West Nile Virus (WNV) is a mosquito-borne pathogen of global significance. It is active on several continents and is responsible for recent outbreaks of fever and fatal encephalitis in humans and horses. While highly virulent strains have been reported in Europe, North, Central and South America, only a benign subtype of WNV (Kunjin virus – KUNV) occurs in Australia. However, virulent, exotic WNV strains are seen as a significant threat to Australia due to the ease with which this virus can move between continents and the presence of suitable vectors and hosts already within Australia. KUNV and WNV subtypes are antigenically and genetically very closely related and cross-react in traditional serological tests. This cross-reactivity makes it very difficult to differentiate between KUNV and WNV infections using standard serological tests. The aim of this thesis was to identify immunogenic epitopes unique to KUNV or WNV and to use these epitopes in the development of a rapid assay that would enable the diagnosis of and surveillance for exotic virulent strains of WNV in Australia. The rapid diagnostic platform chosen was a red blood cell (RBC) agglutination assay that was originally patented and commercialised by AGEN Biomedical Ltd. The RBC agglutination assay reagent consists of the Fab region of a human erythrocyte-specific monoclonal antibody (mAb) conjugated to the epitope of interest (in this instance, a WNV-specific peptide). This bi-functional reagent causes the agglutination of the patient’s erythrocytes in the presence of WNV-specific antibody in the patient’s serum. Traditionally, these RBC agglutination reagents have been produced by chemical conjugation. However, a potentially easier and cheaper method involves the linking of the gene encoding the erythrocyte-specific antibody to that encoding the epitope to create a recombinant version of the bi-functional agglutination reagent through expression using prokaryotic or eukaryotic systems. To identify potential differential epitopes, 18 mAbs to WNV (NY99 strain) prM and envelope (E) proteins were assessed. One mAb (17D7) differentially recognised WNV and KUNV in ELISA and maintained recognition of its corresponding epitope upon reduction and carboxymethylation of the viral antigen, suggesting a continuous (linear) epitope. Using synthetic peptides, the epitope was mapped to a 19 amino acid sequence (WN19: E147-165) encompassing the WNV NY99 E protein glycosylation site at position 154. An amino acid substitution at position E156 of many KUNV strains abolishes this glycosylation moiety. The inability of WNV-positive horse and mouse sera to bind the synthetic peptides indicated that glycosylation was required for recognition of peptide WN19 by WNV-specific antibodies in sera. N-linked glycosylation of WN19 was achieved through expression of the peptide as a C-terminal fusion protein in mammalian cells and specific reactivity of WNV-positive horse sera to the glycosylated WN19 fusion protein was shown by Western blot. Additional sera collected from horses that had been infected with Murray Valley encephalitis virus (MVEV), which is similarly glycosylated at position E154 and exhibits high sequence identity to WNV NY99 in this region, also recognised the recombinant peptide. In contrast, no reactivity with the recombinant peptide was observed by sera from horses infected with the unglycosylated WNV subtype, KUNV. Failure of most WNV- and MVEV-positive horse sera to recognise the epitope as a deglycosylated fusion protein (75% and 100% respectively) confirmed that the N-linked glycan is important for antibody recognition of the peptide. Together, these results suggest that the induction of antibodies to the WN19 epitope during WNV infection of horses is generally associated with E protein glycosylation of the infecting viral strain. To assess the feasibility of using peptide WN19 in a rapid immunoassay, the peptide was recombinantly fused to a RBC (glycophorin)-specific single chain antibody (scFv) using previously published constructs which were developed for the bacterial expression of similar bi-functional reagents. To facilitate glycosylation of peptide WN19, the genes for the bi-functional agglutination reagents were subsequently cloned into eukaryotic expression vectors. An additional set of constructs were also produced in which the genes for the variable regions of the anti-RBC antibody were cloned into a vector for the secreted expression of an intact, humanised IgG1 molecule. Stable cell lines were produced for each of these constructs and secreted up to 700 ng/mL glycophorin-reactive antibody. The secreted recombinant protein could be harvested directly from the cell culture medium and used in RBC agglutination assays, where these bi-functional agglutination reagents could be cross-linked either with mAb 17D7 or by anti-peptide WN19 antibodies present in WNV-positive horse serum. The WNV NY99 prM protein was also identified as a useful marker of WNV-infection in horses, as well as a putative antigen to differentiate equine WNV NY99 and KUNV infections using Western blot. Two anti-WNV prM mAbs were also generated in this study and will be extremely valuable in future studies. Preliminary analysis of the prM epitope(s) bound by these mAbs and WNV-immune sera indicate that the binding site(s) is likely to be localised to pr and is conformational.

