α‐Hemolysin enhances Staphylococcus aureus internalization and survival within mast cells by modulating the expression of β1 integrin

Mast cells (MCs) are important sentinels of the host defence against invading pathogens. We previously reported that Staphylococcus aureus evaded the extracellular antimicrobial activities of MCs by promoting its internalization within these cells via β1 integrins. Here, we investigated the molecular mechanisms governing this process. We found that S. aureus responded to the antimicrobial mediators released by MCs by up‐regulating the expression of α‐hemolysin (Hla), fibronectin‐binding protein A and several regulatory systems. We also found that S. aureus induced the up‐regulation of β1 integrin expression on MCs and that this effect was mediated by Hla‐ADAM10 (a disintegrin and metalloproteinase 10) interaction. Thus, deletion of Hla or inhibition of Hla‐ADAM10 interaction significantly impaired S. aureus internalization within MCs. Furthermore, purified Hla but not the inactive Hla_H35L induced up‐regulation of β1 integrin expression in MCs in a dose‐dependent manner. Our data support a model in which S. aureus counter‐reacts the extracellular microbicidal mechanisms of MCs by increasing expression of fibronectin‐binding proteins and by inducing Hla‐ADAM10‐mediated up‐regulation of β1 integrin in MCs. The up‐regulation of bacterial fibronectin‐binding proteins, concomitantly with the increased expression of its receptor β1 integrin on the MCs, resulted in enhanced S. aureus internalization through the binding of fibronectin‐binding proteins to integrin β1 via fibronectin.     

Rice endosperm is cost‐effective for the production of recombinant griffithsin with potent activity against HIV

Protein microbicides containing neutralizing antibodies and antiviral lectins may help to reduce the rate of infection with human immunodeficiency virus (HIV) if it is possible to manufacture the components in large quantities at a cost affordable in HIV‐endemic regions such as sub‐Saharan Africa. We expressed the antiviral lectin griffithsin (GRFT), which shows potent neutralizing activity against HIV, in the endosperm of transgenic rice plants (Oryza sativa), to determine whether rice can be used to produce inexpensive GRFT as a microbicide ingredient. The yield of ^OSGRFT in the best‐performing plants was 223 μg/g dry seed weight. We also established a one‐step purification protocol, achieving a recovery of 74% and a purity of 80%, which potentially could be developed into a larger‐scale process to facilitate inexpensive downstream processing. ^OSGRFT bound to HIV glycans with similar efficiency to GRFT produced in Escherichia coli. Whole‐cell assays using purified ^OSGRFT and infectivity assays using crude extracts of transgenic rice endosperm confirmed that both crude and pure ^OSGRFT showed potent activity against HIV and the crude extracts were not toxic towards human cell lines, suggesting they could be administered as a microbicide with only minimal processing. A freedom‐to‐operate analysis confirmed that GRFT produced in rice is suitable for commercial development, and an economic evaluation suggested that 1.8 kg/ha of pure GRFT could be produced from rice seeds. Our data therefore indicate that rice could be developed as an inexpensive production platform for GRFT as a microbicide component.     

Structure‐based classification of FAD binding sites: A comparative study of structural alignment tools

A total of six different structural alignment tools (TM‐Align, TriangleMatch, CLICK, ProBis, SiteEngine and GA‐SI) were assessed for their ability to perform two particular tasks: (i) discriminating FAD (flavin adenine dinucleotide) from non‐FAD binding sites, and (ii) performing an all‐to‐all comparison on a set of 883 FAD binding sites for the purpose of classifying them. For the first task, the consistency of each alignment method was evaluated, showing that every method is able to distinguish FAD and non‐FAD binding sites with a high Matthews correlation coefficient. Additionally, GA‐SI was found to provide alignments different from those of the other approaches. The results obtained for the second task revealed more significant differences among alignment methods, as reflected in the poor correlation of their results and highlighted clearly by the independent evaluation of the structural superimpositions generated by each method. The classification itself was performed using the combined results of all methods, using the best result found for each comparison of binding sites. A number of different clustering methods (Single‐linkage, UPGMA, Complete‐linkage, SPICKER and k‐Means clustering) were also used. The groups of similar binding sites (proteins) or clusters generated by the best performing method were further analyzed in terms of local sequence identity, local structural similarity and conservation of analogous contacts with the FAD ligands. Each of the clusters was characterized by a unique set of structural features or patterns, demonstrating that the groups generated truly reflect the structural diversity of FAD binding sites. Proteins 2016; 84:1728–1747. © 2016 Wiley Periodicals, Inc.     

