95 research outputs found
Cycle threshold probability score for immediate and sensitive detection of B.1.351 SARS-CoV-2 lineage
No full textBackground Detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern associated with immune escape is important to safeguard vaccination efficacy. We describe the potential of delayed N gene amplification in the Allplex SARS-CoV-2 Assay (Seegene) for screening of the B.1.351 (20H/501.V2, variant of concern 2 [VOC.V2], South African SARS-CoV-2 variant) lineage. Methods In a study cohort of 397 consecutive polymerase chain reaction-positive samples genotyped by whole-genome sequencing, amplification curves of E/N/S-RdRP targets indicated delayed N vs E gene amplification characteristic of B.1.351. Logistic regression was used to calculate a VOC.V2 probability score that was evaluated as a separate screening test in an independent validation cohort vs sequencing. Results B.1.351 showed a proportionally delayed amplification of the N vs E gene. In logistic regression, only N and E gene cycle thresholds independently contributed to B.1.351 prediction, allowing calculation of a VOC.V2 probability score with an area under the curve of 0.94. At an optimal dichotomous cutoff point of 0.12, the VOC.V2 probability score achieved 98.7% sensitivity at 79.9% specificity, resulting in a negative predictive value (NPV) of 99.6% and a positive predictive value of 54.6%. The probability of B.1.351 increased with an increasing VOC.V2 probability score, achieving a likelihood ratio of 12.01 above 0.5. A near-maximal NPV was confirmed in 153 consecutive validation samples. Conclusions Delayed N vs E gene amplification in the Allplex SARS-CoV-2 Assay can be used for fast and highly sensitive screening of B.1.351
Antigenic profiling of the porcine reproductive and respiratory syndrome virus
No full textPorcine reproductive and respiratory syndrome virus (PRRSV) causes reproductive failure in sows and boars, and is involved in the porcine respiratory disease complex, causing huge economical losses to the swine industry. Despite the development and broad implementation of vaccines, the virus could not be eradicated, and the high variability and fast evolution of the virus compromises the efficacy of the current vaccines in the field. Since PRRSV engages a complex interaction with the host’s immune system, and exploits several strategies to evade protective immunity, the development of new and better PRRSV vaccines is highly relying on a good understanding of the virus-specific immune response. The general goal of this work was to create new insights into one particular hallmark of PRRSV-specific immunity, namely the weak virus-neutralizing antibody response upon infection or vaccination in pigs, in sharp contrast with the robust non-neutralizing antibody response. Chapter 1 summarizes the current literature on general aspects of the virus and the disease, PRRSV-specific immunity, and PRRSV vaccines. Furthermore, the current knowledge on the PRRSV-specific neutralizing antibody response is extensively discussed. In Chapter 2, the particular problem underlying the work in this thesis is described, and a number of aims are stated. The next two chapters consist of research results addressing these aims. In Chapter 3, it was investigated to which extent virus-neutralizing antibodies can be induced by immunization of pigs with inactivated PRRSV. This study addresses the question whether the poor development of virus-neutralizing antibodies is dependent on structural properties of the virus, or rather on replication-dependent mechanisms. Experimental vaccines were produced, based on formerly optimized virus inactivation procedures and by using a number of different adjuvants. Vaccination of PRRSV-negative piglets systematically led to a strong virus-specific antibody response, but the virus-neutralizing antibody response was very weak or absent, indicating that the poor immunogenicity of neutralizing antibody targets in PRRSV is to a large extent inherent to the structural properties of the virus. Still, vaccination with a high dose of inactivated PRRSV in combination with one particular adjuvant induced virus-neutralizing antibodies, and provided a certain level of protection against viremia upon infection. These results formed the basis for further vaccine research at the Laboratory of Virology. Chapter 4 deals with the identification of non-neutralizing and neutralizing antibody targets in the European prototype PRRSV strain Lelystad virus (LV). In a first study (§ 4.1), the antibody response against linear epitopes in the GP4 protein was investigated. It was found that antibodies are to a large extent produced against one particular region of this protein that was earlier described as neutralizing antibody target. Since this region is highly variable, the antibody response against GP4 was also investigated for other virus strain. The conclusion of this study was that the variable region in GP4 of EU-type PRRSV induces neutralizing antibodies upon infection in pigs, but only against strictly homologous virus. In a second study (§ 4.2), the antibody