Molecular findings and approaches spotlighting Mycobacterium bovis persistence in cattle

Mycobacterium tuberculosis (Mtb) and Mycobacterium bovis (M. bovis) are the etiological agents of human and bovine tuberculosis (TB, bTB) respectively, and share genetic identity over 99% at the whole genome level. Progress has been made towards explaining how mycobacteria and their infected hosts remain in balance without producing clinical symptoms of disease, a phenomenon referred to as latency or persistence, which can be mimicked by certain in vitro conditions. Latency/persistence has mainly been studied using Mtb, where the two-component signalling system, dosRS, has been assigned an instrumental role, and even constitutes the current basis for development of new diagnostic methods and treatment addressing this particular stage of TB. M. bovis conserves homolog genes that in Mtb play a role in human latent TB infection and that, by analogy, would allow it to enter a persistent state in infected cattle; nevertheless, little attention has been paid to this stage in bovine hosts. We suggest that many of the advances acquired through the study of Mtb can and should be taken into consideration by research groups and veterinary professionals dealing with bTB. The study of the infection in bovines, paying particular attention to defining the molecular and cellular markers of a M. bovis persistent infection in cattle, presents great opportunities for the development and trial of new diagnostic tests and vaccines, tools that will surely help in promoting eradication of bTB in high-burden settings.     

Extent of Mycobacterium bovis infection in dairy cattle herds subject to partial culling as determined by an interferon-gamma assay

The interferon-gamma (IFN-γ) assay is employed as a complementary diagnostic test for bovine tuberculosis (BTB) in many countries. To simplify this assay, we established a 96-well plate format using the ESAT-6 and CFP-10 antigens and then employed it to determine the extent of Mycobacterium (M.) bovis infection in dairy herds with a history of BTB outbreaks in a country where only selective culling is practiced. The sensitivity and specificity of this IFN-γ assay were 85.9% and 100%, respectively, based on comparison with the conventional single intradermal tuberculin test (SIDT). The IFN-γ assay was also positive in 30.4% and 36.8% of SIDT-negative animals from herds with recent and remote BTB outbreaks, respectively. Of 14 SIDT-negative, IFN-γ positive cattle, five (35.7%) were culture positive and an additional six were positive based on a polymerase chain reaction-based test for M. bovis. Therefore, the IFN-γ assay has the potential to serve as a specific and sensitive test for M. bovis infection in dairy cattle. Further, the results indicated that a substantial portion of SIDT-negative animals in herds with previous BTB outbreaks were actually infected with M. bovis. Accordingly, the present selective-culling strategy may require modifications to include this more sensitive assay.     

Relationship of bovine NOS2 gene polymorphisms to the risk of bovine tuberculosis in Holstein cattle

Many studies suggest significant genetic variation in the resistance of cattle and humans to infection with Mycobacterium bovis, the causative agent of zoonotic tuberculosis. The inducible nitric oxide synthase (iNOS which is encoded by the NOS2 gene) plays a key role in the immunological control of a broad spectrum of infectious agents. This study aimed to investigate the influence of genetic variations in the promoter of the NOS2 gene on bovine tuberculosis (bTB) susceptibility. In this study, the NOS2 genes of 74 bTB-infected Holstein cows and 90 healthy controls were genotyped using PCR followed by nucleotide sequencing. Polymorphisms at rs207692718, rs109279434, rs209895548, rs385993919, rs433717754, rs383366213, rs466730386, rs715225976, rs525673647, rs720757654 and g.19958101T>G in the promoter region of the NOS2 gene were detected. The g.19958101T>G SNP produced two different conformation patterns (TT and TG) and the TG genotype was over-represented in the bTB group (20.27%) compared with the control group (2.22%). The TG genotype frequency of the g.19958101T>G variant was significantly higher in bTB cattle than in healthy controls (OR, 11.19; 95% CI, 2.47–50.73; P=0.0002). The G allele of the g.19958101T>G polymorphism was more frequent in bTB group when compared to control group (10.14% versus 1.11%). Furthermore, the G allele was a risk factor for bTB susceptibility (OR, 10.04; 95% CI, 2.26–44.65; P=0.0002). In conclusion, the g.19958101T>G polymorphism of the NOS2 gene may contribute to the susceptibility of Holstein cattle to bTB.     

