Dissertations / Theses on the topic 'IBV'

Because phosphorylation of the infectious bronchitis virus (IBV) nucleocapsid (N) protein may regulate its multiple roles in viral replication, the dynamics of N phosphorylation were examined. In the infected cell, N was the only viral protein that was phosphorylated as shown by 32P-orthophosphate labeling and Western blot analysis and with IBV specific polyclonal chicken antibody. Using pulse-labeling with 32Porthophosphate, the IBV N protein was found to be phosphorylated in the virion, as well as at all times during infection of Vero cells. One-hour pulse-chase analysis followed by immunoprecipitation of IBV N using rabbit anti-IBV N polyclonal antibody showed that the phosphate on the protein did not fall below 70% of the maximum and remained stable. The small but reproducible drop in phosphorylation could modulate the various functions of the N protein in the infected cell. Simultaneous labeling with 32Porthophosphate and 3H-leucine of infected CEK cells indicated a 3.5-fold increase in the ratio of the 32P:3H counts per minute (cpm) on the virion N protein as compared to the 32P:3H cpm ratio of the N protein from lysates at 7 h p.i. The 32P:3H cpm ratio of the N protein from virion from infected-Vero cell lysates was 10.5X more than the 32P:3H cpm ratio of the N protein obtained at 7 h p.i. It has been shown that the N proteins from the measles and rabies viruses form helical nucleocapsid-like structures when expressed in bacteria (Schoehn et al., 2001; Warnes et al., 1995). The ability of E. coli expressed IBV N protein to form helical-nucleocapsid-like structures was investigated using transmission electron microscopy. Full-length, purified histidine-tagged IBV N protein formed nucleocapsid-like structures when expressed in bacteria. Because E. coli -expressed histidine-tagged fragments of the IBV N protein did not form helical nucleocapsid-like structures, the full-length protein is probably required for assembly of these structures. The highly conserved IBV N protein was also used as a diagnostic tool in an ELISA for detecting anti-IBV antibody in chicken serum using a specialized microwave called the BIOWAVE. The BIOWAVE improves the processing time for an ELISA.

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Poultry production is a very important economic activity in Brazil once the country is among the highest world avian producers. The knowledge of the pathogens epidemiology is essential to the control of infectious diseases and such control is very strict in chickens and turkeys on the avian industry. However, the avian population of the country shows a big diversity of domestic and wild birds and the chicken backyard population is out of this control. To investigate the presence of antibodies against specific viruses in the backyard chicken population, blood was collected of 867 non-vaccinated birds, from 60 farms and 22 counties from the Rio Grande do Sul State, Brazil. Neutralizing antibodies against infectious bronchitis virus (IBV) were detected in 65% (564/867) of the individuals, against avian reovirus (ARV) in 21.6% (187/867) and, against infectious bursal disease virus (IBDV) in 80.2% (695/867). The results presented on the first chapter indicated that the tested viruses are circulating among this population. Among the wild species there is a group of Galliformes, classified at the Cracidae family and commonly known as guans, chachalacas and curassows which deserved a special attention in the present study. To determine the microbiota, the bacterial resistance and the presence of antibodies against specific viruses in these birds, fifty one serum and cloacal swab samples were collected from 10 different cracid species captive in the Rio Grande do Sul State during 2007. Serum samples were submitted to serum-neutralization test and specific antibodies were detected against IBV in 5.9% (3/51) of the cracids, against ARV in 15.7% (8/51) and, against IBDV in 35.3% (18/51). All the samples were found to be negative to fowlpox virus by the AGID test. Bacterial isolation and identification were performed from the cloacal swabs. After that, the isolates were tested for antimicrobial susceptibility. Ninety three bacterial isolates were obtained from 10 different genera. Escherichia spp., Staphylococcus spp. and Streptococcus spp. are among the most prevalent genera. Antimicrobial resistance was observed in several isolates, with Serratia marcescens presenting the highest level of resistance to multi drugs followed by Streptococcus spp., Staphylococcus aureus and Escherichia coli, among others. The results from the second chapter of this dissertation allowed us to conclude that bacterial resistance is spread in the captive cracid microbiota and, most importantly, these species are susceptible to the infection by IBV, IBDV and ARV.
A avicultura é uma das principais atividades econômicas do Brasil, que ocupa posição de destaque entre os exportadores de frango e subprodutos. O conhecimento da epidemiologia de patógenos que podem gerar prejuízos a essa atividade é essencial para o controle das enfermidades infecciosas. Este controle é realizado em larga escala em galinhas e perus criados no sistema industrial. Entretanto a população avícola do país consiste de uma grande diversidade de aves domésticas e selvagens, sendo o grupo conhecido como galinhas de terreiro formado por indivíduos desta espécie criados fora do sistema industrial. Com o objetivo de investigar a presença de anticorpos contra alguns vírus específicos na população de galinhas de terreiro, foram coletadas amostras de sangue de 867 galinhas não vacinadas, em 60 propriedades de 22 municípios do Estado do Rio Grande do Sul, Brasil, que foram testadas pela técnica de soroneutralização. Anticorpos contra o vírus da bronquite infecciosa das galinhas (IBV) foram detectados em 65% (564/867) destas aves, contra o reovírus aviário (ARV) em 21,6% (187/867) e contra o vírus da doença infecciosa da bolsa (IBDV) em 80,2% (695/867). Os descritos no primeiro capítulo desta dissertação permitiram demostrar a circulação dos vírus testados na população descrita. No segundo capítulo trabalhou-se com aves da ordem Galliformes, pertencentes à família Cracidae e conhecidos popularmente como jacus, jacutingas, araquãs e mutuns, com o intuito de conhecer a microbiota, resistência bacteriana e a presença de anticorpos contra vírus de aves, 51 amostras de soro e swab cloacal de 10 diferentes espécies de cracídeos cativos do Estado do Rio Grande do Sul foram coletadas durante o ano de 2007. As amostras de soro foram utilizadas para a detecção de anticorpos neutralizantes contra o IBV, ARV e IBDV, onde obteve os seguintes resultados: contra o IBV em 5,9% (3/51) das amostras positivas, contra o ARV em 15,7% (8/51) e contra o IBDV em 35,3% (18/51). Através do teste de imunodifusão em gel de ágar (IDGA) determinou-se que todas as amostras de soro eram negativas para o vírus da bouba aviária. Para o isolamento e identificação bacteriana foram realizados dos swabs cloacais e, posteriormente, foi testada a susceptibilidade antimicrobiana. Foram obtidos 93 isolados bacterianos, divididos em 10 diferentes gêneros. Entre os gêneros bacterianos mais prevalentes estavam Escherichia spp., Staphylococcus spp. e Streptococcus spp. Foi observada em grande número de isolados resistência à antimicrobianos, sendo que a bactéria Serratia marcescens apresentou o maior índice de resistência múltipla antimicrobiana. Os altos índices de resistência também foram detectados para Streptococcus spp., Staphylococcus aureus e Escherichia coli. Os resultados apresentados permitem concluir que a resistência bacteriana está disseminada na microbiota de cracídeos cativos e que indivíduos destas espécies são suscetíveis à infecção pelo IBV, IBDV e ARV, assim como as galinhas de terreiro.

Using an IBV reverse genetics system a series of recombinant viruses were generated to investigate the potential for utilizing IBV as a vaccine vector. Through the replacement of non-essential regions of the IBV genome, with eGFP or hRluc, factors influencing the stability of recombinant viruses expressing heterologous genes were determined. Expression of heterologous proteins was possible from a variety of virus constructs. The stability of recombinant viruses varied depending on the genome location of the heterologous gene, with replacement of Gene 5 proving to be most stable following passage in cell culture. Stability was strongly influenced by the MOI at which viruses were passaged, with low MOIs resulting in increased stability. The replacement of Gene 5 with a heterologous virus gene may be a suitable target for development of a bivalent vaccine capable of protecting against IBV and a second avian viral disease. Analysis of recombinant IBV mRNA expression profiles led to an investigation into an uncharacterized RNA species, and its link to the IBV intergenic region. A novel subgenomic mRNA was identified associated with the intergenic region that was shown to be transcribed via a non-canonical transcription regulatory sequence. In contradiction to the current model of coronavirus transcription this mRNA has a transcription regulatory sequence derived mainly from the leader, and not the body, transcription regulatory sequence. The non-canonical sequence was shown to be responsible for reduced transcription levels of the intergenic region mRNA. This project proposes the presence of an additional IBV subgenomic mRNA, transcribed via a non-canonical mechanism, and encoding a novel 5th accessory protein of IBV and closely related gammacoronaviruses.

