Search

We present a search for neutrino induced events containing a single, exclusive photon using data from the NOMAD experiment at the CERN SPS where the average energy of the neutrino flux is ≃25 GeV. The search is motivated by an excess of electron-like events in the 200–475 MeV energy region as reported by the MiniBooNE experiment. In NOMAD, photons are identified via their conversion to e+e− in an active target embedded in a magnetic field. The background to the single photon signal is dominated by the asymmetric decay of neutral pions produced either in a coherent neutrino–nucleus interaction, or in a neutrino–nucleon neutral current deep inelastic scattering, or in an interaction occurring outside the fiducial volume. All three backgrounds are determined in situ using control data samples prior to opening the ‘signal-box’. In the signal region, we observe 155 events with a predicted background of 129.2±8.5±3.3. We interpret this as null evidence for excess of single photon events, and set a limit. Assuming that the hypothetical single photon has a momentum distribution similar to that of a photon from the coherent π0 decay, the measurement yields an upper limit on single photon events, <4.0×10−4 per νμ charged current event. Narrowing the search to events where the photon is approximately collinear with the incident neutrino, we observe 78 events with a predicted background of 76.6±4.9±1.9 yielding a more stringent upper limit, <1.6×10−4 per νμ charged current event., Physics

Recent evidence showed that in response to elevated sodium dietary intakes, many body tissues retain Na + ions for long periods of time and can reach concentrations up to 200 mM. This could modulate the immune system and be responsible for several diseases. However, studies brought contrasted results and the effects of external sodium on human dendritic cell (DC) responses to danger signals remain largely unknown. Considering their central role in triggering T cell response, we tested how NaCl-enriched medium influences human DCs properties. We found that DCs submitted to high extracellular Na + concentrations up to 200 mM remain viable and maintain the expression of specific DC markers, however, their maturation, chemotaxis toward CCL19, production of pro-inflammatory cytokines and ROS in response to LPS were also partially inhibited. In line with these results, the T-cell allostimulatory capacity of DCs was also inhibited. Finally, our data indicate that high NaCl concentrations triggered the phosphorylation of SGK1 and ERK1/2 kinases. These results raised the possibility that the previously reported pro-inflammatory effects of high NaCl concentrations on T cells might be counterbalanced by a downregulation of DC activation., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Inc. All rights reserved.)

The induced membrane technique is now well accepted for reconstruction of segmental bone defect. On the other hand, some cases of aseptic non-union are unsuccessfully treated by several surgical attempts for obtaining bone healing. The two stages wrapping induced membrane technique was developed initially for treating atrophic and recalcitrant aseptic non union without bone loss. At the first stage, the site of non-union was firmly fixed and tiles of cement were placed close to the bone on two or three aspects of the bone. At the second stage, after removing the spacers, the induced cavities were filled with cancellous bone autograft. In the two reported cases bone healing was acquired in 4 months. One case was a recalcitrant atrophic non-union of the humeral shaft, the other case concerned the enhancement of an insufficient segmental reconstruction of the femur. The follow up were respectively 3 years and 2 years without complication. The membrane induced by the cement tiles prevents the bone graft resorption and improves the osteogenicity through its biological properties., (Copyright © 2020 Elsevier Masson SAS. All rights reserved.)

Hepatitis B virus (HBV) core protein (HBc) is essential to the formation of the HBV capsid. HBc contains two domains: the N-terminal domain corresponding to residues 1-140 essential to form the icosahedral shell and the C-terminal domain corresponding to a basic and phosphorylated peptide, and required for DNA replication. The role of these two domains for HBV capsid assembly was essentially studied in vitro with HBc purified from mammalian or non-mammalian cell lysates, but their respective role in living cells remains to be clarified. We therefore investigated the assembly of the HBV capsid in Huh7 cells by combining fluorescence lifetime imaging microscopy/Förster's resonance energy transfer, fluorescence correlation spectroscopy and transmission electron microscopy approaches. We found that wild-type HBc forms oligomers early after transfection and at a sub-micromolar concentration. These oligomers are homogeneously diffused throughout the cell. We quantified a stoichiometry ranging from ~170 to ~230 HBc proteins per oligomer, consistent with the visualization of eGFP-containingHBV capsid shaped as native capsid particles by transmission electron microscopy. In contrast, no assembly was observed when HBc-N-terminal domain was expressed. This highlights the essential role of the C-terminal domain to form capsid in mammalian cells. Deletion of either the third helix or of the 124-135 residues of HBc had a dramatic impact on the assembly of the HBV capsid, inducing the formation of mis-assembled oligomers and monomers, respectively. This study shows that our approach using fluorescent derivatives of HBc is an innovative method to investigate HBV capsid formation., (Copyright © 2020. Published by Elsevier Ltd.)

