Precipitation of greigite and pyrite induced by Thermococcales: an advantage to live in Fe- and S-rich environments?

Thermococcales, a major order of archaea inhabiting the iron- and sulfur-rich anaerobic parts of hydrothermal deep-sea vents, have been shown to rapidly produce abundant quantities of pyrite FeS2 in iron-sulfur-rich fluids at 85°C, suggesting that they may contribute to the formation of 'low temperature' FeS2 in their ecosystem. We show that this process operates in Thermococcus kodakarensis only when zero-valent sulfur is directly available as intracellular sulfur vesicles. Whether in the presence or absence of zero-valent sulfur, significant amounts of Fe3 S4 greigite nanocrystals are formed extracellularly. We also show that mineralization of iron sulfides induces massive cell mortality but that concomitantly with the formation of greigite and/or pyrite, a new generation of cells can grow. This phenomenon is observed for Fe concentrations of 5 mM but not higher suggesting that above a threshold in the iron pulse all cells are lysed. We hypothesize that iron sulfides precipitation on former cell materials might induce the release of nutrients in the mineralization medium further used by a fraction of surviving non-mineralized cells allowing production of new alive cells. This suggests that biologically induced mineralization of iron-sulfides could be part of a survival strategy employed by Thermococcales to cope with mineralizing high-temperature hydrothermal environments.

Diagnostic accuracy of a calibrated abdominal compression to predict fluid responsiveness in children.

BACKGROUND: Fluid administration to increase stroke volume index (SVi) is a cornerstone of haemodynamic resuscitation. We assessed the accuracy of SVi variation during a calibrated abdominal compression manoeuvre (ΔSVi-CAC) to predict fluid responsiveness in children. METHODS: Patients younger than 8 yr with acute circulatory failure, regardless of their ventilation status, were selected. SVi, calculated as the average of five velocity-time integrals multiplied by the left ventricular outflow tract surface area, was recorded at four different steps: baseline, after an abdominal compression with a calibrated pressure of 25 mm Hg, after return to baseline, and then after a volume expansion (VE) of 10 ml kg-1 lactated Ringer solution over 10 min. Patients were classified as responders if SVi variation after volume expansion (ΔSVi-VE) increased by at least 15%. RESULTS: The 39 children included had a median [inter-quartile range (IQR)] age of 9 [5-31] months. Twenty patients were fluid responders and 19 were non-responders. ΔSVi-CAC correlated with ΔSVi-VE (r=0.829; P<0.001). The area under the receiver operating characteristic curve (ROCAUC) was 0.94 [95% confidence interval (CI), 0.85-0.99]. The best threshold for ΔSVi-CAC was 11% with a specificity of 95% [95% CI, 84-100] and a sensitivity of 75% [95% CI, 55-95]. ROCAUC of respiratory variation of IVC diameter (ΔIVC) was 0.53 [95% CI, 0.32-0.72]. CONCLUSION: ΔSVi-CAC during abdominal compression was a reliable method to predict fluid responsiveness in children with acute circulatory failure regardless of their ventilation status. CLINICAL TRIALS REGISTRATION: CPP Lyon sud est II: n° ANSM 2015-A00388-41 Clinicaltrial.gov: NCT02505646.

Marine records reveal multiple phases of Toba's last volcanic activity.

The Indonesian Young Toba Tuff (YTT), classically dated around 74 ka BP, is considered as a short-lived explosive cataclysmic super-eruption. The huge amounts of ash and SO2 emitted are likely to have triggered a volcanic winter which accelerated the transition to the last glaciation, and may have induced a human genetic bottleneck. However, the global climatic impact of the YTT or its duration are hotly debated. The present work offers a new interpretation of the Toba volcanic complex eruptive history. Analysing the BAR94-25 marine core proximal to the Toba volcanic center and combining it with high-resolution tephrostratigraphy and δ18O stratigraphy, we show that the Toba complex produced a volcanic succession that consists of at least 17 distinct layers of tephra and cryptotephra. Textural and geochemical analyses show that the tephra layers can be divided in 3 main successive volcanic activity phases (VAP1 to VAP3) over a period of ~ 50 kyr. The main volcanic activity phase, VAP2, including the YTT, is likely composed of 6 eruptive events in an interval whose total duration is ~ 10 ka. Thus, we suggest that the eruptive model of the Toba volcano must be revised as the duration of the Toba volcanic activity was much longer than suggested by previous studies. The implications of re-estimating the emission rate and the dispersion of ashes and SO2 include global environmental reconstitutions, climate change modelling and possibly human migration and evolution.