Orientadores: Clarice Weis Arns, Helena Lage Ferreira
Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia
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Resumo: O vírus da Bronquite Infecciosa (VBI), pertencente à família Coronaviridae, é um importante patógeno à sanidade e fatores econômicos da produção avícola no Brasil e no mundo. O VBI possui múltiplos sorotipos e o frequente surgimento de novas variantes é um dos principais problemas relacionados a este vírus. Este trabalho tem como objetivo a investigação experimental da patogênese de um isolado de pombo (Columba/Brazil/2007/Unicamp/67T), caracterizado molecularmente pelo gene S1 como VBI sorotipo Massachusetts, e seus efeitos in vivo, em galinhas e camundongos. O presente estudo foi dividido em duas partes, na primeira um grupo de aves "specific pathogen free" (SPF) foi inoculado pela via óculo-nasal com a amostra viral proveniente de pombo. Os animais, de um dia de vida, foram sacrificados nos dias 2, 4, 5, 7, 9, 11, 14, 21, 28, 35 e 42 dias pós-inoculação (dpi). Foram coletados suabes de traqueia, seio nasal e cloaca, além de órgãos como pulmão, íleo, pró-ventrículo (coletado entre 7 e 21 dpi), rim, tonsilas cecais (coletada a partir de 4dpi) e testículos (coletado a partir de 5 dpi). Sinais clínicos respiratórios como espirros, estertores, corrimento nasal, além de letargia, diarreia e perda de coordenação foram observados principalmente no 5dpi. A inibição da atividade ciliar ocorreu concomitantemente ao pico de sinais clínicos das aves. Foi analisado tropismo tecidual, através da quantidade de RNA viral detectado, pelo trato digestório. Os maiores títulos de RNA viral foram detectados na tonsila cecal, seguida pelo íleo (ambos no 5dpi) e cloaca (no 2dpi). Além disso, houve detecção de RNA viral no rim e trato respiratório, com maior título de RNA viral na traqueia. Os órgãos que apresentaram maiores danos teciduais através do exame histopatológico foram o rim, íleo e traqueia (todos no 5dpi). Por fim, as aves inoculadas com a amostra do VBI oriundo de pombo produziram anticorpos entre os dias 14 e 21dpi, detectados no soro destes animais através do ELISA. Na segunda parte do trabalho, a capacidade de replicação de diferentes variantes do VBI em camundongos foi avaliada. Para tanto, camundongos das linhagens Balb/C e A/J foram inoculados pela via nasal com duas amostras do sorotipo Massachussets (Mass) e com a variante brasileira (BR-I), e sacrificados no 3, 10 e 14 dpi. Não foram observados sinais clínicos nem lesões macroscópicas graves. O RNA viral foi detectado em todos os órgãos coletados, sendo os principais órgãos de replicação o seio nasal e pulmão (no 3dpi) para os camundongos da linhagem A/J e pulmão e duodeno (ambos no 3dpi) na linhagem de camundongos Balb/C, nos quais os títulos virais detectados foram mais altos. Pneumonia intersticial, edema e infiltrado mononuclear foram as principais alterações histopatológicas observadas no 3dpi em camundongos inoculados com as diferentes variantes. A presença da nucleoproteína viral, pela imunohistoquímica, foi detectada no duodeno, traqueia e pulmão de camundongos no 3dpi nas duas linhagens de camundongos. Os anticorpos contra o coronavírus aviário foram detectados somente no 3dpi. Assim, os resultados do presente estudo demonstraram que a variante Massachussets, com origem de pombo, causa a doença clínica em aves comerciais não vacinadas e pode replicar em modelo mamífero por um curto período de tempo, ressaltando a importância da vacinação e o papel potencial dos roedores como possível reservatório e carreador do vírus
Abstract: Infectious bronchitis virus (IBV) belonging to the family Coronaviridae is an important pathogen to sanity and economics of poultry production in Brazil and worldwide. The VBI has multiple serotypes and the frequent emergence of new variants is one of the main problems related to this virus. This work aims to experimentally investigate the pathogenesis of pigeon sample (Columba/Brazil/2007/Unicamp/67T), molecularly characterized by S1 gene as IBV Massachusetts serotype, and its effects in vivo in chickens and mice. This study was divided into two parts. In the first part, a group of birds "specific pathogen free" (SPF) was inoculated by oculo-nasal route with the viral sample from pigeon. The animals with one-day-old, were sacrificed on 2, 4, 5, 7, 9, 11, 14, 21, 28, 35 and 42 days post-inoculation (dpi). Tracheal swabs, nasal sinus and cloaca were collected, and organs such as lung, ileum, pro-ventricular (collected between 7 and 21dpi), kidney, caecal tonsils (collected from 4dpi) and testes (collected from 5 dpi). Clinical signs such as sneezing, rales, nasal discharge, lethargy, diarrhea, and loss of coordination were observed mainly in the 5dpi. Inhibition of ciliary activity occurred concomitantly with the peak of clinical signs of birds. Tissue tropism was analyzed by the amount of viral RNA detected by the gastrointestinal tract. The higher titers of viral RNA were detected in the cecal tonsil, followed by the ileum (both in 5dpi) and cloaca (in 2dpi). In addition, viral RNA was detected in the kidney and respiratory tract, with highest titer of viral RNA in the trachea. The organs that showed severe tissue damage by histopathology were the kidney, ileum and trachea (all in 5dpi). Finally, the birds inoculated with the sample originated from IBV Pigeon produced antibodies between 14 and 21dpi, detected in the serum of these animals by ELISA. In the second part, the replication capacity of different variants of IBV in mice was evaluated. For this, mice of strains BALB/C and A/J were inoculated intranasally with two strains of Massachusetts (Mass) serotype and the Brazilian variant (BR-I), and sacrificed at 3, 10 and 14 dpi. No clinical signs or severe macroscopic lesions were observed. The viral RNA was detected in all organs collected, higher tittles were detected on sinus and lung (in 3dpi) for mice of strain A/J and on lung and duodenum (both in 3dpi) in the line of Balb/C; in this line the viral titles were higher than the strain A/J. Interstitial pneumonia, edema and mononuclear cell infiltration were the main histopathological changes observed in 3dpi in inoculated mice with different variants. The presence of viral nucleoprotein, immunohistochemistry was detected in the duodenum, trachea and lungs of mice in 3dpi in both mice strains. Antibodies against avian coronaviruses have been detected only in 3dpi. Thus, the results of this study demonstrate that the Massachusetts variant, originating from pigeon, cause clinical disease in commercial poultry unvaccinated and can replicate in mammalian model for a short period of time, emphasizing the importance of vaccination and the potential role of rodents as possible reservoir and the carrier virus
Doutorado
Microbiologia
Doutora em Genética e Biologia Molecular

An infectious cDNA clone of Vero cell adapted Beaudette strain of IBV was constructed using in vitro assembly of cDNA fragments. The entire genome of IBV was RT-PCR amplified into seven fragments, with each piece overlapping about 10 nucleotides. The fragments were ligated and transcribed to synthesize RNA, which was transfected into BHK-21 cells. These cells were then overlaid onto IBV susceptible Vero cells. After five days transfection, the virus was successfully rescued from the transfected cells. The cDNA clone from our laboratory strain has a five nucleotide insertion not present in the originally sequenced virus, resulting in total genome size of 27,613 nucleotides. The infectious cDNA clone was further manipulated to demonstrate its potential as a gene transfer vector, by replacing the ORF5a open reading frame with enhanced green fluorescent protein. The recombinant infectious cDNA clone was also successfully rescued after three days transfection of BHK-21 cells followed by co-culturing with Vero cells. This study showed that the 5a protein, whose function is not known, is not necessary for in vitro IBV replication. This study also showed that the 5a ORF is a good candidate for an insertion site of recombinant genes for the development of IBV infectious cDNA clone as a gene transfer vector.

