Tonsils of the soft palate do not mediate the response of pigs to oral vaccination with heat-inactivated Mycobacterium bovis

Mycobacterium bovis causes animal tuberculosis (TB) in cattle, humans, and other mammalian species, including pigs. The goal of this study was to experimentally assess the responses of pigs with and without a history of tonsillectomy to oral vaccination with heat-inactivated M. bovis and challenge with a virulent M. bovis field strain, to compare pig and wild boar responses using the same vaccination model as previously used in the Eurasian wild boar (Sus scrofa), to evaluate the use of several enzyme-linked immunosorbent assays (ELISAs) and lateral flow tests for in vivo TB diagnosis in pigs, and to verify if these tests are influenced by oral vaccination with inactivated M. bovis. At necropsy, the lesion and culture scores were 20% to 43% higher in the controls than those in the vaccinated pigs. Massive M. bovis growth from thoracic tissue samples was observed in 4 out of 9 controls but in none of the 10 vaccinated pigs. No effect of the presence or absence of tonsils was observed on these scores, suggesting that tonsils are not involved in the protective response to this vaccine in pigs. The serum antibody levels increased significantly only after challenge. At necropsy, the estimated sensitivities of the ELISAs and dual path platform (DPP) assays ranged from 89% to 94%. In the oral mucosa, no differences in gene expression were observed in the control group between the pigs with and without tonsils. In the vaccinated group, the mRNA levels for chemokine (C-C motif) receptor 7 (CCR7), interferon beta (IFN-β), and methylmalonyl coenzyme A mutase (MUT) were higher in pigs with tonsils. Complement component 3 mRNA levels in peripheral blood mononuclear cells (PBMC) increased with vaccination and decreased after M. bovis challenge. This information is relevant for pig production in regions that are endemic for M. bovis and for TB vaccine research.

An inter-laboratory validation of a real time PCR assay to measure host excretion of bacterial pathogens, particularly of Mycobacterium bovis

Advances in the diagnosis of Mycobacterium bovis infection in wildlife hosts may benefit the development of sustainable approaches to the management of bovine tuberculosis in cattle. In the present study, three laboratories from two different countries participated in a validation trial to evaluate the reliability and reproducibility of a real time PCR assay in the detection and quantification of M. bovis from environmental samples. The sample panels consisted of negative badger faeces spiked with a dilution series of M. bovis BCG Pasteur and of field samples of faeces from badgers of unknown infection status taken from badger latrines in areas with high and low incidence of bovine TB (bTB) in cattle. Samples were tested with a previously optimised methodology. The experimental design involved rigorous testing which highlighted a number of potential pitfalls in the analysis of environmental samples using real time PCR. Despite minor variation between operators and laboratories, the validation study demonstrated good concordance between the three laboratories: on the spiked panels, the test showed high levels of agreement in terms of positive/negative detection, with high specificity (100%) and high sensitivity (97%) at levels of 10(5) cells g(-1) and above. Quantitative analysis of the data revealed low variability in recovery of BCG cells between laboratories and operators. On the field samples, the test showed high reproducibility both in terms of positive/negative detection and in the number of cells detected, despite low numbers of samples identified as positive by any laboratory. Use of a parallel PCR inhibition control assay revealed negligible PCR-interfering chemicals co-extracted with the DNA. This is the first example of a multi-laboratory validation of a real time PCR assay for the detection of mycobacteria in environmental samples. Field studies are now required to determine how best to apply the assay for population-level bTB surveillance in wildlife.

