additional explanation for the poor results could be the fact that, frequently, vaccination programs coincide with the initiation of other control measures making it difficult to assess the real effect of the vaccine on paratuberculosis control.

Under the epidemiological effects analyzed, reduction in the proportion of fecal shedders appears to be one of the measurements showing the widest variability. This happens mostly in small studies or in studies carried out in the Netherlands. Besides the qualitative effects in terms of protection conferred, vaccination should be assessed from another, perhaps even more important standpoint, such as is the reduction of amounts of bacteria shed by vaccinated and non-vaccinated animals.

When considering reduction in pathogenetic effects, it should be pointed out that some of the studies had a very short follow up and that the presence of focal lesions of paratuberculosis are weighted the same as the presence of large areas of affected intestine.

Since the vast majority of the studies show a positive effect, the question as to why vaccination has not been given more opportunities comes out with force. Especially, because for years MAP eradication efforts have only shown very moderate success or straight failure due to their enormous costs and frequent relapses of infected animals. Already in the eighties [59] and nineties [37, 60] several studies showed the profitability of vaccination. Over a period of a few years, the economic advantages of vaccination may be up 20 times higher than any testing and culling strategy which, in addition to yielding uncertain results, it results in a much higher economic cost. Other strategies based on certification are compatible with vaccination, and moreover, vaccination might allow a spectrum of other approaches to paratuberculosis control dependant on the financial resources of the farm, region or farmers association, and the actual economic losses sustained by the enterprise. It has been estimated that only a 5% annual clinical incidence of paratuberculosis will justify entering a mixed vaccination and testing and culling strategy [61].

In our opinion, there is a mixture of vested interests on control programs based in testing and culling, simplistic thinking comparing tuberculosis and paratuberculosis, fear of cross-reactions, academic detachment and confusion between ideal objectives and practical needs for the livestock industry. It is clear that the main reason for the opposition to MAP vaccination in cattle has been the problem of expected interference with the diagnosis of tuberculosis and its consequences in trade and national TB programs, however, the availability of an OIE official test -the comparative intradermal tuberculin test- that can very easily solve this problem in the majority of cases, should eliminate this concern on MAP vaccination in cattle. Recent field vaccination trials in cattle with an experimental MAP vaccine (Silirum™, CZV), have shown that less than 0.5% of vaccinated animals will give interference problems when the comparative intradermal tuberculin test is used even if the most restrictive interpretation of results proposed by the OIE is applied (Joseba Garrido, personal communications). The benefits obtained from production increases and reduction in clinical cases of MAP, will largely outweigh the small loss due to culling of these tuberculosis cross-reactive animals. In addition, new plans for the introduction of improved tuberculosis vaccines for cattle [62], will also affect the prospects of MAP vaccination in cattle, since the accompanying DIVA diagnostic test will probably allow for the identification of MAP infected or vaccinated animals.

MAP vaccination concerns in cattle have been further aggravated by the fear of the dairy industry to a crisis of confidence in their products, particularly, if a potential zoonotic link between paratuberculosis and a human disease (IBD/Crohn's disease) is confirmed [63] or if too much discussion and research efforts are focused on this subject. At this moment in which the paratuberculosis scientific community has finally accepted that the key to the paratuberculosis problem might not be eradication, but just control, vaccination offers the solution to this problem, since it not only allows to confine the paratuberculosis problem within the limits of a livestock production issue, while downright calming the worries of farmers, but also provides the perfect cover for doing something against paratuberculosis from a Public Health point of view, without incurring in massive costs. Vaccination might be the beginning of the end of the huge worldwide paratuberculosis problem and might mark the difference between doing nothing and advancing towards global control [64].

Paratuberculosis control poses a though challenge for farmers and veterinarians. Test and cull strategies can be useful in some settings but do not seem to have reached extensive success. Control by vaccination is an alternative that has been longtime in use in some regions and species. A substantial number of vaccination studies where objective information is amenable to meta-analysis treatment have been published in peer reviewed journals or in conference proceedings or other media. The high heterogeneity among reports makes it difficult to accept that the narrow statistical confidence interval obtained in these meta-analyses actually represents the true range of reduction in the whole set of trials. However, the results analyzed here clearly show a general positive effect from vaccination, negative effects only in a few trials, and a positive average balance according to all three variables considered (production, epidemiological or pathogenetic effects). In terms of quantitative reduction, the minimum is an 11% reduction in MAP isolation, which could be considered the worst case average, but with a common outcome at over 50% which is highly profitable from a production point of view. This strategy thus has high chances of have a effect on the overall environmental contamination with MAP, which would mean a significant advance in the fight against paratuberculosis, both in the animal and in the (potential) human public health fields.