Process of designing robust, dependable, safe and secure software for medical devices: Point of care testing device as a case study

This article has been made available through the Brunel Open Access Publishing Fund.Copyright © 2013 Sivanesan Tulasidas et al. This paper presents a holistic methodology for the design of medical device software, which encompasses of a new way of eliciting requirements, system design process, security design guideline, cloud architecture design, combinatorial testing process and agile project management. The paper uses point of care diagnostics as a case study where the software and hardware must be robust, reliable to provide accurate diagnosis of diseases. As software and software intensive systems are becoming increasingly complex, the impact of failures can lead to significant property damage, or damage to the environment. Within the medical diagnostic device software domain such failures can result in misdiagnosis leading to clinical complications and in some cases death. Software faults can arise due to the interaction among the software, the hardware, third party software and the operating environment. Unanticipated environmental changes and latent coding errors lead to operation faults despite of the fact that usually a significant effort has been expended in the design, verification and validation of the software system. It is becoming increasingly more apparent that one needs to adopt different approaches, which will guarantee that a complex software system meets all safety, security, and reliability requirements, in addition to complying with standards such as IEC 62304. There are many initiatives taken to develop safety and security critical systems, at different development phases and in different contexts, ranging from infrastructure design to device design. Different approaches are implemented to design error free software for safety critical systems. By adopting the strategies and processes presented in this paper one can overcome the challenges in developing error free software for medical devices (or safety critical systems).Brunel Open Access Publishing Fund

THE BIOLOGY AND MANAGEMENT OF BRUCELLOSIS IN YELLOWSTONE BISON

Disease management along the boundaries of wildlife reserves is a growing conservation problem worldwide, as infected wildlife can migrate outside protected areas and pose a threat to livestock and human health. The bison Bison bison population in Yellowstone National Park has long been infected with Brucella abortus, the bacterium causing bovine brucellosis. Concern over migratory bison transmitting B. abortus to cattle herds on lands adjacent to Yellowstone has led to proposals for bison vaccination. Model simulations suggest that vaccination is unlikely to eradicate B. abortus from Yellowstone bison but could be an effective tool for reducing the level of infection and eliminating unpopular management practices such as lethal culling. The culling of Yellowstone bison to reduce the risk of brucellosis transmission to cattle is negatively affecting long-term bison conservation because of difficulties in diagnosing actively infected animals. Age-specific serology and B. abortus culture assays from slaughtered bison were used to develop a diagnostic tool to estimate whether particular animals are infective. Findings suggest that active B. abortus infection is age-dependent, which allows true infection probabilities to be estimated based on age and quantitative diagnostic tests. Active brucellosis infection was associated with below-average nutritional condition, with the intensity of B. abortus infection being influenced by seasonal reductions in dietary protein and energy. The reproductive strategy of Yellowstone bison is linked with the seasonal availability of food, which increases bison fitness but may have consequences for B. abortus infection. Seasonal food restriction may also influence the ability of vaccinated bison to recall protective immune responses when later exposed to B. abortus. The rate of fat metabolism was an important factor influencing cell-mediated responses. Thus, individual variation and the seasonal availability of food may reduce vaccine efficacy when vaccination is applied at the population level. Consequently, effective management practices will require a diverse range of integrated methods, which include maintaining separation of livestock and wildlife, managing habitat to reduce brucellosis transmission, and reducing disease prevalence in wildlife. The long-term success of these management practices will depend on sound science and support of the stakeholders involved

Performance evaluation of synergic operation of algorithms enabling opportunistic networks - D4.3

Deliverable D4.3 del projecte OneFITPreprin

Formulation, implementation considerations, and first performance evaluation of algorithmic solutions - D4.1

Deliverable D4.1 del projecte Europeu OneFIT (ICT-2009-257385)This deliverable contains a first version of the algorithmic solutions for enabling opportunistic networks. The presented algorithms cover the full range of identified management tasks: suitability, creation, QoS control, reconfiguration and forced terminations. Preliminary evaluations complement the proposed algorithms. Implementation considerations towards the practicality of the considered algorithms are also included.Preprin

An epidemiological investigation of Coxiella burnetii and Chlamydia spp. as infectious agents causing abortion in dairy cattle in Uruguay