Endemic zoonoses: a one health approach

Endemic zoonotic diseases remain a serious but poorly recognised problem in affected communities in developing countries. Despite the overall burden of zoonoses on human and animal health, information about their impacts in endemic settings is lacking and most of these diseases are continuously being neglected. The non-specific clinical presentation of these diseases has been identified as a major challenge in their identification (even with good laboratory diagnosis), and control. The signs and symptoms in animals and humans respectively, are easily confused with other non-zoonotic diseases, leading to widespread misdiagnosis in areas where diagnostic capacity is limited. The communities that are mostly affected by these diseases live in close proximity with their animals which they depend on for livelihood, which further complicates the understanding of the epidemiology of zoonoses. This thesis reviewed the pattern of reporting of zoonotic pathogens that cause febrile illness in malaria endemic countries, and evaluates the recognition of animal associations among other risk factors in the transmission and management of zoonoses. The findings of the review chapter were further investigated through a laboratory study of risk factors for bovine leptospirosis, and exposure patterns of livestock coxiellosis in the subsequent chapters. A review was undertaken on 840 articles that were part of a bigger review of zoonotic pathogens that cause human fever. The review process involves three main steps: filtering and reference classification, identification of abstracts that describe risk factors, and data extraction and summary analysis of data. Abstracts of the 840 references were transferred into a Microsoft excel spread sheet, where several subsets of abstracts were generated using excel filters and text searches to classify the content of each abstract. Data was then extracted and summarised to describe geographical patterns of the pathogens reported, and determine the frequency animal related risk factors were considered among studies that investigated risk factors for zoonotic pathogen transmission. Subsequently, a seroprevalence study of bovine leptospirosis in northern Tanzania was undertaken in the second chapter of this thesis. The study involved screening of serum samples, which were obtained from an abattoir survey and cross-sectional study (Bacterial Zoonoses Project), for antibodies against Leptospira serovar Hardjo. The data were analysed using generalised linear mixed models (GLMMs), to identify risk factors for cattle infection. The final chapter was the analysis of Q fever data, which were also obtained from the Bacterial Zoonoses Project, to determine exposure patterns across livestock species using generalized linear mixed models (GLMMs). Leptospira spp. (10.8%, 90/840) and Rickettsia spp. (10.7%, 86/840) were identified as the most frequently reported zoonotic pathogens that cause febrile illness, while Rabies virus (0.4%, 3/840) and Francisella spp. (0.1%, 1/840) were least reported, across malaria endemic countries. The majority of the pathogens were reported in Asia, and the frequency of reporting seems to be higher in areas where outbreaks are mostly reported. It was also observed that animal related risk factors are not often considered among other risk factors for zoonotic pathogens that cause human fever in malaria endemic countries. The seroprevalence study indicated that Leptospira serovar Hardjo is widespread in cattle population in northern Tanzania, and animal husbandry systems and age are the two most important risk factors that influence seroprevalence. Cattle in the pastoral systems and adult cattle were significantly more likely to be seropositive compared to non-pastoral and young animals respectively, while there was no significant effect of cattle breed or sex. Exposure patterns of Coxiella burnetii appear different for each livestock species. While most risk factors were identified for goats (such as animal husbandry systems, age and sex) and sheep (animal husbandry systems and sex), there were none for cattle. In addition, there was no evidence of a significant influence of mixed livestock-keeping on animal coxiellosis. Zoonotic agents that cause human fever are common in developing countries. The role of animals in the transmission of zoonotic pathogens that cause febrile illness is not fully recognised and appreciated. Since Leptospira spp. and C. burnetii are among the most frequently reported pathogens that cause human fever across malaria endemic countries, and are also prevalent in livestock population, control and preventive measures that recognise animals as source of infection would be very important especially in livestock-keeping communities where people live in close proximity with their animals.

Clínica e cirurgia em espécies pecuárias

Este relatório foi realizado no âmbito da conclusão do mestrado integrado em medicina veterinária e está estruturado em duas partes. Na primeira parte deste relatório é apresentada a casuística acompanhada ao longo do estágio curricular e a segunda parte é constituída por uma revisão bibliográfica subordinada ao tema, importância da ecografia em reprodução de bovinos, seguida da exposição de um caso clínico sobre orquite em bovinos e a discussão do mesmo; Abstract: clinic and surgery in livestock species This report was carried out under the curriculum of the integrated master in veterinary medicine and is structured in two parts. In the first part of this report is presented the casuistry observed throughout the traineeship and in the second part was made a literature review on the topic ultrasound importance in bovine reproduction, followed by exposure of one clinical case of orchitis in bovine and is respective discussion.

Clínica médica e cirúrgica em espécies pecuárias

O presente relatório foi elaborado com o objetivo de descrever as atividades desenvolvidas no âmbito do estágio curricular do Mestrado Integrado em Medicina Veterinária pela Universidade de Évora. Numa primeira parte é apresentada a casuística acompanhada ao longo do estágio, com referência mais pormenorizada a alguns dos casos clínicos acompanhados nas diversas áreas de intervenção da clínica de espécies pecuarias. A segunda parte deste relatório é composta por uma breve revisão bibliográfica sobre o tema “Cetose em bovinos”. Termina com um estudo que tem como objetivo principal avaliar a relação entre a ocorrência de cetose e os valores do índice de condição corporal no pré-parto e nas primeiras semanas de lactação. Os resultados não evidenciaram qualquer associação estatisticamente significativa entre os valores do índicie de condição corporal e a prevalência de cetose; Clinical medical and surgery in livestock species Abstract: This report was prepared in order to describe the activities developed during the integrated internship of the master's degree in Veterinary Medicine from the University of Evora. The first part describes the casuistics followed along the traineeship with more detailed reference to some of the clinical cases assisted in different areas of intervention of clinic livestock species. The second part of this report consists of a brief literature review on the topic "ketosis in cattle". It ends with a study that aims to evaluate the relationship between the occurrence of ketosis and the values of body condition score in antepartum and during the first weeks of lactation. The results did not show any statistically significant association between the values of body condition score and the prevalence of ketosis.

