One-third of the world’s population is latently infected with Mycobacterium tuberculosis, which causes over 2 million deaths every year. The current live attenuated vaccine, Bacille Calmette-Guerin (BCG), protects against miliary tuberculosis in children, but fails to consistently protect against pulmonary tuberculosis in adults. The global resurgence of tuberculosis, together with the HIV pandemic and emerging multi-drug resistance, has heightened the need for an effective vaccine. Completion of the M. tuberculosis genome sequence paved way for identification of many new candidate antigens for protective vaccine against tuberculosis. This includes the discovery of two multigene families of proteins PE and PPE which constitute 10% of the coding capacity of the M. tuberculosis genome. Members of the PE and PPE protein families are characterized by highly conserved N-terminal domains and the C-terminus, however, exhibit considerable variation in the number of residues as well as in the sequence. Till date, little is known about the functional role of the proteins of PPE or PE family in the biology of M.tuberculosis. Some of the PE_PGRS proteins have been found to be associated with the cell wall and influence interactions with other cells. PE and PPE family of proteins are of potential interest from the point of view of immune response, since they show antigenic variation which may play a role in immune evasion. Very little is known about the immunogenecity of these two classes of proteins and only few proteins have been shown to be potent B or T cell antigens, like Rv3873, Mtb39 and Rv0915c. Two proteins from PE_PGRS subfamily, Rv1759c and Rv3367 are expressed during infection and show antibody response in humans and rabbits, respectively. Rv1196 and Rv0915c from PPE family have been shown to be good T cell antigens. Another study has shown that the PE domain of PE_PGRS protein Rv1818c upon immunization into mice induces good cell mediated immune response in mice, whereas the PGRS domain is responsible for good humoral response. In humans there is increasing evidence to suggest that CD8+ T cells are elicited in response to infection with mycobacteria. CD8+ CTL may play an important role through several mechanisms. They produce potent anti-bacterial cytokines such as IFN-γ and TNF-α in response to antigenic stimulation and IFN-γ is critical for immunity to TB. Thus, identification of antigens and peptides that induce T cell responses could be useful for designing new vaccines to protect against TB. Relatively few epitopes in mycobacterial antigens have so far been identified for human CD8 T cells. In this regard, release of genome sequences of M. tuberculosis has provided an opportunity to identify proteins with vaccine potential that could give immune protection in individuals with different HLA backgrounds. Objectives and scope of the present work 1. Prediction of putative T cell antigens in PE and PPE family of proteins of Mycobacterium tuberculosis through immuno-informatics approach 2. Evaluation of immune response to three of the PE and PPE proteins in mouse model. 3. Evaluation of immune response against chosen PE and PPE proteins of Mycobacterium tuberculosis with Human Peripheral Blood Mononuclear Cells (PBMCs) from PPD positive healthy donors and TB patients. 4. Immune response to multi-epitope DNA vaccine construct for Mycobacterium tuberculosis. Prediction of MHC class I peptides from PE and PPE proteins. In an effort to identify potential T cell antigens from PE and PPE family of proteins, we have carried out a systematic in silico analysis of the 167 different PE and PPE proteins. Employing immuno-informatics approach, a set of HLA class I binding peptides have been identified from these proteins. Further, their binding abilities have been ascertained using independent methods such as molecular modeling and structural analysis methods. The nonameric sequences from PE and PPE families of proteins were predicted to contain high percentage of binding peptides to human class I HLA, whereas PE_PGRS proteins show relatively low level of binding. This difference is seen in spite of PE and PE_PGRS being Sub-families of the same family, PE. Seventy-one high- as well as low-affinity peptides from both PE and PPE proteins have been analyzed for structural compatibility with crystal structures of HLA in terms of intermolecular energies and were found to correlate well with the corresponding affinities predicted by the BIMAS algorithm. Most of the peptides binding to HLA are specific with very few promiscuous binders. Identification of T cell epitopes from three of the PE/PPE proteins using