response against the entire set of envelope proteins (GP2, E, GP3, GP4, GP5 and M) upon infection with LV was investigated. Twenty-one antigenic regions were identified that are capable of inducing antibodies in pigs. Peptide-purified antibodies against four antigenic regions turned out to reduce in vitro virus replication. In addition to the known neutralizing epitope in GP4, two neutralizing antibody targets were identified in GP2, and one in GP3. No neutralizing epitopes were found in E, GP5 and M. Since the neutralizing epitope in GP3 was one of the more immunogenic regions, the antibody response against this epitope was investigated more extensively in different virus strains. Taken together, it can be concluded from Chapter 4 that four linear neutralizing antibody targets are present in the LV envelope proteins: two weakly immunogenic epitopes in GP2, one more immunogenic epitope in GP3, and one strongly immunogenic but highly variable epitope in GP4. Chapter 5 provides a general discussion on the research data generated in this thesis. In § 5.1, it is discussed what can be learned from the vaccination studies in Chapter 3. § 5.2 summarizes the results from Chapter 4, and discusses them in a broader context. First, a synoptic visual overview is given of the non-neutralizing and neutralizing antibody targets that were identified in the viral envelope proteins. Next, some hypotheses are presented on how neutralizing antibodies against different parts of the viral proteins may interfere with viral replication. Subsequently, the current knowledge on PRRSV antigenicity is discussed in light of virus evolution, and some thoughts are made on how the high mutation rate and large variability of the virus may be involved in immune evasion. To conclude, it is critically evaluated to which extent the data generated in this thesis can contribute to the development of new and better PRRSV vaccines.SAMENVATTING
Het porcien reproductief en respiratoir syndroom virus (PRRSV) veroorzaakt reproductiestoornissen bij zeugen en beren, en is betrokken bij het porcien respiratoir ziektecomplex, wat leidt tot enorme economische verliezen in de varkenshouderij. Ondanks de ontwikkeling en brede toepassing van vaccins kon het virus niet worden uitgeroeid, en de grote variabiliteit en snelle evolutie van het virus staan de werkzaamheid van de huidige vaccins in het veld in de weg. Aangezien PRRSV een complexe interactie aangaat met het immuunsysteem van de gastheer en meerdere strategieën gebruikt om de beschermende immuniteit te ontwijken, is de ontwikkeling van nieuwe en betere PRRSV vaccins sterk afhankelijk van een goed inzicht in de virusspecifieke immuunrespons. De algemene doelstelling van deze thesis was het creëren van nieuwe inzichten in één specifiek kenmerk van de PRRSV-specifieke immuniteit, namelijk de zwakke virusneutraliserende antistofrespons na infectie of vaccinatie in varkens, in scherp contrast met de robuuste niet-neutraliserende antistofrespons. Hoofdstuk 1 geeft een samenvatting van de huidige literatuur over algemene aspecten van het virus en de ziekte, de PRRSV-specifieke immuniteit, en PRRSV vaccins. Verder wordt de huidige kennis over de PRRSV-specifieke neutraliserende antistofrespons uitgebreid besproken. In Hoofdstuk 2 wordt het specifiek probleem dat aan de basis ligt van deze thesis beschreven, en worden een aantal doelstellingen vooropgesteld. De volgende twee hoofdstukken omvatten onderzoeksresultaten die aan deze doelstellingen tegemoetkomen. In Hoofdstuk 3 werd onderzocht in welke mate virusneutraliserende antistoffen kunnen worden geïnduceerd door immunisatie van varkens met geïnactiveerd PRRSV. Deze studie komt tegemoet aan de vraag of de zwakke ontwikkeling van virusneutraliserende antistoffen afhankelijk is van structurele eigenschappen van het virus, of eerder van replicatie-afhankelijke mechanismen. Experimentele vaccins werden aangemaakt op basis van eerder geoptimaliseerde virusinactivatiemethodes, en gebruik makend van een aantal verschillende adjuvantia. Vaccinatie van PRRSV-negatieve biggen leidde telkens tot een sterke virusspecifieke antistofrespons, maar de virusneutraliserende antistofrespons was daarentegen erg zwak of afwezig, wat aangeeft dat de zwakke immunogeniciteit van neutraliserende epitopen in PRRSV grotendeels afhankelijk is van structurele eigenschappen van het virus. Toch konden door vaccinatie met een hoge dosis geïnactiveerd PRRSV in combinatie met één specifiek adjuvans virusneutraliserende antistoffen worden geïnduceerd, en er kon een zekere graad van bescherming worden geboden tegen viremie na infectie. Deze resultaten hebben de basis gevormd voor verder vaccinonderzoek aan het Laboratorium voor Virologie. Hoofdstuk 4 handelt over de identificatie van doelwitten voor niet-neutraliserende en neutraliserende antistoffen in de Europese prototype PRRSV stam Lelystad virus (LV). In een eerste studie (§ 4.1) werd de antistofrespons tegen lineaire epitopen in het GP4 eiwit onderzocht. Daaruit bleek dat antistoffen voornamelijk worden gevormd tegen één bepaalde regio in dit eiwit, die eerder werd beschreven als doelwit