Engineering a predatory bacterium as a proficient killer agent for intracellular bio-products recovery:The case of the polyhydroxyalkanoates

12p.-6 fig.-2 tab.This work examines the potential of the predatory bacterium Bdellovibrio bacteriovorus HD100, an obligate predator of other Gram-negative bacteria, as an external cell-lytic agent for recovering valuable intracellular bio-products produced by prey cultures. The bio-product targets to be recovered were polyhydroxyalkanoates (PHAs) produced naturally by Pseudomonas putida and Cupriavidus necator, or by recombinant Escherichia coli strains. B. bacteriovorus with a mutated PHA depolymerase gene to prevent the unwanted breakdown of the bio-product allowed the recovery of up to 80% of that accumulated by the prey bacteria, even at high biomass concentrations. This innovative downstream process highlights how B. bacteriovorus can be used as a novel, biological lytic agent for the inexpensive, industrial scale recovery of intracellular products from different Gram-negative prey cultures.This work was funded by the EU Seventh Framework Programme under grant agreement no. 311815 (SYNPOL), and by grants from the Comunidad de Madrid (P2013/MIT2807) and the Spanish Ministerio de Economía y Competitividad, (BIO2010-21049, BIO2013-44878-R).Peer reviewe

Weak Galactic halo--dwarf spheroidal connection from RR Lyrae stars

We discuss the role that dwarf galaxies may have played in the formation of the Galactic halo (Halo) using RR Lyrae stars (RRL) as tracers of their ancient stellar component. The comparison is performed using two observables (periods, luminosity amplitudes) that are reddening and distance independent. Fundamental mode RRL in six dwarf spheroidals and eleven ultra faint dwarf galaxies (1,300) show a Gaussian period distribution well peaked around a mean period of =0.610+-0.001 days (sigma=0.03). The Halo RRL (15,000) are characterized by a broader period distribution. The fundamental mode RRL in all the dwarf spheroidals apart from Sagittarius are completely lacking in High Amplitude Short Period (HASP) variables, defined as those having P 0.75mag. Such variables are not uncommon in the Halo and among the globular clusters and massive dwarf irregulars. To further interpret this evidence, we considered eighteen globulars covering a broad range in metallicity (-2.3< [Fe/H]< -1.1) and hosting more than 35 RRL each. The metallicity turns out to be the main parameter, since only globulars more metal--rich than [Fe/H] -1.5 host RRL in the HASP region. This finding suggests that dSphs similar to the surviving ones do not appear to be the major building-blocks of the Halo. Leading physical arguments suggest an extreme upper limit of 50% to their contribution. On the other hand, massive dwarfs hosting an old population with a broad metallicity distribution (Large Magellanic Cloud, Sagittarius) may have played a primary role in the formation of the Halo.Comment: accepted for publication in ApJ Lette

The ISLAndS project II: The Lifetime Star Formation Histories of Six Andromeda dSphs

The Initial Star formation and Lifetimes of Andromeda Satellites (ISLAndS) project uses Hubble Space Telescope imaging to study a representative sample of six Andromeda dSph satellite companion galaxies. The main goal of the program is to determine whether the star formation histories (SFHs) of the Andromeda dSph satellites demonstrate significant statistical differences from those of the Milky Way, which may be attributable to the different properties of their local environments. Our observations reach the oldest main sequence turn-offs, allowing a time resolution at the oldest ages of ~ 1 Gyr, which is comparable to the best achievable resolution in the MW satellites. We find that the six dSphs present a variety of SFHs that are not strictly correlated with luminosity or present distance from M31. Specifically, we find a significant range in quenching times (lookback times from 9 to 6 Gyr), but with all quenching times more than ~ 6 Gyr ago. In agreement with observations of Milky Way companions of similar mass, there is no evidence of complete quenching of star formation by the cosmic UV background responsible for reionization, but the possibility of a degree of quenching at reionization cannot be ruled out. We do not find significant differences between the SFHs of the three members of the vast, thin plane of satellites and the three off-plane dSphs. The primary difference between the SFHs of the ISLAndS dSphs and Milky Way dSph companions of similar luminosities and host distances is the absence of very late quenching (< 5 Gyr ago) dSphs in the ISLAndS sample. Thus, models that can reproduce satellite populations with and without late quenching satellites will be of extreme interest.Comment: 24 pages, 11 figures, 3 tables, submitted to the Ap

Functional Divergence of the Arabidopsis Florigen-Interacting bZIP Transcription Factors FD and FDP.