Clínica e cirurgia de espécies pecuárias

O presente relatório, inerente ao estágio curricular do Mestrado Integrado em Medicina Veterinária da Universidade de Évora, tem como objetivo descrever as atividades desenvolvidas durante o estágio na área da clínica e cirurgia de espécies pecuárias. A primeira parte do relatório consiste na descrição do local de estágio e na distribuição casuística das atividades acompanhadas. A segunda parte consiste numa revisão bibliográfica e no desenvolvimento de dois casos clínicos ocorridos em bovinos, relacionados com o tema, “necrobacilose hepática bovina” (NHB). A NHB é uma doença infecciosa de grande impacto económico, principalmente em explorações de bovinos de engorda intensiva. Carateriza-se pela formação de abcessos hepáticos, cujo agente etiológico primário é Fusobacterium necrophorum. É de difícil diagnóstico em vida, raramente demonstra sinais clínicos e o seu tratamento é pouco eficaz. A prevenção baseia-se na adição de antimicrobianos na alimentação, num correto maneio alimentar e eventualmente vacinação; Abstract: Medical and Surgical Pathology of livestock species The present report, inherent of the master in Veterinary Medicine from the University of Évora, aims to describe the activities carried out during the internship in the field of clinic and surgery of livestock species. The first part of the report is the description of the local of internship and the statistical distribution of activities. The second part consists of a literature review and the development of two clinical cases in cattle, related to the theme, “hepatic bovine necrobacillosis "(HBN). The HBN is an infectious disease of great economic impact, especially in feedlot cattle. It is characterized by the formation of liver abscesses, whose primary etiological agent is Fusobacterium necrophorum. It is difficult to diagnose in life, rarely show clinical signs and their treatment is ineffective. Prevention is based on the addition of antimicrobials in food, a correct feed management and eventually vaccination.

Bovine abortion associated with Neospora caninum: Diagnosis and epidemiological aspects of a dairy cattle herd in the northeast region of São Paulo state, Brazil

The objective of this study was to report the presence of Neospora caninum-associated abortion in bovines at a farm in the northeast region of São Paulo State. In January 2010, it was sent to the Department of Pathology, UNESP-Jaboticabal, a bovine fetus with an estimated age of seven months, which was natural of a dairy farm with 300 animals and an average daily production of 3,000 liters of milk, nearly 20 liters per cow. The animals were vaccinated against rabies, foot and mouth disease, carbuncle, brucellosis, leptospirosis, bovine herpes virus type I and bovine viral diarrhea virus. The herd consisted of purebred Holstein animals, Jersey, and mostly by crossbred animals 7-8 (gir x holstein). During necropsy, samples of the serosanguineous liquid present at the thoracic cavity and the heart of the fetus were collected for the detection of anti-Neospora caninum antibodies through Indirect Immunofluorescence Assay (IFA). Fragments of brain, cerebellum, tongue, liver, heart and kidneys were collected for the execution of histopathology (HP), immunohistochemistry (IHC) and Polymerase Chain Reaction. In order that IFA could be performed, the owner was requested blood samples without anticoagulants of the mother and other cows in the farm, with or without a history of abortion. At necropsy, it was verified a severe autolysis of the fetus. The serology of the fetus was 1:25, while the serology of the mother was 1:3,200. At HP, it was observed discrete multifocal non-suppurative encephalomyelitis characterized by gliosis and mononuclear inflammatory infiltration associated with cellular debris. DNA amplification of N. caninum was positive in fragments of brain, tongue, cerebellum, heart and kidneys. At IHC, it has been observed immunoreactivity to a cyst located in the tongue. The owner reported that his herd showed endemic episodes of abortion, while 27.69% (18/65) of the 65 animals sampled were seropositive. Although it has not been a significant difference (p>0.05), a higher seropositivity was observed in animals with a history of abortion (10/26) 38.46%, in comparison with animals without previous abortion (8/39) 20.51%. These findings show that the abortion under study was provoked by the protozoan N. caninum, while this is the first report concerning cattle in the northeast region of São Paulo State.