voor neutraliserende antistoffen. Aangezien deze regio sterk variabel is, werd de antistofrespons tegen GP4 ook onderzocht voor andere virusstammen. Het besluit van deze studie was dat de variabele regio in GP4 van EU-type PRRSV neutraliserende antistoffen induceert na infectie in varkens, maar enkel tegen strikt homoloog virus. In een tweede studie (§ 4.2) werd de antistofrespons tegen de volledige reeks gekende envelop-eiwitten (GP2, E, GP3, GP4, GP5 en M) na LV infectie onderzocht. Eénentwintig antigene regio’s werden geïdentificeerd die antistoffen kunnen induceren bij infectie in varkens. Peptide-opgezuiverde antistoffen tegen vier antigene regio’s bleken in vitro virusreplicatie te reduceren. Naast het gekende neutraliserend epitoop in GP4 werden twee doelwitten voor neutraliserende antistoffen gevonden in GP2, en één in GP3. In E, GP5 en M werden geen neutraliserende epitopen teruggevonden. Daar het neutraliserend epitoop in GP3 één van de meer immunogene regio’s was, werd de antistofrespons tegen dit epitoop verder onderzocht bij verschillende virusstammen. Samenvattend kan uit Hoofdstuk 4 worden besloten dat vier lineaire doelwitten voor neutraliserende antistoffen aanwezig zijn in de LV envelop-eiwitten: twee zwak immunogene epitopen in GP2, één meer immunogeen epitoop in GP3, en één sterk immunogeen maar aanzienlijk variabel epitoop in GP4. Hoofdstuk 5 tenslotte geeft een algemene bespreking van de onderzoeksresultaten in deze thesis. In § 5.1 wordt bediscussieerd wat kan worden geleerd uit de vaccinatie-experimenten in Hoofdstuk 3. § 5.2 vat de resultaten van Hoofdstuk 4 samen, en bespreekt deze in een bredere context. Eerst wordt een beknopt aanschouwelijk overzicht gegeven van de doelwitten voor niet-neutraliserende en neutraliserende antistoffen die werden geïdentificeerd in de virale envelop-eiwitten. Vervolgens worden een aantal hypotheses naar voor geschoven over de manier waarop neutraliserende antistoffen tegen verschillende delen van de virale eiwitten zouden kunnen interfereren met virusreplicatie. Daarna wordt de huidige kennis over PRRSV antigeniciteit bediscussieerd in het licht van virusevolutie, en een aantal bedenkingen worden gemaakt over hoe de hoge mutatiesnelheid en de grote variabiliteit van het virus een rol kunnen spelen bij het ontwijken van het immuunsysteem. Tot slot wordt kritisch geëvalueerd in welke mate de resultaten van deze thesis kunnen bijdragen tot de ontwikkeling van nieuwe en betere PRRSV vaccins
RNA-Sequence analysis of primary alveolar macrophages after in vitro infection with porcine reproductive and respiratory syndrome virus strains of differing virulence
No full textPorcine reproductive and respiratory syndrome virus (PRRSV) mainly infects porcine alveolar macrophages (PAMs), resulting in porcine reproductive and respiratory syndrome (PRRS) in pigs. Most of the transcriptomic studies on PAMs infected with PRRSV conducted thus far have made use of microarray technology. Here, we investigated the transcriptome of PAMs in vitro at 12 h post-infection with two European PRRSV strains characterized by low (Lelystad, LV) and high (Lena) virulence through RNA-Seq. The expression levels of genes, isoforms, alternative transcription start sites (TSS) and differential promoter usage revealed a complex pattern of transcriptional and post-transcriptional gene regulation upon infection with the two strains. Gene ontology analysis confirmed that infection of PAMs with both the Lena and LV strains affected signaling pathways directly linked to the innate immune response, including interferon regulatory factors (IRF), RIG1-like receptors, TLRs and PKR pathways. The results confirmed that interferon signaling is crucial for transcriptional regulation during PAM infection. IFN-beta 1 and IFN-alpha omega, but not IFN-alpha, were up-regulated following infection with either the LV or Lena strain. The down-regulation of canonical pathways, such as the interplay between the innate and adaptive immune responses, cell death and TLR3/TLR7 signaling, was observed for both strains, but Lena triggered a stronger down-regulation than LV. This analysis contributes to a better understanding of the interactions between PRRSV and PAMs and outlines the differences in the responses of PAMs to strains with different levels of virulence, which may lead to the development of new PRRSV control strategies
Monopile installation assessment: A critical assessment of an oscillating monopile during offshore installation
