Proteomic Characterization of Bovine Herpesvirus 4 Extracellular Virions

Gammaherpesviruses are important pathogens in human and animal populations. During early events of infection, these viruses manipulate preexisting host cell signaling pathways to allow successful infection. The different proteins that compose viral particles are therefore likely to have critical functions not only in viral structures and in entry into target cell but also in evasion of the host''s antiviral response. In this study, we analyzed the protein composition of bovine herpesvirus 4 (BoHV-4), a close relative of the human Kaposi''s sarcoma-associated herpesvirus. Using mass spectrometry-based approaches, we identified 37 viral proteins associated with extracellular virions, among which 24 were resistant to proteinase K treatment of intact virions. Analysis of proteins associated with purified capsid-tegument preparations allowed us to define protein localization. In parallel, in order to identify some previously undefined open reading frames, we mapped peptides detected in whole virion lysates onto the six frames of the BoHV-4 genome to generate a proteogenomic map of BoHV-4 virions. Furthermore, we detected important glycosylation of three envelope proteins: gB, gH, and gp180. Finally, we identified 38 host proteins associated with BoHV-4 virions; 15 of these proteins were resistant to proteinase K treatment of intact virions. Many of these have important functions in different cellular pathways involved in virus infection. This study extends our knowledge of gammaherpesvirus virions composition and provides new insights for understanding the life cycle of these viruses.     

Higher genetic diversity in recolonized areas than in refugia of Alnus glutinosa triggered by continent‐wide lineage admixture

Genetic admixture is supposed to be an important trigger of species expansions because it can create the potential for selection of genotypes suitable for new climatic conditions. Up until now, however, no continent‐wide population genetic study has performed a detailed reconstruction of admixture events during natural species expansions. To fill this gap, we analysed the postglacial history of Alnus glutinosa, a keystone species of European swamp habitats, across its entire distribution range using two molecular markers, cpDNA and nuclear microsatellites. CpDNA revealed multiple southern refugia located in the Iberian, Apennine, Balkan and Anatolian Peninsulas, Corsica and North Africa. Analysis of microsatellites variation revealed three main directions of postglacial expansion: (i) from the northern part of the Iberian Peninsula to Western and Central Europe and subsequently to the British Isles, (ii) from the Apennine Peninsula to the Alps and (iii) from the eastern part of the Balkan Peninsula to the Carpathians followed by expansion towards the Northern European plains. This challenges the classical paradigm that most European populations originated from refugial areas in the Carpathians. It has been shown that colonizing lineages have met several times and formed secondary contact zones with unexpectedly high population genetic diversity in Central Europe and Scandinavia. On the contrary, limited genetic admixture in southern refugial areas of A. glutinosa renders rear‐edge populations in the Mediterranean region more vulnerable to extinction due to climate change.     

Effect of a web-based audit and feedback intervention with outreach visits on the clinical performance of multidisciplinary teams: a cluster-randomized trial in cardiac rehabilitation

Background

The objective of this study was to assess the effect of a web-based audit and feedback (A&F) intervention with outreach visits to support decision-making by multidisciplinary teams.

Methods

We performed a multicentre cluster-randomized trial within the field of comprehensive cardiac rehabilitation (CR) in the Netherlands. Our participants were multidisciplinary teams in Dutch CR centres who were enrolled in the study between July 2012 and December 2013 and received the intervention for at least 1 year. The intervention included web-based A&F with feedback on clinical performance, facilities for goal setting and action planning, and educational outreach visits. Teams were randomized either to receive feedback that was limited to psychosocial rehabilitation (study group A) or to physical rehabilitation (study group B). The main outcome measure was the difference in performance between study groups in 11 care processes and six patient outcomes, measured at patient level. Secondary outcomes included effects on guideline concordance for the four main CR therapies.

Results

Data from 18 centres (14,847 patients) were analysed, of which 12 centres (9353 patients) were assigned to group A and six (5494 patients) to group B. During the intervention, a total of 233 quality improvement goals was identified by participating teams, of which 49 (21%) were achieved during the study period. Except for a modest improvement in data completeness (4.5% improvement per year; 95% CI 0.65 to 8.36), we found no effect of our intervention on any of our primary or secondary outcome measures.

Conclusions

Within a multidisciplinary setting, our web-based A&F intervention engaged teams to define local performance improvement goals but failed to support them in actually completing the improvement actions that were needed to achieve those goals. Future research should focus on improving the actionability of feedback on clinical performance and on addressing the socio-technical perspective of the implementation process.