A bronquite infecciosa das galinhas (BIG) é uma doença altamente contagiosa causada por múltiplos genotipos/sorotipos do vírus da bronquite infecciosa das galinhas (IBV), um coronavirus do grupo 3. Embora classicamente associado ao trato respiratório, alguns tipos de IBV têm sido descritos com tropismo pelos rins e pelos tratos reprodutivos e entéricos, o IBV pode ser detectado em diversos tipos de tecidos, e também pode acometer aves de todas as idades. Este estudo tem como objetivo verificar a freqüência do IBV em amostras de diversos órgãos e conteúdo entérico de avós, matrizes, poedeiras comerciais e frangos de corte, genotipar as amostras detectadas e estudar a diversidade molecular entre as amostras brasileiras de IBV. Um total de 844 pools de diversos órgãos e conteúdos entéricos provenientes de 200 lotes de avós, matrizes, poedeiras comerciais e frangos de corte, das regiões Sul, Sudeste, Centro-oeste e Nordeste do Brasil, colhidas durante o período de 2007 a 2009 foram testadas para a presença de IBV com um RT-PCR dirigido à região não traduzida 3′(3′UTR). As aves amostradas apresentaram sinais clínicos compatíveis com a BIG. Todas as amostras de IBV detectadas foram tipificadas utilizando uma RT-PCR dirigida ao gene de espícula do vírus. Dezenove amostras tipificadas como variante foram submetidas ao seqüenciamento parcial da região codificadora da subunidade S1 e à análise genealógica. Considerando os pools de órgãos e de conteúdo entérico, 45,50% foram positivos para a presença de IBV, dos quais, 84,63% pertencem ao genotipo Variante e 9,89% ao sorotipo/genotipo Massachusetts. Considerando os lotes, 73,50% foram positivos para IBV, sendo 77,55% variantes e 6,12% Massachusetts. A análise genealógica revelou quatro linhagens virais, todas agrupadas em um exclusivo grupamento de genotipo brasileiro. Estes resultados demonstram que o IBV está disseminado em todas as regiões avícolas brasileiras, com um predomínio massivo de genotipos não Massachusetts e uma elevada diversidade molecular, que deve ser levada em consideração para desenvolver medidas preventivas contra o IBV.
Infectious bronchitis (IB) is a highly contagious disease of poultry caused by multiple geno/serotypes of avian infectious bronchitis virus (IBV), a group 3 coronavirus. Though classically associated to the respiratory tract, IBV strains also have been described which harbor tropism for the kidneys and the reproductive and enteric tracts, and might be detected in multiple tissues and can also affect birds of all ages. This survey aimed to assess the frequency of in multiple organs and enteric content samples from grandparents, breeders, layers and broilers, to genotype the IBV strains detected and to study the molecular diversity amongst Brazilian IBV strains. A total of 844 pools of multiple organs and enteric contents from 200 flocks of grandparents, breeders, layers and broilers from the Southern, Southeastern, Central-Western and Northeastern Brazilian regions collected between 2007 and 2009 was screened for the presence of IBV with an RT-PCR target to the 3 untranslated region (UTR). The sampled birds presented symptoms compatible with IB. All IBV strains detected were then typed using an RT-PCR target to the spike gene of the virus. Nineteen strains type as variants were submitted to partial sequencing of the S1 coding region and genealogic analysis. Regarding the organs and enteric content pools, 45.50% were positive for the presence of IBV, from which 84.63% were variant and 9.89% Massachusetts. Taking into account the flocks, 73.50% were positive for IBV, being 77.55% variants and 6.12% Massachusetts. Genealogic analysis revealed four viral lineages, all grouped in an exclusive Brazilian genotype cluster. This results shown that IBV is widespread in all Brazilian poultry regions, with a massive predominance of non-Massachusetts genotypes and a high molecular diversity, which must be taken into account in order to develop preventive measures against IB.

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Apesar dos esforços que estão sendo feitos para a inclusão laboral da pessoa com deficiência (PD), observa-se que as deficiências, muitas vezes, podem prejudicar a pessoa na procura de trabalho. O Brasil apresenta cerca de 14,5% da população, ou seja, 24,5 milhões de PDs (IBGE, 2002), porém, de acordo com a RAIS de 2009 somente 288,6 mil estavam em empregos formais no ano de 2009 (BRASIL, 2010). Dessa forma, a ergonomia torna-se imprescindível, uma vez que, através do conhecimento das exigências físicas, cognitivas e organizacionais dos postos de trabalho e a determinação das capacidades funcionais do trabalhador com deficiência, podem ser realizadas adequadamente as adaptações dos postos de trabalho. O objetivo deste estudo foi determinar as exigências físicas da tarefa, indicar o perfil dos trabalhadores com deficiência e definir as adequações necessárias à eficiência do sistema Humano-Tarefa-Máquina, com foco numa obra de construção civil de adutora de abastecimento de água. Foi realizado estudo de campo, onde através do software ErgoDis/IBV foram avaliados três casos reais, postos de trabalho onde há PDs (dois serventes e um armador), para verificar a adequação e/ou a necessidade de alguma adaptação. Além dos casos reais, foram avaliadas as funções de encarregado de turma, pedreiro, carpinteiro e pintor letrista com a finalidade de determinar o perfil dos trabalhadores com deficiência que poderiam exercer essas funções. Para isso foram simulados indivíduos com deficiências no software Ergodis/IBV nas funções avaliadas para determinar se poderiam realizá-las e/ou quais as adaptações necessárias. A partir da análise dos dados foi observado que trabalhadores com deficiência auditiva poderiam exercer as atividades em qualquer uma das funções avaliadas sem nenhuma mudança no ambiente de trabalho e para indivíduos com amputação de perna ou pé, será necessário utilizar próteses adequadas para realizar as atividades nessas funções. Portanto, defende-se que o tema tem um caráter interdisciplinar, devendo ser abordado de forma sistêmica. É fundamental entender as interações entre as pessoas e os elementos dos sistemas de trabalho, a partir dos fundamentos, métodos e técnicas da Ergonomia, de modo a possibilitar a adequação do trabalho às capacidades da PD. Também são importantes os conhecimentos de biomecânica, fisiologia, patologia e dos métodos e técnicas da Fisioterapia, para gerar requisitos e propostas de soluções de Design objetivando facilitar o processo de inclusão laboral das pessoas com deficiência

A bronquite infecciosa é uma doença viral, aguda e altamente contagiosa que afeta as aves da espécie Gallus gallus, de todas as idades, causando grandes perdas econômicas devido à mortalidade, queda na produção e qualidade dos ovos. Variações na sequência de aminoácidos na região hipervariável da subunidadade S1 da proteína de espícula (S) levam à emergência de novos sorotipos do vírus da bronquite infecciosa das galinhas (IBV). Este estudo teve como objetivo seqüenciar parcialmente o gene S1 para determinar a relação filogenética entre amostras vacinais e de campo com o intuito de investigar os genótipos circulantes. Cento e sessenta amostras foram coletadas, na forma de pools, de plantéis sintomáticos durante 2009/2010. Cada pool continha tecidos de 3 a 5 aves e foram compostos por diferentes órgãos (traquéia, pulmões, rins, conteúdo entérico, trato reprodutivo e swabs traqueais). As amostras foram triadas para a presença de IBV utilizando-se uma nested RT-PCR, direcionada a região 3UTR. As amostras positivas, (n=89) foram submetidas a nova nested RT-PCR dirigida para o gene S, resultando em 10 amplicons de 390pb que foram então submetidos a seqüenciamento de DNA e análise filogenética. As amostras brasileiras segregaram-se em dois grupos filogenéticos: Massachusetts (Mass) e tipos marcadamente brasileiros. Seis amostras agruparam-se no grupo Mass, com vacinas vivas atenuadas utilizadas no Brasil. Quatro amostras agruparam com cepas de campo brasileiras previamente descritas (acesso no GeneBank: FJ791257 to FJ791273). A identidade de aminoácidos no grupo Mass variou de 98% a 100%, sugerindo a detecção de vírus vacinal. Enquanto que entre as amostras brasileiras e o grupo Mass, a variabilidade de aminoácidos foi de 79% a 81%. Esses resultados mostram uma variação importante em genótipos virais circulantes nos plantéis industriais avícolas brasileiros. Portanto, o presente estudo reforça a contínua emergência e disseminação de novas linhagens de IBV no Brasil e a importância de constantes pesquisas e monitoramento, para que se compreendam aos fenômenos de emergência de novas linhagens e suas consequências, bem como para otimizar as medidas de controle do vírus.
Infectious bronchitis is a viral disease, acute and highly contagious that affects birds of the Gallus gallus species, of all ages, causing huge economic losses due to mortality, drop in production and quality eggs. Differences between serotypes of IBV are due to variations in the amino acids sequence in S1 hypervariable region subunit of spike protein (S). This study aimed to partially sequence the S1 gene to determine the phylogenetic relationship between field and vaccine viral types, and to investigate circulating genotypes in Brazil. One hundred and sixty pool samples were collected from symptomatic flocks during 2009/2010. Each pool contained tissues from 3 to 5 birds and was composed by different organs (tracheas, lungs, kidneys, enteric contents, tract reproductive and tracheal swabs). Samples were screened for the presence of IBV using a nested RT-PCR targeted to the 3UTR. Positive samples (n=56) were submitted to a nested RT-PCR target to S gene resulting in 10 amplicons of 390pb that were then submitted to DNA sequencing and analysis. Brazilian IBV genotypes segregated in two phylogenetic groups: Massachusetts (Mass) and Brazilian types. Six samples clustered in the Mass group, with live attenuated vaccines used in Brazil. Four samples grouped with field Brazilian strains previously described (GeneBank accession: FJ791257 to FJ791273). In Mass cluster aminoacids identity range from 98% to 100%, suggesting vaccine virus detection, in this last case. Moreover, the verified identity between samples from this study and Mass group ranged 79% to 81%. Therefore the present study reinforces the emergence and circulation of news lineages of IBV in commercial Brazilian flocks, as well as the importance of consistent research and monitoring to better understand this phenomen and its consequences, aiming to improved control measures against the virus.

