Review on Bovine Tuberculosis

: Bovine tuberculosis (BTB) is a chronic infectious disease caused by Mycobacterium bovis . This disease is widely distributed throughout the world and mainly affects animals with occasional human involvement. BTB can have an impact on the national and international economy, affects the ecosystem via transmission to wildlife and is of public health concern due to its zoonotic potential. It is primarily of economic importance as it can have a considerable direct effect on milk and meat production and animal reproduction. Although still present in some industrialized countries, BTB today mostly affects developing countries lacking the resources to apply expensive test and slaughter schemes. In Africa, the disease is present virtually on the whole continent; however, little accurate information on its distribution and prevalence is available. It is a chronic, generally respiratory disease, which is clinically difficult to diagnose although emaciation, loss of appetite, chronic cough and other signs of pneumonia could be symptoms developing at relatively late stages of the infection in cattle. Its pathology is characterized by the formation of granulomatous lesions, which can within the course of the disease regress or exhibit extensive necrosis, calcify or liquefy and subsequently lead to cavity formation. During meat inspection procedures on cattle carcasses in slaughterhouses, tuberculous lesions are primarily found in the upper and lower respiratory tract and associated lymph nodes. However, the bacteria can also develop a systemic infection, disseminate within its host and affect other organs. Aerosol exposure to M. bovis is considered to be the most frequent route of infection in cattle, but infection by ingestion of contaminated material may also occur. However, M. bovis infection in humans can occur through the consumption of contaminated raw or undercooked dairy and/or meat products; meanwhile occupational infection may occur due to exposure through airborne infection among farmers, veterinarians and slaughterhouse workers. Identification of M. bovis by culture, molecular techniques and biochemical methods is important for definitive diagnosis. Evaluations of antemortem tests for the diagnosis of BTB in Africa are scarce but a prerequisite to identify appropriate tools for future disease control programs. Control and prevention of the disease is vaccination and proper management of animals and humans environment.


INTRODUCTION
whose locations depend largely on the route of infection.
Tuberculosis (TB) is an infectious disease caused by lesions involve the mesenteric lymph nodes with possible mycobacteria that has been a major health risk to human spread to other organs [1]. In older cattle, infection is and animals for more than a century. It is widely usually by the respiratory tract with lesions in the lung distributed throughout the world affecting all age groups and dependent lymph nodes [2]. of humans and animals. In humans, it is responsible for Tuberculosis is a chronic infectious disease of more deaths than any other bacterial disease ever today.
animals, birds and humans, caused by members of the Bovine tuberculosis (BTB) is a disease characterized by genus Mycobacterium. In most species it can lead, with formation of granulomatous nodules called tubercles the proliferation of tubercles, to caseation and In calves, it is usually transmitted by ingestion and calcification in the parenchyma of affected organs [3]. The risk of BTB to cattle is dependent on host, Signs like emaciation and weakness are just two of the pathogen and environmental factors and there is a broad cardinal alert symptoms during ante mortem inspection, spectrum of outcomes to infection with M. bovis [12,13] though these two alone do not confirm bovine which are thought to be similar to the effects of the related tuberculosis [4].
pathogen M. tuberculosis Waters et al. [14] in humans. Mycobacterium bovis is the major causative agent of The culling loss due to the disease is estimated to be bovine tuberculosis (BTB) and part of the Mycobacterium 30-50% of the difference between the values of a dairy or tuberculosis complex (MTBC). Although still present in beef breeding cow and its value at slaughter [15]. some industrialized countries, BTB today mostly affects Consumption of raw or unpasteurized animal products developing countries lacking the resources to apply and contact with infected carcasses plays a large role in expensive test and slaughter schemes. In Africa, the zoonotic M. bovis infection of humans in Africa and disease is present virtually on the whole continent; South America [16]. however, little accurate information on its distribution and The occurrence of BTB due to M. bovis in humans is prevalence is available [5].
difficult to determine accurately because of technical Invisible droplets (Aerosols) containing TB bacteria problems in isolating the microorganism. Currently, the may be exhaled or coughed out by infected animals and BTB in humans is becoming increasingly important in then inhaled by susceptible animals or humans. The risk developing countries, as humans and animals are sharing of exposure is greatest in enclosed areas, such as barns.
the same micro-environment and dwelling premises, Inhalation of aerosols is the most common route of especially in rural areas. At present, due to the infection for farm and ranch workers and veterinarians association of mycobacterium with the HIV/AIDS who work with diseased livestock. Livestock also are more pandemic and in view of the high prevalence of HIV/AIDS likely to infect each other when they share a common in the developing world and susceptibility of AIDS watering place contaminated with saliva and other patients to tuberculosis in general, the situation changing discharges from infected animals [6]. Both beef and dairy is most likely. Prevalence data on BTB infection in Africa cattle are susceptible to bovine TB. However, is scarce. There is, however, sufficient evidence to confinement dairies and feedlots are the primary areas of indicate that it is widely distributed in almost all African concern, frequent herd additions increase the opportunity countries and even is found at high prevalence in some for introduction of the disease and high animal densities animal populations [17]. increase the likelihood of the disease spreading among Tuberculosis due to M. bovis is still a neglected herd mates. However, because dairy cattle remain in the disease in animals as well as in human populations in the herd longer and are maintained in relatively dense sub-Saharan countries, where it is less studied [6]. In populations, dairies are at higher risk for heavy herd addition to these bottlenecks to the development of the infection rates [7]. livestock industry, the disease attains much of its It is also a known cause of zoonotic tuberculosis in importance from being zoonotic, causing human humans, which can appear indistinguishable with regard tuberculosis. In humans TB is still a major cause of death to pathogenesis, lesions and clinical findings to that worldwide in general and in the high-burden regions in caused by M. tuberculosis. M. bovis shows a high degree particular [18][19][20]. of virulence for both humans and animals [8].