West Nile Virus (WNV) is a mosquito-borne pathogen of global significance. It is active on several continents and is responsible for recent outbreaks of fever and fatal encephalitis in humans and horses. While highly virulent strains have been reported in Europe, North, Central and South America, only a benign subtype of WNV (Kunjin virus – KUNV) occurs in Australia. However, virulent, exotic WNV strains are seen as a significant threat to Australia due to the ease with which this virus can move between continents and the presence of suitable vectors and hosts already within Australia. KUNV and WNV subtypes are antigenically and genetically very closely related and cross-react in traditional serological tests. This cross-reactivity makes it very difficult to differentiate between KUNV and WNV infections using standard serological tests. The aim of this thesis was to identify immunogenic epitopes unique to KUNV or WNV and to use these epitopes in the development of a rapid assay that would enable the diagnosis of and surveillance for exotic virulent strains of WNV in Australia. The rapid diagnostic platform chosen was a red blood cell (RBC) agglutination assay that was originally patented and commercialised by AGEN Biomedical Ltd. The RBC agglutination assay reagent consists of the Fab region of a human erythrocyte-specific monoclonal antibody (mAb) conjugated to the epitope of interest (in this instance, a WNV-specific peptide). This bi-functional reagent causes the agglutination of the patient’s erythrocytes in the presence of WNV-specific antibody in the patient’s serum. Traditionally, these RBC agglutination reagents have been produced by chemical conjugation. However, a potentially easier and cheaper method involves the linking of the gene encoding the erythrocyte-specific antibody to that encoding the epitope to create a recombinant version of the bi-functional agglutination reagent through expression using prokaryotic or eukaryotic systems. To identify potential differential epitopes, 18 mAbs to WNV (NY99 strain) prM and envelope (E) proteins were assessed. One mAb (17D7) differentially recognised WNV and KUNV in ELISA and maintained recognition of its corresponding epitope upon reduction and carboxymethylation of the viral antigen, suggesting a continuous (linear) epitope. Using synthetic peptides, the epitope was mapped to a 19 amino acid sequence (WN19: E147-165) encompassing the WNV NY99 E protein glycosylation site at position 154. An amino acid substitution at position E156 of many KUNV strains abolishes this glycosylation moiety. The inability of WNV-positive horse and mouse sera to bind the synthetic peptides indicated that glycosylation was required for recognition of peptide WN19 by WNV-specific antibodies in sera. N-linked glycosylation of WN19 was achieved through expression of the peptide as a C-terminal fusion protein in mammalian cells and specific reactivity of WNV-positive horse sera to the glycosylated WN19 fusion protein was shown by Western blot. Additional sera collected from horses that had been infected with Murray Valley encephalitis virus (MVEV), which is similarly glycosylated at position E154 and exhibits high sequence identity to WNV NY99 in this region, also recognised the recombinant peptide. In contrast, no reactivity with the recombinant peptide was observed by sera from horses infected with the unglycosylated WNV subtype, KUNV. Failure of most WNV- and MVEV-positive horse sera to recognise the epitope as a deglycosylated fusion protein (75% and 100% respectively) confirmed that the N-linked glycan is important for antibody recognition of the peptide. Together, these results suggest that the induction of antibodies to the WN19 epitope during WNV infection of horses is generally associated with E protein glycosylation of the infecting viral strain. To assess the feasibility of using peptide WN19 in a rapid immunoassay, the peptide was recombinantly fused to a RBC (glycophorin)-specific single chain antibody (scFv) using previously published constructs which were developed for the bacterial expression of similar bi-functional reagents. To facilitate glycosylation of peptide WN19, the genes for the bi-functional agglutination reagents were subsequently cloned into eukaryotic expression vectors. An additional set of constructs were also produced in which the genes for the variable regions of the anti-RBC antibody were cloned into a vector for the secreted expression of an intact, humanised IgG1 molecule. Stable cell lines were produced for each of these constructs and secreted up to 700 ng/mL glycophorin-reactive antibody. The secreted recombinant protein could be harvested directly from the cell culture medium and used in RBC agglutination assays, where these bi-functional agglutination reagents could be cross-linked either with mAb 17D7 or by anti-peptide WN19 antibodies present in WNV-positive horse serum. The WNV NY99 prM protein was also identified as a useful marker of WNV-infection in horses, as well as a putative antigen to differentiate equine WNV NY99 and KUNV infections using Western blot. Two anti-WNV prM mAbs were also generated in this study and will be extremely valuable in future studies. Preliminary analysis of the prM epitope(s) bound by these mAbs and WNV-immune sera indicate that the binding site(s) is likely to be localised to pr and is conformational.