Impact of flow and temperature on non-dyspnoeic dogs' tolerance undergoing high-flow oxygen therapy.

OBJECTIVES: To prospectively describe the impact of gas flow rate and temperature on dog's tolerance of high-flow nasal oxygen therapy during recovery from anaesthesia, hypothesizing that higher flow rates and temperatures will decrease tolerance. MATERIALS AND METHODS: Twelve non-dyspnoeic client-owned dogs recovering from general anaesthesia were included in this study. After extubation, a nasal cannula was positioned and high-flow nasal oxygen therapy was initiated. Two flow rates (two or four time the theoretical minute ventilation: HF2 and HF4), each of them combined with two temperatures (31 and 37°C: T31 and T37), were randomly applied (four conditions per dog). For each condition, cardiovascular and respiratory parameters (heart rate, respiratory rate, systolic arterial blood pressure and pulse oximeter oxygen saturation), sedation score and tolerance score were recorded at initiation (T0 ) and after 10 minutes of accommodation (T10 ). RESULTS: Sedation scores were not significantly different between the four conditions. Cardiovascular and respiratory parameters were not significantly different between any condition at both T0 and T10 . Tolerance scores were good and not significantly different between any flow rate or temperature (HF2-T31: 4 (2-4), HF4-T31: 4 (2-4), HF2-T37: 4 (2-4), HF4-T37: 4 (1-4)). CLINICAL SIGNIFICANCE: The gas flow rates and temperatures studied have no impact on tolerance during the recovery period of non-dyspnoeic dogs, and high-flow nasal cannula is well tolerated. Further studies are required to confirm these results in dyspnoeic dogs.

Influence of cardiac hemodynamic variables on retinal vessel density measurement on optical coherence tomography angiography in patients with myocardial infarction.

INTRODUCTION: Quantitative measurements of retinal microvasculature by optical coherence tomography angiography (OCT-A) have been used to assess cardiovascular risk profile. However, to date, there are no studies focusing on OCT-A imaging in the setting of the altered hemodynamic status found in high-risk cardiovascular patients. METHODS: To determine the potential association between retinal vascular density on OCT-A and a comprehensive battery of hemodynamic variables in patients with myocardial infarction (MI) using data from the acute phase and at 3 months follow-up after cardiac rehabilitation. This prospective longitudinal study included patients who presented with MI in the cardiology intensive care unit at Dijon University Hospital. Main outcomes and measurements were retinal vessel density on OCT-A, hemodynamic status based on left ventricular ejection fraction (LVEF), and indexed cardiac output during the acute phase of myocardial infarction and at 3 months follow-up. RESULTS: Overall, 30 patients were included in this pilot study. The median (IQR) age was 64 years (55-71) with 87% men. At admission, the mean (SD) LVEF was 53% (11), and the mean indexed cardiac output was 2.70 (0.83) L/min/m2. On OCT-A, the mean inner retinal vascular density was 19.09 (2.80) mm-1. No significant association was found between retinal vascular density and hemodynamic variables. CONCLUSION: We found no significant association between retinal vascular density on OCT-A and hemodynamic variables in the acute phase of a myocardial infarction or after 3 months of cardiac rehabilitation. Therefore, OCT-A findings do not seem to be influenced by the hemodynamic changes associated with myocardial infarction.

[Intra cellular trafficking of influenza virus proteins]. / Le trafic nucléocytoplasmique des protéines et des ribonucléoprotéines du virus de la grippe.