In Ethiopia, M. bovis infection is endemic in cattle On a global scale, this zoonotic pathogen is estimated [21]. The prevalence of BTB in Ethiopia ranged from 3.4% to cause 10-15% of human TB cases in the developing in smallholder production systems to 50% in intensive world [8] and is considered to be the fourth most dairy production system [22,23]. Moreover, a prevalence significant livestock disease in terms of impact on human of 5.15% of BTB was reported in animals slaughtered in health and economics in developing countries, including Nazareth municipality abattoir of central Ethiopia [24]. risks to other livestock and wildlife [9]. Furthermore, there Ethiopia, animals are kept in the same dwelling with their is evidence that subclinical BTB has a negative impact on owners and use of dungs for plastering of wall, floor and productivity in dairy cows, increasing the costs incurred as a source of energy for cooking, doexacerbate chances by the dairy industry [10]. As many parts of the world of spreading the disease human [25]. Thus, it is endemic have no active surveillance programmes and limited and has been reported in different regions of the epidemiological studies, the prevalence and impact of countries, the disease in the countries associated with BTB worldwide is likely to be underestimated [11].
decreased productive efficiency and carcass or organ condemnation in the abattoir, the nationwide distribution more than 99.9% chromosomal identity and they cause of the disease and the economic loss associated with it tuberculosis with similar pathology in various mammalian has not been fully determined due to lack of good hosts [30]. diagnostic facilities. Hence, having the knowledge of M. bovis shows a dysgonic colony shape on distribution, prevalence and risk factors of the disease are Lowenstein-Jensen medium, is negative for niacin fundamental so as to look for effective control strategy accumulation and nitrate reduction, is susceptible to [25].
thiophene-2-carboxylicacid hydrazide (TCH) and shows Though detection of BTB in Ethiopia is most microaerophilic growth on Lebek medium. A further commonly carried out on the basis of tuberculin skin criterion used for differentiation is the intrinsic resistance testing and abattoir inspection [26] regular surveillance to pyrazinamide, which is found in most M. bovis isolates. through skin testing of millions of individual cattle, In contrast, M. tuberculosis shows eugonic growth, is bacteriology and molecular methods are not realistic positive for niacin accumulation and nitrate reduction, is methods for logistic reasons. Abattoir inspection at the resistant to TCH, shows aerophilic growth on Lebek moment remains economically affordable and valuable medium and is usually not monoresistant to pyrazinamide techniques to detect TB lesions in the carcass of More recently, several molecular methods have been slaughtered cattle. The main purpose of post mortem developed that provide clear criteria for the identification examination of carcasses at slaughtered house is for the of M. bovis [31]. protection of the public health, but failure of detecting a Historically, taxonomic segregation of the M. lesion during inspection in cattle with a single lesion will tuberculosis complex has been based on each species' have a huge zoonotic implications. So it is imperative to unique combination of host preference and its evaluate the efficiency of the routine and detailed meat characteristic growth, morphology, physiology and inspection for the detection of suggestive tuberculosis biochemistry [32,33]. No other TB organism has as great lesions [27]. Therefore, the aim this review paper provides a host range as bovine TB, which can infect all warm tips of information on Bovine tuberculosis so that proper blooded vertebrates. M. avium can affect all species of control/preventive measures could be put in place.
birds, as well as hogs and cattle. M. tuberculosis primarily General Characteristics of Mycobacterium Bovis and Its and dogs. Bovine TB has affected animal and human Host Range: Mycobacterium bovis is the organism that causes bovine tuberculosis. The bacteria are acid fast, filamentous, curved rods [28]. The organism does not grow on blood agar plates and requires6-8 weeks of incubation time to see visible growth on Lowenstein-Jensen media. Acid fast staining would yield acid fast positive rod shaped organisms on sputum smears. The tubercle bacillus is about as susceptible to the action of heat and light as any other vegetative organism; but is highly resistant to the action of chemical substances, a fact made use of in obtaining pure cultures from contaminated pathological material. Though drying kills a fair proportion of tubercle bacilli, many may escape this effect. The organism can also survive for long periods in cool shady places, particularly if protected from light by crust formation on infective discharges or dung [29].
Mycobacterium bovis, causative agent of BTB, is a member of the Mycobacterium tuberculosis complex, which also comprises the important human pathogen M. tuberculosis, as well as Mycobacterium canettii, Mycobacterium africanum, Mycobacterium pinnipedii, Mycobacterium microti and Mycobacterium caprae. These phylogenetically closely related bacteria share affects humans but can also be transmitted to hogs, cattle health since antiquity. Once the most prevalent infectious disease of cattle and swine in the United States, bovine TB caused more losses among U.S. farm animals in the early part of this century than all other infectious diseases combined [34].
