5 курс / Пульмонология и фтизиатрия / Clinical_Tuberculosis_Friedman_Lloyd_N_,_Dedicoat

humans, particularly those drinking unpasteurized milk or milk products. Around dairy farms in the Debre Zeit areas, Holsteins were reported to have a prevalence of bovine tuberculosis infection of 55% whereas that in cross-bred cattle was found to be 23%. However, when kept under more intensive feed-lot conditions, tubercular zebu cattle can experience a depression in weight gain and up to a 60% morbidity rate55 demonstrating that under more intensive environmental conditions, zebu cattle have an increased predisposition to clinical symptoms of disease, in comparison to their counterparts reared in less intensive environments.

TRANSMISSION OF INFECTION BY

INGESTION

Milk

Prior to the advent of pasteurization, the main transmission route of M. bovis from cattle to humans, was via ingestion of infected milk. Bovine tuberculosis used to be an endemic disease of humans in the United Kingdom; however, currently, the total annual infection of humans with M. bovis is likely to be between 0.5% and 15% in all cases of tuberculosis confirmed in the United Kingdom, similar to the prevalence of M. bovis infection detected in humans in other industrialized countries. Sale of unpasteurized milk and dairy products is not illegal in England and Wales, as it is in Scotland, as long as it is clearly labelled as such and sold directly from the site where it was produced. It is possible for M. bovis infections to reactivate later on in life and currently the demographics of human M. bovis cases in the United Kingdom is most often the older generation who drank unpasteurized milk as children in the 1940s–1950s and doctors are seeing reactivation of the disease in this population at this late stage of their lives, following incomplete eradication of M. bovis post treatment.

Cheese and other milk products

It has been found that M. bovis can survive well in mature, unpasteurized cheeses as they are less likely than other bacteria to be affected by the potentially more extreme pH of the cheese.29 Not much work has been carried out to look at the impact of the cheese-making process on M. bovis but it has been confirmed that the storage and ripening of certain cheeses does lead to the full inactivation of the bacterium if it was originally present in the raw milk. Currently, there are no validated laboratory techniques that would certify a non-heat-treated dairy product as M. bovis “free”.

Meat and meat products

Every carcass slaughtered at a UK slaughterhouse is inspected postmortem. In reference to tuberculosis infection, particular attention is paid to any lesions in the lungs or lymph nodes, and therefore 59% of cattle with visible tuberculosis lesions but not displaying clinical signs are identified at this point.30 This inspection process complements the National Tuberculosis Testing Scheme, and is especially important in areas of the country where annual tuberculosis testing does not take place.

Occupational exposure of humans is an additional risk factor for contracting M. bovis infection. Time spent within close contact of infected animals offers the opportunity for inhalation of aerosolized bacteria from the lungs of an infected cow. Infection through broken skin, although rare, is also a possibility. It may be possible for a carcass to appear entirely normal on gross postmortem and yet contain hematogenously disseminated bacilli that are able to cause infection post consumption. Cross contamination of lesions may also occur during postmortem processing potentially introducing bacilli into the food chain during unhygienic carcass dressing and contamination of muscle surfaces.31 However, the risk of tuberculous infection from the ingestion of infected meat is minimal and far less efficient than inhalation. The higher doses of tuberculosis bacteria that may cause disease are only found in milk from infected cattle32 and would be almost impossible to find in skeletal muscle tissue with a lower mycobacterial load and no visible tuberculosis lesions. In developing countries where there is potential for undercooking of meat, the risk of infection by ingestion of meat may be slightly higher, especially in areas where meat hygiene inspection occurs only sporadically.33

Infection of other animal species

Wild animals have been implicated in the transmission of M. bovis to livestock, and potentially play a role in zoonotic transmission of the disease to humans. In the United Kingdom and Ireland, the badger (M. meles) is the principal wild animal infected. In other parts of Europe, goats (Capra hircus),40 species of deer, mostly the red deer (C. elaphus), fallow deer (Damam dama), roe deer (Capreolus capreolus), and wild boar41 are affected. In North America and Canada, bison (Bison bison),43 elk (C. canadensis)44, and white-tailed deer (Odocoileus virginianus)45 play an important role in the transmission of M. bovis. In Africa, many species that warrant local conservation are affected, which poses a threat to the survival of local populations.46 The brush-tailed possum (T. vulpecula) and ferret (Mustela furo) in New Zealand,35 the Cape buffalo (Syncerus caffer) in some parts of South Africa, the Kaufe Lechwe antelope (Kobus leche kafuensis), and the white-tailed deer (O. virginianus) in Michigan, USA47,48 are all considered to be maintenance hosts of M. bovis.

In non-maintenance or spillover hosts, or dead-end hosts,48 transmission of M. bovis is self-limiting as a maintenance host is required to be present within the ecosystem for this to occur. These spillover hosts may act as an incidental source of M. bovis for cattle.48 Examples of true dead-end hosts may be horses and sheep. Pigs, goats, farmed wild boar, dogs, cats and camelids in the United Kingdom should be treated as potential amplifier hosts as they have the potential to act as a source of infection for other animals and man.49

BADGERS (M. MELES)

Badgers are an important reservoir and maintenance host for M. bovis in the United Kingdom and Republic of Ireland and there are areas where the disease is endemic in the badger population.20 A greater number of male badgers are infected than female badgers, likely due to the males increased roaming behavior and tendency toward aggression allowing transmission of the disease via bite wounds. However, the majority of infections appear to be via

the respiratory route, with open submandibular abscesses often being the first clinical sign noted. The communal social structure of the badger population, with social grooming and sleeping in groups, provides an excellent environment in which M. bovis can be spread. A large proportion of infected badgers will survive for more than 12 months after the initial infection, and mortality induced by M. bovis infection appears to have only a small part to play in badger population numbers.

Badgers with advanced M. bovis infection shed the bacilli in their urine and this is thought to be the primary infection source for cattle. In some populations where there are recently infected cattle herds, up to 50% infection rate of badgers has been recorded, and the risk of transmission increases as badger density increases. Behavioral studies to determine the degree of interaction between badgers and cattle on pasture has found this to be minimal, and the badgers preferred not to occupy fields occupied by cows and did not approach within 10–15 m of a cow.34 The conclusion was that, rather than direct aerosol transmission between badgers and cattle, if disease spread between badgers and cattle does occur, it is likely through contact with or ingestion of badger urine or feces. The risk of infection to humans directly from badgers is thought to be very low, because close contact between humans and badgers is limited.35 The risk to those humans working more directly with badgers, such as government officials involved in collecting road carcasses, wildlife center employees, veterinary surgeons, farmers,­ and residents of rural areas has been found, unsurprisingly, to be higher.