Influenza virus is a negative strand RNA virus and is one of the rare RNA viruses to replicate in the nucleus. The viral RNA is associated with 4 viral proteins to make ribonucleoprotein particles (RNPs). After cell entry the RNPs are dissociated from the viral matrix protein in the low pH of the endosome and are actively imported into the cell nucleus. After translation of viral mRNAs, the proteins necessary for the assembly of new RNPs (the nucleoprotein and the three subunits of the polymerase complex) are also imported into the nucleus. Apart from these four proteins, part of the newly made matrix protein is also imported and the NEP (nuclear export protein) enters the nucleus probably through diffusion. The nuclear localisation signals on all these viral proteins and their interaction with the cellular transport system are discussed. In the nucleus, the matrix protein binds to the newly assembled RNPs and NEP then binds to the matrix protein. NEP contains the nuclear export signal necessary for transport of the RNPs to the cytoplasm for the budding of new virus particles. There appears to be a intricate ballet in exposing and hiding nuclear transport signals which leads to a unidirectional transport of the RNPs to the nucleus at the start of the infection process and an opposite unidirectional export of RNPs at the end of the infection.

[Treatment and progression of acute communautary osteoarticular infections in healthy children: A retrospective monocentric study of 64 patients]. / Traitement et évolution des infections ostéoarticulaires aiguës communautaires chez l'enfant sain : étude rétrospective monocentrique de 64 cas.

INTRODUCTION: The prognosis of osteoarticular infections has improved over the past 20 years but it still remains potentially severe. The treatment of these infections has been simplified and shortened. In 2008, the Pediatric Infectious Disease Group (GPIP) established new therapeutic guidelines in order to standardize treatment in France. The aim of this study is to analyze practices in a Parisian hospital and assess the efficacy of this treatment in short and medium terms. MATERIALS AND METHODS: This retrospective study focused on patients older than 3 months, without comorbidities, who were hospitalized for an acute osteoarticular infection in 2012 at Trousseau Hospital (Paris), with a follow-up of at least 4 weeks. The patients were selected from the hospital register. RESULTS: The study included 64 patients of 156, who were admitted for examination with a diagnosis of acute osteoarticular infection, bacteriologically confirmed (29/64) or presumed on the basis of bacteriological evidence (35/64). The median age of the patients was 22 months. Of the patients, 78 % were febrile; 35 patients had arthritis (54.7 %), 21 osteomyelitis (32.8 %), seven osteoarthritis (10.9 %), and one spondylitis. Preferential localizations were the knees and hips; 61 % of arthritis cases were diagnosed with ultrasound, 54 % of osteomyelitis cases with scintigraphy. The two main microorganisms found were Kingella kingae (62.1 %) and Staphylococcus aureus (24.1 %). In 98 % of cases, patients were treated by cefamandole, with or without gentamicine, for a median duration of 3 days (1-10) intravenously, with oral relay by amoxicillin-clavulanic acid, for a total duration of 6 weeks, but in association with rifampicin in 40 % of cases without explanation. The median follow-up was 13 weeks, with a treatment success rate of 86 %. CONCLUSION: The study of local practices showed us that the GPIP guidelines are not followed, with the duration of oral treatment being too long. The trend in therapy is toward short treatments of 10-20 days, with a shorter intravenous phase.

[Effectiveness of management of children with severe trauma in a pediatric trauma center and in an adult trauma center: A before-after study]. / Efficacité de la prise en charge des traumatismes graves de l'enfant en salle d'accueil des urgences vitales adulte ou pédiatrique : une étude avant-après.

OBJECTIVE: To compare management of injured children in an adult trauma center (TC) with competencies in pediatric trauma care (2005-2007) and in a pediatric-only trauma center (2010-2012). STUDY DESIGN: A before-after retrospective study. PATIENTS AND METHODS: Fifty-nine children between 1 and 15 years of age admitted to the adult TC (2005-2007) were compared to 56 children admitted to the pediatric TC (2010-2012). Epidemiological data, severity scores, early outcome, and care duration in trauma resuscitation before whole-body CT were collected and compared between the two periods. RESULTS: This study found no significant differences between the two periods in terms of care duration before the whole-body CT scan (28 min [18-40] vs 26.5 min [21-36], P=0.89) and early mortality (eight children [13.5%] vs ten children [17.8%], P=0.35). CONCLUSION: With no differences in early management of injured children demonstrated, this study validates the organization within our pediatric trauma center. The effectiveness of management of children between 1 and 15 years of age with severe trauma seems to be similar in the two contexts.

Involvement of "hinge" nucleotides of Xenopus laevis 5 S rRNA in the RNA structural organization and in the binding of transcription factor TFIIIA.