Susceptible host: M. bovis is of significant importance in livestock and in a wide range of wild animal species worldwide. Bovine species, including bison and buffaloes, are particularly susceptible to the disease, but nearly all warm-blooded animals can be affected. M. bovis is also known to affect humans, causing a serious public health problem where the disease is endemic [9]. Cattle are the usual host for M. bovis, but bovine TB can be transmitted to humans as well as other animals such as swine, bison and cervids (Deer and elk) [35].

Source of Infection and Modes of Transmission: M. bovis
infection is spread to cattle primarily through the inhalation of infectious aerosols, but has also been reported to be spread by ingestion of infectious material from drinking infected milk or ingesting contaminated pasture or feed. But inhalation is the most likely and important route of infection in cattle with TB, since M. bovis is an obligate intracellular parasite and has lesions in field cases predominantly involved the upper a limited survival period outside the host (Depending on and lower respiratory tract and associated lymph nodes the environmental conditions). It is susceptible to drying [36]. Cutaneous, congenital and genital infections have and ultraviolet light, but is relatively resistant to been recorded but are considered rare. Carrier animals are detergents and moderate changes in P . Bovine significant in spreading and perpetuating the infection, tuberculosis (BTB) is a persistent problem among UK but transmission is intermittent and mimics a point source cattle herds. Potential obstacles to BTB control are the epidemic. Aerosol transmission occurs in all environments existence of the badger (Meles meles) as a wildlife and the infective dose by inhalation can be very low.
reservoir [42] and the presence of M. bovis in the However, transmission is only effective over short environment where the organism can survive for months distances, of 1-2 meter and cattle density is therefore a and may remain infectious. Badgers form social groups significant factor in the rate of transmission. Infection is that use communal underground sets where conditions spread more rapidly in intensive animal husbandry are likely to facilitate transmission and provide one focus situations than in extensive or rangeland conditions [37].  [37,39]. mitigate or minimize the risk of further transmission [44]. As is also true of human TB, the risk of M. bovis infection in humans is likely to increase where the Epidemiological Factors Influencing Transmission: prevalence of HIV/AIDS is high due to the susceptibility Consistent with the mostly aerosol spread of M. bovis, of immunosuppressed AIDS patients to TB. Cases of disease prevalence is higher under intensive farming HIV-related human TB due to M. bovis have been practices, such as on dairy farms or where animals are reported in many developed countries. The potential housed indoors. In beef herds, prevalence will generally impact of AIDS/HIV infections in humans on the be lower, but high prevalence (Around 35%) has been transmission of M. bovis to and among humans is of great observed where cattle are overstocked and/or in poor concern and requires careful consideration, wherever condition. In very extensive farming systems, such as in bovine TB is still a major problem [40].
pastoral cattle management, herd prevalence will generally Incubation Period: The incubation period can range from groups can have high prevalence. Under pastoral months to years with the severity depending on the conditions in northern Australia, opportunities for immunity of the host, the size and frequency of the transmission were provided by cattle congregating infectious dose and host genetics. In many cases, around waterholes during the dry season, or on reduced infection will be localized and cleared by the immune amounts of dry land during the wet season [37]. system, such that disease never develops. In humans, The pre-disposing causes that might come into play only 10% of people infected with M. tuberculosis will are listed below: disease of civilization / domestication:develop TB disease in their lifetimes [37]. Infection results herding together facilitates spread; Age: young animals in chronic disease; animals typically present with clinical are more susceptible than older ones; Nutrition: Vitamin signs during times of increased stress or as they age [41].
A & C deficiency predisposes; Housing: Dark, ill Persistence of the Agent.
ventilated, damp dwellings are favorable for the spread; H be lower than in intensive systems, but small family Heredity: Zebu cattle are somewhat more resistant than Despite disease under-reporting in developing exotic or crossbred; Climate: Cold and humid weather is countries, there is, however, sufficient evidence to favorable for spread [29].
indicate that not only the prevalence of disease is higher Geographical Distribution: The geographical distribution any national control and eradication programmes, it is of bovine TB has changed drastically over the past increasing worldwide particularly in Africa [17], Asia and decades. Prior to the introduction of control measures and Latin America [12]. milk pasteurization in developed countries, TB has been In general, the situation is profoundly different in widely distributed throughout the world. Eradication developing countries, which are in general unable to programmes based on test-and-slaughter policies to clear apply expensive test-and-slaughter schemes for the herds of infected animals virtually eliminated TB from control of animal tuberculosis. Although in parts of the livestock in many developed countries. Today, many Latin American and Caribbean countries there has been countries in Europe and North America and Australia are significant progress in bovine tuberculosis control and free of the disease or close to complete eradication in infection rates under 1% have been reported for 30% of livestock. However, the maintenance of M. bovis infection the region's cattle, 70% of cattle are kept in areas where by wildlife species has compromised eradication efforts in rates of infection are higher and where herd prevalence of countries such as in the United Kingdom, Ireland, New up to 56% have been reported [50]. On the African Zealand and parts of the United States of America [12].