No description available.

Made available in DSpace on 2015-04-11T13:54:27Z (GMT). No. of bitstreams: 1 Joquebede.pdf: 1993216 bytes, checksum: 33d4a9881ff6ddc76b2d4122fba69fbf (MD5) Previous issue date: 2013-08-27 / FAPEAM - Fundação de Amparo à Pesquisa do Estado do Amazonas / Em decorrência do alto índice de Tuberculose (TB) no Estado do Amazonas e reconhecendo a adesão ao tratamento como fator relevante para o controle da doença, este trabalho teve como objetivo avaliar o efeito de um programa de atenção farmacêutica na adesão ao tratamento da tuberculose em pacientes da Policlínica Cardoso Fontes na cidade de Manaus- AM. No total 77 pacientes foram randomizados em grupo intervenção (24) e grupo controle (53) sendo comparados posteriormente para avaliação da adesão ao tratamento da TB. O recrutamento dos participantes foi realizado por 3 meses, após a primeira consulta e recebimento dos medicamentos, sendo acompanhados por 6 meses de tratamento. O acompanhamento/seguimento farmacoterapêutico realizado foi baseado no método Dáder ao longo dos seis meses de tratamento, com a utilização de roteiro padronizado que incluiu: educação em saúde; escuta ativa e identificação das necessidades; análise da situação e intervenção/orientação farmacêutica; avaliação e documentação dos resultados Para verificar as associações foi utilizado o teste do quiquadrado de Pearson (χ2) e, para as variáveis que não puderam ser avaliadas pelo o teste χ2, o teste não paramétrico de Mann-Whitney. Para verificação da associaçãodos fatores de risco associados a não adesão ao tratamento mais frequentemente descritos na literatura e a adesão dos pacientes da PCF foi realizada análise multivariada com regressão logística. Também foi realizada regressão logística individual (Odds ratio) para os fatores que se apresentaram associados a não adesão dos pacientes. Para todos os testes o nível de significância foi de 0,05. A atenção farmacêutica demonstrou uma influência positiva na promoção da adesão, com altos índices de adesão no grupo que recebeu a intervenção (atenção farmacêutica). Além disso, houve aumento do conhecimento sobre a doença e sobre o tratamento. O monitoramento do tratamento também auxiliou na baixa ocorrência e prevenção dos Problemas Relacionados ao uso de Medicamentos (PRM) e de Resultados Negativos associados à Medicação (RNM). O estudo demonstrou a importância da Atenção Farmacêutica nos serviços de atendimentos aos pacientes em tratamento da tuberculose, como medida para obter melhores resultados na adesão e na terapia medicamentosa como um todo.

This work deals with the path finding for nonholonomic robots using probabilistic algorithms. The theoretical part analyzes the general problem of finding routes. Subsequently, the work will focus on probabilistic algorithms. The practical part describes design of the applet and web sites that demonstrate probabilistic algorithms to user-specified objects.

This paper deals with the problem of object path planning in 3D space. The goal is to create program which allows users to create a scene used for path planning, perform the planning and finally visualize path in the scene. Work is focused on probabilistic algorithms that are described in the theoretical part. The practical part describes the design and implementation of application. Finally, several experiments are performed to compare the performance of different algorithms and demonstrate the functionality of the program.