Infectious bronchitis virus (IBV) is the causative agent of a highly contagious disease that results in severe economic losses to the global poultry industry. The virus exists in a wide variety of genetically distinct viral types, and both phylogenetic analysis and measures of pairwise similarity among nucleotide or amino acid sequences have been used to classify IBV strains. However, there is currently no consensus on the method by which IBV sequences should be compared, and heterogeneous genetic group designations that are inconsistent with phylogenetic history have been adopted, leading to the confusing coexistence of multiple genotyping schemes. Herein, we propose a simple and repeatable phylogeny-based classification system combined with an unambiguous and rationale lineage nomenclature for the assignment of IBV strains. By using complete nucleotide sequences of the S1 gene we determined the phylogenetic structure of IBV, which in turn allowed us to define 6 genotypes that together comprise 32 distinct viral lineages and a number of inter-lineage recombinants. Because of extensive rate variation among IBVs, we suggest that the inference of phylogenetic relationships alone represents a more appropriate criterion for sequence classification than pairwise sequence comparisons. The adoption of an internationally accepted viral nomenclature is crucial for future studies of IBV epidemiology and evolution, and the classification scheme presented here can be updated and revised novel S1 sequences should become available.
Il virus della bronchite infettiva aviare è l’agente patogeno di una malattia altamente contagiosa responsabile di ingenti perdite economiche nel settore avicolo. Il virus è caratterizzato da una grande variabilità genetica e antigenica che ha favorito la comparsa e la diffusione di molteplici e differenti tipi virali. Analisi filogenetiche e calcolo delle distanze nucleotidiche esistenti tra i ceppi sono state utilizzate per classificare la grande varietà di virus appartenenti a questa famiglia di patogeni. Nonostante ciò, non si è ancora raggiunto un accordo sul metodo con il quale i ceppi dovrebbero essere confrontati. Questo ha generato notevole confusione poiché ha favorito lo sviluppo di numerosi sistemi di classificazione, molto spesso contrastanti l’uno con l’altro, e l’utilizzo di nomenclature eterogenee che in molti casi non riflettevano la filogenesi. Lo scopo di questo lavoro è proporre un sistema di classificazione che sia semplice e ripetibile e basato esclusivamente sulle relazioni filogenetiche esistenti tra i ceppi di bronchite aviaria. Il lavoro si propone inoltre di assegnare una nomenclatura univoca e razionale dei gruppi genici identificati. Mediate l’analisi di tutte le sequenze nucleotidiche della proteina S1 disponibili in banca dati sono stati identificati 32 diversi lineaggi, compresi in 6 genotipi, e un numero di ceppi originati da eventi di ricombinazione tra lineaggi. Poiché la variabilità esistente tra i diversi ceppi di IBV è abbastanza elevata, qui si propone di usare le relazioni filogenetiche piuttosto che il calcolo delle distanze geniche come criterio più adatto per tracciare la storia evolutiva di IBV. L’adozione di una nomenclatura accettata a livello internazionale è di fondamentale importanza per gli studi futuri sull’epidemiologia ed evoluzione del virus della bronchite infettiva aviaria. Inoltre, così com’è stata sviluppata, la classificazione qui proposta può essere revisionata e aggiornata nel momento in cui saranno disponibili nuove sequenze virali della proteina S1.

La production de protéines hétérologues dans les plantes est maintenant possible grâce à la technique de transfert de gènes. Néanmoins, le rendement reste faible et nécessite d'être amélioré. L'adressage et la séquestration dans un compartiment cellulaire particulier est un moyen de favoriser la stabilité des protéines recombinantes et donc d'augmenter leur taux de production. Un nouvel organite observé dans les cellules de tabac transformées par le gène chimérique ibv m- gus pourrait devenir un lieu de stockage intéressant. Cet organite est constitué de tubules de réticulum endoplasmique superposées et enroulées formant une structure concentrique de diamètre d'environ 1,5 pm. Sa formation serait la conséquence d'un mécanisme d'auto-assemblage induit par les propriétés de tétramérisation de la protéine GUS qui permettrait un rapprochement des membranes. Ce système, déjà mis en évidence par une autre équipe, a été reproduit sur le cultivar de tabac SRI. Différents tissus différenciés (feuilles et racines) et indifférenciés (cals) ont été étudiés afin de mettre en évidence la plus forte expression de la protéine 1BV M- GUS. Les tissus indifférenciés, sous forme de cals, semblent accumuler des organites artificiels en grandes quantités. Ce phénomène est particulièrement intéressant sachant la rapidité de croissance des cellules indifférenciées. Après la mise en suspension cellulaire des cals, l'étude de ces cellules au cours d'un cycle de culture a montré une augmentation de l'expression du gène ibv m-gus au cours de la culture avec une forte expression en phase stationnaire de croissance, correspondant à une accumulation des protéines 1BV M-GUS. Celles-ci semblent protégées des dégradations par leur insertion dans la membrane des organites adificiels. L'utilisation de ces compartiments cellulaires serait susceptible d'ouvrir de nouveaux horizons pour la production et la récupération de protéines recombinantes stables dans les plantes.