continent, more than 80% of the human population co-In developed countries, the driving forces for the control exists with cattle in the absence of any organized control and eradication of bovine tuberculosis from the national of bovine tuberculosis [51]. In recent years a growing domestic herd are indisputably of economic and awareness of neglected zoonoses including bovine sociopolitical nature, based mainly on the negative tuberculosis has led to initiative supported by the economic impact of the disease [45]. In large parts of the WHO/FAO/OIE to investigate, calculate and mitigate the developed world, policies regulating the control of bovine unknown risk from these animal diseases on livestock tuberculosis are aimed at complete eradication of the productivity, human health and livelihoods [52]. Overall, disease from its livestock populations as part of an the presence and extent of bovine tuberculosis in the integrated approach to food safety. These policies follow developing world has been poorly investigated in the an expensive test-and slaughter strategy for the control of past, but a number of recent studies have revealed new bovine tuberculosis and significant successes have been data confirming the presence of M. bovis in cattle [53][54][55][56] achieved in many countries [45,46]. On the other hand, and moreover providing insights into the specific risk the benefit and sustainability of such costly programmes factors associated with tuberculosis in cattle in different have been increasingly questioned in the light of the countries and regions. In Africa, high prevalenc rates of rising economic burden and social impacts on and bovine tuberculosis (up to 50% at herd level) were reduced acceptance by farmers [47,48]. However, in reported in areas of Zambia where cattle and Kafue lechwe general, with the exception of a few countries with a shared grazing and water as well as in areas where the wildlife reservoir of M. bovis the prevalence of bovine traditional management of livestock in transhumant herds tuberculosis has reached very low levels, in most (herds which are moved to floodplains for grazing during developed countries [48]. the dry season) prevailed [54,57]. Under these often In developing countries, data on the prevalence of nomadic conditions, the risk of exposure to M. bovis was bovine TB are minimal and the information available may increased significantly by creating multiple herd contacts not represent the true epidemiological status of the and increasing the total herd size. The latter has also been disease. Although bovine TB is notable in many suggested as a driver of the disease prevalence in countries, it is often underreported, particularly in Ethiopia [58] and Ecuador [59]. On the other hand, in countries that lack effective disease surveillance and countries with a rapidly increasing livestock production reporting systems. The insidious nature of the disease, and intensification of production systems such as Iran, which does not cause fulminating outbreaks with high the propagation and insufficient detection of circulating mortality, is likely to decrease reporting of the disease, M. bovis strains may be the most important contributor to leading to a lack of measures for its control [49].
increasing economic losses from bovine tuberculosis, in the developing nations but also that in the absence of rather than the importation of infected cattle, as Many lipids have been implicated in mycobacterial previously suggested [60]. Most importantly, in the virulence which is not found in other bacterial genera. mainly rural livestock producing areas of developing Lipomannan (LM), lipoarabinomannan (LAM), the countries, bovine tuberculosis can have devastating phosphatidylinositol mannosides (PIMs), the cord factors impacts on the livelihood of millions of the world's most trehalose mono-and dimycolate (TMM and TDM) and vulnerable communities as the disease compromises their the phthiocerol dimycocerosates (PDIMs) are all surface sustainable food supply, income and social status [54,60].
bound mycobacterial lipids capable of modulating innate Pathogenesis: After inhalation, the most part of bacilli monomycolyl glycerol (MMG), have been shown to are arrested in the upper respiratory tract. The bacilli modulate host immunity [65] and hyper virulence [66]. which reach to alveoli will be ingested by alveolar The organism replicates intracellularly after it has macrophages. Tubercle bacilli withstand phagocytosis been taken up by the macrophages. A granuloma or (Due to a lot amount of lipids into the cell wall) and tubercle forms as the body tries to wall off the infected multiply in the macrophages. Accumulating macrophages with fibrous tissue. The granuloma is mycobacterium stimulate an inflammatory focus and cell-usually 1-3 cm in diameter, yellow or gray, round and firm. mediated hypersensitivity. Activated macrophages On cut section, the core of the granuloma consists of dry release cytokines which are responsible for specific tissue yellow, caseous, or necrotic cellular debris. The infection lesion, named tubercle. Tubercle is an avascular can spread hematogenously to lymph nodes and other granuloma, composed of a central zone with giant cells areas of the body and cause smaller, 2-3 mm in diameter, and peripheral zone with lymphocytes and fibroblasts tubercles. The formation of these smaller tubercles is (Epithelioid cells) [61]. The ability of mycobacteria to known as "Miliary tuberculosis" [28]. survive and multiply within macrophages determines whether disease will occur within the host. Survival and Clinical Signs: Mycobacterium bovis is considered to be multiplication of the organisms in macrophages at primary a slow growing microbe with a doubling time of around site of infection happens due to prevention of sixteen hours. This may account for why the host which phagosome-lysosome fusion [62].