Prevalence of Mycobacterium bovis infection in traditionally managed cattle at the wildlife-livestock interface in South Africa in the absence of control measures

Cattle are the domestic animal reservoir for Mycobacterium bovis (M. bovis) which also affects other domestic animals, several wildlife species and humans leading to tuberculosis. The study area is in a resource-poor community that is surrounded by several game parks, where M. bovis infection has been previously diagnosed in wildlife. A cross-sectional study was carried out to determine the prevalence of M. bovis infection in 659 cattle from a total of 192 traditionally managed herds using the BOVIGAM® interferon gamma assay (IFN-γ). Infection was confirmed by post mortem examination and M. bovis isolation from three test-positive cattle. Genotyping of the M. bovis isolates was done using spoligotyping and VNTR (variable number of tandem repeats typing). The apparent M. bovis prevalence rate in cattle at animal level was 12% with a true population prevalence of 6% (95% Confidence interval (C.I) 3.8 to 8.1) and a herd prevalence of 28%. Spoligotyping analysis revealed that the M. bovis isolates belonged to spoligotype SB0130 and were shared with wildlife. Three VNTR profiles were identified among the SB0130 isolates from cattle, two of which had previously been detected in buffalo in a game reserve adjacent to the study area. The apparent widespread presence of M. bovis in the cattle population raises a serious public health concern and justifies further investigation into the risk factors for M. bovis transmission to cattle and humans. Moreover, there is an urgent need for effective bTB control measures to reduce infection in the communal cattle and prevent its spread to uninfected herds.

     

Variable number tandem repeat analysis of Mycobacterium bovis isolates from Gyeonggi-do, Korea

Bovine tuberculosis (TB) is a major zoonosis that''s caused by Mycobacterium bovis (M. bovis). Being able to detect M. bovis is important to control bovine TB. We applied a molecular technique, the variable number tandem repeat (VNTR) typing method, to identify and distinguish the M. bovis isolates from Gyeonggi-do, Korea. From 2003 to 2004, 59 M. bovis clinical strains were isolated from dairy cattle in Gyeonggi-do, Korea, and these cattle had tuberculosis-like lesions. Twenty-four published MIRU-VNTR markers were applied to the M. bovis isolates and ten of them showed allelic diversity. The most discriminatory locus for the M. bovis isolates in Korea was QUB 3336 (h = 0.64). QUB 26 and MIRU 31 also showed high discriminative power (h = 0.35). The allelic diversity by the combination of all VNTR loci was 0.86. Six loci (MIRU 31, ETR-A and QUB-18, -26, -3232, -3336) displayed valuable allelic diversity. Twelve genotypes were identified from the 59 M. bovis isolates that originated from 20 cattle farms that were dispersed throughout the region of Gyenggi-do. Two genotypes [designation index (d.i.) = e, g] showed the highest prevalence (20% of the total farms). For the multiple outbreaks on three farms, two successive outbreaks were caused by the same genotype at two farms. Interestingly, the third outbreak at one farm was caused by both a new genotype and a previous genotype. In conclusion, this study suggests that MIRU-VNTR typing is useful to identify and distinguish the M. bovis isolates from Gyeonggi-do, Korea.     