POSSUMS (TRICHOSURUS VULPECULA)

Brush-tailed possums are indigenous to Australia but have colonized New Zealand and provide a significant reservoir of M. bovis for infection of cattle and man in this country, but not in Australia. The first tuberculous lesion detected in a possum was noted in 1967 by a possum trapper. Subsequently, endemic populations of M. bovis- infected possums was found in all areas where tuberculosis from cattle was difficult to eradicate. Transmission between possums is either by aerosol or pseudo-vertical (via the joey) routes. Lesions in possums can be widely disseminated, and the disease is normally fatal after 2–3 months from the onset of clinical disease. Breakdown of infection in previously susceptible cattle herds can occur in the presence of only one infected possum. A cross-sectional survey of possums in New Zealand in 1994 showed a local prevalence of M. bovis infection of up to 20% in clustered areas, but the overall prevalence of infection of possums in New Zealand was 1.2%.36

DEER

Farmed and wild deer are susceptible to infection with M. bovis.37 The disease was first reported in farmed deer in New Zealand and currently tuberculosis is considered to be the most important bacterial disease in farmed deer in New Zealand and the United Kingdom.38 It has been suggested that farmed deer are more susceptible to the disease than wild deer. Griffin found that the immune response of red deer to infection varies,39 and that there was a high heritability of resistance to M. bovis infection in red deer under experimental conditions. Susceptibility to disease varies depending on genotype, previous exposure to disease, nutritional status of the herd and level of sex hormones.38 Pathogenesis of the disease varies with the site of infection. Wild deer or farmed

deer in contact with cattle that are susceptible to disease have the potential to act as a reservoir of infection for cattle. There is the possibility that captive, farmed deer can act as maintenance hosts of M. bovis, and a tuberculosis control programme has been sporadically introduced in the United Kingdom and New Zealand among farmed deer populations to help prevent this.117,118

SMALL RUMINANTS (SHEEP AND GOATS)

Goats are much more susceptible to infection with M. bovis than sheep. However, there is minimal epidemiological significance of goats acting as a reservoir of M. bovis disease.

PIGS

The disease incidence in pigs normally reflects that of the cattle population. The first historical report was made in 1969 by Myers and Steele that in 1921 in the United States 12% of pigs which were slaughtered under Federal Inspection Law were found to have tuberculous lesions. The origin of these pigs was traced and it was found that all of the infected pigs had been fed unpasteurized milk or dairy products, and were kept together with cattle. In fact, this is the most common route of infection in pigs. As with other species, the prevalence of disease in pigs is found to increase with age, and as most intensively farmed pigs are slaughtered for meat before the age of 6 months therefore M. bovis disease does not manifest as too severe a clinical problem in this species. Transmission between pigs has been found to be insignificant and lesions are normally localized. Pulmonary lesions are infrequent, therefore pigs represent an insignificant infection source for cattle.

GOATS

Goats are fairly susceptible to TB and most commonly develop either pulmonary infection or tuberculous mastitis. Bacilli have the potential to be shed in the milk of lactating goats.15 Reports of clinical cases of tuberculosis have been reported in goats in Spain.50 In Spain, goats are considered to be a maintenance host as are cattle. In Germany between 1999 and 2001, a goat-adapted strain, M. bovis subsp caprae, was responsible for a third of all zoonotic tuberculosis cases reported.51

CATS AND DOGS

The incidence of clinical tuberculosis caused by M. bovis in dogs and cats has decreased over the last 100 years, most likely due to the ingestion of milk or offal from tubercular cattle on farms, or from living in close proximity to humans infected with tuberculosis and inhaling sputum particles. Tuberculosis in cats has documented to be caused mainly by M. bovis and M. microti9 with fewer cases of M. avium and non-tuberculous mycobacteria.119 In dogs, M. tuberculosis was found to cause 75% of cases; the remainder was mostly M. bovis and a small proportion of M. avium.52

OTHER MYCOBACTERIA

Members of the M. tuberculosis complex include the following mycobacteria: M. tuberculosis, Mycobacterium africanum, Mycobacterium canetti, M. bovis, M. microti, Mycobacterium orygis, M. caprae, M. pinnipedii, and the recently recognized

Mycobacterium mungi.58 WHO reported that there were 9 million new cases and 1.5 million deaths due to these mycobacteria in 2014.

Non-tuberculous mycobacterial (NTM) infections are present worldwide and are found in many different environmental niches. They are harmless to most individuals and rarely cause human disease. Species of NTM associated with human disease include M. avium, Mycobacterium intracellulare, Mycobacterium kansasii, Mycobacterium fortuitum, Mycobacterium chelonae, Mycobacterium szulgai, M. paratuberculosis, and Mycobacterium scrofulaceum. The majority of NTM infections have been reported in countries in which tuberculosis is not endemic, as the chances of missing NTM infection are higher in countries where tuberculosis is already present. Currently, the individual mycobacteria causing mycobacterial infection are not routinely characterized worldwide. Therefore, some NTM cases in humans with positive Ziehl−Neelsen (ZN) stains will be misclassified as MTB.59 This may result in a patient receiving routine treatment for M. tuberculosis for which the NTM strain may be resistant. Additionally, occasional mixed strain infections of NTM and MTB infections have been reported.60,61

A brief epidemiology of several other mycobacteria are described below:

●●M. tuberculosis: This mycobacterium classically causes disease in humans and is the most common agent to cause tuberculosis. However, although animal infection is uncommon, it has been described among some species (birds, elephants, and other mammals) that have had prolonged and close contact with humans. Transmission of M. tuberculosis between animals and humans has not been reported.

●●M. microti: The name of this serovar of tuberculosis translates directly as “vole,” as it was the first mycobacterium discovered to cause disease in voles during investigations into the cause of cyclic population density changes in this species. Field voles, bank voles, wood mice, and shrews are all susceptible to infection with M. microti. M. microti has also been documented to cause disease in cats62 and other larger animals63 and very occasional reports of human infection are recorded, totalling 13 patients worldwide.64–67

●●Mycobacterium avium complex (MAC): MAC, which includes

M.avium, M. intracellulare and Mycobacterium chimaera, is the most important cause of NTM-related pulmonary disease in humans.68,69 MAC are widely distributed in the environment and rarely cause human disease. Factors such as host susceptibility, pathogen virulence, and environmental risk factors likely play a role in the pathogenesis of the disease. Often patients with MAC also suffer from concurrent immunologic or genetic disorders that predispose to lung infections or bronchiectasis. However, MAC disease can be seen in some patients with no other lung or immune abnormalities.70

●●M. avium: This serovar of tuberculosis causes disease in birds, pigs, cats, humans, and dogs71 and is the most frequently isolated NTM mycobacteria in humans. The clinical signs produced are often indistinguishable from that of the

M.tuberculosis complex and therefore it is often considered to be within the same group.