is infected could take months to show the effects of the Pathogenicity of mycobacteria depends on their infection. TB caused by this strain of Mycobacterium can ability to escape phagocytic killing, mostly imparted by cause different symptoms depending on where the the cell wall constituents: Cord factor (Trehalose infection is taking place. If the infection is in the GI tract dimycolate), surface glycolipid responsible for serpentine for example a main symptoms is extreme abominable growth in vitro; Suphatides, surface glycolipid containing discomfort, an infection in the lungs causes an almost sulphur which prevents fusion of phagosome with uncontrollable cough. It is has been believed that the lysosome and cAMP secreted by the bacteria may also symptoms can be more closely related to the genome of facilitate this; LAM heteropolysaccharide which inhibits this strain than the species. Mycobacteriums all produce macrophage activation by IFNã and induces macrophages mycolic acids in their cell walls. These acids could cause to secrete TNFá which induces fever and IL-10 which the symptoms seen from the infection of M. bovis [67]. suppresses mycobacteria-induced T cell proliferation; the When present, clinical signs can include variable wax of the cell wall, peptidoglycans and other glycolipids pyrexia, weakness, anorexia, emaciation, dyspnea, are responsible for the adjuvant activity attracts antigen enlargement of lymph nodes and coughing, particularly presenting cells [63].
with advanced TB. These signs are not unique to bovine Mycobacteria are released from macrophages and TB. Although normally a chronic debilitating disease, also migrate within macrophages around the body. Waxy bovine TB can assume a more acute, rapidly progressive cell wall contributes to the host immune response to the course [7, 68]. mycobacteria and the development of lesions. The host mounts a cell-mediated immune response with activated Diagnosis of the Disease: Many methods exist to macrophages and sensitised T cells followed by a diagnose cattle suffering from tuberculosis. Among them, delayed-type hypersensitivity response with granuloma In vivo test like tuberculin test, the cellular test based on formation [64].
the quantification of gamma interferon, post mortem immunity [62]. Recently, newly identified lipids, such as diagnosis of macroscopic lesions of tuberculosis, The histological lesions consist of necrotic cells in microscopic examination of lesion, culture, biochemical the center of the tubercle surrounded by epitheloid cells characterization, enzyme linked immunosorbent assay and multinucleated giant cells all encapsulated by (ELISA) testing, tuberculin testing can be performed and collagenous connective tissue. The necrotic core of cells molecular methods [69]. The various symptoms that are can often become calcified as the tubercle matures [28]. observed to be present can all help in diagnosing tuberculosis in cattle [68].

Culture of Mycobacterium
Gross and Histopathology: TB causes abscess-like Jensen and stone brinks media are most commonly used lesions commonly referred to as granulomas or tubercles.
in veterinary bacteriology. Lowenstein-Jensen medium The area of the body affected is usually related to the can be obtained commercially. An agar-based medium route of entry. Because of the frequency of respiratory such as middle brook is used by the bacteria to grow [69]. transfer, lesions are often seen in the lungs and The media are prepared as solid slants in screw-capped associated lymph nodes. Macroscopic lung lesions are bottles. Malachite green dye (0.025g/100ml) is commonly not essential for the spread of TB by the respiratory route.
used as selective agent. Mycobacterium Small and even microscopic lung lesions, which often tuberculosis, Mycobacterium avium and many of the occur concurrently with thoracic lymph node lesions, are atypical mycobacteria require glycerol for growth. often not detected by normal abattoir or field autopsy However, glycerol is inhibitory to Mycobacterium bovis techniques [28].
while sodium pyruvate (0.4%) enhances its growth. Once the organism has entered the bloodstream, Thus, themedia with glycerol and without glycerol lesions may be found in any part of the body and may (But with sodium pyruvate) should be inoculated. result in animals with 'generalized TB. The detection of The mediacan be made more selective by the addition of macroscopic lesions at necropsy is an important aspect of cycloheximide (400ìg/ml), lincomycin (2ìg/ml) andnalidixic the diagnosis of bovine TB. A presumptive diagnosis of acid (35ìg/ml). Each new batch of culture medium should bovine TB is often made on the basis of gross pathology be inoculated with the stock strainsof Mycobacteria to and examination of smears or histological sections made ensure that the medium supports satisfactory growth [71]. from lesions. However, a definitive diagnosis can only be The inoculated media may have to be incubated at 37°C made by isolating M. bovis from animal specimens [70].
for up to 8 weeks and preferably for 10 to 12 weeks with or Lesions in cattle are most frequently seen at necropsy in without carbondioxide for the mycobacteria in the the retropharyngeal, bronchial and mediastinal lymph tuberculosis group [69]. Mycobacterium tuberculosis and nodes, which may be the only affected tissue. The lung, Mycobacteriumavium prefer the caps on the culture liver, spleen and the surface of body cavities may also be media to be loose while Mycobacterium bovis grows best affected. Lesions in other species can differ from the in air tight containers [70]. classical picture seen in cattle [2].