Trial registration

NTR3251

     

Comprehensive identification of LMW-GS genes and their protein products in a common wheat variety

Although it is well known that low-molecular-weight glutenin subunits (LMW-GS) from wheat affect bread and noodle processing quality, the function of specific LMW-GS proteins remains unclear. It is important to find the genes that correspond to individual LMW-GS proteins in order to understand the functions of specific proteins. The objective of this study was to link LMW-GS genes and haplotypes characterized using well known Glu-A3, Glu-B3, and Glu-D3 gene-specific primers to their protein products in a single wheat variety. A total of 36 LMW-GS genes and pseudogenes were amplified from the Korean cultivar Keumkang. These include 11 Glu-3 gene haplotypes, two from the Glu-A3 locus, two from the Glu-B3 locus, and seven from the Glu-D3 locus. To establish relationships between gene haplotypes and their protein products, a glutenin protein fraction was separated by two-dimensional gel electrophoresis (2-DGE) and 17 protein spots were analyzed by N-terminal amino acid sequencing and tandem mass spectrometry (MS/MS). LMW-GS proteins were identified that corresponded to all Glu-3 gene haplotypes except the pseudogenes. This is the first report of the comprehensive characterization of LMW-GS genes and their corresponding proteins in a single wheat cultivar. Our approach will be useful to understand the contributions of individual LMW-GS to the end-use quality of flour.     

Microsatellite DNA evidence for genetic drift and philopatry in Svalbard reindeer

Mainland populations of Arctic reindeer and caribou Rangifer tarandus often undergo extensive movements, whereas populations on islands tend to be isolated and sedentary. To characterize the genetic consequences of this difference, levels of genetic diversity and subdivision of Svalbard reindeer (R. t. platyrhynchus) from two adjacent areas on Nordenskjiöldland, Spitsbergen were estimated using data from up to 14 microsatellites. The mean number of alleles per locus in Svalbard reindeer was 2.4 and mean expected heterozygosity per locus was 0.36. The latter value was significantly lower than in Canadian caribou and Norwegian reindeer but higher than in some other cervid species. Large samples of females (n = 743) and small samples of males (n = 38) from two sites ≈ 45 km apart showed genetic subdivision, which could be due to local population fluctuations or limited gene flow. To our knowledge, this is the first study to report significant differentiation at microsatellite loci in Rangifer at such short geographical distances. Neither population showed genetic evidence for recent population bottlenecks when loci unbiased with respect to heterozygosity were analysed. In contrast, false signals of a recent bottleneck were detected when loci upwardly biased with respect to heterozygosity were analysed. Thus, Svalbard reindeer appeared to conform to the paradigm of island populations made genetically depauperate by genetic drift.     

Phylogenetic classification of transporters and other membrane proteins from Saccharomyces cerevisiae

On the basis of functional and phylogenetic criteria, we have identified a total of 229 subfamilies and 111 singletons predicted to carry out transport or other membrane functions in Saccharomyces cerevisiae. We have extended the Transporter Classification (TC) and created a Membrane Classification (MC) for non-transporter membrane proteins. Using the preliminary phylogenetic digits X, Y, Z (for new families, subfamilies, and clusters, respectively), we allocated a five-digit number to 850 proteins predicted to contain more than two transmembrane domains. Compared with a previous TC of the yeast genome, we classified an additional set of 538 membrane proteins (transporters and non-transporters) and identified 111 novel phylogenetic subfamilies. Electronic Publication     

Effects of a neurotoxin isolated from the sea anemone, Anemonia sulcata, at frog neuromuscular junction (author's transl)]

ATX II is a toxin extracted from tentacles of Anemonia sulcata. It was known that this protein displays neurotoxic effects on frog isolated neuromuscular preparation (Fig. 1, 2) and that muscular contractures observed with ATX II are blocked by d-tubocurarine (Fig. 3) or on a 40-days-denervated gastrocnemius (Fig. 4). Part of these experiments has already appeared. 1. These effects of ATX II depend on calcium concentration in the bathing medium, as is the case for transmitter release. The same results were observed when we substituted strontium to calcium. 2. On an intact sciatic sartorius preparation, ATX II does not act on the amplitude of the miniature endplate potentials (mepps, Fig. 6). The muscular action potential is not modified by this toxin. 3. ATX II increases the frequency of the mepps (Fig. 5). The evoked transmitter release (quantal content) after ATX II is also largely increased (Fig. 7). 4. In conclusion, it is suggested that ATX II acts indirectly on the muscle through an increase in acetylcholine release from the motor nerve terminals.