●●M. avium paratuberculosis (MAP): This mycobacterial infection causes a chronic granulomatous infection of the gastrointestinal tract of domestic ruminants and wildlife worldwide, and is the NTM of greatest importance in the field of

veterinary medicine,72particularly for cattle and sheep. MAP is extremely resistant to destruction and can survive pasteurization, and for up to a year in the environment. Transmission of the disease may occur horizontally via the fecal−oral route, and vertically from infected dams to calves, and the incubation period varies between 2 and 7 years.73 The most common route of introduction of infection into a previously MAP-free herd is via the introduction of previously infected cattle into the herd, or breaks in on-farm biosecurity.

Infection with M. avium paratuberculosis (MAP) is also known as Johne’s disease and is found worldwide. Clinical symptoms of the condition can be attributed to the presence of chronic granulomatous inflammation within the gastrointestinal tract; with diarrhea and weight loss occuring in cattle in advanced stages of MAP infection. Most infected cattle will be in the subclinical stage of disease; less than 5% of the cattle display clinical signs of illness. Fecal culture and antibody detection are the most common diagnostic methods to determine prevalence,74 although there is a decrease in sensitivity of tests for those animals with subclinical disease. In the United States, it was reported in 2009 that up to 68% of the dairy herds contained clinically infected cattle, with the prevalence lower in beef herds. It has been estimated that for every cow showing clinical signs of advanced Johne’s disease, it is likely that 15–25 others in the herd are also infected. Once clinical signs occur there is no known cure for the disease and culling presents the most economic option.

In sheep, the disease is characterized by emaciation, but not chronic diarrhea as in cattle. In many infected flocks, ewe mortality can reach as high as 5%–10% annually due to Johne’s disease.

Although M. avium subsp. paratuberculosis has not been confirmed as a zoonotic agent, there has been suspicion that this bacilli may play some role in the development of Crohn’s disease in humans and one meta-analysis has demonstrated a positive association between the two.75

●●M. caprae: This zoonotic mycobacteria was first characterized in 1999. The major route of human acquisition appears to be from livestock, mainly goats or cattle. No clear evidence of human to human transmission has been reported. M. caprae has only been found in 0.3% of all tuberculosis cases, and its range is almost entirely confined to Europe. The low incidence is thought to be due to the European-wide introduction of public health measures to prevent zoonotic TB transmission, including milk pasteurization and culling infected cattle. Incidence of M. caprae infection is highest in Spain, where this Mycobacteria represents 7.4% of all M. tuberculosis complex isolates from animals.76

●●M. leprae: This bacterium is the etiological agent of leprosy in humans, and gives rise to a chronic granulomatous disease primarily affecting the skin and peripheral nerves. The bacterium can be grown in several species of experimental animals, including the armadillo, nonhuman primates, and rodents. Naturally occurring leprosy has been reported in wild nine-banded armadillos (Dasypus troglodytes) and three species of nonhuman primates (chimpanzees (Pan troglodytes), sooty mangabey monkeys (Cercocebus atys) and cynomolgus macaques (Macaca fascicularis)), therefore qualifying leprosy as a zoonosis.

●●M. lepraemurium: This causes a leprosy-like disease affecting rats and mice, primarily affecting the viscera and skin and very rarely the peripheral nerves. Naturally acquired murine leprosy has been observed in rats, mice, cats and dogs, but not in humans or any other species. Therefore, it appears that murine leprosy is not a zoonosis.77

DIAGNOSTIC TESTS

In humans, the clinical signs observed due to infection by M. tuberculosis or M. bovis are indistinguishable radiographically, pathologically or by direct smear analysis. Prior to 1970, UK laboratories did not make an attempt to distinguish the strains,4 a practice that currently continues in some European countries and, until recently, the United States.78 This is not of grave concern clinically in humans as the treatment of most mycobacterial organisms remains the same. However, M. bovis does have an intrinsic resistance to the drug pyrazinamide, so the use of this drug is not recommended when cases of zoonotic tuberculosis infection are suspected.3 Epidemiologically it would be extremely useful to be aware of the exact cause and the potential transmission route of the clinical disease. Therefore, accessible and cost-effective tests to distinguish these strains of bacilli would be extremely useful. Currently, in order to achieve an accurate diagnosis between strain cultures, biochemical tests and DNA analysis are necessary9 are time-consuming and not cost-effective. Many reports have been made of misdiagnosis of tuberculosis in human patients.79

The accurate use of molecular markers to differentiate M. bovis and M. tuberculosis is also difficult. Organisms of the MTBC complex have been found to have a 99.9% similarity rate between nucleotide levels, and identical 16rRNA sequences.11

Due to the zoonotic potential of some mycobacterial infections affecting animals, care should be taken when handling the potentially affected tissue, and awareness to all involved should be made of the potential zoonotic risk of disease. M. bovis is classified as a Hazard Class 3 biological agent, according to the Control of Substances Hazardous to Health (COSHH) Regulations 1999. Tuberculosis is a notifiable disease in any animal, so if the disease is suspected and the legislation of the country in which the diagnosis is made stipulates, care should be taken that the correct procedures are followed to inform the relevant authorities. Aseptic practice and an aerosol mask when handling samples is normally adequate to reduce the risk of transmission via aerosol or ingestion, and attention should be drawn to wear gloves when handling the biopsy site or biopsy material. The body of any confirmed, euthanized case should be cremated, not buried, so that any infective bacilli present in the carcass can be destroyed.

Diagnostic tests of cattle

Single intradermal comparative cervical tuberculin test

The intradermal skin test is the first-line test of routine surveillance to be carried out in the field in the United Kingdom. In areas

of the United Kingdom where tuberculosis has been discovered in cattle, regular testing is carried out at a predetermined frequency until such a time as eradication from the herd has occurred, if possible.

The intradermal skin test involves subcutaneous injection in the skin of the mid-cervical region of cattle using an antigenic M. bovis purified protein derivative (PPD) mixture, “tuberculin,” which is sourced from heat-killed bacteria.80 Robert Koch was the first to purify this PPD solution in 18823 and bovine PPD remains to this day the most commonly used agent in the field to diagnose bovine tuberculosis. A delayed-type hypersensitivity inflammatory response occurs following injection, causing skin swelling at the inoculation site. Measurement of the diameter of this swelling 72 hours after inoculation correlates with previously determined values; the larger the diameter of the skin swelling, the greater the delayed-type hypersensitivity reaction and therefore the more likely that the cow has previously reacted immunogenically to this stimulus, therefore, the higher the likelihood that a cell-mediated response to M. bovis has already previously occurred.

