Parent-of-origin-specific allelic associations among 106 genomic loci for age at menarche.

Age at menarche is a marker of timing of puberty in females. It varies widely between individuals, is a heritable trait and is associated with risks for obesity, type 2 diabetes, cardiovascular disease, breast cancer and all-cause mortality. Studies of rare human disorders of puberty and animal models point to a complex hypothalamic-pituitary-hormonal regulation, but the mechanisms that determine pubertal timing and underlie its links to disease risk remain unclear. Here, using genome-wide and custom-genotyping arrays in up to 182,416 women of European descent from 57 studies, we found robust evidence (P < 5 × 10(-8)) for 123 signals at 106 genomic loci associated with age at menarche. Many loci were associated with other pubertal traits in both sexes, and there was substantial overlap with genes implicated in body mass index and various diseases, including rare disorders of puberty. Menarche signals were enriched in imprinted regions, with three loci (DLK1-WDR25, MKRN3-MAGEL2 and KCNK9) demonstrating parent-of-origin-specific associations concordant with known parental expression patterns. Pathway analyses implicated nuclear hormone receptors, particularly retinoic acid and γ-aminobutyric acid-B2 receptor signalling, among novel mechanisms that regulate pubertal timing in humans. Our findings suggest a genetic architecture involving at least hundreds of common variants in the coordinated timing of the pubertal transition

Low Temperature-Dependent Salmonid Alphavirus Glycoprotein Processing and Recombinant Virus-Like Particle Formation

Pancreas disease (PD) and sleeping disease (SD) are important viral scourges in aquaculture of Atlantic salmon and rainbow trout. The etiological agent of PD and SD is salmonid alphavirus (SAV), an unusual member of the Togaviridae (genus Alphavirus). SAV replicates at lower temperatures in fish. Outbreaks of SAV are associated with large economic losses of ∼17 to 50 million $/year. Current control strategies rely on vaccination with inactivated virus formulations that are cumbersome to obtain and have intrinsic safety risks. In this research we were able to obtain non-infectious virus-like particles (VLPs) of SAV via expression of recombinant baculoviruses encoding SAV capsid protein and two major immunodominant viral glycoproteins, E1 and E2 in Spodoptera frugiperda Sf9 insect cells. However, this was only achieved when a temperature shift from 27°C to lower temperatures was applied. At 27°C, precursor E2 (PE2) was misfolded and not processed by host furin into mature E2. Hence, E2 was detected neither on the surface of infected cells nor as VLPs in the culture fluid. However, when temperatures during protein expression were lowered, PE2 was processed into mature E2 in a temperature-dependent manner and VLPs were abundantly produced. So, temperature shift-down during synthesis is a prerequisite for correct SAV glycoprotein processing and recombinant VLP production

A Final Move to Your Own House

Objective – What makes a house a home? Many residents of care centres do not feel at home. 'This is not my own house', is what we heard in many of our conversations with residents. What makes the current houses into homes? Probably not the fact that everyone enters through the same door and has to walk through a corridor devoid of any daylight to get to their room. Residential care centre Cornelia by Allévo in the city of Zierikzee is developing a new building that is aimed towards providing the residents with their own home in the last phase of their lives. A sense of ownership and control. Background – A lot of people suffering from dementia are able to reside in their own house in the first phase. Once their situation gets worse or their informal carer becomes unavailable, people often end up in a residential care centre. Unfortunately, these people have to move from living independently to living in a small room in which they lose the feeling of having their own place or being able to manage their own lives. Research question – How can you turn a care home into a home for people with dementia? Every house has a front door, this emphasizes their independence and autonomy. In the new building at Cornelia care centre all residents have their own front door leading to the park. Every visitor can use this front door and residents are able to go in and out of their own volition. The research question examines whether this concept provides a sense of ownership and control to the residents, a feeling of having their own home. Methods – 93 residencies will be built in the care centre. The residencies have a front door to the outside and a back door to the inside, which is the central area of the building where the communal rooms are situated. With this chosen care concept, the residencies are not immediately situated next to a communal room, resulting into more possibilities to place the residents with like-minded people with the same interests, while also making it possible to switch in between. Visitors can park their cars near the front door or put their bicycles against the outside of their loved one's house. The covered area next to the front door makes for a space to sit outside. There is a small bench and a wooden frame offering enough protection from the sun. The residency is a studio in which the sleeping area is separated from the spacious living room. Results & Conclusion – Can having a personal front door provide a feeling of an own home for the residents as well as their visitors? Does it help to choose freely when to go outside? Is it nice to be able to choose between different communal rooms to spend your days? This new building at Cornelia care centre is a search for returning ownership to people with dementia.publishedVersio