Rapidly evolving RNA and ssDNA viruses represent a fascinating field characterized by a strict interconnection between the “speculative” study of viral evolution and its practical implications in everyday veterinary medicine. This thesis has been thought as a collection of manuscripts that aimed to investigate different aspects and levels of viral evolution while still maintaining a focus on practical repercussions. Even if different infectious diseases and etiological agents are considered, they are all functional to the study of different aspects and implications of rapid virus evolution. Considering the heterogeneous nature of the manuscripts, they have been organized according to a “crescent” scale, starting from the lowest scale of viral evolution and progressing to broader scales. The manuscript “Viral subpopulations in aMPV vaccines are unlikely to be responsible for reversion to virulence.” addresses a fine-level analysis of the population structure of the AMPV subtype B live attenuated vaccine and its potential role in the previously demonstrated phenomenon of reversion to virulence. The widespread administration of live attenuated vaccines, despite their obvious advantages in terms of reducing disease prevalence, clinical signs and economic losses, is associated with costs that are not limited to the risk of reversion to virulence or to direct economic costs. Based on a wide collection of Italian samples, “Continued use of IBV 793B vaccine needs reassessment after its withdrawal led to the genotype’s disappearance” reports the impact of these vaccines in complicating the diagnostic process and, as a consequence, the interpretation of the epidemiological scenario in the absence of known vaccine markers. Obviously, updated knowledge of the strains currently circulating in a particular area is of great relevance for the implementation of proper control strategies. With this aim in mind, a field survey, which is published in “Field survey of Avian Metapneumovirus in Northern Italy”, was conducted on hundreds of Italian farms to estimate and characterize the AMPV strains circulating in our country. To further support frequent and extensive surveys, an assay that is able to detect, quantify and genotype the two AMPV subtypes currently circulating in Italy was developed and validated. Because economic constraints often represent a major limit, efforts were made to reduce the assay costs compared with other real-time RT-PCR methods while still guaranteeing comparable or superior performances (“A Sensitive, Reproducible, and Economic Real-Time Reverse Transcription PCR Detecting Avian Metapneumovirus Subtypes A and B”). Unfortunately, the diagnosis of rapidly evolving RNA viruses is itself an arduous task that requires a continuous evaluation and updating of diagnostic tools, even in laboratory that receive samples from limited geographic areas. The manuscripts entitled “Observation of high recombination occurrence of Porcine reproductive and respiratory syndrome virus in field condition” and “Phylodynamic analysis of Porcine reproductive and respiratory syndrome virus (PRRSV) in Italy: action of selective pressures and interactions between different clades.” address the study of the molecular epidemiology of Porcine reproductive and respiratory syndrome virus (PRRSV) in Italy considering the evolutionary forces driving PRRSV evolution at the local scale (i.e., high substitution rate, recombination, interaction between different clades and action of selective pressures). The high heterogeneity of PRRSV in the national contest was then evaluated with respect to the challenges that it poses in the development and validation of RT-PCR- and qRT-PCR-based diagnostic methods (“Validation and comparison of different end point and real time RT-PCR assays for detection and genotyping of porcine reproductive and respiratory syndrome virus”) and assessing its impact on diagnostic accuracy (“The impact of porcine reproductive and respiratory syndrome virus genetic heterogeneity on molecular assay performances”). Similarly, “International trades, local spread and viral evolution: the case of Porcine circovirus type 2 (PCV2) strains heterogeneity in Italy” investigates the genetic variability of PCV2 within national borders and compares it with the knowledge of its molecular epidemiology available from other countries. This study provides evidence regarding the role of both “in loco” evolution and importation of different genotypes and strains from foreign countries in determining the Italian PCV2 genetic heterogeneity. The crescent amount of PCV2 sequences deposited in publically available databases has revealed its marked variability and challenged the current classification criteria. Nevertheless, at least a superficial knowledge of the PCV2 molecular epidemiology is mandatory for the planning and evaluation of control strategies. “Revisiting the Taxonomical classification of PCV2: still a real challenge” proposes new criteria for the classification of PCV2 into different genotypes. Our aim was to provide a scheme that both accounts for the constraint imposed by the biological proprieties of this virus and allows a rapid, practical and easy way to classify PCV2 strains even during routine diagnostic activity. Last, “Genetic characterisation of porcine circovirus type 2 (PCV2) strains from feral pigs in the Brazilian Pantanal: an opportunity to reconstruct the history of PCV2 evolution” investigates more speculative issues inherent to the PCV2 origin. The discovery of a PCV2c genotype, which, to date, was believed to be extinct, in a feral pig population characterized by a peculiar population history and by a complex, and still partially known, relationship network with other PCV2-susceptible species opens exciting scenarios concerning the history and origin of PCV2
I virus a RNA e a ssDNA rappresentano un affascinante campo di ricerca caratterizzato da una stretta interconnessione fra lo studio di aspetti più “speculativi”, inerenti all’evoluzione virale, e le implicazioni che questi comportano nella pratica veterinaria. La presente tesi è stata concepita come una raccolta di articoli che hanno esplorato diversi aspetti e livelli dell’evoluzione virale senza per questo trascurarne le ripercussioni pratiche. Diverse malattie infettive ed agenti eziologici sono stati selezionati onde investigare i diversi aspetti e implicazioni della rapida evoluzione di questi virus. In considerazione della natura eterogena dei manoscritti prodotti, questi sono stati organizzati secondo una “scala crescente”, dal livello più basso dell’evoluzione virale, procedendo verso scale via via maggiori. L’elaborato “Viral subpopulations in aMPV vaccines are unlikely to be responsible for reversion to virulence” affronta un analisi ad alta risoluzione della presenza di sottopopolazioni virali nei vaccini vivi attenuati basati sul sottotipo B del metapneumovirus aviare e del loro potenziale ruolo nel determinare fenomeni di reversione a virulenza. La somministrazione su vasta scala di vaccini vivi attenuati, nonostante gli ovvi vantaggi in termini di riduzione della prevalenza virale, dei segni clinici e delle perdite economiche, è associata a costi che non sono limitati a quelli di tipo meramente pecuniario. Sulla base di un ampia raccolta di campioni italiani, l’articolo “Continued use of IBV 793B vaccine needs reassessment after its withdrawal led to the genotype’s disappearance” descrive l’impatto di questi vaccini, in assenza di marker vaccinali noti, nel complicare il processo diagnostico e, conseguentemente, l’interpretazione dello scenario epidemiologico. Ovviamente, una conoscenza aggiornata degli stipiti virali circolanti in una particolare area è di fondamentale importanza per l’implementazione di adeguate misure di controllo. Con quest’obiettivo in mente, uno studio di campo , pubblicato in “Diffusione dell’infezione da metapneumovirus aviare in allevamenti di tacchini e broiler nel Nord Italia”, è stato condotto su centinaia di allevamenti al fine di stimare e caratterizzare i ceppi di AMPV circolanti nel nostro territorio. In aggiunta, al fine di favorire studi sempre più frequenti e estesi, è stato sviluppato un test diagnostico in grado di rilevare, quantificare e tipizzare i sottotipi di AMPV attualmente circolanti in Italia. In considerazione del fatto che le spese relative a questi test rappresentano spesso uno dei maggiori limiti all’attività diagnostica e di ricerca, si è cercato di ridurre i costi rispetto ad altre metodiche comunemente utilizzate, garantendo nel contempo le medesime o migliori performance diagnostiche(“A Sensitive, Reproducible, and Economic Real-Time Reverse Transcription PCR Detecting Avian Metapneumovirus Subtypes A and B”). Sfortunatamente la diagnosi dei virus a RNA, essendo caratterizzati da una rapida evoluzione, rappresenta di per se stessa un ardua sfida e richiede una continua dedizione alla rivalutazione e aggiornamento delle metodiche diagnostiche esistenti. Gli articoli Observation of high recombination occurrence of Porcine reproductive and respiratory syndrome virus in field condition” e “Phylodynamic analysis of Porcine reproductive and respiratory syndrome virus (PRRSV) in Italy: action of selective pressures and interactions between different clades.” sono incentrati sullo studio dell’epidemiologia molecolare del Porcine reproductive and respiratory syndrome virus (PRRSV) in Italia. In particolare sono state studiate le forze evolutive che ne modellano e condizionano l’evoluzione (i.e., alto tasso di sostituzione nucleotidica, interazione fra differenti clade e azione di diverse pressioni selettive). L’elevata eterogeneità genetica di PRRSV nel nostro territorio è stata poi analizzata alla luce delle sue ripercussioni nello sviluppo e validazione di metodiche diagnostiche basate sull’uso della RT-PCR e qRT-PCR (“Validation and comparison of different end point and real time RT-PCR assays for detection and genotyping of porcine reproductive and respiratory syndrome virus”) e ne è stato valutato l’impatto sull’accuratezza diagnostica (“The impact of porcine reproductive and respiratory syndrome virus genetic heterogeneity on molecular assay performances”). Similmente, “International trades, local spread and viral evolution: the case of Porcine circovirus type 2 (PCV2) strains heterogeneity in Italy”, indaga la variabilità genetica di PCV2 all’interno dei confine nazionali comparandola altresì con le informazioni di epidemiologia molecolare disponibili per altri Stati. Il presente studio ha permesso di fornire significative evidenze sul ruolo sia dei commerci internazionali che dell’evoluzione “in loco” nel determinare l’eterogeneità di PCV2 riscontrata in Italia. Il progressivo aumento delle sequenza di PCV2 depositate in database pubblici ne ha dimostrato la grande variabilità su scala mondiale e ha evidenziato i limiti dei criteri di classificazione attualmente in uso. Tuttavia, una conoscenza quantomeno superficiale dell’epidemiologia molecolare di PCV2 è di basilare importanza per la pianificazione e la valutazione delle strategie di controllo. “Revisiting the Taxonomical classification of PCV2: still a real challenge” propone dei nuovi criteri per la classificazione di questo virus in diversi genotipi. Il nostro intento è stato quello di fornire uno schema che, pur tenendo conto dei limiti imposti dalle caratteristiche biologiche del virus, permettesse una rapida, pratica e facile genotipizzazione dei ceppi di PCV2 e che potesse quindi rispondere alle esigenze imposte dalla routinaria l’attività diagnostica. Infine, “Genetic characterisation of porcine circovirus type 2 (PCV2) strains from feral pigs in the Brazilian Pantanal: an opportunity to reconstruct the history of PCV2 evolution”, si confronta con il campo, più specualtivo, dell’origine di PCV2. La scoperta di un ceppo appartenente al genotipo PCV2c, sino ad oggi ritenuto estinto, in una popolazione di suini selvatici caratterizzata da una peculiare storia e da complesse, a ancora solo parzialmente conosciute, relazioni con altre specie suscettibili a questo patogeno, apre nuovi ed eccitanti scenari concernenti la storia e l’origine di PCV2