Nucleotides in the bifurcation region of the 5 S rRNA, the junction of the three helical domains, play a central role in determining the coaxial stacking interactions and tertiary structure of the RNA. We have used site-directed mutagenesis of Xenopus laevis oocyte 5 S rRNA to make all possible nucleotide substitutions at three positions in loop A (10, 11 and 13) and at the G66.U109 base-pair at the beginning of helix V. Certain double point mutations were constructed to ascertain the relationship between loop A nucleotides and the G.U base-pair. The importance of the size of the bifurcation region was tested by the creation of a single nucleotide deletion mutant and two single nucleotide insertion mutants. The effects of these mutations on the structure and function of the 5 S rRNA were determined by solution structure probing of approximately half of the mutants with chemical reagents, and by measuring the relative binding affinity of each mutant for transcription factor TFIIIA. Proposed structural rearrangements in the bifurcation region were tested by using a graphic modeling method combining stereochemical constraints and chemical reactivity data. From this work, several insights were obtained into the general problem of helix stacking and RNA folding at complex bifurcation regions. None of the mutations caused an alteration of the coaxial stacking of helix V on helix II proposed for the wild-type 5 S rRNA. However, the formation of a Watson-Crick pair between nucleotide 13 of loop A and nucleotide 66 at the top of helix V does cause a destabilization of the proximal part of this helix. Also, nucleotide 109 at the top of helix V will preferentially pair with nucleotide 10 of loop A rather than nucleotide 66 when both possibilities are provided, without affecting the stability of helix V, even though the G.U pair is disrupted. The effects of these mutations on TFIIIA binding indicate that the bifurcation region is critical for protein recognition. One important feature of the relationship between 5 S rRNA structure and TFIIIA recognition resulting from this study was the observation that any mutation that constrains the bifurcation loop results in a reduced affinity of the RNA for TFIIIA, unless it is compensated for by an increased flexibility elsewhere.

Functional sites in the 5' region of human immunodeficiency virus type 1 RNA form defined structural domains.

The 5' region of HIV-1 RNA contains functional elements involved in key steps of the retroviral cycle, such as genomic RNA transcription, splicing, translation, dimerization or initiation of reverse transcription. In the present work, we investigated the conformation of the first 500 nucleotides covering the RNA leader and the 5' gag coding sequences of HIV-MAL, using chemical probing. We provide detailed information on almost each nucleotide at one of their Watson-Crick positions and on position N-7 of purines. Experiments were conducted on two in vitro transcribed RNA fragments (1 to 707 and 1 to 311). A secondary structure model was derived by combining the experimental data, computer predictions and sequence comparison. Under conditions favoring dimerization (high salt concentration), HIV-1 RNA folds into independent structural domains that can be related to defined functional regions. The first domain corresponds to TAR forming a stable stem-loop. Intrinsic structural features are found to stabilize the TAR hairpin loop. The second domain (nucleotides 56 to 299) contains the PBS sequence, which is located in a stable subdomain constrained by a four stem junction (nucleotides 139 to 218). Although the MAL isolate has an insertion near the PBS, probably resulting from the duplication of a 23-nucleotide sequence, the structural organization of this subdomain is conserved in all other HIV-1 isolates. The third domain (nucleotides 300 to 404) contains the splice donor site, packaging and dimerization elements and the AUG initiation codon of gag. A major result is the structural versatility of this region. Two mutually exclusive structures, both equally in agreement with probing data, could modulate the different functions involving this domain. The reduced accessibility of the gag translational initiation site possibly accounts for the low efficiency of the in vitro translation of the dimer. Finally, the 5' gag coding sequences form a metastable domain.

Functional determinants of the Epstein-Barr virus protease.

Herpesvirus proteases are essential for the production of progeny virus. They cleave the assembly protein that fills the immature capsid in order to make place for the viral DNA. The recombinant protease of the human gamma-herpesvirus Epstein-Barr virus (EBV) was expressed in Escherichia coli and purified. Circular dichroism indicated that the protein was properly folded with a secondary structure content similar to that of other herpesvirus proteases. Gel filtration and sedimentation analysis indicated a fast monomer-dimer equilibrium of the protease with a K(d) of about 60 microM. This value was not influenced by glycerol but was lowered to 1.7 microM in the presence of 0.5 M sodium citrate. We also developed an HPLC-based enzymatic assay using a 20 amino acid residue synthetic peptide substrate derived from one of the viral target sequences for the protease. We found that conditions that stabilised the dimer also led to a higher enzymatic activity. Through sequential deletion of amino acid residues from either side of the cleavage site, the minimal peptide substrate for the protease was determined as P5-P2'. This minimal sequence is shorter than that for other herpesvirus proteases. The implications of our findings are discussed with reference to the viral life-cycle. These results are the first ever published on the EBV protease and represent a first step towards the development of protease inhibitors.