Testing the cranial evolutionary allometric ‘rule’ in Galliformes

Recent comparative studies have indicated the existence of a common cranial evolutionary allometric (CREA) pattern in mammals and birds, in which smaller species have relatively smaller faces and bigger braincases than larger species. In these studies, cranial allometry was tested using a multivariate regression between shape (described using landmarks coordinates) and size (i.e. centroid size), after accounting for phylogenetic relatedness. Alternatively, cranial allometry can be determined by comparing the sizes of two anatomical parts using a bivariate regression analysis. In this analysis, a slope higher or lower than one indicates the existence of positive or negative allometry, respectively. Thus, in those species that support the CREA ‘rule’, positive allometry is expected for the association between face size and braincase size, which would indicate that larger species have disproportionally larger faces. In this study, I applied these two approaches to explore cranial allometry in 83 Galliformes (Aves, Galloanserae), ranging in mean body weight from 30 g to 2.5 kg. The multivariate regression between shape and centroid size revealed the existence of a significant allometric pattern resembling CREA, whereas the second analysis revealed a negative allometry for beak size and braincase size (i.e. contrary to the CREA ‘rule’, larger galliform species have disproportionally shorter beaks than smaller galliform species). This study suggests that the presence of CREA may be overestimated when using cranium size as the standard measurement.     

Amphibian skin fungal communities vary across host species and do not correlate with infection by a pathogenic fungus

Amphibian population declines caused by the fungus Batrachochytrium dendrobatidis (Bd) have prompted studies on the bacterial community that resides on amphibian skin. However, studies addressing the fungal portion of these symbiont communities have lagged behind. Using ITS1 amplicon sequencing, we examined the fungal portion of the skin microbiome of temperate and tropical amphibian species currently coexisting with Bd in nature. We assessed cooccurrence patterns between bacterial and fungal OTUs using a subset of samples for which bacterial 16S rRNA gene amplicon data were also available. We determined that fungal communities were dominated by members of the phyla Ascomycota and Basidiomycota, and also by Chytridiomycota in the most aquatic amphibian species. Alpha diversity of the fungal communities differed across host species, and fungal community structure differed across species and regions. However, we did not find a correlation between fungal diversity/community structure and Bd infection, though we did identify significant correlations between Bd and specific OTUs. Moreover, positive bacterial–fungal cooccurrences suggest that positive interactions between these organisms occur in the skin microbiome. Understanding the ecology of amphibian skin fungi, and their interactions with bacteria will complement our knowledge of the factors influencing community assembly and the overall function of these symbiont communities.     

Local extinctions may be evidenced by the holes of the morphometric hypervolume in dung beetle communities

The body shape of a species is associated with its evolutionary history and can reflect behavioural peculiarities related to the ecological niche of each species. Morphology can characterise the morphometric niche of species and can be represented as body shape points within a morphometric universe. This information can be to calculate the morphometric diversity of communities through hypervolume metrics, and the hole sizes that remain in the morphometric hypervolume, which are empty areas with no species. Such holes may be ‘natural’ or caused by a local extinction. In this study, we evaluate the ecological community of dung beetles through the lens of morphometric diversity. We evaluated 38 dung beetle species from 30 subtropical communities in southern Brazil sampled in the summer of 2015, including 15 forest remnant communities from the Atlantic Forest and 15 communities from adjacent maize cultivations. The shape of 495 dung beetle specimens was measured using geometric morphometric and hypervolume techniques to calculate the morphometric diversity and the hole size of each of the 30 communities. We found that the taxonomic diversity positively correlated with the morphometric diversity and negatively correlated with the size of the holes. We also found that forest communities had higher values of morphometric diversity and smaller holes in the hypervolume than the maize cultivation communities, suggesting that local extinction may reduce community body shape spaces.     

Eight new microsatellite markers in Atlantic cod (Gadus morhua L.) derived from an enriched genomic library

One hundred and fifty random clones from an enriched genomic library of Atlantic cod were sequenced. Primer pairs were designed for 15 microsatellites containing perfect di‐, tri‐, tetra‐ and hexanucleotide repeats and characterized in 96 unrelated fish. Eight markers were successfully amplified, with the number of alleles ranging from two to nine per locus and observed heterozygosity ranging from 0.341 to 0.977. Loci Gmo‐G13 and Gmo‐G14 had a significant excess of homozygotes. All loci conformed to the Hardy–Weinberg equilibrium. Genetic linkage disequilibrium analysis between all pairs of the loci showed no significant departure from the null hypothesis between any of the loci.     

Challenges of antiangiogenic cancer therapy: trials and errors, and renewed hope

Angiogenesis inhibition has been proposed as a general strategy to fight cancer. However, in spite of the promising preclinical results, a first generation of antiangiogenic compounds yielded poor results in clinical trials. Conceptual errors and mistakes in the design of trials and in the definition of clinical end-points could account for these negative results. In this context of discouraging results, a second generation of antiangiogenic therapies is showing positive results in phases II and III trials at the beginning of the twenty-first century. In fact, several combined treatments with conventional chemotherapy and antiangiogenic compounds have been recently approved. The discovery and pharmacological development of future generations of angiogenesis inhibitors will benefit from further advances in the understanding of the mechanisms involved in human angiogenesis. New styles of trials are necessary, to avoid missing potential therapeutic effects. Different clinical end-points, new surrogate biomarkers and methods of imaging will be helpful in this process. Real efficacy in clinical trials may come with the combined use of antiangiogenic agents with conventional chemotherapy or radiotherapy, and combinations of several antiangiogenic compounds with different mechanisms of action. Finally, the existing antiangiogenic strategies should include other approaches such as vascular targeting or angioprevention.