A Final Move to Your Own House

Objective – What makes a house a home? Many residents of care centres do not feel at home. 'This is not my own house', is what we heard in many of our conversations with residents. What makes the current houses into homes? Probably not the fact that everyone enters through the same door and has to walk through a corridor devoid of any daylight to get to their room. Residential care centre Cornelia by Allévo in the city of Zierikzee is developing a new building that is aimed towards providing the residents with their own home in the last phase of their lives. A sense of ownership and control. Background – A lot of people suffering from dementia are able to reside in their own house in the first phase. Once their situation gets worse or their informal carer becomes unavailable, people often end up in a residential care centre. Unfortunately, these people have to move from living independently to living in a small room in which they lose the feeling of having their own place or being able to manage their own lives. Research question – How can you turn a care home into a home for people with dementia? Every house has a front door, this emphasizes their independence and autonomy. In the new building at Cornelia care centre all residents have their own front door leading to the park. Every visitor can use this front door and residents are able to go in and out of their own volition. The research question examines whether this concept provides a sense of ownership and control to the residents, a feeling of having their own home. Methods – 93 residencies will be built in the care centre. The residencies have a front door to the outside and a back door to the inside, which is the central area of the building where the communal rooms are situated. With this chosen care concept, the residencies are not immediately situated next to a communal room, resulting into more possibilities to place the residents with like-minded people with the same interests, while also making it possible to switch in between. Visitors can park their cars near the front door or put their bicycles against the outside of their loved one's house. The covered area next to the front door makes for a space to sit outside. There is a small bench and a wooden frame offering enough protection from the sun. The residency is a studio in which the sleeping area is separated from the spacious living room. Results & Conclusion – Can having a personal front door provide a feeling of an own home for the residents as well as their visitors? Does it help to choose freely when to go outside? Is it nice to be able to choose between different communal rooms to spend your days? This new building at Cornelia care centre is a search for returning ownership to people with dementia

Current and next-generation bleutongue vaccines : Requirements, strategies, and prospects for different field situations

Bluetongue virus (BTV) causes the hemorrhagic disease bluetongue (BT) in ruminants. The best way to control outbreaks is vaccination. Currently, conventionally modified-live and inactivated vaccines are commercially available, which have been successfully used to control BT, but nonetheless have their specific shortcomings. Therefore, there is a need for improved BT vaccines.The ideal BT vaccine is efficacious, safe, affordable, protective against multiple serotypes and enables the differentiation of infected from vaccinated animals. Different field situations require specific vaccine profiles. Single serotype outbreaks in former BT-free areas need rapid onset of protection against viremia of the respective serotype. In contrary, endemic multiple serotype situations require long-lasting protection against all circulating serotypes. The ideal BT vaccine for all field situations does not exist and balancing between vaccine properties is needed.Many new vaccines candidates, ranging from non-replicating subunits to replicating next-generation reverse genetics based vaccines, have been developed. Some have been tested extensively in large numbers of ruminants, whereas others were developed recently and have only been tested in vitro and in mice models. Most vaccine candidates are promising, but have their specific shortcomings and advantages. In this review, current and next-generation BT vaccines are discussed in the light of prerequisites for different field situations.<br/

Non-structural protein NS3/NS3a is required for propagation of bluetongue virus in Culicoides sonorensis

Background: Bluetongue virus (BTV) causes non-contagious haemorrhagic disease in ruminants and is transmitted by Culicoides spp. biting midges. BTV encodes four non-structural proteins of which NS3/NS3a is functional in virus release. NS3/NS3a is not essential for in vitro virus replication. However, deletion of NS3/NS3a leads to delayed virus release from mammalian cells and largely reduces virus release from insect cells. NS3/NS3a knockout BTV in sheep causes no viremia, but induces sterile immunity and is therefore proposed to be a Disabled Infectious Single Animal (DISA) vaccine candidate. In the absence of viremia, uptake of this vaccine strain by blood-feeding midges would be highly unlikely. Nevertheless, unintended replication of vaccine strains within vectors, and subsequent recombination or re-assortment resulting in virulent phenotypes and transmission is a safety concern of modified-live vaccines. Methods: The role of NS3/NS3a in replication and dissemination of BTV1, expressing VP2 of serotype 2 within colonized Culicoides sonorensis midges was investigated. Virus strains were generated using reverse genetics and their growth was examined in vitro. A laboratory colony of C. sonorensis, a known competent BTV vector, was fed or injected with BTV with or without expressing NS3/NS3a and replication in the midge was examined using RT PCR. Crossing of the midgut infection barrier was examined by separate testing of midge heads and bodies. Results: Although the parental NS3/NS3a expressing strain was not able to replicate and disseminate within C. sonorensis after oral feeding, this virus was able to replicate efficiently when the midgut infection barrier was bypassed by intrathoracic injection, whereas the NS3/NS3a knockout mutant was unable to replicate. This demonstrates that NS3/NS3a is required for viral replication within Culicoides. Conclusion: The lack of viremia and the inability to replicate within the vector, clearly demonstrate the inability of NS3/NS3a knockout DISA vaccine strains to be transmitted by midges.</p

Balance of RNA sequence requirement and NS3/NS3a expression of segment 10 of orbiviruses