This thesis describes two proposed schemes that could be used to secure mobile messaging (SMS/MMS) as well as one scheme that could be used to secure mobile content. The security services we considered in securing the mobile messages are confidentiality, authentication, non-repudiation and integrity. We used Identity Based Cryptography in order to secure the mobile messaging and Blowfish algorithm to secure the mobile content. Due to some of the disadvantages imposed by the Identity Based Cryptography, we recommended using it along with the RSA algorithm. The proposed schemes were implemented in java and tested on an actual device, Nokia N70. In addition, we measured the time required by each of the algorithms we used to encrypt/decrypt a certain number of bytes. We found that the time taken by RSA and Blowfish algorithms will not be noticeable by the user. However, since the implementation of the Identity Based Cryptography we used was not meant to run on mobile devices, we encountered a noticeable delay whenever encrypting/decrypting the data using this algorithm. Securing the SMS messages will make it to be considered as one of the proposed means that could be used to conduct m-commerce. In addition, securing the MMS messages and the mobile content will increase the usability and the reliability of the mobile phones especially to the users on the move.
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The first aim of this research was to determine the prevalence of IBV in broilers within the Canterbury province, New Zealand, in late winter and to search for associations with management or environmental factors. The second aim was to study how ambient stressors affect the immune system in birds, their adaptive capacity to respond, and the price that they have to pay in order to return to homeostasis. In a case control study, binary logistic regression analyses were used to seek associations between the presence of IBV in broilers and various risk factors that had been linked in other studies to the presence of different avian pathogens: ambient ammonia, oxygen, carbon dioxide, humidity and litter humidity. Pairs of sheds were selected from ten large broiler farms in Canterbury. One shed (case) from each pair contained poultry that had a production or health alteration that suggested the presence of IBV and the other was a control shed. Overall, IBV was detected by RT-PCR in 50% of the farms. In 2 of the 5 positive farms (but none of the control sheds) where IBV was detected there were accompanying clinical signs that suggested infectious bronchitis (IB). Ambient humidity was the only risk factor that showed an association (inverse) with the prevalence of IBV (p = 0.05; OR = 0.92). It was concluded within the constraints of the totally enclosed management systems described, that humidity had an influence on the presence of IBV, but temperature, ammonia, carbon dioxide, oxygen or litter humidity had no effect. In another study environmental temperatures were changed in order to affect the biological function and adaptive capacity of chickens following infection with IBV. The 'affective states' of the animal were assessed by measuring levels of corticosterone (CORT) in plasma and tonic immobility (TI). It was found that low (10 +/- 2°C) and high (30 +/- 2°C) temperatures exacerbated the respiratory signs and lesions in birds infected with IBV as compared to those housed at moderate (20 +/- 2°C) temperatures. The chickens housed at high temperatures showed significantly decreased growth, a higher proportion of hepatic lesions (principally haemorrhages) and a longer tonic immobility period, but there was no significant alteration in the plasma levels of CORT. The birds housed at low temperatures developed a higher proportion of heart lesions (hydropericardium, ventricular hypertrophy) and had significantly higher levels of plasma CORT than birds housed under moderate and/or high temperatures. The specific antibody response to IBV decreased in birds housed under high temperatures. Interestingly the birds housed at high temperatures developed significantly higher levels of haemagglutinin antibodies to sheep red blood cells (SRBC) than those birds housed under low or moderated temperatures. Cell mediated immunity was not significantly affected by heat or cold stress in the first 13 days of treatment but at 20 days the levels of interferon gamma in the birds subjected to low temperatures were lower than in the high temperature group. In other trials, the exogenous administration of low physiological doses of oral CORT (as compared to high pharmacological doses typically used in such experiments) to birds resulted in suppression or enhancement of the immune response depending on duration of treatment and/or dose and nature of the antigen. To our knowledge, this is the first study to show that exogenous CORT can produce an enhancement in the immune response in chickens. iv In conclusion, environmental stressors such as high or low temperatures do affect the physiology of the fast-growing broiler. The adjustments the birds have to make to maintain homeostasis impacts on the course of common infectious diseases, such as IB, that normally is mild in the New Zealand poultry industry. The administration of exogenous CORT showed that this hormone may be part of the physiological stress response and acts as a messenger to prepare the immune system for potential challenges (e.g., infection).

This thesis analyses the concept of 'chance' as it is understood by two Muslim philosophers, Ibn Sīnā (Lat. Avicenna, CE 980-1037) and Ibn Rushd (Lat. Averroes, CE 1126-1198). On the philosophical plane, I seek to ascertain whether they are determinists, i.e., whether they hold that everything that happens is necessarily conditioned by its causes so that it could not have been otherwise. This analysis discusses chance from a physical and a metaphysical perspective. Physics is here understood in the Aristotelian sense as the study of nature and change, and metaphysics as the study of being qua being (ontology) and of the divine (theology). Hence a particular stress on natural causation and on divine providence and causation. On the historical-philosophical plane I endeavour to determine the historical/philosophical sources of their views, namely the Graeco-Arabic philosophical tradition - Aristotelian and Neoplatonic on the one Band, and the tradition of Islamic theology (kalām) on the other. Particular emphasis is laid upon the original way in which Ibn Sīnā and Ibn Rushd combine these two traditions.

Les protéases sont impliquées dans de nombreux processus biologiques et ont des origines très variées (cellules immunitaires, cellules épithéliales...). Leur activité est régulée par des inhibiteurs de protéases. Cette étude s'intéresse a` la balance entre les protéases épithéliales et leurs inhibiteurs dans le contexte de pathologies affectant l'intégrité´ de l'épithélium intestinal. Les conséquences d'un déséquilibre de la balance protéolytique ont été étudiées dans deux maladies chroniques aux composantes différentes, les Maladies Inflammatoires Chroniques de l'Intestin (MICI) (cycles de poussées inflammatoires) et le Syndrome du Côlon Irritable (SCI) (cycles de douleur). La muqueuse colique de patients atteints de SCI sécrète une activité de type trypsine dont l'origine et les fonctions sont mal définies. Cette étude s'intéresse à la source de cette activité trypsique dans le côlon des patients atteints de SCI, sa nature et son rôle dans l'activation neuronale. L'activité trypsique chez les patients atteints de SCI est augmentée majoritairement dans les cellules épithéliales (CEI). La stimulation de monocouches de cellules épithéliales avec du LPS ou de l'épinéphrine induit une augmentation de la quantité de trypsine-3 et de sa sécrétion au pôle basal de la cellule épithéliale stimulée, en corrélation avec une augmentation de l'activité trypsique. L'hyperactivité de la trypsine-3 au pôle basal entraine une perte de la fonction barrière de l'épithélium, anomalie retrouvée aussi chez les patients atteints de SCI. De plus, nous avons mis en évidence que la trypsine-3 est capable d'activer les neurones humains myentériques et les neurones sensitifs murins. In vivo, son administration intra-rectale à des souris induit une hypersensibilité viscérale dépendante du récepteur PAR2 (Protease Activated Receptor-2). Ainsi, les cellules épithéliales intestinales des patients atteints de SCI produisent et sécrètent de la trypsine-3 spécifiquement au pôle basal. Cette activité trypsique active des neurones sensitifs, participant à l'hypersensibilité viscérale, symptôme majeur dont souffrent les patients atteints de SCI. Les pathologies inflammatoires peuvent aussi être source d'un dérèglement protéolytique. Les patients atteints de MICI présentent une dérégulation de la balance élastolytique colique. Notre équipe a montré que l'apport d'ELAFINE (un inhibiteur d'élastase) par la bactérie génétiquement modifiée L.lactis au contact de la muqueuse inflammatoire protège les souris traitées en réduisant l'inflammation intestinale. Cependant les mécanismes protecteurs médiés par L'Elafine restent encore à déterminer. L'ELAFINE est un inhibiteur d'élastases mais possède aussi des propriétés antimicrobiennes. Afin de mettre en lumière quelles fonctions de L'Elafine portent les propriétés anti-inflammatoires des mutants de L'Elafine ont été générés et insérés dans la bactérie L.lactis: une mutation pour annuler sa fonction inhibiteur de protéase, un second mutant de son domaine antimicrobien et un dernier mutant ayant perdu ces 2 fonctions. Dans des monocouches de cellules intestinales épithéliales, l'apport d'ELAFINE protège de l'inflammation médiée par un déséquilibre de l'activité élastolytique : on observe une restauration de la fonction barrière de l'épithélium et une diminution des cytokines pro-inflammatoires (CXCL8 et IP10). La mutation du domaine antimicrobien n'affecte pas ces propriétés. Cependant, l'absence de la boucle inhibitrice annihile les propriétés anti-inflammatoires de L'Elafine. Ces travaux ont pu mettre en évidence l'importance de l'équilibre protéolytique au sein de la cellule épithéliale dans les pathologies intestinales. La balance entre les protéases et leurs inhibiteurs joue donc un rôle dans l'homéostasie épithéliale et les pathologies inflammatoires
Proteases are involved in some biologic processes and their origins are variable (immune cells, epithelial cells...). Their activity is regulated by antiproteases. This study investigates the balance between proteases and their inhibitors in pathologies which modify epithelium integrity. Consequences of an unbalance in proteolytic activity was studied in two chronic pathologies with different components: Inflammatory Bowel Disease (IBD) (cycles of inflammatory boost) and Irritable Bowel Syndrome (IBS) (cycles of pain symptoms). Colonic mucosa from IBS patients releases trypsin activity. The origin and the functions of this activity are not well defined. This study investigated the source of this trypsin activity in the côlon of IBS patients, its nature and its role in neuronal activation. Trypsin activity from IBS patients is increased mostly in epithelial cells. Stimulation of epithelial cell monolayers with LPS or epinephrine induces an increase of trypsin-3 quantity and its secretion specifically in the basal side of epithelial cells. This is in correlation with the increase of trypsin activity. Trypsin-3 hyperactivity at the basal side provokes a loss of epithelium barrier function, which is also found in colons of IBS patients. Then, we have highlighted that trypsin-3 is able to activate human myenteric neurons and murine sensitive neurons. In vivo, its intra-rectal administration to mice induces a visceral hypersensitivity dependent of PAR2 (Protease Activated Receptors 2). Thus, intestinal epithelial cells from IBS patients produce and release trypsin-3 specifically on their basal side. This trypsin activity activates sensitive neurons which participate to visceral hypersensitivity, a major symptom of IBS patients. Inflammatory pathologies could be a source of proteolytic malfunction. IBD patients have a dysregulation of elastolytic balance in the colon. Our team has shown that ELAFIN (an elastase inhibitor) delivered by the bacteria genetically modified L.lactis near the inflamed mucosa, protects mice from intestinal inflammation. However, the protective mechanisms induced by ELAFIN need to be investigate. ELAFIN is an elastase inhibitor but have also antimicrobial properties. With the aim to highlight what function of ELAFIN owns anti-inflammatory properties, mutants of ELAFIN have been generated and were insered into L.lactis: a first mutant lacked its antiprotease function, a second lacked antimicrobial properties and a last mutant lacked both properties. In intestinal epithelial monolayers, ELAFIN delivered by L.lactis protects against inflammation: a restauration of epithelial barrier function and a decrease of pro-inflammatory cytokines (CXCL8 and IP10) are observed. Mutation of antimicrobial domain doesn't affected these properties. Nevertheless, the absence of inhibitory loop annihilates anti-inflammatory functions of ELAFIN. This work highlights the importance of proteolytic balance inside the epithelial cell in intestinal pathologies. The balance between proteases and antiproteases plays an important role in epithelial homeostasis