M. bovis (PPD) antigenic compounds are shared by many other mycobacteria,81 including members of the M. avium intracellulare complex.82 Therefore, M. avium antigens are injected as a control agent, separately to either the mid-cervical region, at the same time as the M. bovis antigen injection, making the intradermal skin test comparative. These antigens are often found in the environment, and may confound the outcome of the M. bovis diagnostic tests.83 Comparing the difference in the diameter of the skin reactions between the M. bovis and M. avium antigens determines the difference in size of the delayed-type hypersensitivity and therefore allows for the discrimination to be made whether the cow is infected with M. bovis, or solely sensitized to environmental mycobacteria.

Established standards differentiate the diameter of the skin reaction, so the cattle are classified with the largest diameter of M. bovis skin reaction as “reactors,” medium sized as “inconclusive,” or “negative” for those that match or have a smaller diameter than the control, M. avium.

The sensitivity of the intradermal skin test has been reported to be 80%.84 Although some animals may present with physical signs of tuberculosis infection or postmortem changes, they may not respond to the intradermal skin test85 due to the presence of advanced disease, very initial disease, i.e., prior to 6 weeks postinfection, testing of a cow within 6 weeks of calving, or the administration of a variable dose using a multidrug syringe. Therefore, currently in the United Kingdom, tissue samples are still submitted for culture at postmortem from cattle with a negative single intradermal comparative cervical tuberculin test (SICCT) that have shown physical clinical signs of disease or have suspicious postmortem findings.

Alternatively, specificity of the intradermal skin test can also be considered to be on the lower side, with false positives recorded with evidence of a delayed-type hypersensitivity reaction and no visible lesions discovered on postmortem examination.85 However, as a public health measure, under Annex B of European Directive 64/432/EEC, all skin test reactors are considered to be affected by tuberculosis and are therefore slaughtered, regardless of the bacterial culture results.

An increased frequency of the SICCT test affects test performance. In the United Kingdom, tests are not validated if they are carried out more frequently than 30 days apart. The antigens injected during the test may modulate the cell-mediated immunity of the cow and depress the response to subsequent skin tests.86 Therefore, should disease confirmation be required within this time period, an alternative test should be performed.

Interferon-gamma test

The interferon-gamma test was first documented and initially used in Australia during the Brucellosis and Tuberculosis Eradication Program.87,88 This ELISA (enzyme-linked immunosorbent assay) detected cell-mediated resistance of interferon-gamma to tuberculosis using whole blood following incubation with PPD-b.89 Individual sensitivity of this test is higher than the tuberculin skin test described previously, which is felt to be superior to detect an infected herd. With a higher sensitivity, the interferon-gamma test is thought to be better served to more accurately detect infected individuals within herds,89 and if there is detection of the cytokine interferon-gamma, a preferentially positive result is indicated.90 The SICCT and inter- feron-gamma tests detect infection at different stages of disease. The tuberculin skin test is more widely used as a first-line detection test in cattle in the United Kingdom, one other benefit of this test being that the results of the test are quick, more cost-effective to gather and do not require a well-equipped laboratory to generate results.

Unfortunately, a negative interferon-gamma reaction and absence of visible lesions on postmortem examination with a positive intradermal skin test does not confirm that the cow is free of infection.

Postmortem examination

All carcasses following slaughter in the United Kingdom are routinely grossly inspected for the presence of visible tuberculosis lesions. Routine postmortem examination detects approximately 9%–12% of newly diagnosed tuberculosis cases in the United Kingdom annually, with the remainder detected on antemortem testing. Between 1998 and 2006, 30%–53% of slaughterhouse cases reported as suspected tuberculosis to the Meat Hygiene Service have yielded a final diagnosis of M. bovis.

Following slaughter, all confirmed tuberculosis reactors are submitted for postmortem examination with the aim not just to validate the antemortem testing, but to assess the severity of disease and provide epidemiological information about the nature of the bacilli present. Genotyping tests are carried out on the tissues to determine the latest strain responsible for tuberculosis herd breakdown. In the United Kingdom, postmortem examination of cattle is carried out at The Veterinary Laboratories Agencies, which are government-run institutions serving the country. The isolation rate of tuberculosis bacilli from reactors with visible lesions is normally approximately 90%, although gross lesions on postmortem cannot be considered confirmatory of disease.91 Postmortem testing determines the risk of infection to other animals within the same herd as well as to human contact, and following on from this to ascertain the number of follow-up tests likely required to restore the herd’s tuberculosis-free status, as

well as to what degree contact animals and humans need to be monitored for the development of the disease in the future.

Where cases of tuberculosis have been confirmed by antemortem diagnosis but no visible lesions are present or grow on postmortem examination, the Animal and Plant Health Agency (APHA) will carry out histological diagnosis to provide a presumptive answer within 2 weeks.

Laboratory culture

Definitive diagnosis of M. bovis can be made microbiologically from tissue from the lesion.30 Samples suitable to submit for culture include sputum, pleural fluid, or prepared tissues, for example, a lung or lymph node biopsy. Selective laboratory culture using an acid-fast stain such as ZN can provide confirmation of this slow-growing bacteria after 6–8 weeks of incubation at 37°C. Should acid-fast organisms grow that do not show the typical growth pattern of M. bovis, a multiplex PCR (polymerase chain reaction) test is carried out to help in the differentiation of MAC or other organisms of the Mycobacterium genus. This primary technique has currently replaced bio-typing in the United Kingdom for tuberculosis identification.

Bacterial culture methods are not currently found to be highly sensitive, although they are very specific. Sensitivity is particularly lost when pooled samples of lymph nodes from a suspected cow with no visible lesions are submitted.30 European Directive 64/432/ EEC, Section 1, Annex B, describes techniques to identify M. bovis from clinical and biological specimens but does not provide specific directives to diagnose and confirm M. bovis infections in cattle, although methods used may conform to those specified in Chapter 2.3.3 (Bovine TB of the World Organisation for Animal Health [OIE] Manual of Standards for Diagnostic Tests and Vaccines).

