Expression and characterization of the assimilatory NADH-nitrite reductase from the phototrophic bacterium Rhodobacter capsulatus E1F1

A nas gene region from Rhodobacter capsulatus E1F1 containing the putative nasB gene for nitrite reductase was previously cloned. The recombinant His₆-NasB protein overproduced in E. coli showed nitrite reductase activity in vitro with both reduced methyl viologen and NADH as electron donors. The apparent K _m values for nitrite and NADH were 0.5 mM and 20 μM, respectively, at the pH and temperature optima (pH 9 and 30°C). The optical spectrum showed features that indicate the presence of FAD, iron-sulfur cluster and siroheme as prosthetic groups, and nitrite reductase activity was inhibited by sulfide and iron reagents. These results indicate that the phototrophic bacterium R. capsulatus E1F1 possesses an assimilatory NADH-nitrite reductase similar to that described in non-phototrophic organisms.     

Evolutionary history of the European free-tailed bat,a tropical affinity species spanning across the Mediterranean Basin

The Mediterranean Basin is a global biodiversity hotspot, hosting a number of native species belonging to families that are found almost exclusively in tropical climates. Yet, whether or not these taxa were able to survive in the Mediterranean region during the Quaternary climatic oscillations remains unknown. Focusing on the European free-tailed bat (Tadarida teniotis), we aimed to (a) identify potential ancient populations and glacial refugia; (b) determine the post-glacial colonization routes across the Mediterranean; and (c) evaluate current population structure and demography. Mitochondrial and nuclear markers were used to understand T. teniotis evolutionary and demographic history. We show that T. teniotis is likely restricted to the Western Palearctic, with mitochondrial phylogeny suggesting a split between an Anatolian/Middle East clade and a European clade. Nuclear data pointed to three genetic populations, one of which is an isolated and highly differentiated group in the Canary Islands, another distributed across Iberia, Morocco, and France, and a third stretching from Italy to the east, with admixture following a pattern of isolation by distance. Evolutionary and demographic reconstruction supports a pre-Last Glacial Maximum (LGM) colonization of Italy and the Anatolian/Middle East, while the remaining populations were colonized from Italy after the Younger Dryas. We also found support for demographic expansion following the Iberian colonization. The results show that during the LGM T. teniotis persisted in Mediterranean refugia and has subsequently expanded to its current circum-Mediterranean range. Our findings raise questions regarding the physiological and ecological traits that enabled species with tropical affinities to survive in colder climates.     

Macrophage-specific responses to human- and animal-adapted tubercle bacilli reveal pathogen and host factors driving multinucleated cell formation

The Mycobacterium tuberculosis complex (MTBC) is a group of related pathogens that cause tuberculosis (TB) in mammals. MTBC species are distinguished by their ability to sustain in distinct host populations. While Mycobacterium bovis (Mbv) sustains transmission cycles in cattle and wild animals and causes zoonotic TB, M. tuberculosis (Mtb) affects human populations and seldom causes disease in cattle. The host and pathogen determinants underlying host tropism between MTBC species are still unknown. Macrophages are the main host cell that encounters mycobacteria upon initial infection, and we hypothesised that early interactions between the macrophage and mycobacteria influence species-specific disease outcome. To identify factors that contribute to host tropism, we analysed blood-derived primary human and bovine macrophages (hMϕ or bMϕ, respectively) infected with Mbv and Mtb. We show that Mbv and Mtb reside in different cellular compartments and differentially replicate in hMϕ whereas both Mbv and Mtb efficiently replicate in bMϕ. Specifically, we show that out of the four infection combinations, only the infection of bMϕ with Mbv promoted the formation of multinucleated giant cells (MNGCs), a hallmark of tuberculous granulomas. Mechanistically, we demonstrate that both MPB70 from Mbv and extracellular vesicles released by Mbv-infected bMϕ promote macrophage multinucleation. Importantly, we extended our in vitro studies to show that granulomas from Mbv-infected but not Mtb-infected cattle contained higher numbers of MNGCs. Our findings implicate MNGC formation in the contrasting pathology between Mtb and Mbv for the bovine host and identify MPB70 from Mbv and extracellular vesicles from bMϕ as mediators of this process.     