DNA immunization This work describes the evaluation of immune responses to three of the PE and PPE proteins in mouse model. Three of PE and PPE proteins, coded by Rv1818c, Rv3812 and Rv3018c genes were chosen based on immuno-informatics approach. They were cloned, expressed in prokaryotic and mammalian expression vectors and recombinant protein expressing stable cell lines were made. T lymphocytes from DNA immunized mice recognize synthetic peptides from chosen proteins in vitro, indicating that these peptides are being processed and presented by MHC molecules to T cells. By MHC stabilization assay, 5 of the synthetic peptides were found to stabilize the MHC class I molecules on the cell surface for more than 6 hrs, validating the computational prediction. Recognition of T cell epitopes derived from PE/PPE proteins by human PBMCs This work describes the evaluation of immune response against three of PE and PPE proteins of Mycobacterium tuberculosis with Human Peripheral Blood Mononuclear Cells (PBMCs) from PPD positive Healthy donors and TB patients. Proliferation response of PBMCs from ten PPD positive healthy donors as well as from ten TB patients, indicated that the peptides from PE and PPE proteins of Mtb can sensitize naive T cells and induce peptide specific IFN-γ and also the T cell response to the chosen peptides was both HLA class I restricted and CD8 mediated. After the peptide specific expansion, significant percentage of CD8+ T cells were shown to secrete IFN-γ and stained positive for perforin. Antigen specific CD8+ T cells were found to have cytolytic potential in addition to their cytokine function. Immune response to a multiepitope DNA vaccine in mouse model Minigene poly-epitope vaccine constructs coding for nine peptides derived from identified T cell antigens of PE and PPE proteins and three of the experimentally mapped epitopes from M tuberculosis was designed and constructed. The minigene was used to immunize mice and the immune response was tested. The DNA primed splenocytes recognized the full length poly-epitope protein as well as the individual peptides. T cell response to epitopes was enhanced by mere presence in multi-epitope construct compared to full length antigens. Human PBMCs derived from both PPD+ve and TB patients also recognized the peptides in vitro. It is thus obvious that a large cocktail of proteins are required to achieve reasonable population coverage. Besides, this work suggests the feasibility of designing haplotype specific subunit vaccine, which can be given to individuals with known HLA haplotype. The haplotype specific vaccines can be combined to target a population where the distribution of HLA alleles is known. This work also indicates that use of single or limited number of genes in a DNA vaccine may not be suitable to cover a given population.

Bakgrund: Pandemin orsakad av Covid-19 har tvingat intensivvården till stora omställningar. Som del av restriktioner för att reducera risk för smittspridning måste skyddsutrustning användas vid patientkontakt med smittad patient. Användandet av skyddsutrustning har introducerat tankar på hur den vårdande relationen mellan patient och intensivvårdssjuksköterska påverkats.  Syfte: Syftet med studien var att belysa intensivvårdssjuksköterskans upplevelse av hur arbetet i skyddsutrustning under pågående pandemi påverkat den vårdande relationen med patienten. Metod: Semistrukturerade intervjuer genomfördes med sex intensivvårdssjuksköterskor. Intervjuerna låg vidare till grund för analys enligt tolkande fenomenologisk analys (IPA).  Resultat: Fyra överordnade teman framträdde ur analysen. I temat fokus på kommunikation framkom olika sätt skyddsutrustningen påverkat kommunikationen. Upplevelsen av avskärmning framkom i temat fokus på distans där närhet uttrycktes som avgörande för en vårdande relation. I temat fokus på lidande beskrevs hur skyddsutrustningen inverkat på lindrande beröring men också att den kan orsaka lidande. Känslor av otillräcklighet i den vårdande relation och maktlöshet inför att inte förmå förändra eller påverka framkom i temat fokus på känslor.  Slutsats: Skyddsutrustningen påverkar den vårdande relationen. Skyddsutrustningen reducerar möjligheten till etablering av vårdande relation vilket riskerar minska patientens möjlighet till delaktighet. Behov föreligger av ytterligare forskning om påverkan med fokus på prevention. Ett organisatoriskt medvetande om skyddsutrustningen inverkan på den vårdande relationen kan ligga till grund för interventioner ämnade att reducera dess negativa påverkan. / Background: The pandemic caused by COVID-19 has forced the intensive care to make several adjustments. As part of restrictions to prevent spread of infection personal protective equipment (PPE) has to be used when in contact with infected patients. The use of PPE has introduced questions of its impact on the caring relationship between the patient and intensive care nurse.  