Gene Content and Diversity of the Loci Encoding Biosynthesis of Capsular Polysaccharides of the 15 Serovar Reference Strains of Haemophilus parasuis

Haemophilus parasuis is the causative agent of Glässer''s disease, a systemic disease of pigs, and is also associated with pneumonia. H. parasuis can be classified into 15 different serovars. Here we report, from the 15 serotyping reference strains, the DNA sequences of the loci containing genes for the biosynthesis of the group 1 capsular polysaccharides, which are potential virulence factors of this bacterium. We contend that these loci contain genes for polysaccharide capsule structures, and not a lipopolysaccharide O antigen, supported by the fact that they contain genes such as wza, wzb, and wzc, which are associated with the export of polysaccharide capsules in the current capsule classification system. A conserved region at the 3′ end of the locus, containing the wza, ptp, wzs, and iscR genes, is consistent with the characteristic export region 1 of the model group 1 capsule locus. A potential serovar-specific region (region 2) has been found by comparing the predicted coding sequences (CDSs) in all 15 loci for synteny and homology. The region is unique to each reference strain with the exception of those in serovars 5 and 12, which are identical in terms of gene content. The identification and characterization of this locus among the 15 serovars is the first step in understanding the genetic, molecular, and structural bases of serovar specificity in this poorly studied but important pathogen and opens up the possibility of developing an improved molecular serotyping system, which would greatly assist diagnosis and control of Glässer''s disease.     

A sphingosine 1-phosphate receptor 2 selective allosteric agonist

Molecular probe tool compounds for the Sphingosine 1-phosphate receptor 2 (S1PR2) are important for investigating the multiple biological processes in which the S1PR2 receptor has been implicated. Amongst these are NF-κB-mediated tumor cell survival and fibroblast chemotaxis to fibronectin. Here we report our efforts to identify selective chemical probes for S1PR2 and their characterization. We employed high throughput screening to identify two compounds which activate the S1PR2 receptor. SAR optimization led to compounds with high nanomolar potency. These compounds, XAX-162 and CYM-5520, are highly selective and do not activate other S1P receptors. Binding of CYM-5520 is not competitive with the antagonist JTE-013. Mutation of receptor residues responsible for binding to the zwitterionic headgroup of sphingosine 1-phosphate (S1P) abolishes S1P activation of the receptor, but not activation by CYM-5520. Competitive binding experiments with radiolabeled S1P demonstrate that CYM-5520 is an allosteric agonist and does not displace the native ligand. Computational modeling suggests that CYM-5520 binds lower in the orthosteric binding pocket, and that co-binding with S1P is energetically well tolerated. In summary, we have identified an allosteric S1PR2 selective agonist compound.     

Do foods imported into the UK have a greater environmental impact than the same foods produced within the UK?

Purpose

This study of seven foods assessed whether there are modes or locations of production that require significantly fewer inputs, and hence cause less pollution, than others. For example, would increasing imports of field-grown tomatoes from the Mediterranean reduce greenhouse gas (GHG) emissions by reducing the need for production in heated greenhouses in the UK, taking account of the additional transport emissions? Is meat production in the UK less polluting than the import of red meat from the southern hemisphere?

Methods

We carried out a life-cycle inventory for each commodity, which quantified flows relating to life-cycle assessment (LCA) impact categories: primary energy use, acidification, eutrophication, abiotic resource use, pesticide use, land occupation and ozone depletion. The system boundary included all production inputs up to arrival at the retail distribution centre (RDC). The allocation of production burdens for meat products was on the basis of economic value. We evaluated indicator foods from which it is possible to draw parallels for foods whose production follows a similar chain: tomatoes (greenhouse crops), strawberries (field-grown soft fruit), apples (stored for year-round supply or imported during spring and summer), potatoes (early season imports or long-stored UK produce), poultry and beef (imported from countries such as Brazil) and lamb (imported to balance domestic spring–autumn supply).

Results and discussion

Total pre-farm gate global warming potential (GWP) of potatoes and beef were less for UK production than for production in the alternative country. Up to delivery to the RDC, total GWP were less for UK potatoes, beef and apples than for production elsewhere. Production of tomatoes and strawberries in Spain, poultry in Brazil and lamb in New Zealand produced less GWP than in the UK despite emissions that took place during transport. For foods produced with only small burdens of GWP, such as apples and strawberries, the burden from transport may be a large proportion of the total. For foods with inherently large GWP per tonne, such as meat products, burdens arising from transport may only be a small proportion of the total.

Conclusions

When considering the GWP of food production, imports from countries where productivity is greater and/or where refrigerated storage requirement is less will lead to less total GWP than axiomatic preference for local produce. However, prioritising GWP may lead to increases in other environmental burdens, in particular leading to both greater demands on and decreasing quality of water resources.     

Risk Factors and Screening for Trypanosoma cruzi Infection of Dutch Blood Donors

Background

Blood donors unaware of Trypanosoma cruzi infection may donate infectious blood. Risk factors and the presence of T. cruzi antibodies in at-risk Dutch blood donors were studied to assess whether specific blood safety measures are warranted in the Netherlands.

Methodology

Birth in a country endemic for Chagas disease (CEC), having a mother born in a CEC, or having resided for at least six continuous months in a CEC were considered risk factors for T. cruzi infection. From March through September 2013, risk factor questions were asked to all donors who volunteered to donate blood or blood components. Serum samples were collected from donors reporting one or more risk factors, and screened for IgG antibodies to T. cruzi by EIA.

