Pyrosequencing Unveils Cystic Fibrosis Lung Microbiome Differences Associated with a Severe Lung Function Decline

Chronic airway infection is a hallmark feature of cystic fibrosis (CF) disease. In the present study, sputum samples from CF patients were collected and characterized by 16S rRNA gene-targeted approach, to assess how lung microbiota composition changes following a severe decline in lung function. In particular, we compared the airway microbiota of two groups of patients with CF, i.e. patients with a substantial decline in their lung function (SD) and patients with a stable lung function (S). The two groups showed a different bacterial composition, with SD patients reporting a more heterogeneous community than the S ones. Pseudomonas was the dominant genus in both S and SD patients followed by Staphylococcus and Prevotella. Other than the classical CF pathogens and the most commonly identified non-classical genera in CF, we found the presence of the unusual anaerobic genus Sneathia. Moreover, the oligotyping analysis revealed the presence of other minor genera described in CF, highlighting the polymicrobial nature of CF infection. Finally, the analysis of correlation and anti-correlation networks showed the presence of antagonism and ecological independence between members of Pseudomonas genus and the rest of CF airways microbiota, with S patients showing a more interconnected community in S patients than in SD ones. This population structure suggests a higher resilience of S microbiota with respect to SD, which in turn may hinder the potential adverse impact of aggressive pathogens (e.g. Pseudomonas). In conclusion, our findings shed a new light on CF airway microbiota ecology, improving current knowledge about its composition and polymicrobial interactions in patients with CF.     

Comparison between Yeasts from Grape and Agave Musts for Traits of Technological interest

Summary In Mexico there are different alcoholic beverages produced from agave juices from different agave plants, which are cooked, fermented and distilled. For tequila production only Agave tequilana is allowed. In this study we compared yeast strains of different species from different origin (agave and grape juice) for parameters of technological interest, such as SO₂ and copper resistance, ethanol tolerance and enzymatic activities. All agave strains were found to be more resistant to SO₂ and agave non-Saccharomyces yeasts were more tolerant to ethanol, whereas grape strains exhibited positive results for β-glucosidase and β-xylosidase activities. As regards fermentations of Agave tequilana juice with ethanol added at different concentrations, only agave Saccharomyces strains were more tolerant to ethanol than grape strains.     

Cell Survival and Polarity of Drosophila Follicle Cells Require the Activity of Ecdysone Receptor B1 Isoform

Proper assembly and maintenance of epithelia are critical for normal development and homeostasis. Here, using the Drosophila ovary as a model, we identify a role for the B1 isoform of the ecdysone receptor (EcR-B1) in this process. We performed a reverse genetic analysis of EcR-B1 function during oogenesis and demonstrate that silencing of this receptor isoform causes loss of integrity and multilayering of the follicular epithelium. We show that multilayered follicle cells lack proper cell polarity with altered distribution of apical and basolateral cell polarity markers including atypical-protein kinase C (aPKC), Discs-large (Dlg), and Scribble (Scrib) and aberrant accumulation of adherens junctions and F-actin cytoskeleton. We find that the EcR-B1 isoform is required for proper follicle cell polarity both during early stages of oogenesis, when follicle cells undergo the mitotic cell cycle, and at midoogenesis when these cells stop dividing and undergo several endocycles. In addition, we show that the EcR-B1 isoform is required during early oogenesis for follicle cell survival and that disruption of its function causes apoptotic cell death induced by caspase.     

High-level HIV-1 Nef transient expression in Nicotiana benthamiana using the P19 gene silencing suppressor protein of Artichoke Mottled Crinckle Virus

Background  

In recent years, different HIV antigens have been successfully expressed in plants by either stable transformation or transient expression systems. Among HIV proteins, Nef is considered a promising target for the formulation of a multi-component vaccine due to its implication in the first steps of viral infection. Attempts to express Nef as a single protein product (not fused to a stabilizing protein) in transgenic plants resulted in disappointingly low yields (about 0.5% of total soluble protein). In this work we describe a transient expression system based on co-agroinfiltration of plant virus gene silencing suppressor proteins in Nicotiana benthamiana, followed by a two-step affinity purification protocol of plant-derived Nef.     

Stability of telomeric G-quadruplexes

In most eukaryotes, telomeric DNA consists of repeats of a short motif that includes consecutive guanines and may hence fold into G-quadruplexes. Budding yeasts have telomeres composed of longer repeats and show variation in the degree of repeat homogeneity. Although telomeric sequences from several organisms have been shown to fold into G-quadruplexes in vitro, surprisingly, no study has been dedicated to the comparison of G-quadruplex folding and stability of known telomeric sequences. Furthermore, to our knowledge, folding of yeast telomeric sequences into intramolecular G-quadruplexes has never been investigated. Using biophysical and biochemical methods, we studied sequences mimicking about four repetitions of telomeric motifs from a variety of organisms, including yeasts, with the aim of comparing the G-quadruplex folding potential of telomeric sequences among eukaryotes. G-quadruplex folding did not appear to be a conserved feature among yeast telomeric sequences. By contrast, all known telomeric sequences from eukaryotes other than yeasts folded into G-quadruplexes. Nevertheless, while G(3)T(1-4)A repeats (found in a variety of organisms) and G(4)T(2,4) repeats (found in ciliates) folded into stable G-quadruplexes, G-quadruplexes formed by repetitions of G(2)T(2)A and G(2)CT(2)A motifs (found in many insects and in nematodes, respectively) appeared to be in equilibrium with non-G-quadruplex structures (likely hairpin-duplexes).     