The interest for Business Process Management (BPM) is increasing in Sweden. Government agencies such as the Swedish Nation Board of Student Aid (CSN), the Swedish Companies Registration Office (Bolagsverket) and the Swedish Social Insurance Agency (Försäkringskassan) are implementing BPM into their organizations. Sogeti is an IT-consulting company that has employees that works at CSN with BPM, and one of the tools they use for integration is IBM Integration Designer. Since this technology is new and is gaining popularity, there is a need for increased knowledge about it. This thesis report explores the tool IBM Integration Designer, regarding its different ways to integrate with systems and compares the different options for exposing the integration solu- tions. By researching documentation from IBM, and by learning how to use the tool itself, knowledge could be acquired about IBM Integration Designer. The result was an implementation of the five different export bindings: SCA, HTTP, SOAP over HTTP, Enterprise JavaBeans (EJB) and Java Messaging Service (JMS), a comparison of these export bindings and finally a service in IBM Inte- gration Designer that accesses a database and uses an external SMS API to send text messages. The result has been satisfactory to the purpose of this project in giving insight into IBM Integration Designer, one of the popular tools for inte- grating BMP.

This thesis is an attempt to study Ibn Jama'ah's approach to educational reform as illustrated in his Tadhkirat al-Sami' wa al-Mutakallim fi Adab al-'Alim wa al Muta'allim. Ibn Jama'ah (639-733 A.H. / 1241-1333 A.D.) was a distinguished Shafi'i Chief Judge and prominent scholar of Islamic studies during the Bahri Mamluk's Sultanate in Egypt and Syria. Ibn Jama'ah's theory of education reflects an emphasis on the Qur'an and hadith as primary sources of knowledge. Specifically, his suggestions for curriculum composition are designed to facilitate the evolution of a pious, religiously oriented generation of scholars. Ibn Jama'ah emphasizes the need for the teacher to foster motivation among his student body. Furthermore, he recommends that the teacher carefully gauge his students' abilities and limits. Another critical component of Ibn Jama'ah's educational program is the need for the teacher to realize the influence he has on his students; consequently, he must carry himself in a respectable and pious manner. He also addresses the various duties needed for a student to excell in his studies. In his evaluation of the teacher and student relationship, one can detect his affiliation with the Sufi khanqah tradition.

This thesis explores the epistemology and theology of a Sufi theorist Ibn al-ʿArabī (d. 1240) and one of the most controversial Muslim scholars Ibn Taymiyya (d. 1328) comparatively. Although scholars have long emphasized the differences between these two influential and significant figures, the purpose of this thesis is to focus on their potential similarities. The present work at the outset shows that both Ibn al-ʿArabī and Ibn Taymiyya regard revelation, the human intellect, and inspiration – unveiling (ilhām – kashf) to be the main epistemological means for acquiring religious knowledge. However, they differ over the relative value of these means or tools. Both Ibn al-ʿArabī and Ibn Taymiyya place revelation at the top of the hierarchy, but after that Ibn al-ʿArabī places inspiration – unveiling ahead of reason while Ibn Taymiyya reverses the order. The other main subject elaborated in this thesis is the theological issue of the divine names and attributes. This subject is connected to many other theological problems, and both Ibn al-ʿArabī and Ibn Taymiyya pay considerable attention to it. Just as many similar approaches, ideas, and interpretations may be seen concerning their epistemological stances, it is possible to see similarities in their theological positions although there are crucial distinctions as well. The unique way of finding out this similarity is to keep in mind the existence of the duality found in the thought and expression of Ibn al-ʿArabī. Briefly, the main finding of the thesis is that Ibn al-ʿArabī and Ibn Taymiyya are more similar in their epistemologies and theologies than many scholars have thought. They are benefitting from the same epistemological means while putting them in different order. Also, they share similar theological points concerning the divine names and attributes, creation, the divine will, and the incomparability of God while expressing them through different conceptions and styles.

Public Health
Ph.D.
Annually, more than 15 million U.S. children witness intimate partner violence (IPV) in their homes between caregivers. Witnessing IPV during childhood has been associated with increased risk of adulthood IPV victimization, although this relationship has most commonly been examined among older populations of white adults (McKinney, Caetano, Ramisetty-Mikler, & Nelson, 2009; Straus, 1989; Whitfield, Anda, Dube, & Felitti, 2003), not minority women who experience a disproportionately high burden of IPV victimization (Breiding, Chen, & Black, 2014). It also is unclear whether there are specific characteristics of childhood witnessing of IPV that may aid in understanding the increased probability of IPV victimization observed among young, minority women. This study examined associations between four characteristics of childhood witnessing of IPV (direction of violence witnessed, academic disruption, fearfulness, and frequency) and adulthood physical and sexual IPV victimization. Further, psychosocial moderators of these associations were examined. Existing data collected from 315 women residing in North Philadelphia were analyzed. Overall, 28% of women witnessed IPV during childhood and nearly one in four women reported IPV victimization. Among these women there was diversity with regards to the characteristics of witnessing reported. Unadjusted associations observed that women who witnessed bidirectional IPV, experienced academic disruption, or reported fearful witnessing were significantly more likely to experience adulthood IPV victimization than women who did not witness IPV during childhood. After adjusting for covariates, these witnessing characteristics were no longer significantly associated with adulthood IPV victimization; however, the risk of adulthood IPV victimization was consistently related to current depressive symptoms and substance use. Modest support for effect modification by low self-esteem was observed. A significant difference in the predicted probability of adulthood IPV victimization was observed among women reporting bidirectional childhood witnessing of IPV and high self-esteem in adulthood compared to women reporting bidirectional witnessing and low self-esteem (difference=0.51, 95% CI: 0.06, 0.95, p=0.027). Childhood abuse was associated with greater probability of adulthood IPV victimization regardless of the frequency of childhood witnessing of IPV (aOR=5.8, 95% CI: 2.2, 13.6). Given the high prevalence of childhood witnessing of IPV observed in this study, these findings highlight the importance of screening for childhood IPV witnessing. In addition, these results suggest particular groups of urban women who may be at highest risk for adulthood IPV victimization, including those experiencing depressive symptoms, substance use, and a history of childhood abuse. Identifying adolescents and young adults with these characteristics and providing early prevention services may reduce the probability of future IPV victimization among young, sexually active urban women. Additionally, examining the role of low self-esteem during child- and adulthood may inform efforts to prevent IPV victimization. These findings highlight the need for prospective, longitudinal studies of urban children and adolescents to examine the complex causal pathways between childhood witnessing of IPV, psychosocial characteristics, and adulthood IPV victimization.
Temple University--Theses

Durch die enge Verknüpfung von Konstruktion, Berechnung und Versuch in Verbindung mit einem mathematischen Ersatzmodell ist es möglich den optimalen Einlasskanal für einen bestimmten Zylinderkopf zu entwerfen.