Structural studies on site-directed mutants of domain 3 of Xenopus laevis oocyte 5 S ribosomal RNA.

Base substitutions have been introduced into the highly conserved sequences of loops D and E within domain 3 of Xenopus laevis oocyte 5 S rRNA. The effects of these mutations on the solution structure of this 5 S rRNA have been studied by means of probing with nucleases, and with chemical reagents under native and semi-denaturing conditions. The data obtained with these mutants support the graphic model of Xenopus oocyte 5 S rRNA proposed by Westhof et al. In particular, our results rule out the existence of long-range base-pairing interactions between loop C and either loop D or loop E. The data also confirm that loops D and E in the wild-type 5 S RNA adopt unusual secondary structures and illustrate the importance of nucleotide sequence in the formation of intrinsic local loop conformations via non-canonical base-pairs and specific base-phosphate contacts. Consistent with this conclusion is our observation that the domain 3 fragment of Xenopus oocyte 5 S rRNA adopts the same conformation as the corresponding region in the full-length 5 S rRNA.

Ribosomal 5S RNA from Xenopus laevis oocytes: conformation and interaction with transcription factor IIIA.

This review describes extensive studies on 5S rRNA from X laevis oocytes combining conformational analyses in solution (using a variety of chemical and enzymatic probes), computer modeling, site-directed mutagenesis, crosslinking and TFIIIA binding. The proposed 3-dimensional model adopts a Y-shaped structure with no tertiary interactions between the different domains of the RNA. The conserved nucleotides are not crucial for the tertiary folding but they maintain an intrinsic structure in the loop regions. The model was tested by the analysis of several 5S rRNA mutants. A series of 5S RNA mutants with defined block sequence changes in regions corresponding to each of the loop regions was constructed by in vitro transcription of the mutated genes. Our results show that none of the mutations perturbs the Y-shaped structure of the RNA, although they induce conformational changes restricted to the mutated regions. The interaction of the resulting 5S rRNA mutants with TFIIIA was determined by a direct binding assay. Only the mutations in the hinge region between the 3 helical domains have a significant effect on the binding for the protein. Finally, TFIIIA was crosslinked by the use of trans-diamminedichloroplatinum (II) to a region covering the fork region. Our results show that (i) the tertiary structure does not involve long-range interactions; (ii) the intrinsic structures in loops are strictly sequence-dependent; (iii) the hinge nucleotides govern the relative orientation of the 3 helical domains; (iv) TFIIIA recognizes essentially specific features of the tertiary structure of 5S rRNA.

Higher-order structure of domain III in Escherichia coli 16S ribosomal RNA, 30S subunit and 70S ribosome.

We have investigated in detail the conformation of domain III of 16S rRNA (nucleotides 913-1408), using a variety of chemical and enzymatic structure probes. The sites of reaction were identified by primer extension with reverse transcriptase using appropriate oligodeoxyribonucleotide primers. This study has been done on 16S rRNA in its naked form, in the 30S subunit and in the 70S ribosome. Data obtained with naked RNA broadly confirm the secondary structure model proposed essentially by comparative sequence analysis, and allow identification of nucleotides involved in tertiary interactions. Our results are in reasonably good agreement with structure probing experiments of Moazed et al. [1]. However, several discrepancies have been observed. Within the 30S subunit, a high number of nucleotides become unreactive whereas other nucleotides show an enhanced reactivity. This probably reflects local conformational changes. Interestingly, they are located in strategic regions of the RNA, e.g. around C1400 (involved in tRNA binding) and C1192 (involved in spectinomycin recognition). Results are also discussed together with the topographical localization of the ribosomal proteins in this area. The study on the 70S particle allows identification of regions at the interface of subunits or exposed at the surface of the ribosome.

Multidrug-resistant bacteria infection control: study of compliance with isolation precautions in a Paris university hospital.

