Cat-to-Human Transmission of Mycobacterium bovis, United Kingdom.

Human infection with Mycobacterium bovis is reported infrequently in the United Kingdom. Most cases involve previous consumption of unpasteurized milk. We report a rare occurrence of 2 incidents of cat-to-human transmission of M. bovis during a cluster of infection in cats.

Ageing is associated with molecular signatures of inflammation and type 2 diabetes in rat pancreatic islets.

AIMS/HYPOTHESIS: Ageing is a major risk factor for development of metabolic diseases such as type 2 diabetes. Identification of the mechanisms underlying this association could help to elucidate the relationship between age-associated progressive loss of metabolic health and development of type 2 diabetes. We aimed to determine molecular signatures during ageing in the endocrine pancreas. METHODS: Global gene transcription was measured in pancreatic islets isolated from young and old rats by Ilumina BeadChip arrays. Promoter DNA methylation was measured by Sequenom MassArray in 46 genes that showed differential expression with age, and correlations with expression were established. Alterations in morphological and cellular processes with age were determined by immunohistochemical methods. RESULTS: Age-related changes in gene expression were found at 623 loci (>1.5-fold, false discovery rate [FDR] <5%), with a significant (FDR < 0.05) enrichment in genes previously implicated in islet-cell function (Enpp1, Abcc8), type 2 diabetes (Tspan8, Kcnq1), inflammatory processes (Cxcl9, Il33) and extracellular matrix organisation (Col3a1, Dpt). Age-associated transcriptional differences negatively correlated with promoter DNA methylation at several loci related to inflammation, glucose homeostasis, cell proliferation and cell-matrix interactions (Il33, Cxcl9, Gpr119, Fbp2, Col3a1, Dpt, Spp1). CONCLUSIONS/INTERPRETATION: Our findings suggest that a significant proportion of pancreatic islets develop a low-grade 'chronic' inflammatory status with ageing and this may trigger altered functional plasticity. Furthermore, we identified changes in expression of genes previously linked to type 2 diabetes and associated changes in DNA methylation that could explain their age-associated dysregulation. These findings provide new insights into key (epi)genetic signatures of the ageing process in islets.

Maternal diet and aging alter the epigenetic control of a promoter-enhancer interaction at the Hnf4a gene in rat pancreatic islets.

Environmental factors interact with the genome throughout life to determine gene expression and, consequently, tissue function and disease risk. One such factor that is known to play an important role in determining long-term metabolic health is diet during critical periods of development. Epigenetic regulation of gene expression has been implicated in mediating these programming effects of early diet. The precise epigenetic mechanisms that underlie these effects remain largely unknown. Here, we show that the transcription factor Hnf4a, which has been implicated in the etiology of type 2 diabetes (T2D), is epigenetically regulated by maternal diet and aging in rat islets. Transcriptional activity of Hnf4a in islets is restricted to the distal P2 promoter through its open chromatin configuration and an islet-specific interaction between the P2 promoter and a downstream enhancer. Exposure to suboptimal nutrition during early development leads to epigenetic silencing at the enhancer region, which weakens the P2 promoter-enhancer interaction and results in a permanent reduction in Hnf4a expression. Aging leads to progressive epigenetic silencing of the entire Hnf4a locus in islets, an effect that is more pronounced in rats exposed to a poor maternal diet. Our findings provide evidence for environmentally induced epigenetic changes at the Hnf4a enhancer that alter its interaction with the P2 promoter, and consequently determine T2D risk. We therefore propose that environmentally induced changes in promoter-enhancer interactions represent a fundamental epigenetic mechanism by which nutrition and aging can influence long-term health.

Splitting of a prevalent Mycobacterium bovis spoligotype by variable-number tandem-repeat typing reveals high heterogeneity in an evolving clonal group.