Genetic work

In the United Kingdom, the APHA routinely determines the genotype of tuberculosis isolates from cattle in order to provide more information about epidemiology of this disease across the country while monitoring for new tuberculosis herd breakdowns and their spread. Spoligotyping, a technique to analyze polymorphisms in various repeats of DNA sequences, was the principal technique used for molecular typing in the 1990s and earlier 2000s.92,93 In 1998, Frothingham94 provided a method to analyze the highly polymorphic DNA loci of M. bovis to analyze finer discrimination between strains, which is used in tandem with spoligotyping by the APHA. The main feature of this is the ability to differentiate the isolates of the most prevalent and widespread tuberculosis strain, SB0140, found in cattle and badgers in the United Kingdom.95 The APHA has amassed spoligotyping data from over 31,000 M. bovis strains detected in the United Kingdom between 1997 and 2004 from cattle, badgers, deer, and other mammals (APHA, unpublished). Of these, 6500 isolates also have variable number tandem repeat (VNTR) data, and therefore comprise the largest collection of M. bovis genotype isolates within Europe. This epidemiologic resource has helped scientists to investigate the spread and incidence of M. bovis isolates across the United Kingdom over this period, and the evolution of the MTBC complex.11 VNTR typing

was used to good effect in the 13-year outbreak of bovine tuberculosis in Switzerland, which occurred after a 15-year period of time during which no bovine tuberculosis was diagnosed in the country. The VNTR data found that the 17 isolates tested from outbreaks across the country were identical, caused by the M. bovis spoligotype SB0120, indicating a single source of infection.97 Comparison of one previously archived sample of bovine tuberculosis from infection in Switzerland 15 years earlier using MIRUVNTR testing showed an identical VNTR profile, insinuating that the infectious agent had persisted in the dairy herd for nearly 15 years prior to reactivation.

Several candidate genes are being assessed for their use as genetic markers of M. bovis. Once these genes are discovered, the aim is to assess whether they have the potential to alter the disease phenotype of cattle. One marker gene includes the bovine natural resistance-associated macrophage protein gene (NRAMP1), which has been ubiquitously identified in mice models and humans. The survival of M. bovis BCG appears to be linked to the presence of a resistant allele of this gene.98 Small comparative epidemiologic studies have been carried out to examine plausible genes within cattle populations, however these are normally scientifically underpowered to determine significant associations.

In the future, it is likely that a whole genome approach will be used to investigate the single-nucleotide polymorphism (SNP) numbers and determine the degree of disease-associated variation. A whole genome sequencing approach will determine the presence of particular disease-associated variants and in humans has the potential to identify the genotype present in cases of drug-resistant tuberculosis and mechanisms that influence the phenotype. This initial whole genome approach will identify the most polymorphic areas of the genome, and the chromosome that corresponds to the genetic information of the phenotype of interest. Potentially, findings show that a specific allelic combination occurs more commonly in the diseased population. The linkage disequilibrium, or occurrence in members of a population of linked genes in nonrandom proportions, of M. bovis infections in cattle appears to be much smaller than that of M. tuberculosis affecting humans, likely due to the occurrence of nonrandom mating in cattle and their smaller population size,99,100 which acts to reduce the overall number of SNPs. There are currently 800,000 SNPs in current bovine genome SNP arrays, and this may be sufficient to effectively map the entirety of the bovine genome.

Diagnostic tests in dogs and cats

As previously mentioned, the potential for zoonotic transmission of mycobacterial infections from dogs and cats is a concern. Should a dog and cat infected with a zoonotic strain be detected but the threat to human health be found to be minimal, diagnostic tests in dogs and cats can be carried out with a slightly different focus to that of cattle and other herd animals where containment of the spread of this zoonotic disease within the herd is of extreme importance.

Diagnosis of infection with the Mycobacterium tuberculosis complex relies on a combination of histopathology, mycobacterial culture and molecular testing. Suggestive historical and clinical findings will prompt testing for the infection.

The dog or cat should be thoroughly evaluated to determine the location of infection and severity of the disease, extent of systemic involvement and local infection, in order to determine the likely prognosis for this specific animal. The most common findings on physical examination of cats infected with M. bovis and M. microti are single or multiple cutaneous nodular lesions. Multifocal peripheral lymphadenopathy may be present. The results of serum biochemistry and hematology are often non-specific. Biochemical and hematological tests, if performed, will normally identify nonspecific changes. The presence of ionized hypercalcemia is a poor prognostic indicator101 as it suggests a higher burden of granulomatous disease.

Radiography can be used to assess if there is respiratory involvement. Changes may be variable and potentially not be specific or diagnostic for the disease. They may include tracheobronchial lymphadenopathy, interstitial to alveolar lung infiltration, lobar consolidation, and pleural effusion. Abdominal radiography or ultrasound may demonstrate hepatomegaly or splenomegaly, lymphadenopathy, mineralization of the mesenteric lymph nodes, or ascites. Assessment of bones by radiography may also reveal areas of bony lysis and sclerosis, osteoarthritis, discospondylitis, or periosteitis.

Specific tests that can be undertaken are similar to those available for other animals, and include the interferon-gamma test, which is available from the APHA in the United Kingdom.103 Intradermal skin testing results are often unreliable in cats, as they do not tend to show a strong reaction.104,105 False-negative results can occur in both cats and dogs.

To generate confirmation that mycobacteria are involved, fineneedle aspiration or biopsy samples of affected tissues should be obtained and stained with ZN stain or similar. On microscopy of stained tissue, the number of acid-fast organisms present within macrophages may vary and depend on the specific mycobacteria present, the location of the granuloma, and the nature and strength of the host immune response. A mixed population of often heavily vacuolated histiocytes and smaller numbers of degenerate or nondegenerate neutrophils and small lymphocytes may also be seen. Once the organisms have been viewed microscopically, it is important to perform a culture as the gold-stan- dard method to determine the mycobacterial species involved and establish the potential zoonotic risk of the infection, the source of the infection, and options available for treatment. However, often tissues in which acid-fast staining organisms are seen may fail to grow mycobacteria in the laboratory, and culture is continued for as long as 8–12 weeks as the organisms grow very slowly, to maximize the opportunity for diagnosis.

Other diagnostic techniques available include molecular PCR, which is particularly useful when bacteriological culture results are not available,106–109 mycobacterial interspersed repetitive unit- variable-number tandem repeat (MIRU-VNTR)120 and spoligotyping. Using this technique, bacteria can be identified within tissue aspirates or buffy coat preparations separated from the blood.

Histopathology

There are several presentations of tuberculous masses that may be viewed on biopsy or postmortem examination. Large, solid, tumor-like masses or multiple small disseminated masses

throughout the body may lead to a similar diagnosis. Often the gross appearance of the mass is a grayish-white color, with haemorrhagic edges and a soft purulent center due the presence of caseous necrosis. Lesions in the lungs are often a grayish red color, and may be associated with serosanguinous pleural fluid. Renal lesions typically occur as infarcts within the cortex, and intestinal lesions include ulceration of Peyer’s patches, with the presence of submucosal tubercles.