The Interactive Highway Safety Design Model (IHSDM) is a suite of software developed by the Federal Highway Administration (FHWA) for monitoring and analyzing two-lane rural highways in the United States. As IHSDM is a fairly "young" program a limited amount of research has been conducted to evaluate its practicability and reliability. To determine if IHSDM can be adopted into the engineering decision making process in Utah, a study was conducted under the supervision of the Utah Department of Transportation (UDOT) to evaluate its applicability to audit safety of two-lane rural highways in Utah. IHSDM consists of six modules: Policy Review Module (PRM), Crash Prediction Module (CPM), Design Consistency Module (DCM), Traffic Analysis Module (TAM), Intersection Review Module (IRM), and Driver/Vehicle Module (DVM) (still under construction). Among the six modules, two were chosen for evaluation because of their applicability to audit safety of the two-lane rural highways in Utah, namely CPM and IRM. For the evaluation of the CPM, three two-lane rural highway sections were selected. The results of this evaluation show that the CPM can produce reasonably reliable crash predictions if appropriate input data, especially alignment data, reflect the existing conditions at reasonable accuracy and engineering judgment is used. Using crash records available from UDOT's crash database and CPM's crash prediction capability, UDOT's traffic and safety engineers can locate "hot spots" for detailed safety audit, thus making the safety audit task more focused and effective. Unlike the CPM, the outputs of the IRM are qualitative and include primarily suggestions and recommendations. They will help the traffic and safety engineers identify what to look for as they visit the sites, such as a lack of stopping sight distance and a lack of passing sight distance. The interpretation of the IRM requires knowledge of various aspects of highway design, familiarity with A Policy on Geometric Design of Highways and Streets by the American Association of State Highway and Transportation Officials (AASHTO), and experience in traffic engineering. Based on the findings of the study, it is concluded that the CPM and IRM of IHSDM could be a useful tool for engineering decision-making during safety audits of two-lane rural highways. But the outputs from these modules demand knowledge and experience in highway design. It is recommended that the other modules of IHSDM be tested to fully appreciate the capability of IHSDM. The software can be a knowledgebased program that can help novice engineers to learn how to design safe two-lane rural highways.

The designer proline-rich antimicrobial peptide (PrAMP) Chex1-Arg20 amide (ARV-1502) is active against Gram-negative and Gram-positive pathogens in differentmurine infection models when administered parenterally and possesses a wide therapeutic index. Here we studied the pharmacokinetics of ARV-1502 for the first time when administered intramuscularly or intravenously (IV) in Sprague Dawley rats and Beagle dogs. First, a specific and robust quantitation method relying on parallel reaction monitoring (PRM) using a high-resolution hybrid quadrupole-Orbitrap mass spectrometer coupled on-line to reversed-phase uHPLC was established and validated. The limit of detection was 2 ng/mL and the limit of quantitation was 4 ng/mL when spiked to pooled rat and dog plasma. When ARV-1502 was administered IV at doses of 75 and 250 μg/kg in dogs and rats, the plasma concentrations were 0.7 and 3.4μg/mL 2min post-administration, respectively. ARV-1502 plasma concentrations declined exponentially reaching levels between 2 and 4 ng/mL after 2 h. Intramuscular administration of 0.75 mg/kg in dogs and 2.5 mg/kg in rats resulted in a different pharmacokinetics profile. The plasma concentrations peaked at 15min post-injection at 1μg/mL (dogs) and 12μg/mL (rats) and decreased exponentially within 3 h to 4 and 16 ng/mL, respectively. The initial plasma concentrations of ARV-1502 and the decay timing afterwards indicated that the peptide circulated in the blood stream for several hours, at some point above the minimal inhibitory concentration against multidrug-resistant Enterobacteriaceae, with blood concentrations sufficient to suppress bacterial growth and to modulate the immune system.