Mycobacterium bovis populations in countries with persistent bovine tuberculosis usually show a prevalent spoligotype with a wide geographical distribution. This study applied mycobacterial interspersed repetitive-unit-variable-number tandem-repeat (MIRU-VNTR) typing to a random panel of 115 M. bovis isolates that are representative of the most frequent spoligotype in the Iberian Peninsula, SB0121. VNTR typing targeted nine loci: ETR-A (alias VNTR2165), ETR-B (VNTR2461), ETR-D (MIRU4, VNTR580), ETR-E (MIRU31, VNTR3192), MIRU26 (VNTR2996), QUB11a (VNTR2163a), QUB11b (VNTR2163b), QUB26 (VNTR4052), and QUB3232 (VNTR3232). We found a high degree of diversity among the studied isolates (discriminatory index [D] = 0.9856), which were split into 65 different MIRU-VNTR types. An alternative short-format MIRU-VNTR typing targeting only the four loci with the highest variability values was found to offer an equivalent discriminatory index. Minimum spanning trees using the MIRU-VNTR data showed the hypothetical evolution of an apparent clonal group. MIRU-VNTR analysis was also applied to the isolates of 176 animals from 15 farms infected by M. bovis SB0121; in 10 farms, the analysis revealed the coexistence of two to five different MIRU types differing in one to six loci, which highlights the frequency of undetected heterogeneity.

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.

African 2, a clonal complex of Mycobacterium bovis epidemiologically important in East Africa.

We have identified a clonal complex of Mycobacterium bovis isolated at high frequency from cattle in Uganda, Burundi, Tanzania, and Ethiopia. We have named this related group of M. bovis strains the African 2 (Af2) clonal complex of M. bovis. Af2 strains are defined by a specific chromosomal deletion (RDAf2) and can be identified by the absence of spacers 3 to 7 in their spoligotype patterns. Deletion analysis of M. bovis isolates from Algeria, Mali, Chad, Nigeria, Cameroon, South Africa, and Mozambique did not identify any strains of the Af2 clonal complex, suggesting that this clonal complex of M. bovis is localized in East Africa. The specific spoligotype pattern of the Af2 clonal complex was rarely identified among isolates from outside Africa, and the few isolates that were found and tested were intact at the RDAf2 locus. We conclude that the Af2 clonal complex is localized to cattle in East Africa. We found that strains of the Af2 clonal complex of M. bovis have, in general, four or more copies of the insertion sequence IS6110, in contrast to the majority of M. bovis strains isolated from cattle, which are thought to carry only one or a few copies.

Poor maternal nutrition leads to alterations in oxidative stress, antioxidant defense capacity, and markers of fibrosis in rat islets: potential underlying mechanisms for development of the diabetic phenotype in later life.

Low birth weight is associated with glucose intolerance, insulin resistance, and type 2 diabetes (T2D) in later life. Good evidence indicates that the environment plays an important role in this relationship. However, the mechanisms underlying these relationships are defined poorly. Islets are particularly susceptible to oxidative stress, and this condition combined with fibrosis is thought to be instrumental in T2D pathogenesis. Here we use our maternal low-protein (LP) rat model to determine the effect of early diet on oxidative stress and fibrosis in pancreatic islets of male offspring at 3 and 15 mo of age. Islet xanthine oxidase (XO) expression was increased in 15-mo LP offspring, which suggests increased oxidative-stress. Manganese superoxide-dismutase (MnSOD), copper-zinc superoxide dismutase (CuZnSOD), and heme oxygenase-1 (HO-1) (antioxidant enzymes) were reduced significantly in LP offspring, which indicated impairment of oxidative defense. Expression of fibrosis markers collagen I and collagen III also increased in 15-mo LP offspring. Angiotensin II receptor type I (AT(II)R(1)), induced by hyperglycemia and oxidative-stress, was significantly up-regulated in 15-mo LP offspring. Lipid peroxidation was also increased in 15-mo LP animals. We conclude that maternal protein restriction causes age-associated increased oxidative stress, impairment of oxidative defense, and fibrosis. These findings provide mechanisms by which suboptimal early nutrition can lead to T2D development later in life.

African 1, an epidemiologically important clonal complex of Mycobacterium bovis dominant in Mali, Nigeria, Cameroon, and Chad.