Isolation practices in a university hospital were analyzed for 137 patients with multidrug-resistant bacteria. Isolation was ordered in writing by physicians for 40% and instituted by nurses for 60%; 74% were isolated. Compliance depended on physician ordering in writing (odds ratio, 36.3; 95% confidence interval, 4.8-274.9). Nurses complied best with hand washing.

Use of lead(II) to probe the structure of large RNA's. Conformation of the 3' terminal domain of E. coli 16S rRNA and its involvement in building the tRNA binding sites.

The present work shows that lead(II) can be used as a convenient structure probe to map the conformation of large RNA's and to follow discrete conformational changes at different functional states. We have investigated the conformation of the 3' domain of the E. coli 16S rRNA (nucleotides 1295-1542) in its naked form, in the 30S subunit and in the 70S ribosome. Our study clearly shows a preferential affinity of Pb(II) for interhelical and loop regions and suggests a high sensitivity for dynamic and flexible regions. Within 30S subunits, some cleavages are strongly decreased as the result of protein-induced protection, while others are enhanced suggesting local conformational adjustments. These rearrangements occur at functionally strategic regions of the RNA centered around nucleotides 1337, 1400, 1500 and near the 3' end of the RNA. The association of 30S and 50S subunits causes further protections at several nucleotides and some enhanced reactivities that can be interpreted in terms of subunits interface and allosteric transitions. The binding of E. coli tRNA-Phe to the 70S ribosome results in message-independent (positions 1337 and 1397) and message-dependent (1399-1400, 1491-1492 and 1505) protections. A third class of protection (1344-1345, 1393-1395, 1403-1409, 1412-1414, 1504, 1506-1507 and 1517-1519) is observed in message-directed 30S subunits, which are induced by both tRNA binding and 50S subunit association. This extensive reduction of reactivity most probably reflects an allosteric transition rather than a direct shielding.

[Cytomegalovirus infections in patients after liver transplantation: determination of risk groups]. / Infections à cytomégalovirus chez les transplantés hépatiques: détermination de groupes à risque.

The goal of this study was to identify high-risk groups for cytomegalovirus infection after liver transplantation. Sixty-one patients were evaluated. Twenty-five patients (41 percent) had infection. Among the 16 patients who were seronegative for the virus before transplantation, 11 received a liver graft and blood products from seronegative donors and none of them developed infection. All seronegative recipients of a liver from seropositive donors (5/5) developed primary infection. Among the 45 patients seropositive before transplantation, 20 developed a cytomegalovirus infection, whatever the donor serologic status. The incidence of symptomatic reactivation or reinfection was high (14/20), and, for 12/14 of them, associated with early acute rejection. Two high-risk groups of patients, eligible for cytomegalovirus prophylaxis, were identified: seronegative recipients of seropositive donors and seropositive recipients with early acute rejection.

[Acute hepatic artery thrombosis in pediatric liver transplantation: surgical thrombectomy and in situ fibrinolysis]. / Thromboses aiguës de l'artère hépatique en transplantation hépatique pédiatrique: désobstruction chirurgicale et fibrinolyse in situ.

In pediatric liver transplantation, hepatic artery thrombosis usually leads to graft loss, early due to hepatic necrosis when it occurs during the first week following the transplant procedure, or later due to biliary complications. Liver retransplantation is the usual attitude. However, urgent surgical hepatic arterial thrombectomy to restore the blood flow can be successful when early diagnosis is made with Doppler ultrasound examination and angiography. Four hepatic arterial thrombectomies were performed as an emergency with additional intra-hepatic arterial fibrinolytic treatment, in three children, 1.5, 3 and 5.5 years of age. Mean duration between the first signs of hepatic artery thrombosis and thrombectomy was 16 hours. None of the children had an urgent liver retransplantation. A complete success was obtained in one case, with normal liver function tests and patent hepatic artery on the Doppler ultrasound examination at the present time. In the two other cases, hepatic artery thrombosis recurred, in spite of repeated thrombectomy in one case; following this attempt complications of hepatic artery thrombosis occurred in the two patients: ischemic necrosis of the left lobe (1 case), biliary leak (1 case) and stenosis of the common bile duct (2 cases). A complete success in one case and a partial success in the two others lead us to advocate urgent thrombectomy and in situ fibrinolytic treatment when early diagnosis of hepatic artery thrombosis is made.