Par rapport aux exigences de plus en plus importantes relatives au Maintien en Condition Opérationnelle d'un système industriel, le processus de maintenance joue un rôle fondamental pour l'amélioration de la disponibilité, de la productivité, etc. Pour essayer de contrôler au mieux ces performances, les responsables de maintenance doivent donc être capables de choisir les stratégies de maintenance et les ressources à mettre en oeuvre les plus adaptées aux besoins. Dans un objectif d'aide à la prise de décisions en maintenance, les travaux présentés dans ce mémoire ont pour objet de proposer une méthodologie pour l'élaboration d'un modèle support permettant par simulation d'évaluer les différentes stratégies. La valeur ajoutée de la méthodologie réside dans l'unification, à base de modèles relationnels probabilistes (PRM), des différents types de connaissance nécessaires à la construction de ce modèle d'évaluation. Ce dernier est ainsi construit à partir de motifs génériques et modulables représentatifs des variables décisionnels du système industriel (système principal) et de son système de maintenance. Ces motifs, par instanciation, facilitent la construction des modèles d'applications spécifiques. Cette méthodologie, issue du projet ANR SKOOB, est testée sur le cas applicatif de la maintenance d'un système de production de ferment. / Nowadays, the importance of the maintenance function has increased, due to the requirements on the maintain in operational conditions phase (MCO) of the system-of-interest (SI). As well as for the relevant role of maintenance in improving availability, performance efficiency, total plant availability, etc. To control performances, maintenance managers should be able to make some choices about the maintenance strategies and the resources that can fulfil the requirements. Within this context, we propose a methodology to formalize a model allowing to perform simulation to assess maintenance strategies. The scientific contribution of our work is that this approach unify by using a probabilistic relational model (PRM), different kind of knowledge needed to assess maintenance strategies. Knowledge is presented as generic and modular patterns based on PRM. These patterns integrate relevant decisional variables of the system of interest and of its maintenance system. This approach eases the modeling phase for a specific application. This methodology is one of the results of the project ANR SKOOB. This approach was tested on an industrial case for the maintenance of a harvest production process

由於全球化的浪潮，將市場競爭帶向新的局面，企業不再只尋求短期 的利潤，更傾向與他們的外部競爭者以及內部人員維持長期的合作關係， 以替企業帶來更大利益，這種共生共存的關係，亦即夥伴關係。已有許多 研究發現，發展夥伴關係能為企業帶來益處，學者也找出建立夥伴關係和 夥伴關係演化過程中的關鍵成功因素，或者針對形成夥伴關係的結合力進 行研究，然而，很少有學者提及如何管理夥伴關係，本研究以此觀點出發， 目的在找出供應商夥伴關係管理的重要因素並提出實踐的執行項目，以供 企業評估並改進其供應商夥伴關係管理的流程，另外提出夥伴關係度量 (Partner Relationship Metrics)之概念，讓企業據此分數來選擇供應商夥伴。 本研究首先由17 篇關於夥伴關係的文獻中找出18 個夥伴關係的關鍵 成功因素(Critical Success Factor, CSF)，並在每一關鍵成功因素下，定義若 干個關鍵執行項目(Key Practice Items, KPI)。這些關鍵執行項目都代表著供 應商合作關係中執行的行動，以關鍵成功因素及文獻中學者所提的關鍵結 合力(Bond)形成訪談內容，採取企業訪談法挑選台灣資訊電子製造業供應 鏈中分別屬於零組件供應商、加工製造者以及系統組裝商的六家企業進行 訪談。研究結果顯示，六家企業都認為順應性(Adapatibility)、信任(Trust)、 參與度(Participation)是重要的因素，另外也發現，訪談對象依其企業規模、 屬於買方或賣方市場等因素乃訪談結果有差異的原因之一。 關鍵字：B2B、供應商夥伴關係、關鍵成功因素、夥伴關係度量 / Over the past few years, companies have faced increasing challenges in their relationships with external competitors as well as internal employees. Recent years, managers tend to build a long term, ongoing relationships which are called partnerships. Researches have found that developing partnerships can contribute to the profit of a company and reduce the risk. Some researchers focused on the factors of the successful partnership; others on the bonds between partners. However, studies is lacking on how to manage a partnership. The purpose of this project is to find factors that make a successful supplier partnership and propose some key practice items (KPI) as well. Furthermore, we define partner relationship metrics (PRM) by combing bonds and success factors of partnership to help managers evaluate their supplier partners. This project is based on partnership literatures and business interviews. First, we chose 18 critical success factors (CSF) of the partnership from 17 literatures. Then, we surveyed 6 businesses of Taiwan Information & Electronic Industry and divided them into 3 categories to get the empirical data. We found all companies agree that Adaptability, Trust and Participation are the top 3 important factors in managing their supplier partner relationships. In addition, we discovered that the size of the companies and the one who holds the power in the supplier-buyer relationship influence the research results. Key words: B2B, supplier partner relationship, critical success factor (CSF), partner relationship metrics (PRM)