We have identified a clonal complex of Mycobacterium bovis present at high frequency in cattle in population samples from several sub-Saharan west-central African countries. This closely related group of bacteria is defined by a specific chromosomal deletion (RDAf1) and can be identified by the absence of spacer 30 in the standard spoligotype typing scheme. We have named this group of strains the African 1 (Af1) clonal complex and have defined the spoligotype signature of this clonal complex as being the same as the M. bovis BCG vaccine strain but with the deletion of spacer 30. Strains of the Af1 clonal complex were found at high frequency in population samples of M. bovis from cattle in Mali, Cameroon, Nigeria, and Chad, and using a combination of variable-number tandem repeat typing and spoligotyping, we show that the population of M. bovis in each of these countries is distinct, suggesting that the recent mixing of strains between countries is not common in this area of Africa. Strains with the Af1-specific deletion (RDAf1) were not identified in M. bovis isolates from Algeria, Burundi, Ethiopia, Madagascar, Mozambique, South Africa, Tanzania, and Uganda. Furthermore, the spoligotype signature of the Af1 clonal complex has not been identified in population samples of bovine tuberculosis from Europe, Iran, and South America. These observations suggest that the Af1 clonal complex is geographically localized, albeit to several African countries, and we suggest that the dominance of the clonal complex in this region is the result of an original introduction into cows naïve to bovine tuberculosis.

Adverse effects of reduced oxygen tension on the proliferative capacity of rat kidney and insulin-secreting cell lines involve DNA damage and stress responses.

Standard cell culture conditions do not reflect the physiological environment in terms of oxygen tension (20% vs 3%). The effects of lowering oxygen tension on cell proliferation in culture can be beneficial as well as detrimental depending on the cell line studied, but the molecular mechanism underlying such effects is not fully understood. We observed that the proliferative capacity of the rat cell lines NRK and INS-1 was inhibited when cultured under 3% oxygen as compared to 20% oxygen. Suppression of proliferation in NRK cells was accompanied by induction of DNA double strand breaks whereas in INS-1 cells it was accompanied by up-regulation of p53 and p27. Although Sirt1 was up-regulated in both cell lines by 3% oxygen the effects on antioxidant enzymes (MnSOD, CuZnSOD and catalase) were cell line specific. Marked up-regulation of heme oxygenase-1 (HO-1) was detected in both NRK and INS-1 cells when cultured in 3% oxygen. HO-1 expression can be readily induced by exposure to hydrogen peroxide in culture. These results suggest that reduced oxygen tension suppresses the proliferative capacity of these two cell lines through a stress response that is similar to an oxidative stress response but the molecular events that lead to the reduced cell proliferation are cell line specific.

Molecular characterization of Mycobacterium bovis strains isolated from cattle slaughtered at two abattoirs in Algeria.

BACKGROUND: Bovine Tuberculosis is prevalent in Algeria despite governmental attempts to control the disease. The objective of this study was to conduct, for the first time, molecular characterization of a population sample of Mycobacterium bovis strains isolated from slaughter cattle in Algeria. Between August and November 2007, 7250 animals were consecutively screened at the abattoirs of Algiers and Blida. In 260 animals, gross visible granulomatous lesions were detected and put into culture. Bacterial isolates were subsequently analysed by molecular methods. RESULTS: Altogether, 101 bacterial strains from 100 animals were subjected to molecular characterization. M. bovis was isolated from 88 animals. Other bacteria isolated included one strain of M. caprae, four Rhodococcus equi strains, three Non-tuberculous Mycobacteria (NTM) and five strains of other bacterial species. The M. bovis strains isolated showed 22 different spoligotype patterns; four of them had not been previously reported. The majority of M. bovis strains (89%) showed spoligotype patterns that were previously observed in strains from European cattle. Variable Number of Tandem Repeat (VNTR) typing supported a link between M. bovis strains from Algeria and France. One spoligotype pattern has also been shown to be frequent in M. bovis strains from Mali although the VNTR pattern of the Algerian strains differed from the Malian strains. CONCLUSION: M. bovis infections account for a high amount of granulomatous lesions detected in Algerian slaughter cattle during standard meat inspection at Algiers and Blida abattoir. Molecular typing results suggested a link between Algerian and European strains of M. bovis.

Cluster of human tuberculosis caused by Mycobacterium bovis: evidence for person-to-person transmission in the UK.

