BSA Hydrogel Beads Functionalized with a Specific Aptamer Library for Capturing Pseudomonas aeruginosa in Serum and Blood

Systemic blood stream infections are a major threat to human health and are dramatically increasing worldwide. Pseudomonas aeruginosa is a WHO-alerted multi-resistant pathogen of extreme importance as a cause of sepsis. Septicemia patients have significantly increased survival chances if sepsis is diagnosed in the early stages. Affinity materials can not only represent attractive tools for specific diagnostics of pathogens in the blood but can prospectively also serve as the technical foundation of therapeutic filtration devices. Based on the recently developed aptamers directed against P. aeruginosa, we here present aptamer-functionalized beads for specific binding of this pathogen in blood samples. These aptamer capture beads (ACBs) are manufactured by crosslinking bovine serum albumin (BSA) in an emulsion and subsequent functionalization with the amino-modified aptamers on the bead surface using the thiol- and amino-reactive bispecific crosslinker PEG(4)-SPDP. Specific and quantitative binding of P. aeruginosa as the dedicated target of the ACBs was demonstrated in serum and blood. These initial but promising results may open new routes for the development of ACBs as a platform technology for fast and reliable diagnosis of bloodstream infections and, in the long term, blood filtration techniques in the fight against sepsis

Remodeling of the Streptococcus agalactiae Transcriptome in Response to Growth Temperature

BACKGROUND: To act as a commensal bacterium and a pathogen in humans and animals, Streptococcus agalactiae (group B streptococcus, GBS) must be able to monitor and adapt to different environmental conditions. Temperature variation is a one of the most commonly encountered variables. METHODOLOGY/PRINCIPAL FINDINGS: To understand the extent to which GBS modify gene expression in response to temperatures encountered in the various hosts, we conducted a whole genome transcriptome analysis of organisms grown at 30 degrees C and 40 degrees C. We identified extensive transcriptome remodeling at various stages of growth, especially in the stationary phase (significant transcript changes occurred for 25% of the genes). A large proportion of genes involved in metabolism was up-regulated at 30 degrees C in stationary phase. Conversely, genes up-regulated at 40 degrees C relative to 30 degrees C include those encoding virulence factors such as hemolysins and extracellular secreted proteins with LPXTG motifs. Over-expression of hemolysins was linked to larger zones of hemolysis and enhanced hemolytic activity at 40 degrees C. A key theme identified by our study was that genes involved in purine metabolism and iron acquisition were significantly up-regulated at 40 degrees C. CONCLUSION/SIGNIFICANCE: Growth of GBS in vitro at different temperatures resulted in extensive remodeling of the transcriptome, including genes encoding proven and putative virulence genes. The data provide extensive new leads for molecular pathogenesis research

Extensive Adaptive Changes Occur in the Transcriptome of Streptococcus agalactiae (Group B Streptococcus) in Response to Incubation with Human Blood

To enhance understanding of how Streptococcus agalactiae (group B streptococcus, GBS) adapts during invasive infection, we performed a whole-genome transcriptome analysis after incubation with whole human blood. Global changes occurred in the GBS transcriptome rapidly in response to blood contact following shift from growth in a rich laboratory medium. Most (83%) of the significantly altered transcripts were down-regulated after 30 minutes of incubation in blood, and all functional categories of genes were abundantly represented. We observed complex dynamic changes in the expression of transcriptional regulators and stress response genes that allow GBS to rapidly adapt to blood. The transcripts of relatively few proven virulence genes were up-regulated during the first 90 minutes. However, a key discovery was that genes encoding proteins involved in interaction with the host coagulation/fibrinolysis system and bacterial-host interactions were rapidly up-regulated. Extensive transcript changes also occurred for genes involved in carbohydrate metabolism, including multi-functional proteins and regulators putatively involved in pathogenesis. Finally, we discovered that an incubation temperature closer to that occurring in patients with severe infection and high fever (40°C) induced additional differences in the GBS transcriptome relative to normal body temperature (37°C). Taken together, the data provide extensive new information about transcriptional adaptation of GBS exposed to human blood, a crucial step during GBS pathogenesis in invasive diseases, and identify many new leads for molecular pathogenesis research

Dual Role for Pilus in Adherence to Epithelial Cells and Biofilm Formation in Streptococcus agalactiae