Histopathological reports for the disease will often report the presence of foamy macrophages and granulomatous inflammation, some necrotic areas, and a variable number of acid-fast bacilli within and outside macrophages.107 Lymphocytes and fibroblasts may also be present, but multinucleate giant cells are often absent, or rarely seen.105,101

Calcification may be seen within larger tubercles, potentially surrounded by zones of histiocytic cells and a well-defined fibrous capsule.105

Feline leprosy may generate either lepromatous or tuberculous lesions, while a diagnosis of tuberculosis may generate only tuberculous lesions.107

TREATMENT

Cattle

M. bovis infections detected in cattle are not suitable for treatment due to the zoonotic and infectious nature of the disease. The public health and risk of infection to other herd members is too great and the treatment period is too long and uneconomic to make treatment of cattle a viable option. The treatment period required to effectively cure a cow would be 4–12 months of daily dosing, with the potential for relapse of infection once treatment is ceased. An additional problem that is posed by the concept of treating cattle is the potential for drug residues to be left in milk, meat and other products consumed by humans. Given that human drug resistance to standard tuberculosis treatment is a serious concern, the use of large proportions of the same drugs in cattle may serve to exacerbate this problem, and increase the degree of multidrugresistant infections among the human population.

Currently, however, drug-resistant isolates of M. bovis have rarely been reported. Due to the infrequency of treatment of the disease in cattle, the selective pressure for resistance is likely to be low and so apart from the innate resistance of this mycobacteria to pyrazinamide, it would be expected currently that most M. bovis infections be susceptible to routine mycobacterial treatment, if required.

Companion animals

Consideration can be given to treatment of companion animals under the guidance of the treating veterinary surgeon. The potential for dissemination of zoonotic bacteria is lower in these species than in production animals, as generally companion animals are housed in much smaller groups, and can be contained indoors, with less opportunity for dissemination of infection among numerous animals as would occur in a herd situation. However,

regardless of this, the zoonotic potential of the infection should be carefully considered, and public health authorities should be notified if a diagnosis of MTBC infection is made in a cat or dog.

Prior to the onset of treatment of a confirmed case of a mycobacterial infection in a dog or a cat, several factors should be taken into consideration with all of the human members of the household where the animal resides.

i.In cases where there is any human individual suffering any degree of immunocompromise within a household (e.g., the recipient of an organ transplant or an individual undergoing chemotherapy treatment) euthanasia of the dog or cat, rather than treatment, would be advised to limit the risk of harm to the human.

ii.The nature of the symptoms of disease displayed by the dog or cat. If the disease appears to be generalized, respiratory tract involvement is present, or there are cutaneous lesions that are continually draining fluid, the risk of zoonotic transmission is increased, and euthanasia rather than treatment should be considered.

iii.The owners of suitable candidates for treatment should understand that the treatment is long term and a challenge to maintain should the patient be noncompliant. The drugs used to treat tuberculosis have some degree of inherent toxicity and the financial cost of the ongoing drug treatment and monitoring can be very high. The owner should also be made aware that while some drug therapies may act to suppress the tuberculosis infection, eradication may not take place, and once treatment finishes the infection may recrudesce. Therefore, treatment for an indefinite period of time may be required.

However, having considered these points, treatment of cutaneous uncomplicated forms of tuberculosis in dogs and cats does often offer a good prognosis. Treatment protocols normally involve an initial and continuation phase of treatment with antibiotics. Currently, the initial recommendation would be treatment with three drugs for 2 months, followed by a continuation phase of two drugs for at least 4 months. The exact regime will depend on the type and severity of disease. Although antimicrobial susceptibility testing can be carried out on affected tissues, it has been found that the sensitivity results in vitro do not always match those in vivo. Surgical excision of affected tissues can be carried out to submit for analysis and further culture work, but again, the use of these tissue specimens for an accurate culture and sensitivity result may not always provide a successful result. There is a risk of wound dehiscence and local recurrence of infection should attempts be made to surgically remove large portions of infected tissue. However for the local cutaneous signs of disease surgical excision could be considered.

Culture and sensitivity can take up to 6 weeks or more to generate a result, and in the interim time, treatment with a fluoroquinolone antibiotic has been advised, although consideration should be offered to the reported side effects of treatment with this drug to cats. Currently, this route of treatment is only advised in cases of localized cutaneous infection and generally it is more sensible to institute the recommended double or triple therapy once a diagnosis has been made as this therapy offers the best chance

of clinical resolution. Treatment with a single fluoroquinolone increases the opportunity for the bacteria to develop resistance to this single antibiotic. If triple or double therapy is initiated prior to culture results being known, it is strongly advised to continue the same treatment once the diagnosis of a mycobacterial infection has been made. Ideally, the exact mycobacteria should be known prior to treatment. In many cases, culture results of suspected M. bovis cases in cats are negative, but ZN positive stains may be seen on cytology or histopathology. In these cases where a very high suspicion of infection with tuberculosis exists, the owners should be counselled about the risks and complications of treatment prior to initiation.

In cats, treatment with three oral tablets daily may present a compliance issue and the owner may struggle with this regimen for a protracted period of time. Should this be the case, treatment should continue with two drugs alone, normally for a longer period of 6–9 months.64 The most effective combination for treatment in cats is considered to be a combination of rifampicin, isoniazid, and ethambutol. There are some newer, less toxic drugs available that may be worth considering. The fluoroquinolones have the potential to treat infection, e.g., marbofloxacin and clarithromycin can be used effectively to treat tuberculosis in animals, especially in combination with rifampicin. Clinical experience denotes the most effective treatment regime to be an initial phase of treatment with rifampicin−fluoroquinolone combination and clarithromycin or azithromycin, followed by continuation of the treatment with rifampicin and a fluoroquinolone or an azithromycin or clarithromycin alternative. To ease administration, medication can be administered as a liquid in a single syringe, or all three tablets together in a gelatin capsule. Should resistance to drug therapy develop, a rifampicin, isoniazid, ethambutol combination may be considered. There is natural resistance to the treatment of M. bovis with pyrazinamide, as previously mentioned.

The prognosis following treatment of infection in dogs and cats depends on the mycobacteria involved and the extent and severity of infection. M. microti has often shown a favorable response to treatment. Many cases can obtain a long-term remission of infection, although response to treatment should be guarded.

CONTROL

General principles of control

From a pathogenic and economic perspective, of all the mycobacteria that can infect animals, the most important to control is M. bovis. An important consideration is the route of disease transmission, which in the case of M. bovis can be either by aerosol or ingestion, and the linkages and interfaces by which effective transmission of the mycobacteria is achieved.