No full textAn offshore wind turbine requires a stable support structure to support the tower and nacelle. The support structure in the form of a monopile is lowered to the seabed and hammered in the soil. During the lowering process the monopile may show excessive motions, that occur on an irregular basis. The cause for this is unknown. This thesis is devoted to find the cause, to prevent this from happening in future operations. First off a literature study and analysis of the environmental conditions is carried out. A study in the mechanical behaviour of the system will provide a good perspective on the situation. Finally, the cause is verified with model tests and a numerical model is validated. From the environmental conditions wind and current are expelled as (main) contributor. Data analysis and system analysis conclude that waves are the main cause for the problem. In the model tests this cause is validated together with the mechanical behaviour (natural frequency) of the system. The natural frequency of the monopile is depending on the waterlevel. Wave frequencies (in the spectrum) close to the natural frequency cause a resonance phenomenon, with large monopile motions as consequence. A numerical model, based on a mechanical system and the Morison's equation is validated. The numerical model is a good and fast solution for modeling monopile motions, from wave conditions. The model can be used to predict future scenarios to prevent excessive monopile motions to occur, a (potential) tool for engineering purposes (determining workability)
Analysing the Relation Between Gaze Location and Gap Acceptance Decisions During Highway Merges
No full textBackground: Merging on a highway is a complex driving task that requires a lot of interaction with other road users. During these tasks, a driver is required to evaluate gaps in space and time between the themselves and other road users and obstacles in order to arrive at the right moment to merge onto the highway. To improve safety and increase road efficiency, it is necessary to understand the decision process during merging decisions. A way of achieving this, is to understand what visual information humans use during this decision process. This study investigated the relation between gaze location and gap acceptance decisions during highway merges.Methods: An experiment was performed in which 26 participants monitored an automated vehicle (AV) that was driving on a highway on-ramp. The participants were given the task to train the AV in whether or not to merge in front of an upcoming vehicle that was already driving on the highway. An eye tracker was used to measure gaze data, which was used to find the relation between gaze behaviour and decision outcomes and response times. A mixed-effects logistic model was used for a statistical analysis with decision outcomes as a dependent variable and different gap sizes as predictor variables. A mixed-effects linear model was used to find the relation between response times and dwell times and the different gap sizes and decision outcomes as predictor variables. For both the decision outcome and response time model, dwell time was later included to find the effect on the predictive validity.Results: The results show that a larger time and distance gap to the upcoming vehicle relate to a higher merging probability. For larger time gaps to the on-ramp, the probability of merging was found to be smaller. It was also found that time gaps to the end of the on-ramp significantly relate to response times, with an increase of 55ms per 1s. Larger time gaps to the upcoming vehicle significantly relates to larger response times, with an increase of 64ms per 1s. No significant relation was found between response time and distance gaps to the upcoming vehicle. The response time was found to be 0.60s longer for rejected gap decisions. The time gap to the end of the on-ramp significantly relates to dwell time, with an increase of 0.56% per 1s. The distance gap to the upcoming vehicle significantly relates to dwell time, with an increase of 0.60% per 10m. The time gap to the upcoming vehicle significantly relates to dwell time, with an increase of 0.52% per 1s. The presented results show as well that a significant relation exists between gaze behaviour and decision outcomes and response times. When analysing decision outcomes and response times, the interaction between dwell time and gap sizes should be taken into account. This improved the predictive validity of the used regression models.Conclusion: Several pieces of evidence suggest that gaze behaviour assist in understanding the human decision making process during merging. This study can serve as a basis for cognitive models that can investigate how the relation between gaze behaviour and gap sizes, decision outcomes and response times can help to understand and potentially predict gap acceptance decisions.Mechanical Engineering | Vehicle Engineering | Dynamics and Control
The Whale: Cultural centre Holesovice
No full textThe cultural centre will provide the city of Prague with a community centre combined with several large scale amenities (a cinema, a club, a big event hall and a skybar) to complement and, at the same time, relieve pressure on the inner city. The building is situated in a green, livable, accessible and diverse new area (outside of the clogged inner city) for both tourists and Praguers to experience and enjoy. Pedestrians are placed on a pedestal and find themselves in an unprecedented new inviting and exiting public space. The cultural centre will work on a city scale and a quarter scale. The goal should be to bring together different social groups. It will function as a community centre for the to be developed urban area as well as the existing neighborhoods (Letná and Holesovice). A place where they can meet, relax and use the wide variety of amenities. Concomitantly the building will give the Praguer an additional place to relax/go out during the weekend