Kin-informative recognition cues in ants

Although social groups are characterized by cooperation, they are also often the scene of conflict. In non-clonal systems, the reproductive interests of group members will differ and individuals may benefit by exploiting the cooperative efforts of other group members. However, such selfish behaviour is thought to be rare in one of the classic examples of cooperation--social insect colonies--because the colony-level costs of individual selfishness select against cues that would allow workers to recognize their closest relatives. In accord with this, previous studies of wasps and ants have found little or no kin information in recognition cues. Here, we test the hypothesis that social insects do not have kin-informative recognition cues by investigating the recognition cues and relatedness of workers from four colonies of the ant Acromyrmex octospinosus. Contrary to the theoretical prediction, we show that the cuticular hydrocarbons of ant workers in all four colonies are informative enough to allow full-sisters to be distinguished from half-sisters with a high accuracy. These results contradict the hypothesis of non-heritable recognition cues and suggest that there is more potential for within-colony conflicts in genetically diverse societies than previously thought.     

Pseudomonas syringae pv. actinidiae draft genomes comparison reveal strain-specific features involved in adaptation and virulence to Actinidia species

A recent re-emerging bacterial canker disease incited by Pseudomonas syringae pv. actinidiae (Psa) is causing severe economic losses to Actinidia chinensis and A. deliciosa cultivations in southern Europe, New Zealand, Chile and South Korea. Little is known about the genetic features of this pathovar. We generated genome-wide Illumina sequence data from two Psa strains causing outbreaks of bacterial canker on the A. deliciosa cv. Hayward in Japan (J-Psa, type-strain of the pathovar) and in Italy (I-Psa) in 1984 and 1992, respectively as well as from a Psa strain (I2-Psa) isolated at the beginning of the recent epidemic on A. chinensis cv. Hort16A in Italy. All strains were isolated from typical leaf spot symptoms. The phylogenetic relationships revealed that Psa is more closely related to P. s. pv. theae than to P. avellanae within genomospecies 8. Comparative genomic analyses revealed both relevant intrapathovar variations and putative pathovar-specific genomic regions in Psa. The genomic sequences of J-Psa and I-Psa were very similar. Conversely, the I2-Psa genome encodes four additional effector protein genes, lacks a 50 kb plasmid and the phaseolotoxin gene cluster, argK-tox but has acquired a 160 kb plasmid and putative prophage sequences. Several lines of evidence from the analysis of the genome sequences support the hypothesis that this strain did not evolve from the Psa population that caused the epidemics in 1984-1992 in Japan and Italy but rather is the product of a recent independent evolution of the pathovar actinidiae for infecting Actinidia spp. All Psa strains share the genetic potential for copper resistance, antibiotic detoxification, high affinity iron acquisition and detoxification of nitric oxide of plant origin. Similar to other sequenced phytopathogenic pseudomonads associated with woody plant species, the Psa strains isolated from leaves also display a set of genes involved in the catabolism of plant-derived aromatic compounds.     

Systemic administration of high-molecular weight hyaluronan stimulates wound healing in genetically diabetic mice

Hyaluronic acid (HA), an essential component of the extracellular matrix, is an efficient space filler that maintains hydration, serves as a substrate for assembly of proteoglycans and is involved in wound healing. Although numerous pieces of evidence demonstrate beneficial effects in promoting wound healing in diabetes, a systemic approach has never been tested. We used an incisional wound healing model in genetically diabetic mice to test the effects of systemic injection of HA. Diabetic (n = 56) and normoglycemic (n = 56) mice were subjected to incision and randomized (8 groups of 7 animals each) to receive HA at different doses, 7.5, 15 and 30 mg/kg/i.p., or vehicle (0.9% NaCl solution) for 12 days. At the end of the experiment animals were sacrificed and skin wounds were excised for histological, biochemical and molecular analysis. Histology revealed that the most effective dose to improve wound repair and angiogenesis in diabetic mice was 30 mg/kg. Furthermore HA injection (30 mg/kg) improved the altered healing pattern in diabetic animals, increased skin remodeling proteins TGF-β and transglutaminase-II and restored the altered expression of cyclin B1/Cdc2 complex. Evaluation of skin from diabetic animals injected with HA revealed also an increase in HA content, suggesting that systemic injection may be able to restore the reduced intracellular HA pool of diabetic mice. Finally HA markedly improved skin mechanical properties. These promising results, if confirmed in a clinical setting, may improve the care and management of diabetic patients.     

Characterization of a Small Family (CAIII) of Microsatellite-Containing Sequences with X-Y Homology

Four X-linked loci showing homology with a previously described Y-linked polymorphic locus (DYS413) were identified and characterized. By fluorescent in situ hybridization (FISH), somatic cell hybrids, and YAC screening, the X-linked members of this small family of sequences (CAIII) all map in Xp22, while the Y members map in Yq11. These loci contribute to the overall similarity of the two genomic regions. All of the CAIII loci contain an internal microsatellite of the (CA)_n type. The microsatellites display extensive length polymorphism in two of the X-linked members as well as in the Y members. In addition, common sequence variants are found in the portions flanking the microsatellites in two of the X-linked members. Our results indicate that, during the evolution of this family, length variation on the Y chromosome was accumulated at a rate not slower than that on the X chromosome. Finally, these sequences represent a model system with which to analyze human populations for similar X- and Y-linked polymorphisms. Received: 29 July 1996 / Accepted: 15 January 1997