Orbiviruses are insect-transmitted, non-enveloped viruses with a ten-segmented dsRNA genome of which the bluetongue virus (BTV) is the prototype. Viral non-structural protein NS3/NS3a is encoded by genome segment 10 (Seg-10), and is involved in different virus release mechanisms. This protein induces specific release via membrane disruptions and budding in both insect and mammalian cells, but also the cytopathogenic release that is only seen in mammalian cells. NS3/NS3a is not essential for virus replication in vitro with BTV Seg-10 containing RNA elements essential for virus replication, even if protein is not expressed. Recently, new BTV serotypes with distinct NS3/NS3a sequence and cell tropism have been identified. Multiple studies have hinted at the importance of Seg-10 in orbivirus replication, but the exact prerequisites are still unknown. Here, more insight is obtained with regard to the needs for orbivirus Seg-10 and the balance between protein expression and RNA elements. Multiple silent mutations in the BTV NS3a ORF destabilized Seg-10, resulting in deletions and sequences originating from other viral segments being inserted, indicating strong selection at the level of RNA during replication in mammalian cells in vitro. The NS3a ORFs of other orbiviruses were successfully exchanged in BTV1 Seg-10, resulting in viable chimeric viruses. NS3/NS3a proteins in these chimeric viruses were generally functional in mammalian cells, but not in insect cells. NS3/NS3a of the novel BTV serotypes 25 and 26 affected virus release from Culicoides cells, which might be one of the reasons for their distinct cell tropism

Requirements and comparative analysis of reverse genetics for bluetongue virus (BTV) and African horse sickness virus (AHSV)

<p>Background: Bluetongue virus (BTV) and African horse sickness virus (AHSV) are distinct arthropod borne virus species in the genus Orbivirus (Reoviridae family), causing the notifiable diseases Bluetongue and African horse sickness of ruminants and equids, respectively. Reverse genetics systems for these orbiviruses with their ten-segmented genome of double stranded RNA have been developed. Initially, two subsequent transfections of in vitro synthesized capped run-off RNA transcripts resulted in the recovery of BTV. Reverse genetics has been improved by transfection of expression plasmids followed by transfection of ten RNA transcripts. Recovery of AHSV was further improved by use of expression plasmids containing optimized open reading frames. Results: Plasmids containing full length cDNA of the 10 genome segments for T7 promoter-driven production of full length run-off RNA transcripts and expression plasmids with optimized open reading frames (ORFs) were used. BTV and AHSV were rescued using reverse genetics. The requirement of each expression plasmid and capping of RNA transcripts for reverse genetics were studied and compared for BTV and AHSV. BTV was recovered by transfection of VP1 and NS2 expression plasmids followed by transfection of a set of ten capped RNAs. VP3 expression plasmid was also required if uncapped RNAs were transfected. Recovery of AHSV required transfection of VP1, VP3 and NS2 expression plasmids followed by transfection of capped RNA transcripts. Plasmid-driven expression of VP4, 6 and 7 was also needed when uncapped RNA transcripts were used. Irrespective of capping of RNA transcripts, NS1 expression plasmid was not needed for recovery, although NS1 protein is essential for virus propagation. Improvement of reverse genetics for AHSV was clearly demonstrated by rescue of several mutants and reassortants that were not rescued with previous methods. Conclusions: A limited number of expression plasmids is required for rescue of BTV or AHSV using reverse genetics, making the system much more versatile and generally applicable. Optimization of reverse genetics enlarge the possibilities to rescue virus mutants and reassortants, and will greatly benefit the control of these important diseases of livestock and companion animals.</p

Bluetongue Disabled Infectious Single Animal (DISA) vaccine: Studies on the optimal route and dose in sheep

Bluetongue (BT) is a disease of ruminants caused by bluetongue virus (BTV) transmitted by biting midges of the Culicoides genus. Outbreaks have been controlled successfully by vaccination, however, currently available BT vaccines have several shortcomings. Recently, we have developed BT Disabled Infectious Single Animal (DISA) vaccines based on live-attenuated BTV without expression of dispensable non-structural NS3/NS3a protein. DISA vaccines are non-pathogenic replicating vaccines, do not cause viremia, enable DIVA and are highly protective. NS3/NS3a protein is involved in virus release, cytopathogenic effect and suppression of Interferon-I induction, suggesting that the vaccination route can be of importance. A standardized dose of DISA vaccine for serotype 8 has successfully been tested by subcutaneous vaccination. We show that 10 and 100 times dilutions of this previously tested dose did not reduce the VP7 humoral response. Further, the vaccination route of DISA vaccine strongly determined the induction of VP7 directed antibodies (Abs). Intravenous vaccination induced high and prolonged humoral response but is not practical in field situations. VP7 seroconversion was stronger by intramuscular vaccination than by subcutaneous vaccination. For both vaccination routes and for two different DISA vaccine backbones, IgM Abs were rapidly induced but declined after 14 days post vaccination (dpv), whereas the IgG response was slower. Interestingly, intramuscular vaccination resulted in an initial peak followed by a decline up to 21 dpv and then increased again. This second increase is a steady and continuous increase of IgG Abs. These results indicate that intramuscular vaccination is the optimal route. The protective dose of DISA vaccine has not been determined yet, but it is expected to be significantly lower than of currently used BT vaccines. Therefore, in addition to the advantages of improved safety and DIVA compatibility, the novel DISA vaccines will be cost–competitive to commercially available live attenuated and inactivated vaccines for Bluetongue