Epidemiology of Brucellosis and Q Fever in Linked Human and Animal Populations in Northern Togo

BACKGROUND: Although brucellosis (Brucella spp.) and Q Fever (Coxiella burnetii) are zoonoses of global importance, very little high quality data are available from West Africa. METHODS/PRINCIPAL FINDINGS: A serosurvey was conducted in Togo's main livestock-raising zone in 2011 in 25 randomly selected villages, including 683 people, 596 cattle, 465 sheep and 221 goats. Additionally, 464 transhumant cattle from Burkina Faso were sampled in 2012. The serological analyses performed were the Rose Bengal Test and ELISA for brucellosis and ELISA and the immunofluorescence assay (IFA) for Q Fever Brucellosis did not appear to pose a major human health problem in the study zone, with only 7 seropositive participants. B. abortus was isolated from 3 bovine hygroma samples, and is likely to be the predominant circulating strain. This may explain the observed seropositivity amongst village cattle (9.2%, 95%CI:4.3-18.6%) and transhumant cattle (7.3%, 95%CI:3.5-14.7%), with an absence of seropositive small ruminants. Exposure of livestock and people to C. burnetii was common, potentially influenced by cultural factors. People of Fulani ethnicity had greater livestock contact and a significantly higher seroprevalence than other ethnic groups (Fulani: 45.5%, 95%CI:37.7-53.6%; non-Fulani: 27.1%, 95%CI:20.6-34.7%). Appropriate diagnostic test cut-off values in endemic settings requires further investigation. Both brucellosis and Q Fever appeared to impact on livestock production. Seropositive cows were more likely to have aborted a foetus during the previous year than seronegative cows, when adjusted for age. This odds was 3.8 times higher (95%CI: 1.2-12.1) for brucellosis and 6.7 times higher (95%CI: 1.3-34.8) for Q Fever. CONCLUSIONS: This is the first epidemiological study of zoonoses in Togo in linked human and animal populations, providing much needed data for West Africa. Exposure to Brucella and C. burnetii is common but further research is needed into the clinical and economic impact.

A note on inbreeding in dairy cattle breeding

Starting from the study at the beginning of the East German "Heterosisfeldversuch", where PANICKE et al. (1975) considered the possibilities of a targeted use of inbreeding and heterotic effects, we show and discuss results of inbreeding studies in the USA dairy cattle breeding. Several research groups worldwide presented effective tools for managing inbreeding in dairy cattle. Their efforts underline the need of inbreeding studies. Contemplating inbreeding is necessary for any breeding decision to avoid inbreeding depression and for improved genetic analyses, e.g. in QTL- estimation. A novel methodology (HERNANDEZ-SANCHEZ et al., 2004a and b) is suggested for estimating inbreeding at the three levels of population, individual and locus.

Multilocus sequence analysis provides insights into molecular epidemiology of Chlamydia pecorum infections in Australian sheep, cattle and koalas

Chlamydia pecorum is a significant pathogen of domestic livestock and wildlife. We have developed a C. pecorum-specific multilocus sequence analysis (MLSA) scheme to examine the genetic diversity of and relationships between Australian sheep, cattle, and koala isolates. An MLSA of seven concatenated housekeeping gene fragments was performed using 35 isolates, including 18 livestock isolates (11 Australian sheep, one Australian cow, and six U.S. livestock isolates) and 17 Australian koala isolates. Phylogenetic analyses showed that the koala isolates formed a distinct clade, with limited clustering with C. pecorum isolates from Australian sheep. We identified 11 MLSA sequence types (STs) among Australian C. pecorum isolates, 10 of them novel, with koala and sheep sharing at least one identical ST (designated ST2013Aa). ST23, previously identified in global C. pecorum livestock isolates, was observed here in a subset of Australian bovine and sheep isolates. Most notably, ST23 was found in association with multiple disease states and hosts, providing insights into the transmission of this pathogen between livestock hosts. The complexity of the epidemiology of this disease was further highlighted by the observation that at least two examples of sheep were infected with different C. pecorum STs in the eyes and gastrointestinal tract. We have demonstrated the feasibility of our MLSA scheme for understanding the host relationship that exists between Australian C. pecorum strains and provide the first molecular epidemiological data on infections in Australian livestock hosts.