Flowering of many plant species depends on interactions between basic leucine zipper (bZIP) transcription factors and systemically transported florigen proteins. Members of the genus Arabidopsis contain two of these bZIPs, FD and FDP, which we show have largely complementary expression patterns in shoot apices before and during flowering. CRISPR-Cas9-induced null mutants for FDP flower slightly earlier than wild-type, whereas fd mutants are late flowering. Identical G-box sequences are enriched at FD and FDP binding sites, but only FD binds to genes involved in flowering and only fd alters their transcription. However, both proteins bind to genes involved in responses to the phytohormone abscisic acid (ABA), which controls developmental and stress responses. Many of these genes are differentially expressed in both fd and fdp mutant seedlings, which also show reduced ABA sensitivity. Thus, florigen-interacting bZIPs have distinct functions in flowering dependent on their expression patterns and, at earlier stages in development, play common roles in phytohormone signaling

Gut Microbiota Associated with Clostridioides difficile Carriage in Three Clinical Groups (Inflammatory Bowel Disease, C. difficile Infection and Healthcare Workers) in Hospital Field

peer reviewedClostridioides difficile is an anaerobic spore-forming Gram-positive bacterium. C. difficile carriage and 16S rDNA profiling were studied in three clinical groups at three different sampling times: inflammatory bowel disease (IBD) patients, C. difficile infection (CDI) patients and healthcare workers (HCWs). Diversity analysis was realized in the three clinical groups, the positive and negative C. difficile carriage groups and the three analysis periods. Concerning the three clinical groups, β-diversity tests showed significant differences between them, especially between the HCW group and IBD group and between IBD patients and CDI patients. The Simpson index (evenness) showed a significant difference between two clinical groups (HCWs and IBD). Several genera were significantly different in the IBD patient group (Sutterella, Agathobacter) and in the CDI patient group (Enterococcus, Clostridioides). Concerning the positive and negative C. difficile carriage groups, β-diversity tests showed significant differences. Shannon, Simpson and InvSimpson indexes showed significant differences between the two groups. Several genera had significantly different relative prevalences in the negative group (Agathobacter, Sutterella, Anaerostipes, Oscillospira) and the positive group (Enterococcus, Enterobacteriaceae_ge and Enterobacterales_ge). A microbiota footprint was detected in C. difficile-positive carriers. More experiments are needed to test this microbiota footprint to see its impact on C. difficile infection

Impact of Baseline Steroids on Efficacy of Programmed Cell Death-1 and Programmed Death-Ligand 1 Blockade in Patients With Non-Small-Cell Lung Cancer