BACKGROUND: Despite a recent resurgence in the incidence of bovine tuberculosis in UK cattle herds, no associated rise in the number of cases in man has been noted. Disease due to human Mycobacterium bovis infection usually occurs in older patients, in whom drinking unpasteurised milk in the past is the probable source of infection. Person-to-person transmission is very rare. METHODS: After identification of two epidemiologically-linked cases of human M bovis infection through routine laboratory and surveillance activities, all patients identified with M bovis infection in the Midlands from 2001-05 (n=20) were assessed by DNA fingerprinting (MIRU-VNTR and spoligotyping), with additional interviews for patients with a clustered strain. FINDINGS: A cluster of six cases was identified. All clustered cases were young and UK-born; five patients had pulmonary disease, and one patient died due to M bovis meningitis, with four patients possessing factors predisposing to tuberculosis. All patients had common social links through visits to bars in two different areas. With the exception of the first case, there was an absence of zoonotic links or consumption of unpasteurised dairy products, suggesting that person-to-person transmission had occurred. INTERPRETATION: This report of several instances of M bovis transmission between people in a modern urban setting emphasises the need to maintain control measures for human and bovine tuberculosis. Transmission and subsequent disease was probably due to a combination of host and environmental factors. Prospective surveillance and DNA fingerprinting identified the cluster, enabling health protection teams to set up control measures and prevent further transmission.

Molecular characterisation of Mycobacterium bovis isolated from cattle slaughtered at the Bamako abattoir in Mali.

BACKGROUND: Mali is one of the most important livestock producers of the Sahel region of Africa. A high frequency of bovine tuberculosis (BTB) has been reported but surveillance and control schemes are restricted to abattoir inspections only. The objective of this study was to conduct, for the first time, molecular characterisation of Mycobacterium bovis strains isolated from cattle slaughtered at the Bamako abattoir. Of 3330 animals screened only 60 exhibited gross visible lesions. From these animals, twenty strains of M. bovis were isolated and characterised by spoligotyping. RESULTS: Organ lesions typical of BTB were most often detected in the liver, followed by the lung and the peritoneum. M. bovis was isolated from 20 animals and 7 different spoligotypes were observed among these 20 strains; three of the patterns had not been previously reported. Spoligotype patterns from thirteen of the strains lacked spacer 30, a characteristic common in strains of M. bovis found in Chad, Cameroon and Nigeria. However, unlike the other three Central African countries, the majority of spoligotype patterns observed in Mali also lacked spacer 6. Of the remaining seven strains, six had spoligotype patterns identical to strains commonly isolated in France and Spain. CONCLUSION: Two groups of M. bovis were detected in cattle slaughtered at the Bamako abattoir. The spoligotype pattern of the first group has similarities to strains previously observed in Chad, Cameroon and Nigeria. The additional absence of spacer 6 in the majority of these strains suggests a Mali specific clone. The spoligotype patterns of the remaining strains suggest that they may have been of European origin.

Molecular characterisation of Mycobacterium caprae strains isolated in Poland.

Bovine tuberculosis (bovine TB, bTB) is caused by bovine bacilli: Mycobacterium bovis and M caprae The studies conducted in Poland, in the National Bovine Tuberculosis Reference Laboratory in the Department of Microbiology of the National Veterinary Research Institute in Pulawy, show that animal tuberculosis in Poland is also caused by M caprae We here describe the identification and genotypic assessment of 52 isolates of M caprae obtained from Polish cattle and wild animals over the last five years. We show that strains isolated from bison have significant genotypic diversity and are distinct compared with the genotypes of strains isolated from cattle. Similarly, isolates from cattle herds can be highly genotypically variable. Formal designation of the members of the Mycobacterium tuberculosis complex is controversial in Poland; there is a gap in veterinary legislation with regard to bTB and no explicit mention of M caprae causing tuberculosis in animal.

Recent advances in our knowledge of Mycobacterium bovis: a feeling for the organism.

Significant and rapid progress has been made in our knowledge and understanding of Mycobacterium bovis since the last international M. bovis conference 5 years ago. Much of this progress has been underpinned by the completion of the genome sequence. This important milestone has catalysed research into the development of a number of improved tools with which to combat bovine tuberculosis. In this article we will review recent progress made in the development of these tools and in our understanding of the organism, its evolution and spread. Comparison of the genome sequence with those of other members of the Mycobacterium tuberculosis complex has enabled insights into the evolution of M. bovis. This analysis also indicates that the M. tuberculosis complex have the propensity to adapt to new host species. The use of high throughput molecular typing methods has revealed that the recent bovine tuberculosis epidemic in Great Britain is being driven by a number of clonal expansions, which cannot be explained by random mutation and drift alone. Completion of a number of mycobacterial genome sequences has allowed the development of antigen mining techniques that rapidly identify M. bovis-specific genes. These can then be used as reagents in the gamma interferon assay to increase the specificity of the assay and also to discriminate between Bacillus of Calmette and Guérin (BCG) vaccinated animals and those infected with M. bovis. In the longer term, comparisons between the genomes of M. bovis and BCG will allow insight into how BCG became attenuated following serial passage on artificial growth media and reveal clues into how to improve the vaccine efficacy of BCG.