Lesions in cattle may vary in size from 1 mm to more Colonial Morphology: The luxuriant growth of than 10 cm in diameter. There may be single lesions in Mycobacterium tuberculosis on glycerol containing lymph nodes or a primary complex that is, lesions in a media, giving the characteristic 'Rough, tough and parenchymatous organ and a lymph node draining the buff'colonies is known as eugenic while the growth of organ. Most lesions appear as firm or hard, white, grey or Mycobacterium avium on media containing glycerol yellow nodules. The cut surface usually shows a isalso described as eugenic. Mycobacterium bovis has yellowish, caseous centere, which is dry and firm [2]. sparse, thin growth on glycerol containing media that is Calcification is common, particularly in lymph nodes and called dysgenic. Mycobacterium bovis, however, grow on sectioning the lesion, a gritty sensation and grating well on pyruvate-containing media without glycerol [72]. sound indicate its occurrence. Conglomerate tubercles, formed by the growth and coalescence of one or more Histology and Acid-fast Staining: During necropsy of adjacent tubercles, may occur over the pleural or cattle suspected of being infected with BTB, tissue peritoneal surfaces. Metastases give rise to myriad samples are collected and examined for histopathological tubercles of the same size, usually 2-3 mm in diameter. Old (Microscopic) lesions that are compatible with M. bovis. lesions may be encapsulated by connective tissue, In addition to looking for specific lesions under the heavily calcified and inspissated (Very dense) [28,29,41]. microscope, pathologist can use special stain to identify

Media for Mycobacteria:
The egg based Lowenstein- where, + = positive, _= negative, TCH= thiophen-2-carbonic acid hydrazide Source: Quinn and Markey [70] organisms that are compatible with M. bovis, the Biochemical Tests: The definitive identification of bacterium that causes BTB. The high lipid content, which ranges from 20 -40% of the dry cell weight, is largely responsible for the ability of these bacteria to resist decolonization with acidified organic solvents [73]. The bacteria that take up this stain, including M. bovis, will appear as short red or pink rods when examined under a microscope [72].
Preliminary examination of tissues suspected of being tuberculous should include the preparation of suitably stained smears. The identifiable smear can be made on a new slide from scrapings of the cut surface of tissue. The smear should be air dried and fixed by flaming for one to two seconds. The kinyoum modification of the Zeihl-Neelsen stain is recommended because no heat is required [69]. The Zeihl-Neelsen method is commonly used to stain the mycobacteria. The smears are treated as with concentrated carbol fuchsin by heating and then decolorized with a sulfuric acid and alcohol solution. Malachite green or methylene blue is commonly used counter stains [73]. The stained slides are observed with an ordinary light microscope for the presence of acid-fast bacilli, which appear as red, colloidal or bacillary cells 1-3 microns in length occurring singly or in clumps [72].

Pigment Production and Response to Light:
The Mycobacteria that produce yellowish-orange carotenoid pigments are called chromogenic [70,74]. The term photo chromogenic is applied to those mycobacteria that produce pigment only if exposed to light. The scotochromogenic Mycobacteria produce pigment when incubated either in light or in the dark. Pigment formation is tested with young, well-developed colonies on Lowenstein-Jensen medium. The cultures are exposed to a 100 Watt, clear electric light bulb, at adistance of 50 cm, for at least an hour and then incubated again in darkness for a further 1-3 days. After this treatment the photochromogens will develop pigment. Older colonies of mycobacteria in the tuberculosis group often have a yellowish hue but they are described as nonchromogenic [70]. thespecies of mycobacteria is largely based on biochemical criteria [73]. These biochemical tests are niacin production test, nitrate reduction, deamination of pyrazinamide, urease test, inhibition and tolerance test [70].

Immunological Diagnostic Tests Tuberculin Skin Test:
The tuberculin test based on adelayed type hypersensitivity to mycobacterial is the standard ante mortem test in cattle. It is convenient, cost effective method for assessing cell mediated responses to a variety of antigens and it is "gold standard" for diagnostic screening for detection of new or asymptomatic Mycobacterium tuberculosis complex infection [75]. The reaction in cattle is usually detectable 30-50 days after infection. The injection site should be examined for a reaction 72 hours post inoculation [41]. The tuberculin is prepared from cultures of M. tuberculosis or M. bovis grown on synthetic media. The tuberculin test is usually performed between the mid necks, but the test can also be performed in the caudal fold of the tail. The skin of the neck is more sensitive to tuberculin than the skin of the caudal fold. To compensate for this difference, higher doses of tuberculin may be used in the caudal fold of the tail [69].
Bovine tuberculin is more potent and specific and the potency of tuberculins must be estimated by biological methods, based on comparison with standard tuberculins and potency is expressed in the international unit (IU) [2]. In several countries, bovine tuberculin is considered to be of acceptable potency if its estimated potency guarantees per bovine dose at least 2000 IU in cattle. In cattle with diminished allergic sensitivity, a higher dose of bovine tuberculin is needed and the volume of each injection dose must not exceed 0.2m. Cell mediated hypersensitivity, acquired through infection can be demonstrated systematically by fever or ophthalmically by conjunctivitis, or dermally by local swelling, when tuberculin test or its purified protein derivative (PPD) is given by the subcutaneous conjuctival or intradermal route, respectively [69]. Molecular Diagnosis of the Disease: Following methods have been evaluated for the detection of the M. preliminary screening of suspected samples using acid tuberculosis complex in fresh and fixed tissues [69]. fast staining, isolation can be carried out in a Various primers have been used, as described above. bacteriological medium. However, cross-contamination Amplification products have been analysed by among bovine carcasses, improper decontamination hybridisation with probes or by gel electrophoresis. procedure and duration of isolation procedure (often 3 Commercial kits and the in-house methods, in fresh, frozen weeks and up to 8-10 weeks in liquid medium) jeopardizes or boric acid-preserved tissues, have shown variable and the isolation of M. bovis. The lengthy duration of less than satisfactory results in interlaboratory isolation procedure imposes an unavoidable delay in comparisons [80]. False-positive and false negative important decisions about outbreaks and of suspected results, particularly in specimens containing low numbers herds put under restriction. Shorter time-span diagnostic of bacilli, have reduced the reliability of this test. procedures are required for quicker decision [69].