Spatial predictors of bovine tuberculosis infection and Brucella spp. exposure in pastoralist and agropastoralist livestock herds in the Ruaha ecosystem of Tanzania

While many studies investigate animal-related risk factors for disease, few have considered environmental or spatial risk factors in the dynamics of bovine tuberculosis (bTB) and brucellosis. In the Ruaha ecosystem of Tanzania, we investigated the role of household location as a predictor for infection with Mycobacterium bovis and exposure to Brucella in pastoralist and agropastoralist cattle herds in a typical African wildlife-livestock-human interface. ArcGIS was utilized to calculate Euclidian distances between households and the nearest river, village center, protected area, and other infected households, followed by multivariate logistic regression to assess the association between risk factors and herd-level bTB and Brucella outcomes. Global and local spatial clustering of bTB-infected and Brucella-exposed herds was explored using the Cuzick-Edward’s test and SaTScan spatial scan statistics. Households located farther from rivers and closer to village centers and herds belonging to agropastoralists were more likely to have bTB-positive cattle. Risk of Brucella exposure increased with proximity to protected areas. One spatial cluster of households with Brucella spp. seropositive cattle was identified. Spatial factors may be useful for assessing disease risk and for formulating intervention and control strategies for households that manage cattle in ecosystems characterized by seasonally limited resources and intense wildlife-livestock interfaces.     

Bartonella and Babesia infections in cattle and their ticks in Taiwan

Bartonella and Babesia infections and the association with cattle breed and age as well as tick species infesting selected cattle herds in Taiwan were investigated. Blood samples were collected from 518 dairy cows and 59 beef cattle on 14 farms and 415 ticks were collected from these animals or in a field. Bartonella and Babesia species were isolated and/or detected in the cattle blood samples and from a selected subset (n = 254) of the ticks either by culture or DNA extraction, PCR testing and DNA sequence analysis. Bartonella bovis was isolated from a dairy cow and was detected in 25 (42.4%) beef cattle and 40 (15.7%) tick DNA samples. This is the first isolation of B. bovis from cattle in Asia and detection of a wide variety of Bartonella species in Rhipicephalus microplus. Babesia spp. were detected only on one farm from dairy cows either infected by Babesia bovis (n = 10, 1.9%) or B. bigemina (n = 3, 0.6%).     

Serological reactivity to Mycobacterium bovis protein antigens in cattle

The serological response to 12 purified Mycobacterium bovis antigens were examined in an ELISA assay. These antigens included the majority of M. bovis protein antigens described to date and in most cases they were very similar to the M. tuberculosis antigens of the same molecular mass.The purified antigens were tested against sera from M. bovis infected cattle, M. bovis culture-negative cattle from infected herds and animals infected with related microorganisms, mainly other mycobacterial species. All the antigens gave strong reactions with at least some sera from the M. bovis infected group and showed cross-reactivity with some of the sera from the other two groups. The antigen with the highest specificity reacted strongly with only 60% of the M. bovis infected sera. Antigens that reacted with most or all of the M. bovis infected sera also gave the highest cross-reactivity with sera from the other two groups. These results indicate that a serological test based on any one or a combination of these antigens, without removal of the cross-reacting epitopes, would be unsatisfactory.     

The prevalence and distribution of Mycobacterium bovis infection in European badgers (Meles meles) as determined by enhanced post mortem examination and bacteriological culture

The accurate diagnosis of Mycobacterium bovis infection in badgers is key to understanding the epidemiology of tuberculosis in this species and has significant implications for devising strategies to limit spread of the disease. In this study, badgers (n = 215) in the Republic of Ireland were examined at post mortem and tissues were collected from a range of anatomical locations and pooled into groups for bacterial culture of M. bovis. By assessing confirmed gross visible lesions (VL) alone, infection was detected in 12.1% of badgers. However, by including the results of all culture positive pooled samples, the overall infection prevalence increased significantly to 36.3%. Two-thirds (66.7%) of infected animals had no visible lesions (NVL). While the thoracic cavity (lungs and pulmonary lymph nodes) was found to be the most common site of infection, in a proportion of animals infection was absent from the lungs and draining lymph nodes and was confined to the lymph nodes of the carcase or the head. This may indicate an early extrapulmonary dissemination of infection or alternatively, in the case of the head lymph nodes, a secondary pathogenic pathway involving the lymphoid tissues of the upper respiratory tract (URT).     