Investigation of the cause of geographic disparities in IDEXX ELISA sensitivity in serum samples from Mycobacterium bovis-infected cattle.

Accurately identifying Mycobacterium bovis-infected cattle is critical for bovine tuberculosis prevention and control. One method for identifying infected cattle is an ELISA developed by IDEXX laboratories, which detects antibodies to two M. bovis proteins, MPB70 and MPB83. The assay's sensitivity varies by geographic region, with sensitivities of 77%, 45%, and 9% in bovine serum samples from the United Kingdom (n = 126), the United States (n = 146), and Mexico (n = 128), respectively. We hypothesized that geographically-biased sequence variation in mpb70 and mpb83, or in the genes that regulate their expression (sigK and rskA), may explain these differing sensitivities. This hypothesis was tested by comparing the sequences of these four genes in 455 M. bovis strains isolated from cattle in the aforementioned countries. For each gene, a single, common sequence was identified in most genomes of the M. bovis strains collected in all three countries. Twelve of the 455 strains were isolated from infected cattle for which the IDEXX ELISA was also performed. Five of the seven ELISA-positive genomes and three of the five ELISA-negative genomes contained the most common sequence of all four genes. Thus, sequence variation in mpb70, mpb83, sigK, and rskA does not explain the geographic disparities in IDEXX ELISA sensitivity.

Why doesn't bovine tuberculosis transmit between humans?

Tuberculosis caused by Mycobacterium tuberculosis is an important bacterial pathogen of man. This human-adapted pathogen was ancestral to a lineage of animal-adapted strains which cause similar disease in many different mammals but are unable to transmit between humans. How did the animal-adapted strains lose the ability to transmit between humans?

Overview and phylogeny of Mycobacterium tuberculosis complex organisms: implications for diagnostics and legislation of bovine tuberculosis.

Members of the Mycobacterium tuberculosis complex (MTBC) cause a serious disease with similar pathology, tuberculosis; in this review, bovine tuberculosis will be considered as disease caused by any member of the MTBC in bovids. Bovine tuberculosis is responsible for significant economic loss due to costly eradication programs and trade limitations and poses a threat to both endangered and protected species as well as to public health. We here give an overview on all members of the MTBC, focusing on their isolation from different animal hosts. We also review the recent advances made in elucidating the evolutionary and phylogenetic relationships of members of the MTBC. Because the nomenclature of the MTBC is controversial, its members have been considered species, subspecies or ecotypes, this review discusses the possible implications for diagnostics and the legal consequences of naming of new species.

Mycobacterium bovis infection in domestic pigs in Great Britain.

Mycobacterium bovis, the causative agent of bovine tuberculosis (TB), infects a wide range of wild and domestic mammals. Despite a control programme spanning decades, M. bovis infection levels in cattle in Great Britain (GB) have continued to rise over recent years. As the incidence of infection in cattle and wildlife may be linked to that in swine, data relating to infection of pigs identified at slaughter were examined in this study. Between 2007 and 2011, almost all M. bovis-infected pigs originated from farms in the South-West and West-Midland regions of England. The data suggest that pigs raised outdoors or on holdings with poor biosecurity may be more vulnerable to infection with M. bovis. In the majority of cases, the same strains of M. bovis were found in pigs and cattle, despite that fact that direct contact between these species was rarely observed. Genotyping and geographical mapping data indicated that some strains found in pigs may correlate better with those present in badgers, rather than cattle. In consequence, it is proposed that pigs may represent a useful sentinel for M. bovis infection in wildlife in GB. Given the potential implications of this infection for the pig industry, and for the on-going effort to control bovine TB, the importance of understanding the epidemiology and pathogenesis of M. bovis infection, as well as monitoring its prevalence, in pigs should not be underestimated.