Variability in results has been attributed to the low copy Therefore, there is an urgent need for a rapid, safe and number of the target sequence per bacillus combined with reliable method to diagnose of bovine TB. The most a low number of bacilli. Variability has also been attributed promising technique for approaching this diagnostic to decontamination methods, DNA extraction procedures, dilemma is polymerase chain reaction (PCR). PCR has techniques for the elimination of polymerase enzyme been used to amplify different regions of the inhibitors, internal and external controls and procedures mycobacterial genome, making it a good candidate for for the prevention of cross-contamination. Improvement assisting with species identification in a variety of in the reliability of PCR as a practical test for the detection specimens [77]. of M. tuberculosis complex in fresh clinical specimens will Rapid identification of isolates to the level of M.
require the development of standardized and robust tuberculosis complex can be made by Gen Probe TB procedures [69]. Cross contamination is the greatest complex DNA probe or polymerase chain reaction (PCR) problem with this type of application and this is why targeting 16S-23S rRNA, the insertion sequences IS6110 proper controls have to be set up with each amplification. and IS1081 and genes coding for M.-tuberculosis-However, PCR is now being used on a routine basis in complex-specific proteins, such as MPB70 and the 38 kDa some laboratories to detect the M. tuberculosis group in antigen b have been used. Specific identification of an paraffin embedded tissues [81,82]. Although direct PCR isolate as M. bovis can be made using PCR targeting a can produce a rapid result, it is recommended that culture mutation at nucleotide positions 285 in the oxyR gene, 169 be used in parallel to confirm a viable M. bovis infection in the pncA gene, 675/756/1311/1410 and 1450 of the gyrB [82]. gene and presence/absence of RDs (Regions of A variety of DNA-fingerprinting techniques has been Difference) [78]. Alternatively molecular typing developed to distinguish the M. tuberculosis complex techniques, such as spoligotyping will identify M. bovis isolates for epidemiological purposes. These methods can isolates and provide some molecular-typing information distinguish between different strains of M. bovis and will on the isolate that is of epidemiological value [79]. enable patterns of origin, transmission and spread of M. PCR has been widely evaluated for the detection of bovis to be described [83,84]. The most widely used M. tuberculosis complex in clinical samples (mainly method is spoligotyping (from 'spacer oligotyping'), sputum) in human patients and has recently been used for which allows the differentiation of strains inside each the diagnosis of tuberculosis in animals. A number of species belonging to the M. tuberculosis complex, commercially available kits and various 'in-house' including M. bovis and can also distinguish M. bovis from M. tuberculosis [79]. The use of a standard from retention being a fraction of the value of a carcass. nomenclature for the spoligotypes according to the database Mbovis.org (http://www.mbovis.org) is encouraged to allow international comparison of profiles.
Other techniques include restriction endonuclease analysis (REA) and restriction fragment length polymorphism (RFLP) using IS6110 probe (especially where there are >3-4 copies of IS6110 in the isolate), the direct repeat (DR) region probe, the PGRS (polymorphic GC repeat sequence) probe [85] and the Pucd probes [86]. The mycobacterial interspersed repetitive units (MIRU)-variable number tandem repeat (VNTR) typing has also been developed to increase the discrimination of the M. tuberculosis complex species [69]. Often a combination of techniques may be used to gain the maximum discrimination between strains [87].
The genome of M. bovis has been sequenced [88] and this information has contributed to improved methods of genetic fingerprinting and to the development of PCR assays that define the subspecies of the M. tuberculosis complex [69].

Economic and Public Health Importance of the Disease:
The economic impact of bovine TB on livestock production is extremely difficult to determine accurately. The disease reduces livestock productivity in general and may be economically devastating for the cattle industry, especially the dairy sector [89]. Most important is the impact of the risk of infection to humans, particularly for women and children who appear to be more susceptible to the disease in countries with poor socio-economic conditions and weak veterinary and public health services. Although estimates of the costs associated with bovine TB and its control refer only to specific countries, all data suggest that worldwide economic losses due to the disease are significant. These losses include those related to animal production, markets and trade as well as the costs of implementating surveillance and control programmes. Losses to TB are also extremely important when endangered wildlife species are involved [89,90].
In general BTB affects the national and international economy in different ways. The most obvious losses from BTB in cattle are direct productivity losses (Reduced benefit), which can be categorized into slaughter and "On-farm" losses. Slaughter losses comprise the cost of cattle condemnation and retention, with the loss from condemnation being essentially the purchased value of a slaughter animal and the loss On-farm losses comprise the losses from decreased milk and meat production, the increased reproduction efforts and replacement costs for infected cattle [90].