Western Kentucky University (WKU) offered a course entitled Biochemistry for the Health Sciences in the spring semester of 2007. The course was taught in Interactive Television (ITV) format, and was the first time a class had been taught in the format enabling two-way communication by the Chemistry Department. One section was present in an ITV broadcast classroom on the main campus in Bowling Green, Kentucky. Another section received the broadcast on the campus of Western Kentucky University-Owensboro (WKU-O), at Owensboro Community and Technical College (OCTC), located in Owensboro, Kentucky. The purpose of this thesis is to evaluate the delivery of the chemistry class and to ascertain if ITV is a suitable format for chemistry classes. Professor Larry Byrd taught the class in person to twenty-nine students in Bowling Green. David Martin, the author of this thesis, attended the class at WKU-0 along with one student, who was required to take the class for her Health Sciences major. The thesis addresses the technical aspects of an ITV class, the logistical challenges faced at both locations, the subject matter covered, and the reactions of students. A survey was administered to the Bowling Green class and the Owensboro student was interviewed. Suggestions for improvements were made for future classes. The conclusion is that ITV can certainly be used successfully for the delivery of chemistry classes. The student in Owensboro received an "A" for the semester. This result compared to 50% "A's", 32 % "B's", 9% "C's", and 9% "D's" and "W's" for the total class. The most needed improvement for the class is the inclusion of demonstrations of laboratory experiments on video. The technical and logistical challenges encountered can be overcome with more experience on the part of the professor and improvements in the courier transportation system. This particular class is scheduled to be offered via ITV in the fall 2007 semester and will be broadcast to multiple remote locations. This limited study of an ITV chemistry class shows that the delivery method is appropriate and reasonable. To broaden the statistical base for comparison of ITV classes to traditional classes, another series of experiments should be performed. A proposal for a study between a traditional class, an ITV main campus class, an ITV remote class, and a web-based class is included. This study is designed to compare student performance over a range of delivery methods for the same course.

This thesis highlights a new interpretation of the writings of Ibn Taymiyya. Previous scholarship has stressed the legalistic, social or religious aspect, often at the expense of the philosophical content of Ibn Taymiyya's works. The explanatory insight of a study on the theory of knowledge, hitherto neglected, is evidenced by its capability to demonstrate the convergence of elementary, religious, intuitive and rational principles. The theory illustrates the concerns of a synthetic mind whose attempt was to broaden and not restrict the domain of knowledge vis-a-vis the Divine. Knowledge is not man's privilege and is available to all of creation. All created entities have the capacity to know their Creator. This thesis attempts to shed light on the mechanisms of the acquisition of knowledge about the Divine in their modes of availability to the creatures and to man. Tensions of the human predicament thus participate in the logical framework of the discussion. The attempt is to define the domain of knowledge, its components, and its parameters in the quest for a perfect acquisition of knowledge.

International Telemetering Conference Proceedings / October 30-November 02, 1989 / Town & Country Hotel & Convention Center, San Diego, California
The Voice Main Card (VMC) functions as an enhanced telephone answering machine and is designed as a plug-in card for the IBM PC and compatibles. In addition to standard answering machine functions, the VMC features programmable outgoing message selection, message routing, response to caller's touch tone signals, and remote programming ability. The VMC will answer incoming telephone calls, deliver outgoing messages which are Programmably selectable from as many as 16 digitized audio messages stored on the PC's hard disk, record incoming messages to the hard disk or optionally to an external tape recorder, route messages to a specified receiver, respond to a caller's touch tone signals, and enter a remote programming mode as a result of a special code sent by the caller. Audio messages are processed digitally via A/D and D/A converters which receive and send 8-bit data to and from the IBM PC through a selectable port address. The A/D conversion is implemented with the ADC0802 which is operated at a clock rate of 512 kHz. The D/A conversion is implemented with the DAC0830. Interface with the telephone line is accomplished with the speech circuit TP5700 coupled through opto-couplers. Messages are recorded using an external 600-ohm dynamic microphone and played back through an external 8-ohm speaker. A full-featured, pull-down menu program is provided with the VMC and implements all programmable functions. Data In/Out (I/O) with the card is interrupt-driven to allow apparent simultaneous disk I/O.

Bakgrund: IBS (Irritabel Bowel Syndrome) är en funktionell mag-tarmsjukdom, med en oklar patofysiologi och etiologi. IBS är en vanlig åkomma hos den västerländska befolkningen och karakteriseras av återkommande buksmärta/obehag, uppblåsthet, diarré och/eller förstoppning samt ökad gasbildning. Det finns idag inget botemedel för IBS, men intresset för probiotika som behandling vid IBS har på senare tid ökat. Probiotika, vilket definieras som levande mikroorganismer, som när de intas i adekvata mängder ger en hälsovinst hos värden tros ha en symptomatisk effekt vid IBS, och har därmed varit av intresse för behandling av IBS. Syfte: Syftet med denna litteraturstudie är att undersöka om probiotika har någon terapeutisk effekt och kan användas som behandling vid IBS. Metod: Fem randomiserade, dubbelblindade, placebo-kontrollerade studier granskades. De undersökte den terapeutiska effekten av varierande probiotika på IBS-patienter. Artiklarna erhölls genom sökningar i den medicinska databasen PubMed, under februari månad 2016. Resultat: Av dessa fem studier visade fyra på en signifikant symptomförbättring av framförallt buksmärta/obehag. Bäst resultat sågs i en studie som undersökte probiotikan Lactobacillus plantarum 299v. Tre av fem studier visade på en signifikant förbättring av testpersonernas livskvalité. I endast en av studierna, som undersökte effekten av Escherichia coli Nissle 1917, påvisades ingen signifikant skillnad mellan E.coli (probiotika) och placebo. Slutsats: Baserat på de fem studierna föreligger det i nuläget skäliga bevis för att en behandling med specifika probiotika kan ge symtomförbättring av buksmärta/obehag hos IBS-patienter, och förbättrar patienternas livskvalité. Vidare studier krävs för att fastställa den mest effektiva probiotikan, dosen och behandlingslängden.
Background: IBS (Irritable Bowel Syndrome) is a functional gastrointestinal disorder, with an unclear etiology and pathophysiology. IBS is a common disorder in the Western population and is characterized by recurrent abdominal pain/ discomfort, bloating, flatulence, diarrhea and/ or constipation. There is currently no cure for IBS, but the interest in probiotics as an option of treatment has recently increased. Probiotics have been defined as live microorganisms that, when administered in adequate amounts, provide a health benefit on the host, and are believed to have a symptomatic effect in IBS. Probiotics have therefore been of interest for the treatment in IBS. Purpose: The purpose of this study is to examine whether probiotics have a therapeutic effect and if it could be used as a treatment for IBS. Method: Five randomized, double-blind, placebo-controlled studies were reviewed that examined the therapeutic effect of different probiotics in the IBS-patients. Articles were obtained through searches in the medical database PubMed, during the month of February 2016. Results: Four of the five studies showed a significant improvement of symptoms especially in abdominal pain/ discomfort. The best results were seen in a study that investigated the probiotic Lactobacillus plantarum 299v. Three of the five studies showed a significant improvement in quality of life (QOL) of the subjects in the study. One of the five studies which examined the effect of Escherichia coli Nissle 1917, there was no significant difference between E.coli (probiotics) and placebo. Conclusion: There is reasonable evidence that treatment with certain probiotics might provide improvement in symptoms of abdominal pain/ discomfort, and increase patients quality of life based on the five studies. Further studies are required to determine the most effective probiotic, dose and duration of IBS-treatment.

This paper describes the IBM Power8 system in comparison to the Intel Xeon processors, widely used in today’s solutions. The performance is not evaluated only on the whole system level but also on the level of threads, cores and a memory. Different metrics are demonstrated on typical optimized algorithms. The benchmarked Power8 processor provides extremely fast memory providing sustainable bandwidth up to 145 GB/s between main memory and processor, which Intel is unable to compete. Computation power is comparable (Matrix multiplication) or worse (N-body simulation, division, more complex algorithms) in comparison with current Intel Haswell-EP. The IBM Power8 is able to compete Intel processors these days and it will be interesting to observe the future generation of Power9 and its performance in comparison to current and future Intel processors.

This diploma thesis presents Big Data technologies and their possible use cases and applications. Theoretical part is initially focused on definition of term Big Data and afterwards is focused on Big Data technology, particularly on Hadoop framework. There are described principles of Hadoop, such as distributed storage and data processing, and its individual components. Furthermore are presented the largest vendors of Big Data technologies. At the end of this part of the thesis are described possible use cases of Big Data technologies and also some case studies. The practical part describes implementation of demo example of Big Data technologies and it is divided into two chapters. The first chapter of the practical part deals with conceptual design of demo example, used products and architecture of the solution. Afterwards, implementation of the demo example is described in the second chapter, from preparation of demo environment to creation of applications. Goals of this thesis are description and characteristics of Big Data, presentation of the largest vendors and their Big Data products, description of possible use cases of Big Data technologies and especially implementation of demo example in Big Data tools from IBM.

Zugl.: Berlin, Techn. Universiẗat, Habil.-Schr., 1994 u.d.T.: Steinbach, Jörg: Theoretische Untersuchungen reaktionstechnischer Probleme in der Syntheseoptimierung und der chemischen Sicherheitstechnik.