Increased prevalence of Mycoplasma bovis in the Netherlands

Summary

The epidemiology, therapy, and prevention of M. bovis infections are briefly reviewed In a survey begun in 1982 M. bovis was found frequently in the respiratory of veal calves and beef cattle with respiratory problems. In replacement calves infected with respiratory disease in dairy herds, however, the organism has only been detected since 1986. Respiratory tract specimens collected from calves with respiratory disease were submitted for examination for M. bovis from 1986 to 1991 and originated from 83 herds. Mycoplasma bovis was detected in specimens from 59 of the herds, 20% of which were dairy herds and 80% fattening herds. Arthritis caused by M. bovis was observed in 12 herds until July 1991. Since 1976 when the first mastitis outbreak caused by M. bovis was diagnosed M. bovis has caused 14 more outbreaks. The number of diseased cattle varied from 1 tot 16 per farm, and clinical signs of mastitis varied from mild to severe. In all instances the infection has been eradicated from the herds. Because M. bovis can cause great losses in intensively reared cattle herds, it is advisable to separate purchased veal calves and beef cattle from dairy cattle to prevent further spread of M. bovis.     

Molecular Epidemiology of Mycobacterium bovis in Humans and Cattle

Bovine tuberculosis (bTB), caused by Mycobacterium bovis (M. bovis), is a serious re‐emerging disease in both animals and humans. The evolution of the Multi‐ and Extensively drug‐resistant M. bovis strains (MDR‐TB and XDR‐TB) represents a global threat to public health. Worldwide, the disease is responsible for great economic losses in the veterinary field, serious threat to the ecosystem, and about 3.1% of human TB cases, up to 16% in Tanzania. Only thorough investigation to understand the pathogen's epidemiology can help in controlling the disease and minimizing its threat. For this purpose, various tools have been developed for use in advanced molecular epidemiological studies of bTB, either alone or in combination with standard conventional epidemiological approaches. These techniques enable the analysis of the intra‐ and inter‐species transmission dynamics of bTB. The delivered data can reveal detailed insights into the source of infection, correlations among human and bovine isolates, strain diversity and evolution, spread, geographical localization, host preference, tracing of certain virulence factors such as antibiotic resistance genes, and finally the risk factors for the maintenance and spread of M. bovis. They also allow for the determination of epidemic and endemic strains. This, in turn, has a significant diagnostic impact and helps in vaccine development for bTB eradication programs. The present review discusses many topics including the aetiology, epidemiology and importance of M. bovis, the prevalence of bTB in humans and animals in various countries, the molecular epidemiology of M. bovis, and finally applied molecular epidemiological techniques.     

Performance of the SD Bioline TB Ag MPT64 Rapid test for quick confirmation of Mycobacterium bovis isolates from animals

Mycobacterium (M.) bovis, a bacterium in the M. tuberculosis complex, is a causative agent of bovine tuberculosis, a contagious disease of animals. Mycobacterial culture is the gold standard for diagnosing bovine tuberculosis, but this technique is laborious and time-consuming. In the present study, performance of the SD Bioline TB Ag MPT4 Rapid test, an immunochromatographic assay, was evaluated using reference bacterial strains and M. bovis field isolates collected from animals. The SD MPT64 Rapid test produced positive results for 95.5% (63/66) of the M. bovis isolates from cattle and 97.9% (46/47) of the isolates from deer. Additionally, the test had a sensitivity of 96.5% (95% CI, 91.2-99.0), specificity of 100% (95% CI, 96.7-100.0), positive predictive value of 100% (95% CI, 96.7-100.0), and negative predictive value of 92.9% (95% CI, 82.7-98.0) for M. bovis isolates. In conclusion, the SD MPT64 Rapid test is simple to use and may be useful for quickly confirming the presence of M. bovis in animals.     