Treatment with programmed cell death-1 or programmed death ligand 1 (PD-(L)1) inhibitors is now standard therapy for patients with lung cancer. The immunosuppressive effect of corticosteroids may reduce efficacy of PD-(L)1 blockade. On-treatment corticosteroids for treatment of immune-related adverse events do not seem to affect efficacy, but the potential impact of baseline corticosteroids at the time of treatment initiation is unknown. Clinical trials typically excluded patients who received baseline corticosteroids, which led us to use real-world data to examine the effect of corticosteroids at treatment initiation. We identified patients who were PD-(L)1-naïve with advanced non-small-cell lung cancer from two institutions-Memorial Sloan Kettering Cancer Center and Gustave Roussy Cancer Center-who were treated with single-agent PD-(L)1 blockade. Clinical and pharmacy records were reviewed to identify corticosteroid use at the time of beginning anti-PD-(L)1 therapy. We performed multivariable analyses using Cox proportional hazards regression model and logistic regression. Ninety (14%) of 640 patients treated with single-agent PD-(L)1 blockade received corticosteroids of ≥ 10 mg of prednisone equivalent daily at the start of the PD-(L)1 blockade. Common indications for corticosteroids were dyspnea (33%), fatigue (21%), and brain metastases (19%). In both independent cohorts, Memorial Sloan Kettering Cancer Center (n = 455) and Gustave Roussy Cancer Center (n = 185), baseline corticosteroids were associated with decreased overall response rate, progression-free survival, and overall survival with PD-(L)1 blockade. In a multivariable analysis of the pooled population, adjusting for smoking history, performance status, and history of brain metastases, baseline corticosteroids remained significantly associated with decreased progression-free survival (hazard ratio, 1.3; P = .03), and overall survival (hazard ratio, 1.7; P Baseline corticosteroid use of ≥ 10 mg of prednisone equivalent was associated with poorer outcome in patients with non-small-cell lung cancer who were treated with PD-(L)1 blockade. Prudent use of corticosteroids at the time of initiating PD-(L)1 blockade is recommended

Organised Genome Dynamics in the Escherichia coli Species Results in Highly Diverse Adaptive Paths

The Escherichia coli species represents one of the best-studied model organisms, but also encompasses a variety of commensal and pathogenic strains that diversify by high rates of genetic change. We uniformly (re-) annotated the genomes of 20 commensal and pathogenic E. coli strains and one strain of E. fergusonii (the closest E. coli related species), including seven that we sequenced to completion. Within the ∼18,000 families of orthologous genes, we found ∼2,000 common to all strains. Although recombination rates are much higher than mutation rates, we show, both theoretically and using phylogenetic inference, that this does not obscure the phylogenetic signal, which places the B2 phylogenetic group and one group D strain at the basal position. Based on this phylogeny, we inferred past evolutionary events of gain and loss of genes, identifying functional classes under opposite selection pressures. We found an important adaptive role for metabolism diversification within group B2 and Shigella strains, but identified few or no extraintestinal virulence-specific genes, which could render difficult the development of a vaccine against extraintestinal infections. Genome flux in E. coli is confined to a small number of conserved positions in the chromosome, which most often are not associated with integrases or tRNA genes. Core genes flanking some of these regions show higher rates of recombination, suggesting that a gene, once acquired by a strain, spreads within the species by homologous recombination at the flanking genes. Finally, the genome's long-scale structure of recombination indicates lower recombination rates, but not higher mutation rates, at the terminus of replication. The ensuing effect of background selection and biased gene conversion may thus explain why this region is A+T-rich and shows high sequence divergence but low sequence polymorphism. Overall, despite a very high gene flow, genes co-exist in an organised genome

Insight into cross-talk between intra-amoebal pathogens

Abstract: Background: Amoebae are phagocytic protists where genetic exchanges might take place between amoeba-resistant bacteria. These amoebal pathogens are able to escape the phagocytic behaviour of their host. They belong to different bacterial phyla and often show a larger genome size than human-infecting pathogens. This characteristic is proposed to be the result of frequent gene exchanges with other bacteria that share a sympatric lifestyle and contrasts with the genome reduction observed among strict human pathogens.Results: We sequenced the genome of a new amoebal pathogen, Legionella drancourtii, and compared its gene content to that of a Chlamydia-related bacterium, Parachlamydia acanthamoebae. Phylogenetic reconstructions identified seven potential horizontal gene transfers (HGTs) between the two amoeba-resistant bacteria, including a complete operon of four genes that encodes an ABC-type transporter. These comparisons pinpointed potential cases of gene exchange between P. acanthamoebae and Legionella pneumophila, as well as gene exchanges between other members of the Legionellales and Chlamydiales orders. Moreover, nine cases represent possible HGTs between representatives from the Legionellales or Chlamydiales and members of the Rickettsiales order.Conclusions: This study identifies numerous gene exchanges between intracellular Legionellales and Chlamydiales bacteria, which could preferentially occur within common inclusions in their amoebal hosts. Therefore it contributes to improve our knowledge on the intra-amoebal gene properties associated to their specific lifestyle