Control of the disease in cattle

Cattle are considered the maintenance hosts of M. bovis, able to maintain the infection within their own population. On-farm control of M. bovis in cattle is extremely important. In order to eliminate the disease in an endemic area, regular herd surveillance

is essential, both for noninfected herds to monitor for new breakouts of infection, and currently infected herds to monitor the progression of disease within the herd. This can best be achieved by regular herd testing, which normally takes place using the SICCT or potentially interferon-gamma assay, and rigorous monitoring of carcasses during meat hygiene inspection.110 Treatment of infected cattle is not economically feasible, and the positive animal is slaughtered with the aim to eliminate the disease and reduce the spread of infection.84

On-farm hygiene and biosecurity considerations are also extremely important to prevent the spread of infection. This will decrease any indirect spread via fomites if feed and water troughs are regularly disinfected and appropriate personal protective clothing is utilized when handling animals or infected carcasses. Ideally, infected carcasses should be cremated or, if this is not a possibility, buried at least 4 ft below the ground. The rodent population should be kept under control to reduce spread of infection.

Diagnostic tests including DNA fingerprinting techniques such as variable number of tandem repeat (VNTR) typing and spoligotyping help to determine the strain of M. bovis present within a herd and have epidemiological value to control disease by helping to trace the route of infection and ensuring that adequate control strategies are instigated.15,92,111

Currently, the UK Government is midway through implementing a 25-year plan for the eradication of bovine tuberculosis from this country which was first unveiled in 2013. The Department for the Environment, Food and Rural Affairs (DEFRA) has divided the country into three areas; “low-risk area,” “edge area,” and “high-risk area,” to reflect differences in how the disease is spread and the number of infected cattle detected upon routine testing in each region. In the high-risk areas of the South West and Midlands, the focus of control is to contain the disease in cattle and wildlife and reverse the spread of the disease northwards. The final goal of this scheme is to move the status of low-risk areas to that of “OTF,” thereby easing the economic burden of TB control in the United Kingdom and facilitating trade with other countries. The target is for most of England to achieve OTF status by 2025, and the whole country by 2038. This strategy leans heavily on the experience of New Zealand.

Attempts are made to control tuberculosis in the United Kingdom by testing every cattle herd at least every 1–4 years, funded by the UK taxpayer. The exact frequency is calculated according to the specific numbers of reactors in a certain geographical area recorded over the last 2, 4, or 6 years in compliance with the Directive 64/432/EEC. Increased surveillance can be instigated at a local level as necessary and any herd that represents a local or public health risk will be tested annually, in addition to herds where new breakouts of bovine tuberculosis are detected. Herds at a higher risk of TB infections include, for example, those which produce unpasteurized milk for sale or those that hire out bulls for mating, will be tested annually.

Should macroscopic tuberculosis lesions be detected at the slaughterhouse during carcass processing, a report will be made by the Meat Hygiene Service to the Local State Veterinary Service. Tracings will be carried out to determine the origin of the infected cattle and bacteriological and pathological samples will be submitted from in-contact herd members to the APHA.

As described earlier, the primary test used for screening for tuberculosis in herds in the United Kingdom is the SICCT. This uses inoculation with M. avium subsp. avium and M. bovis tuberculin to assess previous sensitization to these allergens. The test is carried out in accordance with the procedure described by the Commission Regulation 1226/2002 (Annex B to Directive 64/432/ EEC). The interferon-gamma test is used as a second-line test to supplement the SICCT test, particularly in areas where there is protracted and extensive breakdown of tuberculosis control.113

Milk from reactor animals has been banned from sale under any circumstances following instigation of the EU Food Hygiene Regulation 853/2004 in England, Wales and Scotland. This regulation states, however, that sale of milk from nonreactor animals in the same herd is permitted as long as it has been correctly heattreated. Herds from which raw milk is sold should be registered under Regulation 4 of DEFRA’s Dairy Hygiene Inspectorate. Testing of the dairy premises for M. bovis are carried out free of charge with a frequency according to the appreciated risk within the local area and for the specific herd. The producer must pay for the microbiological testing of the milk for M. bovis, but not for more general bacterial contaminants.

Unpasteurized milk from herds that have been declared as OTF can be sold by dairy farmers directly from the holding where it is produced, as long as the milk is clearly labelled as raw, as defined by the Council Directive 64/432/EEC. The Animal and Plant Health Agency has advised that annual testing should be performed on all herds from which unpasteurized milk is sold for human consumption. This is more frequent than in OTF herds where milk is pasteurized before consumption or use in order to catch any tuberculosis-reactor cattle before the bacilli become well established in the udder and shed into the milk.

Should tuberculosis be suspected during meat inspection in any part of the carcass or offal, a more detailed postmortem examination is carried out than normal, following specific governmental requirements. Should generalized tuberculosis lesions be detected, the entire offal and carcass are condemned and do not enter the human food chain. If a single lesion is found on any organ or the associated lymph nodes, only the affected part of the animal is removed from the food chain, as any residual contamination of muscle should be deactivated during the cooking process. A tuberculin reactor cow can only be deemed fit for human consumption should it be entirely free of tuberculosis lesions following a thorough postmortem inspection. Reactors, if present, should be slaughtered at the end of the production line, in order to minimize cross contamination. During the early studies of investigation of the pathogenesis of tuberculosis in animals it was quickly realized by Francis114 that surveillance and control at postmortem was an important requirement to minimize zoonotic transmission of disease. The very first link between infected meat and human tuberculosis infection had been postulated years before, in 1893, by Brehrend,115 who called for effective meat inspection. This call was reinforced by Ostertag116 who claimed that humans could contract tuberculosis from eating infected meat. Actually, the current evidence of this route of transmission from cattle to humans is weak or nonexistent96 and the risk for consumption in industrialized countries is extremely low. The route of ingestion causing infection in animals has not been documented in many

countries over decades. In the United Kingdom, there has never been documented evidence of gastrointestinal transmission of M. bovis to humans due to consumption of infected meat, however, given the infectious and zoonotic nature of the disease, caution is warranted.

Control of disease in wildlife maintenance hosts

Wildlife such as badgers or brush-tailed possums can also be considered to be maintenance hosts of M. bovis, as these species are able to maintain the infection within their own population, and control among these groups should be considered in order to eliminate the disease. Eradicating any possible contact between wildlife and cattle limits the spread of infection, although this is not always feasible. Culling or vaccinating of an adequate level of affected wildlife species are strategies that are currently employed in an effort to effect control of bovine tuberculosis in the UK.

In New Zealand, following recognition that possums infected with bovine tuberculosis were a significant source of persistent infection in cattle in the early 1970s, the Ministry of Agriculture and Fisheries began to carry out possum control operations in areas where there was a persistent tuberculosis problem in cattle. Possum control proved to be difficult to achieve and in 1991–1992 there remained 20 areas in New Zealand where M. bovis infection was present in the possum populations. Eighty-three percent of test reactor cattle and those under movement restriction were found to live in these regions. From a financial perspective, the budget for tuberculosis control in New Zealand in the 1992–1993 financial year was NZ$21.82 million, and a total of NZ$5.1 million was spent on possum control. These methods helped to decrease the spread of M. bovis in cattle in this country, but the disease in New Zealand still has not been eradicated.