Background: Urinary tract infections (UTI) are the second cause of community-acquired bacterial infections in the elderly. Distinguishing symptomatic UTI from asymptomatic bacteriuria is problematic, as older adults are less likely to present with localized urinary symptoms. We evaluated characteristics of patients presenting UTI among elderly with sepsis. Moreover, we aimed to evaluate the sensibility and specificity of urine dipstick tests in the diagnosis of UTI in geriatric population., Patients and Method: We led a prospective, monocentric, observational study between April 2017 and January 2018. We included patients hospitalized in geriatric wards, who were prescribed urine culture for UTI symptoms or/and infection without primary sites for which a urine culture was prescribed. Dipstick urinalyses were performed for all patients. Clinical and biological characteristics of all patients were compared according to the final diagnosis of UTI. Moreover, results of dipstick tests were evaluated for the diagnosis of UTI in this population., Results: Among 165 patients, 67 (40.6 %) had a UTI and 98 (59.4 %) had another diagnosis. These two groups were comparable for age and daily-living activities. In the UTI group, the proportion of women was higher than in the other group (P<0.05), and mean MMSE score was lower (P<0.05). Positive urine dipstick test for leukocytes and/or nitrites had high sensitivity (92 %), but low specificity (50 %). Negative predictive value of this test was high (91 %)., Conclusion: For suspicion of UTI among elderly, few criteria are specific. Negative dipstick tests can suggest an absence of UTI due to its high negative predictive value., (Copyright © 2019 Société Nationale Française de Médecine Interne (SNFMI). Published by Elsevier Masson SAS. All rights reserved.)

Urinary tract infections (UTIs) in patients with neurogenic bladder are a major public health issue due to their high incidence and major consequences. Despite their frequency and potential severity, their physiopathology and management are poorly known. We provide a narrative literature review on the epidemiology, physiopathology, diagnostic criteria, microbiology, antimicrobial management, and prevention. UTIs among patients with neurogenic bladder are associated with high morbidity and healthcare utilization. Risk factors for UTI among this population are: indwelling catheter, urinary stasis, high bladder pressure, and bladder stones. Their diagnosis is a major challenge as clinical signs are often non-specific and rare. A urinary sample should be analyzed in appropriate conditions before any antibiotic prescription. According to most guidelines, a bacterial threshold≥103CFU/ml associated with symptoms is acceptable to define UTI in patients with neurogenic bladder. The management of acute symptomatic UTI is not evidence-based. A management with a single agent and a short antibiotic treatment of 10 days or less seems effective. Antibiotic selection should be based on the patient's resistance patterns. Asymptomatic bacteriuria should not be treated to avoid the emergence of bacterial resistance. Regarding preventive measures, use of clean intermittent catheterization, intravesical botulinum toxin injection, and prevention using antibiotic cycling are effective. Bacterial interference is promising but randomized controlled trials are needed. Large ongoing cohorts and randomized controlled trials should soon provide more evidence-based data., (Copyright © 2019 Elsevier Masson SAS. All rights reserved.)

