Modeling and analysis of ArsR genetic circuits

Please contact the authors for the abstract

Genetic and physiological insights into the diazotrophicactivity of a non-cyanobacterial marine diazotroph

International audienceNitrogen (N2) fixation, or diazotrophy, supports a large part of primary production in oceans. Culture-independent approaches highlighted the presence in abundance of marine non-cyanobacterial diazotrophs (NCD), but their ecophysiology remains elusive, mostly because of the low number of isolated NCD and because of the lack of available genetic tools for these isolates. Here, a dual genetic and functional approach allowed unveiling the ecophysiology of a marine NCD affiliated to the species Vibrio diazotrophicus. Physiological characterization of the first marine NCD mutant obtained so far was performed using a soft-gellan assay, demonstrating that a ΔnifH mutant is not able to grow in nitrogen-free media. Furthermore, we demonstrated that V. diazotrophicus produces a thick biofilm under diazotrophic conditions, suggesting biofilm production as an adaptive response of this NCD to cope with the inhibition of nitrogen fixation by molecular oxygen. Finally, the genomic signature of V. diazotrophicus is essentially absent from metagenomic data of Tara Ocean expeditions, despite having been isolated from various marine environments. We think that the genetically tractable V. diazotrophicus strain used in this study may serve as an ideal model to study the ecophysiology of these overlooked procaryotic group

Can selenium‐enriched spirulina supplementation ameliorate sepsis outcomes in selenium‐deficient animals?

In intensive care units, sepsis is the first cause of death. In this pathology, inflammation and oxidative status play a crucial role in patient outcomes. Interestingly, 92% of septic patients exhibit low selenium plasma concentrations (a component of antioxidant enzymes). Moreover, Spirulina platensis, a blue-green algae, demonstrated anti-inflammatory effects. In this context, the main purpose of our study was to analyze the effect of a selenium-enriched spirulina after a selenium deficiency on sepsis outcome in rats. Sixty-four rats were fed 12 weeks with a selenium-deficient food. After 8 weeks, rats were supplemented (via drinking water) for 4 weeks with sodium selenite (Se), spirulina (Spi), or selenium-enriched spirulina (SeSp). Sepsis was then induced by cecal ligature and puncture, and survival duration was observed. The plasma selenium concentration was measured by ICPMS. Expression of GPx1 and GPx3 mRNA was measured by RT-PCR. Blood parameters (lactates and HCO3− concentrations, pH, PO2, and PCO2) were analyzed at 0, 1, and 2 h as well as inflammatory cytokines (IL-6, TNF-α, IL-10). Sodium selenite and SeSP supplementations restored plasma selenium concentration prior to sepsis. The survival duration of SeSP septic rats was significantly lower than that of selenium-supplemented ones. Gpx1 mRNA was increased after a selenium-enriched spirulina supplementation while Gpx3 mRNA levels remained unchanged. Furthermore, sodium selenite prevented sepsis-induced acidosis. Our results show that on a basis of a Se deficiency, selenium-enriched spirulina supplementations significantly worsen sepsis outcome when compared to Se supplementation. Furthermore, Se supplementation but not selenium-enriched spirulina supplementation decreased inflammation and restored acid–base equilibrium after a sepsis induction

Genetic and physiological insights into the diazotrophic activity of a non‐cyanobacterial marine diazotroph

Nitrogen (N2) fixation, or diazotrophy, supports a large part of primary production in oceans. Culture-independent approaches highlighted the presence in abundance of marine non-cyanobacterial diazotrophs (NCD) but their ecophysiology remains elusive, mostly because of the low number of isolated NCD and because of the lack of available genetic tools for these isolates. Here, a dual genetic and functional approach allowed unveiling the ecophysiology of a marine NCD affiliated to the species Vibrio diazotrophicus. Physiological characterization of the first marine NCD mutant obtained so far was performed using a soft-gellan assay, demonstrating that a ΔnifH mutant in not able to grow in nitrogen-free media. Furthermore, we demonstrated that V. diazotrophicus produces a thick biofilm under diazotrophic conditions, suggesting biofilm production as an adaptive response of this NCD to cope with the inhibition of nitrogen-fixation by molecular oxygen. Finally, the genomic signature of V. diazotrophicus is essentially absent from metagenomic data of Tara Ocean expeditions, despite having been isolated from various marine environments. We think that the genetically tractable V. diazotrophicus strain used in this study may serve as an ideal model to study the ecophysiology of these overlooked procaryotic group

Genetic and physiological insights into the diazotrophicactivity of a non-cyanobacterial marine diazotroph

International audienceNitrogen (N2) fixation, or diazotrophy, supports a large part of primary production in oceans. Culture-independent approaches highlighted the presence in abundance of marine non-cyanobacterial diazotrophs (NCD), but their ecophysiology remains elusive, mostly because of the low number of isolated NCD and because of the lack of available genetic tools for these isolates. Here, a dual genetic and functional approach allowed unveiling the ecophysiology of a marine NCD affiliated to the species Vibrio diazotrophicus. Physiological characterization of the first marine NCD mutant obtained so far was performed using a soft-gellan assay, demonstrating that a ΔnifH mutant is not able to grow in nitrogen-free media. Furthermore, we demonstrated that V. diazotrophicus produces a thick biofilm under diazotrophic conditions, suggesting biofilm production as an adaptive response of this NCD to cope with the inhibition of nitrogen fixation by molecular oxygen. Finally, the genomic signature of V. diazotrophicus is essentially absent from metagenomic data of Tara Ocean expeditions, despite having been isolated from various marine environments. We think that the genetically tractable V. diazotrophicus strain used in this study may serve as an ideal model to study the ecophysiology of these overlooked procaryotic group

Th cell subset distribution is altered in SLE patients.