Technical feasibility of a quick bridge replacement strategy with minimal traffic hindrance: On the retainment of existing foundations and the application of Advanced Cementitious Materials in an Accelerated Bridge Construction method
No full textBridges are a crucial part of the dense Dutch highway network. Almost 50% of the current bridges are built between 1960 and 1980 for an intended service life of 50 years and thus they might reach the end of their lifetime in the near future. This strongly indicates that a large challenge can be expected regarding infrastructural replacement.In current practice, the replacement of bridges within the highway network often leads to substantial traffic hindrance, which has a large negative impact on the Dutch economy. Therefore reduction of traffic hindrance – together with sustainability – is generally an important quality criteria in MEAT-procedures (Most Economic Advantageous Tender) for tender assignments. In order to prepare for the upcoming replacement challenge, there is a need for contractors, the government and product suppliers to invest in the development of a sustainable tender strategy now.The main objective of this thesis was to propose a tender strategy that incorporates sustainability and in particular the reduction of traffic hindrance into a technically feasible design. An extensive literature review has led to a quick bridge replacement strategy consisting of three time-reducing actions, listed according to their potential profit in construction time:1. Select the Accelerated Bridge Construction (ABC) approach:By means of lateral sliding or transportation by SPMTs (Self-Propelled Modular Transporters) entire superstructures can be constructed off-site and transported to their final location in a matter of hours.2. Retain the existing foundations:In terms of on-site construction time it would be beneficial to retain the existing foundations. Additionally, theory suggests that a profit in bearing capacity can be obtained compared to the current design codes, which might lead to the total elimination of additional foundation elements required.3. Avoid intermediate supports:Less elements are required if intermediate supports are avoided in the new bridge design. This does however lead to longer spans, and to prevent additional groundwork activities from an increase in deck height a higher slenderness must be obtained. Furthermore, the new superstructure design must be as light as possible, not only to allow for foundation retainment, but also to facilitate transportation and speed of erection.In particular, the technical feasibility of these actions - both separately and combined - required more research. A case study considering a two-span plate bridge was used to investigate these strategy actions. Research was done into the obtainable profit in retained pile foundations and the application of UHPC in a slender and lightweight bridge concept.It was concluded that the proposed tender strategy has a high potential. Not only is the on-site reduction time diminished but the retainment of the existing foundations and the application of UHPC in a slender and lightweight superstructure may also lead to a highly sustainable design.<br/
Finding the Operating Limits and Optimal Configuration of an Electrically Assisted Turbofan
No full textHybrid electric propulsion is a promising new technology to reduce emissions and fuel consumption of an aircraft. In this study a parallel hybrid electric configuration is considered. Since electric motors supply power in the most demanding take-off and climb phases to achieve the required thrust, the turbofan can be redesigned.It was found that the cruise thrust specific fuel consumption could be improved, even though no electric power is supplied in that phase. The hybrid-electric aircraft shows potential to save fuel over a complete mission. Most of these savings are however achieved due to fully electric taxiing. The weight of the electrical system largely offsets the efficiency improvements of the gas turbine during flight. Given that optimistic technology levels are used for these results, parallel hybrid electric propulsion is not expected to be used in the coming decade on short haul flights with narrow-body aircraft.Aerospace Engineering | Flight Performance and Propulsio
Nieuwenhuysenbuurt Futureproof: Minimizing the construction waste released by the demolition of Dutch post-war social housing
No full textMoving towards the Dutch circularity goals of 2050, this project aims to diminish the amount of construction waste released by the demolition of 282 post-war dwellings in the Nieuwenhuysenbuurt in Amsterdam West (1954). Through the use of a flexible and demountable building system, construction waste is diminished now and in the future.Architecture, Urbanism and Building Sciences | Building Technolog
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