Evaluation of Spoligotyping and MIRU-VNTR for Mycobacterium bovis in Xinjiang, China

Mycobacterium bovis, the causative agent of bovine tuberculosis (bTB) in cattle, is also a pathogen for human and other mammals. In this study, 406 cows were screened for bTB by both single intradermal comparative cervical tuberculin (SICCT) test and IFN-γ assay. 135 M. bovis were isolated from 31 SICCT and IFN-γ double-positive cows in Xinjiang Uygur Autonomous Region of China. Spoligotyping and MIRU-VNTR were evaluated for genotyping, and 4 and 7 genotypes were identified, respectively. A new combination of nine MIRU-VNTR loci was most discriminative for M. bovis clones from Xinjiang. Interestingly, two new spoligotypes (SB1903 and SB1904) and special repeat numbers of three loci (ETR-D, QUB 1895 and QUB 3336) were discovered in this study. These results indicated a specific epidemic conservation in Xinjiang, China. M. bovis strains with the unique genotypes were isolated from the herds maintaining parent cows imported from the bTB-free countries, suggesting a possible transmission from the local breed of Xinjiang brown cattle.     

Towards harmonised procedures in wildlife epidemiological investigations: A serosurvey of infection with Mycobacterium bovis and closely related agents in wild boar (Sus scrofa) in Switzerland

Bovine tuberculosis (bTB) is a (re-)emerging disease in European countries, including Switzerland. This study assesses the seroprevalence of infection with Mycobacterium bovis and closely related agents in wild boar (Sus scrofa) in Switzerland, because wild boar are potential maintenance hosts of these pathogens. The study employs harmonised laboratory methods to facilitate comparison with the situation in other countries. Eighteen out of 743 blood samples tested seropositive (2.4%, CI: 1.5–3.9%) by ELISA, and the results for 61 animals previously assessed using culture and PCR indicated that this serological test was not 100% specific for M. bovis, cross-reacting with M. microti. Nevertheless, serology appears to be an appropriate test methodology in the harmonisation of wild boar testing throughout Europe. In accordance with previous findings, the low seroprevalence found in wild boar suggests wildlife is an unlikely source of the M. bovis infections recently detected in cattle in Switzerland. This finding contrasts with the epidemiological situation pertaining in southern Spain.     

Prevalence of Mycobacterium bovis skin positivity and associated risk factors in cattle from Western Uganda

Purpose

A cross-sectional study was undertaken to determine the prevalence of Mycobacterium bovis skin positivity and associated risk factors in cattle in western Uganda.

Methods

Herds were selected using multi-stage cluster sampling. The comparative cervical intradermal tuberculin test (CCT) was used to determine cattle tuberculosis status using US Department of Agriculture protocols. Risk factor data were collected from cattle owners through questionnaires collected by in-person interviews. Multivariable logistic regression models were used to measure the association between risk factors and herd CCT reactor prevalence.

Results

A total of 525 cattle from 63 herds were screened for M. bovis infection. Of the 525 cattle tested, 2.1 % were CCT reactors and 15.43 % were CCT suspects. Of herds tested, 14.28 % had at least 1 CCT reactor. Using a private water source for cattle and not introducing new cattle into the farm were associated with lower prevalence of M. bovis skin positivity. The herd-level prevalence of M. bovis reactors in Kashaari County of Mbarara District was 14.5 %, and the individual cattle prevalence was low (2.1 %).

Conclusions

Using communal sources of drinking water for cattle and introducing new cattle on the farm were farm management practices associated with increased risk of M. bovis exposure in cattle. Despite the low prevalence of bovine tuberculosis (TB), there is a need to educate the populace on the possibility of human infection with zoonotic TB and for educating farmers on practices to reduce the risk of acquiring M. bovis in the Mbarara District.     