966. CLAVIJA DOMINGENSIS

Clavija domingensis Urb. & Ekman was one of the many Haitian endemics that were described based on collections made by the great Swedish botanist Leonard Ekman between 1924 and 1928. The species is Critically Endangered sensu IUCN (criteria c2a(i); D) and it is currently the focus of conservation initiatives in Jardin Botanique des Cayes (Haiti), Jardín Botánico Nacional Dr. Rafael M. Moscoso (Dominican Republic), and Fairchild Tropical Botanic Garden (U.S.A.). Now known from only six localities from southern Haiti, each locality only represents a single individual. The species is illustrated based on plants grown in Fairchild Tropical Botanic Garden.     

Further aspects on the hemolytic activity of the antibiotic lipopeptide iturin A

The bacterial lipopeptide iturin A is able to cause hemolysis of human erythrocytes in a dose-dependent manner. Hemolysis takes place at iturin concentrations below its critical micellar concentration. Relative kinetics determinations clearly show that K⁺ leakage occurs prior to hemoglobin release. Furthermore, hemolysis can be prevented by addition to the outer solution of osmotic protectants of appropriate size. Altogether these results indicate that iturin A-induced hemolysis follows a colloid-osmotic mechanism, with the formation of a membrane pore of average diameter 32 Å. Iturin A is capable of inducing leakage of an aqueous fluorescent probe trapped in human erythrocyte ghosts, but not in large unilamellar liposomes made of various lipid compositions. The different permeabilizing effects of iturin A on model and biological membranes are discussed on the light of the presented results.     

The SNPlex genotyping system: a flexible and scalable platform for SNP genotyping.

We developed the SNPlex Genotyping System to address the need for accurate genotyping data, high sample throughput, study design flexibility, and cost efficiency. The system uses oligonucleotide ligation/polymerase chain reaction and capillary electrophoresis to analyze bi-allelic single nucleotide polymorphism genotypes. It is well suited for single nucleotide polymorphism genotyping efforts in which throughput and cost efficiency are essential. The SNPlex Genotyping System offers a high degree of flexibility and scalability, allowing the selection of custom-defined sets of SNPs for medium- to high-throughput genotyping projects. It is therefore suitable for a broad range of study designs. In this article we describe the principle and applications of the SNPlex Genotyping System, as well as a set of single nucleotide polymorphism selection tools and validated assay resources that accelerate the assay design process. We developed the control pool, an oligonucleotide ligation probe set for training and quality-control purposes, which interrogates 48 SNPs simultaneously. We present performance data from this control pool obtained by testing genomic DNA samples from 44 individuals. in addition, we present data from a study that analyzed 521 SNPs in 92 individuals. Combined, both studies show the SNPlex Genotyping system to have a 99.32% overall call rate, 99.95% precision, and 99.84% concordance with genotypes analyzed by TaqMan probe-based assays. The SNPlex Genotyping System is an efficient and reliable tool for a broad range of genotyping applications, supported by applications for study design, data analysis, and data management.     

Understory spider diversity in two remnants of tropical montane cloud forest in Chiapas, Mexico

We evaluated the spider diversity of a tropical montane cloud forest understory in two nearby sites with different degree of human disturbance at the Biosphere Reserve Volcán Tacaná, Chiapas, Mexico. The study was conducted over a 24 days period distributed in 6 months in 2009, covering dry and rainy seasons. A total of 8,370 spiders (1,208 adults and 7,162 juveniles) were collected. Determined specimens (7,747) represented 112 species and morphospecies, 71 genera and 22 families. The results showed that human disturbance has an influence on spider communities: species richness was significantly higher in the preserved site as regards to the disturbed site. Despite their proximity, the composition of spider communities showed only a moderate similarity between the two sites. No differences in abundance were found among sites when considering the whole sample, but sites differed clearly when seasons were analyzed separately. The rainy season had a negative effect on the abundance of spiders in the preserved site. Although the spider community structure was very similar between sites, there was a trend towards a greater species evenness in the preserved site for the whole sampling period and for the dry season.