<p>Th cell subset frequencies were determined according to the expression of CXCR3 and CCR6 on CD4⁺CD45RA^-CXCR5^- T cells. The frequency of each Th cell subset was defined in samples from inactive SLE patients (n = 13), active SLE patients (n = 6) and sex and age-matched healthy controls (n = 19). Results are expressed as the mean ± sem. *<i>p</i> < 0.05, **<i>p</i> < 0.01, ***<i>p</i> < 0.001 (one-way ANOVA test). ns: not significant.</p

IgE levels are increased in the serum of SLE patients.

<p>The quantification of IgG and IgE levels in the serum of inactive SLE patients (n = 17), active SLE patients (n = 6) and sex and age-matched healthy controls (n = 21) were determined by a sandwich ELISA assay. Results are expressed as the mean IgG concentration (g/L; A) or IgE concentration (ng/ml; B) from duplicate wells and correlation between IgG levels and T_FH2 cell frequency is represented (A). Each data point represents an individual subject and horizontal lines show the mean ± sem. * <i>p</i> < 0.05, **<i>p</i> < 0.01 (unpaired Student’s t test and Pearson correlation coefficient). ns: not significant. ANA IgE in sera were detected using Hep-2 cells as substrate (C). Compared to healthy individual sera (HC, shown as an example), some IgG-depleted sera from lupus patients (4/12) (SLE, 1 out of 4 representative picture) yielded homogeneous nuclear staining.</p

Circulating T_FH Subset Distribution Is Strongly Affected in Lupus Patients with an Active Disease

<div><p>Follicular helper T cells (T_FH) represent a distinct subset of CD4⁺ T cells specialized in providing help to B lymphocytes, which may play a central role in autoimmune diseases having a major B cell component such as systemic lupus erythematosus. Recently, T_FH subsets that share common phenotypic and functional characteristics with T_FH cells from germinal centers, have been described in the peripheral blood from healthy individuals. The aim of this study was to analyze the distribution of such populations in lupus patients. Circulating T_FH cell subsets were defined by multicolor flow cytometry as T_FH17 (CXCR3^-CCR6⁺), T_FH1 (CXCR3 ⁺ CCR6^-) or T_FH2 (CXCR3^-CCR6^-) cells among CXCR5 ⁺ CD45RA^-CD4⁺ T cells in the peripheral blood of 23 SLE patients and 23 sex and age-matched healthy controls. IL-21 receptor expression by B cells was analyzed by flow cytometry and the serum levels of IL-21 and Igs were determined by ELISA tests. We found that the T_FH2 cell subset frequency is strongly and significantly increased in lupus patients with an active disease (SLEDAI score>8), while the T_FH1 cell subset percentage is greatly decreased. The T_FH2 and T_FH1 cell subset frequency alteration is associated with the presence of high Ig levels and autoantibodies in patient’s sera. Moreover, the T_FH2 cell subset enhancement correlates with an increased frequency of double negative memory B cells (CD27^-IgD^-CD19⁺ cells) expressing the IL-21R. Finally, we found that IgE levels in lupus patients’ sera correlate with disease activity and seem to be associated with high T_FH2 cell subset frequency. In conclusion, our study describes for the first time the distribution of circulating T_FH cell subsets in lupus patients. Interestingly, we found an increased frequency of T_FH2 cells, which correlates with disease activity. Our results suggest that this subset might play a key role in lupus pathogenesis.</p> </div

The frequency of memory B cells expressing IL-21R is enhanced in SLE patients and correlates with T_FH2 cell increase.

<p>IL-21 concentration was measured in the serum of SLE patients (n = 88) and healthy controls (n = 44) by a sandwich ELISA assay (A). Results are expressed as the mean IL-21 concentration (pg/ml) from duplicate wells. Each data point represents an individual subject and horizontal lines show the mean ± sem. Circulating B cell subsets were defined according to the expression of IgD and CD27 on CD19⁺IL-21R⁺ cells allowing the definition of 5 populations: Ab-secreting cells (ASC), non switched memory cells (NSmem), switched memory cells (Smem) and double negative memory cells (DNmem). Representative dot plots obtained with samples from one healthy control and one SLE patient are shown as an example (B). The distribution of each B cell subset among IL-21R⁺ cells in healthy controls (n = 14) and SLE patients (n = 14) is represented (C). The frequency of CD27⁺ memory (left) and DN memory (right) B cells expressing IL-21R is compared between healthy controls (n = 14) and SLE patients (n = 14). Correlation between the IL-21R⁺ DN memory cells and T_FH2 cell subset frequencies in SLE patients (n= 14) is represented (E). Each data point represents an individual subject; horizontal lines show the mean ± sem. *<i>p</i> < 0.05, **<i>p</i> < 0.01 (Mann-Whitney U test and Pearson correlation coefficient).</p

T_FH2 cell frequency is significantly increased in active SLE patients.

<p>Correlations between the SLEDAI score and the percentage of each T_FH cell subset from SLE patients (n = 19) are shown (A). Dot plots of CXCR3 and CCR6 expression on gated CD3⁺CD4⁺CD45RA^-CXCR5⁺ T cells from an healthy individual, an inactive SLE patient and an active patient are shown as examples (B). T_FH2 and T_FH1 cell subset frequencies were analyzed according to the presence or the absence of anti-dsDNA Abs (C). Each data point represents an individual subject; horizontal lines show the mean ± sem. *<i>p</i> < 0.05, **<i>p</i> < 0.01, ***<i>p</i> < 0.001 (unpaired Student’s t test and Spearman’s rank correlation test). ns: not significant.</p