The Escherichia coli O157:H7 EhaB autotransporter protein binds to laminin and collagen I and induces a serum IgA response in O157:H7 challenged cattle

Enterohaemorrhagic Escherichia coli (EHEC) are a subgroup of Shiga toxin-producing E. coli that cause gastrointestinal disease with the potential for life-threatening sequelae. Cattle serve as the natural reservoir for EHEC and outbreaks occur sporadically as a result of contaminated beef and other farming products. While certain EHEC virulence mechanisms have been extensively studied, the factors that mediate host colonization are poorly defined. Previously, we identified four proteins (EhaA,B,C,D) from the prototypic EHEC strain EDL933 that belong to the autotransporter (AT) family. Here we characterize the EhaB AT protein. EhaB was shown to be located at the cell surface and overexpression in E. coli K-12 resulted in significant biofilm formation under continuous flow conditions. Overexpression of EhaB in E. coli K12 and EDL933 backgrounds also promoted adhesion to the extracellular matrix proteins collagen I and laminin. An EhaB-specific antibody revealed that EhaB is expressed in E. coli EDL933 following in vitro growth. EhaB also cross-reacted with serum IgA from cattle challenged with E. coli O157:H7, indicating that EhaB is expressed in vivo and elicits a host IgA immune response.

Ecology of uncultivated oscillospira species in the rumen of cattle, sheep, and reindeer as assessed by microscopy and molecular approaches

The ecology of the uncultured, but large and morphologically conspicuous, rumen bacterium Oscillospira spp. was studied. Oscillospira-specific 16S rRNA gene sequences were detected in North American domestic cattle, sheep from Australia and Japan, and Norwegian reindeer. Phylogenetic analysis of the sequences obtained allowed definition of three operational taxonomic units within the Oscillospira clade. Consistent with this genetic diversity, we observed atypical smaller morphotypes by using an Oscillospira-specific fluorescence in situ hybridization probe. Despite the visual disappearance of typical large Oscillospira morphotypes, the presence of Oscillospira spp. was still detected by Oscillospira-specific PCR in the rumen of cattle and sheep. These observations suggest the broad presence of Oscillospira species in various rumen ecosystems with the level, and most likely the morphological form, dependent on diet. An ecological analysis based on enumeration of the morphologically conspicuous, large-septate form confirms that the highest counts are associated with the feeding of fresh forage diets to cattle and sheep and in two different subspecies of reindeer investigated.

A Genetic Analysis of Lysosomal Enzyme Activities in Brahman Cattle

Analyses of variance and co variance were carried out on the activities of three lysosomal enzymes in mononuclear blood cells from Brahman cattle. These were hexosaminidase (HEX), beta-D-galacto-sidase (GAL) and acid alpha-glucosidase (GLU) which had been measured in blood mononuclear cells from 1752 cattle from 6 herds in a Pompe's disease control programme. Herd of origin and date of bleeding significantly affected the level of activity of all enzymes. In addition, HEX and GAL were affected by age and HEX by the sex of the animal bled. Estimates of heritability from sire variances were 0.29:t 0.09 for HEX, 0.31 :t 0.09 for GAL and 0.44:t 0.09 for GLU. Genetic correlations between all enzymes were positive. The data indicate the existence of a major gene causing Pompe's disease and responsible for 16% of the genetic variation in GLU. One standard deviation of selection differential for high GLU should almost eliminate Pompe's disease from the population. The effi-ciency of selection would be aided by estimating the breeding value for GLU using measurements of HEX and GLU and taking account of an animal's sex, age, date of bleeding and herd of origin.

Scrotal Circumference at Puberty and its Relationship to Testes Mass in Three Genotypes of Beef Cattle

Scrotal circumference (SC) is a simple, non-invasive measurement commonly used to evaluate bull breeding potential although its validity as a predictor of fertility is questionable (Holroyd, 1998). SC is highly heritable but varies with breed and animal factors such as condition, live weight and age. As an indicator of fertility, recommended SC values range broadly from 30cm to 38cm (Miller, 1992). It is assumed that SC accurately reflects testes mass (TM) which may be related to direct measures of fertility such as spermatogenesis (Entwistle, 1992). The SC measurements made here test the assumption that SC, used to estimate testes volume (TV), is directly related to TM. Miller (1992) reported a value of 261mm as the SC threshold for puberty. We have studied serial SC measurements so as to devise a more accurate means of using SC to determine puberty.