Aim: The aim with this study was to illustrate the intensive care nurses experience of how working in PPE during the pandemic has affected the caring relationship with the patient.  Method: Semi- structured interviews was conducted with six intensive care nurses. The interviews served as data for an interpretative phenomenological analysis (IPA).  Results: Four superior themes emerged from the analysis. In the theme focus on communication emerged different way of which the PPE has affected communication. The experience of being shielded emerged in the theme focus on distance where closeness was expressed as essential. In the theme focus on suffering was the PPE´s impact on touch described and also its potential cause of suffering. Emotions of inadequacy in regards to the caring relationship and powerlessness in regards of not being able to change or affect emerged in the theme focus on emotions.  Conclusion: The caring relationship has been affected by the use of PPE. PPE reduces the possibility of establishing a caring relationship which endangers the patient’s active participation. There is need for further research about affect with focus on prevention. An organizational awareness regarding PPE´s effect on the caring relationship could lay the foundation for interventions supposed to reduce its negative impact.

Workplace injuries are common in today's society due to a lack of adequately worn safety equipment. A system that only admits appropriately equipped personnel can be created to improve working conditions and worker safety. The goal is thus to develop a system that will improve construction workers' safety. Building such a system necessitates computer vision, which entails object recognition, facial recognition, and human recognition, among other things. The basic idea is first to detect the human and remove the background to speed up the process and avoid potential interferences. After that, the cropped image is subjected to facial and object recognition. The code is written in Python and includes libraries such as OpenCV, face_recognition, and CVZone. Some of the different algorithms chosen were YOLOv4 and Histogram of Oriented Gradients. The results were measured at three respectively five-meter distances. As a result of the system’s pipeline, algorithms, and software, a mean average precision of 99% and 89% was achieved at the respective distances. At three and five meters, the model achieved a precision rate of 100%. The recall rates were 96% - 100% at 3m and 54% - 100% at 5m. Finally, the fps was measured at 1.2 on a system without GPU. / Skador på arbetsplatsen är vanliga i dagens samhälle på grund av att skyddsutrustning inte används eller används felaktigt. Målet är därför att bygga ett robust system som ska förbättra säkerhet. Ett system som endast ger tillträde till personal med rätt skyddsutrustning kan skapas för att förbättra arbetsförhållandena och arbetarsäkerheten. Att bygga ett sådant system kräver datorseende, vilket bland annat innebär objektigenkänning, ansiktsigenkänning och mänsklig igenkänning. Grundidén är att först upptäcka människan och ta bort bakgrunden för att göra processen mer effektiv och undvika potentiella störningar. Därefter appliceras ansikts- och objektigenkänning på den beskurna bilden. Koden är skriven i Python och inkluderar bland annat bibliotek som: OpenCV, face_recognition och CVZone. Några av de algoritmer som valdes var YOLOv4 och Histogram of Oriented Gradients. Resultatet mättes på tre, respektive fem meters avstånd. Systemets pipeline, algoritmer och mjukvara gav en medelprecision för alla klasser på 99%, och 89% för respektive avstånd. För tre och fem meters avstånd uppnådde modellen en precision på 100%. Recall uppnådde värden mellan 96% - 100% vid 3 meters avstånd och 54% - 100% vid 5 meters avstånd. Avslutningsvis uppmättes antalet bilder per sekund till 1,2 på ett system utan GPU.

This master’s thesis focus on the analysis of work conditions and risk assessment for specific company and specific project. It brings better condition to current action in health and safety area during all working activities. Emphasis is placed on safety of work and health protection of workers. The work provides information about theory of security work explores its practical application in the organization and gives a view into risk assess and risk management.