Streptococcus agalactiae is a common human commensal and a major life-threatening pathogen in neonates. Adherence to host epithelial cells is the first critical step of the infectious process. Pili have been observed on the surface of several gram-positive bacteria including S. agalactiae. We previously characterized the pilus-encoding operon gbs1479-1474 in strain NEM316. This pilus is composed of three structural subunit proteins: Gbs1478 (PilA), Gbs1477 (PilB), and Gbs1474 (PilC), and its assembly involves two class C sortases (SrtC3 and SrtC4). PilB, the bona fide pilin, is the major component; PilA, the pilus associated adhesin, and PilC, are both accessory proteins incorporated into the pilus backbone. We first addressed the role of the housekeeping sortase A in pilus biogenesis and showed that it is essential for the covalent anchoring of the pilus fiber to the peptidoglycan. We next aimed at understanding the role of the pilus fiber in bacterial adherence and at resolving the paradox of an adhesive but dispensable pilus. Combining immunoblotting and electron microscopy analyses, we showed that the PilB fiber is essential for efficient PilA display on the surface of the capsulated strain NEM316. We then demonstrated that pilus integrity becomes critical for adherence to respiratory epithelial cells under flow-conditions mimicking an in vivo situation and revealing the limitations of the commonly used static adherence model. Interestingly, PilA exhibits a von Willebrand adhesion domain (VWA) found in many extracellular eucaryotic proteins. We show here that the VWA domain of PilA is essential for its adhesive function, demonstrating for the first time the functionality of a prokaryotic VWA homolog. Furthermore, the auto aggregative phenotype of NEM316 observed in standing liquid culture was strongly reduced in all three individual pilus mutants. S. agalactiae strain NEM316 was able to form biofilm in microtiter plate and, strikingly, the PilA and PilB mutants were strongly impaired in biofilm formation. Surprisingly, the VWA domain involved in adherence to epithelial cells was not required for biofilm formation

Effects of vegetation structure and human impact on understory honey plant richness: implications for pollinator visitation

This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made.Background Though the biomass of floral vegetation in understory plant communities in a forested ecosystem only accounts for less than 1% of the total biomass of a forest, they contain most of the floral resources of a forest. The diversity of understory honey plants determines visitation rate of pollinators such as honey bee (Apis mellifera) as they provide rich food resources. Since the flower visitation and foraging activity of pollinators lead to the provision of pollination service, it also means the enhancement of plant-pollinator relationship. Therefore, an appropriate management scheme for understory vegetation is essential in order to conserve pollinator population that is decreasing due to habitat destruction and disease infection. This research examined the diversity of understory honey plant and studied how it is related to environmental variables such as (1) canopy density, (2) horizontal heterogeneity of canopy surface height, (3) slope gradient, and (4) distance from roads. Vegetation survey data of 39 plots of mixed forests in Chuncheon, Korea, were used, and possible management practices for understory vegetation were suggested. Results This study found that 113 species among 141 species of honey plant of the forests were classified as understory vegetation. Also, the understory honey plant diversity is significantly positively correlated with distance from the nearest road and horizontal heterogeneity of canopy surface height and negatively correlated with canopy density. Conclusions The diversity of understory honey plant vegetation is correlated to vegetation structure and human impact. In order to enhance the diversity of understory honey plant, management of density and height of canopy is necessary. This study suggests that improved diversity of canopy cover through thinning of overstory vegetation can increase the diversity of understory honey plant species

On the Determinants of Social Capital in Greece Compared to Countries of the European Union

Social capital refers to the stock of social relations, based on norms and networks of cooperation and trust that spill over to the market and state to enhance collective action between actors and achieve improved social efficiency and economic growth. The aim of the present paper is to discuss the implications of contemporary literature and empirical findings on social capital for the growth prospects of Greece, compared to the member-states of the European Union. In order to examine the potential of social capital to enhance growth, we must look into the factors that determine the nature and context of trust, norms and networks that have emerged in our multinational, multiethnic and multicultural Europe.The contribution of this paper is to offer insight on the determinants of social capital in Greece, compared to the European Union (EU - former 15 member-states). For this purpose, we regress an index of individual group membership, derived from the European Community Household Panel (ECHP), on a set of individual as well as aggregate factors of social capital. Regression results provide evidence of the impact of both individual and institutional characteristics on group membership. Differences on the extent of group membership between countries might be indicative of the historical and cultural differences that have affected the evolution of social capital across Europe. Particularly in Greece, the relatively low level of group membership compared to the other EU countries might provide further evidence of its low levels of civicness. Historically, its weak civil society has been a result of a prior civic tradition of clientelism under arbitrary rule, the interference of special-interest groups and the lack of credibility and impartiality from the part of the state. And these factors might be responsible for the slow pace in reform and growth observed compared to the rest of the EU. Nevertheless, the findings on the determinants of social capital may direct us to possible means of rebuilding patterns of participatory and cooperative behavior, especially in countries with low levels of trust and civicness, such as Greece