Since the discovery that badgers are maintenance hosts for M. bovis in the 1970s, limitation of bovine tuberculosis in the badger population of the United Kingdom and Republic of Ireland has been an important consideration to aid control of the disease among cattle populations. The Kreb’s report, published in 1997,5 concluded that despite there being compelling evidence that badgers were involved in transmitting the infection to cattle, the effectiveness of badger culling could not be quantified using available data. A recommendation was made to establish the Randomised Badger Culling Trial (RBCT) in order to quantify the impact of culling badgers on incidence of TB in cattle and determine the effectiveness of strategies to reduce the risk of a TB cattle herd breakdown. This took place from 1998 to 2005 and was overseen by the Independent Scientific Group (ISG). During this period, nearly 8,900 badgers were culled across large areas where there was a high risk of bovine TB. The conclusions of this trial, published in June 2007, were that although badgers were a clear source of cattle TB, badger culling could make no meaningful contribution to cattle TB control in Britain. In addition, weaknesses in the cattle testing regimes mean that cattle themselves contributed significantly to the persistence of TB in all areas where the disease occurs, and in some parts of Britain cattle were likely to be the main source of infection. However, a different review of the basic data presented in this report made by the then Government Chief

Scientific Advisor Sir David King in 2007 at the government’s request produced a different interpretation; that badger culling “would have a significant effect on reducing TB in cattle”. It would appear that the reasons for the differences in the conclusions in the two reports occurred as the ISG’s report took economic feasibility into account when making its calculations, while Sir David King’s group of experts did not include practicalities or costs in their considerations. DEFRA published a (bovine) Tuberculosis Eradication Programme for England in 2011* and the pilot badger cull in South West England was announced in December 2011. Licences to cull badgers were granted to land owners by Natural England under the Protection of Badgers Act 1992 and the Wildlife and Countryside Act 1981 with provisions to cover culling for four years. This pilot cull was initiated in 2013, with an aim to eradicate 70% of the badger population in each 150 km2 the culling areas over a six-week period. To minimize perturbation (movement of badgers disturbed by culling out of their own territories and consequently spreading disease), land owners were requested to identify natural barriers to badger movements, and establish the culling areas with these at their edge.† Badgers were culled using a controlled shooting method, as well as cage trapping and shooting. In 2015, the British Veterinary Association (BVA) concluded that it could not be demonstrated that controlled shooting could be carried out humanely and effectively. Although the Association remained supportive of the cull as a necessary part of a comprehensive strategy for controlling and eradicating bovine tuberculosis, they called on the government to revert only to the method of cage trapping and shooting method to deliver the badger cull by a safer and more humane and effective route. The long-term effect of badger culling on the prevalence of tuberculosis in the English cattle population remains to be seen. Some limited assessment was performed in 2016 and concluded that “reductions in TB incidence were associated with culling in the first two years in the intervention areas, when compared to areas with no culling.” However, the authors did acknowledge that the interim data analysed originated only from two years” worth of data from only two of the culling sites; further analysis was necessary.121

In 2018, the Godfray Report identified that it would be desirable to move from a lethal to a non-lethal control strategy for badger control.122 The government has since advised that the control strategy will move towards use of the BCG vaccine in badgers to vaccinate against tuberculosis, retaining the option of culling in areas where the bovine tuberculosis is rife and epidemiological evidence points to a significant disease reservoir in badgers. Over time it is thought that this will reduce the number of new cases and increase the herd immunity of badgers to the disease. A pilot study was undertaken between 2010 and 2014,123 the results of which suggest that badger vaccination had no effect on the incidence of TB in cattle, although this study had limitations and bias and further studies need to be carried out. Badgers have a lifespan124 of 3−5 years, so a cyclical plan of either culling or vaccination control methods every four years is predicted to be effective to reduce the disease in the badger population. Plans for a roll out of badger vaccination in the UK are to be carried out by DEFRA as

* Defra, bTB Eradication Programme for England, July 2011.

† Defra, Update on measures to tackle bTB, 14 December 2011.

part of its drive to eradicate bovine TB, with the aim to wind down the current badger cull programme by the midto late 2020s.

Vaccination

A successful vaccine targeting M. bovis in cattle and other animals has not yet been developed, although efforts are underway. The bacillus Calmette−Gùerin (BCG) vaccine, a live-attenuated strain of M. bovis, is recommended to be administered to babies, children and adults under the age of 35 who are at risk of catching tuberculosis. The safety of this live vaccine for humans is well investigated and documented and the efficacy of live vaccines is always greater than killed. However, the use of the BCG vaccine is not permitted in the United States as field trials have repeatedly demonstrated that the protection offered does not appear to protect all members of a vaccinated population, and delayed-type hypersensitivity induced by the vaccine causes humans to react positively to the tuberculin skin test used to diagnose active tuberculosis infection, therefore reducing the value of this test, should it be required as part of a diagnostic investigation, although the interferon-gamma test for latent infection does not cross react with BCG or MAC.

Used alone, the BCG vaccine does not provide cost-effective or useful protection against infection of M. bovis in cattle. The current BCG vaccine does not have a high enough efficacy to provide full protection against tuberculosis, and is currently administered to cattle to provide a “Ring Vaccination” initiative in exposed herds where a breakout of TB infection has occurred. The same concern with this vaccine exists in cattle as in humans, as cattle that have received the BCG vaccine may offer a false-positive reaction to the tuberculin SICCT and therefore cattle infected with tuberculosis, and those which have been vaccinated, cannot be differentiated. The rational development of an effective vaccine for cattle involves the development of the optimal combination of M. bovis antigens that are required for protective immunity. Diagnostic tests based on antigens not incorporated into the vaccine would allow differentiation of the immune response generated by either vaccination or infection.

The BCG vaccination is licensed for intramuscular use in badgers in the United Kingdom, although an oral formulation, which would be easier and more cost-effective to administer, has also been trialed.55 Wales is the only country currently implementing a BCG vaccination trial in this species, and the results of this trial are yet to be published.

Control of disease spread to “spillover” hosts

In the domestic situation, cats and dogs can be considered the main spillover hosts of M. bovis, i.e., they are not very effective at transmitting the disease to other members of their own species or to other species, including humans. Consideration needs to be given to mechanisms to reduce transmission of mycobacteria from “maintenance” hosts to “spillover” hosts, as by definition no further transmission of disease will occur from a “spillover” host. An important consideration to limit the spread of disease is to limit the degree of contact of “spillover” hosts with any infected