Infectious diseases are the leading aetiological cause of bovine abortion globally as well as in Uruguayan dairy farms. Studies conducted in this country showed that a proportion of aborted fetuses without diagnosis present histological lesions compatible with the action of an infectious agent. These undiagnosed abortion cases could be due to infectious agents not currently included in the battery of diagnostic tests routinely applied. Infectious agents such as Coxiella burnetii and Chlamydia spp., typically considered to have a minor or relative marginal role in causing abortion in cattle, have never been systematically investigated in Uruguay. This thesis challenged the current diagnostic protocols, seeking evidence supporting the inclusion of these agents to dismiss the number of undiagnosed cases. This work investigated the association of these pathogens with cattle abortion in commercial dairy herds and the potential as zoonotic threats in the local context. The thesis will first introduce the main characteristics of dairy production in Uruguay (Chapter One) and thoroughly review Coxiella burnetii and Chlamydia abortus (Chapter Two). After this, two main sections will be presented. The first section including a systematic review and meta-analysis (Chapter Three), and the second section comprising the observational fieldwork studies conducted out on dairy farms in Uruguay, which includes a cross-sectional study of pooled milk (PM) samples from commercial herds (Chapter Four), a case-series sampling of placentas (Chapter Five), a case-control sampling of aborted and non-aborted dairy animals (Chapter Six), and finally a retrospective cohort evaluation of workers exposed to a local bovine Q fever outbreak (Chapter Seven). A final discussion (Chapter Eight) will consider the main findings' implications and integrate these results into the general body of knowledge. The systematic review and meta-analysis evidenced bulk tank milk (BTM) sampling as a broadly used epidemiological methodology for large-scale investigations of C. burnetii and provided a global framework of the prevalence of C. burnetii in collective milk samples from commercial farms. This study showed a widespread herd-level circulation of C. burnetii in bovine dairy farms by reporting a high overall global prevalence of 37.0% (CI95% 25.2-49.5%). A meta-regression showed the herd size as the most relevant moderator, with the odds of a BTM sample testing positive doubling with every unit increase. This C. burnetii meta-prevalence roles as a benchmark for comparison with the findings of the molecular investigation on PM samples obtained from local dairy farms. The mass-scale molecular evaluation on PM samples evidenced a low incidence (1.7%) of C. burnetii DNA. The findings showed that clinically healthy (asymptomatic) cows might shed the bacterium, raising awareness of Q fever as potential food safety and public health concern considering the C. burnetii survival as a milk-borne pathogen in unpasteurised milk and raw dairy products. Due to the exceptionally high infectivity, low infective doses, and aerosol transmission, the culture of C. burnetii and Chlamydiales should be done on BSL-3 laboratories not currently operating in Uruguay. Bacterial cultivation was not attempted in any of the studies conducted in this thesis. Hitherto the lack of diagnostic tools and the impossibility of doing culture have restricted local epidemiological investigation of these agents. Therefore, developing diagnostic tests to be used routinely in domestic laboratories is imperative to save costs and optimise currently available facilities and work with higher autonomy. For this purpose, a published protocol targeting well-evaluated genes was adapted to provide an available tool for local laboratories. Coxiella burnetii, C. abortus, and C. pecorum were investigated for the first time using an m-PCR in placentas from aborted dairy cows. Coxiella burnetii-DNA was detected and quantified in those samples, which supported this bacterium as an abortifacient agent in Uruguay. No co-infections of these pathogens were found. Evidence supporting Chlamydiales as a source of cattle abortion remain blurred. Coxiella burnetii was detected on the aborted placenta from a cow from an artisanal cheese-producing farm. Consumption of raw milk and dairy products represent a potential source for human infection. This finding underlined that the public health risk posed by C. burnetii should not be neglected and should be emphasised the need for on-farm milk pasteurisation by local artisanal cheesemakers. Molecular investigation of C. abortus in vulvo-vaginal swabs samples showed no evidence of this bacterium neither in aborted nor in control animals. Difficulties in identifying low-grade infection and evaluating a single sample per animal would have constrained the detection. The first attempted studies conducted so far support C. abortus as a no substantial abortifacient agent in cattle from Uruguay. Serological evidence confirmed the local bovine population as a potential reservoir for C. burnetii infection in humans. Anti-C. burnetii phase II IgM and IgG immunoglobulins were detected in a group of farmworkers and laboratory technicians exposed to aborted dairy cattle or aborted materials (fetuses and placenta) by indirect immunofluorescence. Molecular approaches were assessed, optimised and validated on veterinary clinical samples such as aborted placentas, vulvo-vaginal swabs or collective milk samples, providing valuable alternatives beyond the bacterial culture and isolation. The thesis presents original research studies that utilise different epidemiological strategies to search for evidence of an association between the infection by the pathogens and the occurrence of bovine abortion.Agencia Nacional de Investigación e InnovaciónInstituto Nacional de Investigación Agropecuari

The 3′ processing factor CstF functions in the DNA repair response

Following DNA damage, mRNA levels decrease, reflecting a coordinated interaction of the DNA repair, transcription and RNA processing machineries. In this study, we provide evidence that transcription and polyadenylation of mRNA precursors are both affected in vivo by UV treatment. We next show that the polyadenylation factor CstF, plays a direct role in the DNA damage response. Cells with reduced levels of CstF display decreased viability following UV treatment, reduced ability to ubiquitinate RNA polymerase II (RNAP II), and defects in repair of DNA damage. Furthermore, we show that CstF, RNAP II and BARD1 are all found at sites of repaired DNA. Our results indicate that CstF plays an active role in the response to DNA damage, providing a link between transcription-coupled RNA processing and DNA repair

Mechanisms of Endocytic Sorting: A Dissertation

Endocytosis is important for the regulation of signal transduction and for the movement of essential cellular components from outside the cell to their appropriate intracellular compartment(s). Two established mechanisms of endocytosis are clathrinmediated (CME) and clathrin-independent endocytosis, and they are responsible for internalization of different ligands. In this study, the newly established technique of total internal reflection fluorescent microscopy (TIRF-M) was used, along with standard biochemical and molecular biological tools, to systematically study the sorting and early trafficking of two established ligands of endocytosis, transferrin (Tf) and epidermal growth factor (EGF). TIRF-M studies revealed that Tf binds its receptor that is located in large clathrin arrays positioned just below the surface of the cell and that these large clathrin platforms serves as the major site of CME at the plasma membrane. EGF endocytosis is very different and occurs as follows 1) the liganded EGFR recruits Rab5 to the plasma membrane, 2) Rab5 concentrates around vesicles containing liganded EGFR and 3) these vesicles co-localize with EEA1 enriched endosomes. EEA1 was shown to play a pivotal role in EGF endocytosis, establishing a new role for EEA1 in vesicle trafficking in addition to its role in tethering and fusion. Finally, WDFY2, a new FYVE domain protein was shown to decorate a specific subset of vesicles, upstream of the EEA1 vesicle pool that appear to participate in Tf endocytosis. These studies establish new functions and components of endocytosis that enhances our understanding of this complex process