Results

Risk factors for T. cruzi infection were reported by 1,426 of 227,278 donors (0.6%). Testing 1,333 at-risk donors, none (0.0%; 95%, CI 0.0–0.4%) was seroreactive for IgG antibodies to T. cruzi. A total of 472 donors were born in a CEC; 553 donors reported their mother being born in a CEC; and 1,121 donors reported a long-term stay in a CEC. The vast majority of reported risk factors were related to Suriname and Brazil. Overall, the participants resided for 7,694 years in CECs, which equals 2.8 million overnight stays. Of those, 1.9 million nights were spent in Suriname.

Conclusions/Significance

Asymptomatic T. cruzi infection appears to be extremely rare among Dutch blood donors. Blood safety interventions to mitigate the risk of T. cruzi transmission by transfusion would be highly cost-ineffective in the Netherlands, and are thus not required.     

A targeted capture approach to generating reference sequence databases for chloroplast gene regions

Metabarcoding has improved the way we understand plants within our environment, from their ecology and conservation to invasive species management. The notion of identifying plant taxa within environmental samples relies on the ability to match unknown sequences to known reference libraries. Without comprehensive reference databases, species can go undetected or be incorrectly assigned, leading to false‐positive and false‐negative detections. To improve our ability to generate reference sequence databases, we developed a targeted capture approach using the OZBaits_CP V1.0 set, designed to capture chloroplast gene regions across the entirety of flowering plant diversity. We focused on generating a reference database for coastal temperate plant species given the lack of reference sequences for these taxa. Our approach was successful across all specimens with a target gene recovery rate of 92%, which was achieved in a single assay (i.e., samples were pooled), thus making this approach much faster and more efficient than standard barcoding. Further testing of this database highlighted 80% of all samples could be discriminated to family level across all gene regions with some genes achieving greater resolution than others—which was also dependent on the taxon of interest. Thus, we demonstrate the importance of generating reference sequences across multiple chloroplast gene regions as no single loci are sufficient to discriminate across all plant groups. The targeted capture approach outlined in this study provides a way forward to achieve this.     

Identification of sodC encoding periplasmic [Cu,Zn]-superoxide dismutase in Salmonella

Abstract sodC , encoding [Cu,Zn]-cofactored Superoxide dismutase, once thought to be virtually confined to eukaryotes, has now been described in many Gram-negative pathogens that have their primary niche of colonization in the upper respiratory tract. Their role in host-parasite interactive biology is unknown. We here show that members of the major human and animal enteric pathogenic species Salmonella harbour a version of sodC most closely resembling that found in Brucella abortus . The enzyme it encodes is a novel candidate determinant of virulence in Salmonella , an intracellular pathogen potentially exposed to toxic oxygen free radicals within its intracellular niche.     

GIT1 activates p21-activated kinase through a mechanism independent of p21 binding

p21-activated kinases (PAKs) associate with a guanine nucleotide exchange factor, Pak-interacting exchange factor (PIX), which in turn binds the paxillin-associated adaptor GIT1 that targets the complex to focal adhesions. Here, a detailed structure-function analysis of GIT1 reveals how this multidomain adaptor also participates in activation of PAK. Kinase activation does not occur via Cdc42 or Rac1 GTPase binding to PAK. The ability of GIT1 to stimulate alphaPAK autophosphorylation requires the participation of the GIT N-terminal Arf-GAP domain but not Arf-GAP activity and involves phosphorylation of PAK at residues common to Cdc42-mediated activation. Thus, the activation of PAK at adhesion complexes involves a complex interplay between the kinase, Rho GTPases and protein partners that provide localization cues.     

A mammalian artificial chromosome engineering system (ACE System) applicable to biopharmaceutical protein production, transgenesis and gene-based cell therapy

Mammalian artificial chromosomes (MACs) provide a means to introduce large payloads of genetic information into the cell in an autonomously replicating, non-integrating format. Unique among MACs, the mammalian satellite DNA-based Artificial Chromosome Expression (ACE) can be reproducibly generated de novo in cell lines of different species and readily purified from the host cells' chromosomes. Purified mammalian ACEs can then be re-introduced into a variety of recipient cell lines where they have been stably maintained for extended periods in the absence of selective pressure. In order to extend the utility of ACEs, we have established the ACE System, a versatile and flexible platform for the reliable engineering of ACEs. The ACE System includes a Platform ACE, containing >50 recombination acceptor sites, that can carry single or multiple copies of genes of interest using specially designed targeting vectors (ATV) and a site-specific integrase (ACE Integrase). Using this approach, specific loading of one or two gene targets has been achieved in LMTK⁻ and CHO cells. The use of the ACE System for biological engineering of eukaryotic cells, including mammalian cells, with applications in biopharmaceutical production, transgenesis and gene-based cell therapy is discussed.     

Linking performance development to new roles and expectations of the employee

After downsizing, organizations need to look at new methods with which to develop employees. When monetary incentives have been stripped away and greater productivity is expected of workers, the challenge is to get the work of the organization accomplished while maintaining human dignity and meeting everyone's needs. This article ties in these issues with a nontraditional view of the performance management process that can help keep employees motivated to achieve new organizational goals.     

The Influence of Spike Rate and Stimulus Duration on Noradrenergic Neurons

We model spiking neurons in locus coeruleus (LC), a brain nucleus involved in modulating cognitive performance, and compare with recent experimental data. Extracellular recordings from LC of monkeys performing target detection and selective attention tasks show varying responses dependent on stimuli and performance accuracy. From membrane voltage and ion channel equations, we derive a phase oscillator model for LC neurons. Average spiking probabilities of a pool of cells over many trials are then computed via a probability density formulation. These show that: (1) Post-stimulus response is elevated in populations with lower spike rates; (2) Responses decay exponentially due to noise and variable pre-stimulus spike rates; and (3) Shorter stimuli preferentially cause depressed post-activation spiking. These results allow us to propose mechanisms for the different LC responses observed across behavioral and task conditions, and to make explicit the role of baseline firing rates and the duration of task-related inputs in determining LC response.