Apart from direct productivity losses, BTB has profound economic consequences for national and international trade. On an international scale, BTB affects access to foreign markets due to import bans on animals and animal products from countries where the disease is enzootic. This situation has also major implications for other economic sectors, which are linked to livestock production. Moreover, BTB can create inefficiencies in the world market as e.g. economically inefficient but disease free exporting countries will receive more revenues than economically efficient countries, which cannot export animal products due to enzootic BTB [2]. Presence of the disease in wildlife has considerable economic consequences. Not only is disease eradication more difficult and costly but BTB can theoretically affect entire ecosystems with unpredictable impact on many areas of private interest such as e.g. tourism [90].
The economic impact of BTB in Africa is exacerbated through a number of factors. First, the fast growing population, especially in urban areas, causes an increase in demand especially for dairy products and meat and promotes the intensification of livestock production in peri-urban areas [90]. Importantly, intensive livestock production systems show generally a higher prevalence of BTB than extensive production systems. Second, developing countries lack the financial resources for disease control. This leads to a vicious cycle in which increased poverty affects the means for disease control and vice versa. Third, wildlife reservoirs in Africa are difficult to control; also, contact between transhumant cattle herds and wildlife may be particularly difficult to prevent in Africa. Forth, African countries have little access to the international trade and sanitary measures in industrialized countries may be used for protectionist purposes. Fifth, the public and political awareness are very low [89].
Bovine TB is a zoonotic disease that can have serious consequences for public health. The transmission of M. bovis from cattle to humans was once common in industrialized countries, but human infections were virtually eliminated in countries with effective programmes for eradicating the disease in cattle and high standards of food safety, particularly the pasteurization of milk. The incidence of human TB due to M. bovis varies considerably among countries depending on the prevalence of the disease in cattle, socio-economic conditions, consumer habits and practiced food hygiene.  [68]. One way to ensure that cattle do not become infected cases [94] while other estimates range from 0.4 to 8 is to eliminate any possible interaction with deer. The percent, demonstrating that M. bovis is an important indirect contact is usually a result of cattle ingesting feed factor in human TB [40]. that has been contaminated by deer saliva. It is The proportion of which BTB contributes to the total recommended that any feed for cattle be protected and of tuberculosis cases in humans depends on the stored away from deer. Other programs to control the deer prevalence of the disease in cattle, socioeconomic population, such as hunting and banning feeding, have conditions, consumer habits, practiced food hygiene and been implemented to decrease the density of deer and the medical prophylaxis measures. In countries where BTB in population of affected deer [41]. There is long-term drug cattle is still highly prevalent, pasteurization is not widely therapies that could, in principle, be used to treat the practiced and/or milk hygiene is insufficient, usually condition. Although the anti-tuberculosis drug estimated to be about 10% to 15% of human tuberculosis Pyrazinamide is ineffective against M. Bovis, the use of is considered to be caused by BTB [95].
isoniazid and rifampicin could be used effectively [97]. Regassa [96] demonstrated the association of M.
In Ethiopiaon government owned dairy farms, test tuberculosis and M. bovis in causing tuberculosis and isolation of reactors combined with pasteurization of between humans and cattle. The cattle owned by milk are the current undergoing control practices. tuberculous patients had a higher prevalence (24.3%) However, these measures, as compared to the cattle than cattle owned by non-tuberculous owners with 8.6%. population of the country, are found to be insignificant The author also noted that 73.8% and 16.7% of 42 human [26]. (In general terms, control measures in the traditional isolates were identified as M. tuberculosis and M. bovis extensive production systems are more difficult and and from cattle isolates 18.1% and 45.5% of 11 were found complex. In Ethiopia so far, control of BTB through the to be M. tuberculosis and M. bovis species, respectively. test-and-slaughter policy is not yet established. Most This showed that the role of M. bovis in causing human commonly culling of infected animals (Especially in tuberculosis seemed to be significantly important. On the government: owned farms) and improving sanitary and other hand, in Ethiopia, consuming raw meat is a welcome hygienic standards in other dairy farms is the actual tradition, thus meat may also remain to be another area of control measure of BTB infection [98]. concern or threat to be a source of BTB infection [96].

Control and Prevention: Control methods have served to reduce the number of infections. The rate of bovine
Mycobacterium bovis is the causative agent of tuberculosis in the United States is so low that the bovine tuberculosis (BTB) and belongs to the disease is considered to be practically eradicated.
Mycobacterium tuberculosis complex (MTBC) of bacterial Solutions of phenol, iodine, glutaraldehyde and strains. The most prominent member of the MTBC is M.

CONCLUSIONS
tuberculosis, the principle causative agent of tuberculosis REFERENCES in humans, causing each year more than 1.5 million deaths and having experienced a recent re-emergence through 1. Terefe, D., 2012.Gross pathological lesions of bovine the advent of HIV/AIDS and the appearance of multi drug tuberculosis and efficiency of meat inspection resistant strains. Moreover, wildlife reservoirs of M. bovis procedure to detect-infected cattle in Adama hamper disease eradication schemes in several countries. municipal abattoir: Department of Pathology and It also bears a zoonotic potential and it is of public health Parasitology, College of Veterinary Medicine, concern. It affects the national and international economy Haramaya University, Ethiopia.