Potential use of lymphocyte blastogenic responses in diagnosis of bovine tuberculosis

A comparison of in vitro lymphocyte responses and delayed type tuberculin skin test responses was made in an animal experimentally exposed to a Mycobacterium bovis-infected animal and in cattle naturally infected with M. bovis. Tuberculin skin tests did not suppress in vitro lymphocyte responses to M. bovis PPD and to M. avium PPD tuberculin. The whole blood test used in these studies provided for considerable savings in time as compared to use of purified lymphocytes for evaluating in vitro cellular responses. Variations in the responsiveness of lymphocytes to specific mycobacterial antigens was observed, therefore, it is recommended that profiles be established using three or more tests conducted at 14-day intervals.     

Trace micro-nutrients may affect susceptibility to bovine tuberculosis in cattle

Bovine tuberculosis (bTB) is a continuing problem in British herds. Micro-nutrients are important for the maintenance of well-functioning immune system. The aim of this study was to determine whether the selenium, copper and vitamin B12 status of cattle was associated with Mycobacterium bovis (M. bovis) infection. Between 2002 and 2005, 200 cattle (43% dairy, mean age 4.6 years), reactors according to the standard interpretation of the tuberculin test, and 200 in-contacts (41% dairy, mean age 4.4 years) non-reactors, which had been in contact with cattle with bTB, were selected from herds in England and Wales. Levels of the seleno enzyme glutathione peroxidase (GSHPx), copper and vitamin B12 were measured in blood. Confirmation of bTB infection was made by bacteriological culture and histopathology following a detailed postmortem. Levels of selenium and copper were also measured in a random sample of 63 livers. bTB was confirmed by culture/histology in 23/200 (11.5%) of in-contacts and 110/200 (55%) of reactors. In blood drawn at recruitment, GSHPx was lower in cattle with confirmed bTB compared to other cattle (geometric means 59.7 u/mL versus 78.9 u/mL red blood cells (RBC), p < 0.01). Vitamin B12 was similar (geometric means 161.5 pmol/L versus 165.5 pmol/L, p = 0.62) and copper was similar (geometric means 14.4 μmol/L versus 14.1 μmol/L, p = 0.55). In logistic regression models including all micro-nutrients simultaneously and controlling for age, sex, animal production class, herd size, number of reactors, postmortem laboratory and seasonal trends, lower levels of GSHPx (adjusted OR 0.42, 95% CI 0.21–0.81 per 100 u/mL RBC, p = 0.01) and higher levels of copper (adjusted OR 1.69, 95% CI 1.21–2.36 per 5 μmol/L, p < 0.01) were associated with an increased risk of confirmed bTB but there was no association with vitamin B12. There was evidence for a stronger association between confirmed bTB and GSHPx in in-contacts (adjusted OR 0.21, 95% CI 0.06–0.79 per 100 u/mL RBC) compared to reactors (adjusted OR 0.50, 95% CI 0.21–1.23 per 100 u/mL RBC) (p = 0.08 for interaction). Lower liver copper was associated with a higher risk of confirmed bTB (adjusted OR 0.15, 95% CI 0.02–1.0 per 5000 μmol/kg dry mass, p = 0.05) but there was no association between liver selenium and bTB. Trace micro-nutrient status may affect susceptibility to M. bovis infection in cattle. Further studies are needed.     