Structure and diversity trends at Fagus timberline in central Italy

Structure and diversity trends (β-diversity and species richness) across the Fagus sylvatica timberline in the central Apennines were investigated. Twenty-three belt transects were laid out across the upper forest line in the Simbruini Mountains. Number of species, plant cover, and height of different layers were recorded in each quadrat. The moving split-window method was used to detect ecological discontinuities across beech timberlines. We show how β-diversity changes along timberlines and we put forward some hypotheses about the possible dynamics of these transitions. Fourmodels resulted from the analysis of β-diversity trends: two β-diversity peaks indicated a transition where shrubs, mainly Juniperus communis ssp. alpina, (two high peaks) or beech scrub (two small peaks) formed a mantle that could allow forest expansion. One high β-diversity peak referred to an anthropo-zoogenic boundary maintained by disturbance, without the presence of a mantle. A little peak indicated a gradual transition at the upper potential timberline limit where beech forest had lost its typical floristical composition and structural characteristics

Ruxolitinib for Glucocorticoid-Refractory Acute Graft-versus-Host Disease

BACKGROUND: Acute graft-versus-host disease (GVHD) remains a major limitation of allogeneic stem-cell transplantation; not all patients have a response to standard glucocorticoid treatment. In a phase 2 trial, ruxolitinib, a selective Janus kinase (JAK1 and JAK2) inhibitor, showed potential efficacy in patients with glucocorticoid-refractory acute GVHD. METHODS: We conducted a multicenter, randomized, open-label, phase 3 trial comparing the efficacy and safety of oral ruxolitinib (10 mg twice daily) with the investigator's choice of therapy from a list of nine commonly used options (control) in patients 12 years of age or older who had glucocorticoid-refractory acute GVHD after allogeneic stem-cell transplantation. The primary end point was overall response (complete response or partial response) at day 28. The key secondary end point was durable overall response at day 56. RESULTS: A total of 309 patients underwent randomization; 154 patients were assigned to the ruxolitinib group and 155 to the control group. Overall response at day 28 was higher in the ruxolitinib group than in the control group (62% [96 patients] vs. 39% [61]; odds ratio, 2.64; 95% confidence interval [CI], 1.65 to 4.22; P<0.001). Durable overall response at day 56 was higher in the ruxolitinib group than in the control group (40% [61 patients] vs. 22% [34]; odds ratio, 2.38; 95% CI, 1.43 to 3.94; P<0.001). The estimated cumulative incidence of loss of response at 6 months was 10% in the ruxolitinib group and 39% in the control group. The median failure-free survival was considerably longer with ruxolitinib than with control (5.0 months vs. 1.0 month; hazard ratio for relapse or progression of hematologic disease, non-relapse-related death, or addition of new systemic therapy for acute GVHD, 0.46; 95% CI, 0.35 to 0.60). The median overall survival was 11.1 months in the ruxolitinib group and 6.5 months in the control group (hazard ratio for death, 0.83; 95% CI, 0.60 to 1.15). The most common adverse events up to day 28 were thrombocytopenia (in 50 of 152 patients [33%] in the ruxolitinib group and 27 of 150 [18%] in the control group), anemia (in 46 [30%] and 42 [28%], respectively), and cytomegalovirus infection (in 39 [26%] and 31 [21%]). CONCLUSIONS: Ruxolitinib therapy led to significant improvements in efficacy outcomes, with a higher incidence of thrombocytopenia, the most frequent toxic effect, than that observed with control therapy