The effect of various Mycoplasma bovis isolates on bovine leukocyte responses

Mycoplasma bovis (M. bovis) contributes to a number of clinical syndromes in cattle; in particular, chronic pneumonia that is poorly responsive to therapy has been increasingly recognized as an important cause of morbidity, mortality, and financial loss. M. bovis impairs host immune function, but little is known about whether field isolates vary significantly in their effect on immune function. This research tested the hypothesis that different field isolates vary in their ability to suppress cellular metabolism and cellular production of radical oxygen species (ROS) by bovine leukocytes. Total blood leukocytes from 6 cattle were exposed to six field isolates, two diagnostic lab isolates, and two high passage laboratory isolates of M. bovis, and ROS production was measured by oxidation of dihydrorhodamine 123 (DHR-123). Cellular metabolism was measured by reduction of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT). Significant differences in the response to some field isolates were identified. Three field isolates and both diagnostic lab isolates significantly decreased ROS production by leukocytes from multiple cattle, while the high pass laboratory isolates did not. In contrast, MTT reduction was not significantly impaired by any of the M. bovis strains tested. M. bovis impairs ROS production by bovine leukocytes; the magnitude of the effect appears to be isolate-dependent, and is not related to a general impairment of cellular metabolism. Chronic M. bovis infection in some cattle may be related to impaired ability of leukocytes to produce ROS when exposed to M. bovis.     

Mycoplasma bovis infections in Swiss dairy cattle: a clinical investigation

Mycoplasma bovis causes mastitis in dairy cows and is associated with pneumonia and polyarthritis in cattle. The present investigation included a retrospective case–control study to identify potential herd-level risk factors for M. bovis associated disease, and a prospective cohort study to evaluate the course of clinical disease in M. bovis infected dairy cattle herds in Switzerland. Eighteen herds with confirmed M. bovis cases were visited twice within an average interval of 75 d. One control herd with no history of clinical mycoplasmosis, matched for herd size, was randomly selected within a 10 km range for each case herd. Animal health data, production data, information on milking and feeding-management, housing and presence of potential stress- factors were collected. Composite quarter milk samples were aseptically collected from all lactating cows and 5% of all animals within each herd were sampled by nasal swabs. Organ samples of culled diseased cows were collected when logistically possible. All samples were analyzed by real-time polymerase chain reaction (PCR). In case herds, incidence risk of pneumonia, arthritis and clinical mastitis prior to the first visit and incidence rates of clinical mastitis and clinical pneumonia between the two visits was estimated. Logistic regression was used to identify potential herd-level risk factors for M. bovis infection. In case herds, incidence risk of M. bovis mastitis prior to the first visit ranged from 2 to 15%, whereas 2 to 35% of the cows suffered from clinical pneumonia within the 12 months prior to the first herd visit. The incidence rates of mycoplasmal mastitis and clinical pneumonia between the two herd visits were low in case herds (0–0.1 per animal year at risk and 0.1-0.6 per animal year at risk, respectively). In the retrospective-case-control study high mean milk production, appropriate stimulation until milk-let-down, fore-stripping, animal movements (cattle shows and trade), presence of stress-factors, and use of a specific brand of milking equipment, were identified as potential herd-level risk factors. The prospective cohort study revealed a decreased incidence of clinical disease within three months and prolonged colonization of the nasal cavity by M. bovis in young stock.     

Molecular genotyping of Mycobacterium bovis isolated from cattle tissues in the North West Region of Cameroon

An epidemiological study was carried out to determine the Mycobacterium bovis strains causing bovine tuberculosis (TB) in cattle in North West Cameroon. Suspected TB lesions from slaughtered cattle were cultured on Lowenstein–Jensen and Middlebrook 7 H9 media to isolate mycobacteria agents for molecular genotyping using deletion analysis and spoligotyping. PCR-based genomic deletion typing showed that 54 of 103 tubercle bacilli isolated from cattle tissue were M. bovis strains and the African 1 clonal complex was widespread in affected cattle. Spoligotyping analysis revealed a closely related group of five M. bovis strains. SB0953, the dominant spoligotype pattern, and four new patterns identified as SB2161, SB2162, SB2663 and SB2664 according to the www.Mbovis.org international spoligotype database were identified. These spoligotypes were similar to other M. bovis strains recovered from bordering regions and other parts of Africa. The findings provided useful facts on the zoonotic risks of bovine TB and overwhelming evidence of the significance of M. bovis infection to human TB in the North West Region of Cameroon. The study revealed that bovine TB was widespread in cattle destined for human consumption and also has important implications for the control of TB in animals and humans in Cameroon.