HBc is a small protein essential for the formation of the icosahedral HBV capsid. Its multiple roles in the replication cycle make this protein a promising target for the development of antiviral molecules. Based on the structure of HBc, a series of HBV assembly inhibitors, also known as capsid assembly modulators, were identified. We investigated the effect of BAY 41-4109, a heteroaryldihydropyrimidine derivative that promotes the assembly of a non-capsid polymer. We showed, by confocal microscopy, that BAY 41-4109 mediated HBc aggregation, mostly in the cytoplasm of Huh7 cells. Image analysis revealed that aggregate size depended on BAY 41-4109 concentration and treatment duration. Large aggregates in the vicinity of the nucleus were enclosed by invaginations of the nuclear envelope. This deformation of the nuclear envelope was confirmed by transmission electron microscopy (TEM) and immuno-TEM. These two techniques also revealed that the HBc aggregates were accumulations of capsid-like shells with an electron-dense material consisting of HBV core fragments. These findings, shedding light on the ultrastructural organization of HBc aggregates, provide insight into the mechanisms of action of BAY 41-4109 against HBV and will serve as a basis for comparison with other HBV capsid assembly inhibitors., (Copyright © 2019. Published by Elsevier B.V.)

The perforators of the fibular artery provide a well vascularised supra fascial network which allows to raise a proximally or a distally based island fascio cutaneous flap with an adipo-fascial pedicle. We present a short series of five cases of this flap for coverage of soft tissue defects involving the region of the knee, the distal third of the leg and the lateral aspect of the heel. All flaps healed entirely without venous congestion. The advantages of the fascio cutaneous fibular island flap are the supine operative position, the preservation of the sural nerve and the lesser saphenous vein and a pivot point which can be located at the middle third of the leg. According to our experience, the fascio cutaneous fibular island flap is especially indicated for repairing defects of the distal leg., (Copyright © 2018 Elsevier Masson SAS. All rights reserved.)

Acromioclavicular (AC) joint arthropathy remains one of the most common causes of shoulder pain. In the case of AC joint arthropathy resistant to conservative treatment, most authors have recognized distal clavicle resection as the gold-standard treatment. However, some challenges remain to be solved. One is the difficulty in visualization of the superior and posterior part of the distal clavicle from the midlateral portal, causing an incomplete resection of the distal clavicle. This could potentially lead to unresolved pain and therefore surgical failure. We propose a technique for arthroscopic resection of the distal clavicle and the medial portion of the acromion, without any added portal: bipolar AC joint resection. The term "bipolar" is used because both the acromion and the clavicle are resected, without injuring the superior capsule.

Objectives: The proper use of antibiotics is a public health priority to preserve their effectiveness. Little data is available on outpatient antibiotic prescriptions, especially in the emergency department. We aimed to assess the quality of outpatient antibiotic prescriptions in our hospital., Patients and Methods: Retrospective monocentric study of antibiotic prescriptions written to adult patients managed at the emergency department without hospitalization (November 15th, 2012-November 15th, 2013). Prescriptions were evaluated by an infectious disease specialist and an emergency physician on the basis of local recommendations compiled from national and international guidelines., Results: A total of 760 prescriptions were reviewed. The most frequent indications were urinary tract infections (n=263; 34.6%), cutaneous infections (n=198; 26.05%), respiratory tract infections (n=101; 13.28%), and ENT infections (n=62; 8.15%). The most frequently prescribed antibiotics were fluoroquinolones (n=314; 40.83%) and amoxicillin-clavulanic acid (n=245; 31.85%). Overall, 455 prescriptions (59.86%) did not comply with guidelines. The main reasons for inadequacy were the absence of an indication for antibiotic therapy (n=197; 40.7%), an inadequate spectrum of activity, i.e. too broad, (n=95; 19.62%), and excessive treatment duration (n=87; 17.97%). Rates of inadequate prescriptions were 82.26% for ENT infections, 71.2% for cutaneous infections, 46.53% for respiratory tract infections, and 38.4% for urinary tract infections., Conclusion: Antibiotic prescriptions written to outpatients in the emergency department are often inadequate. Enhancing prescribers' training and handing out guidelines is therefore necessary. The quality of these prescriptions should then be re-assessed., (Copyright © 2016 Elsevier Masson SAS. All rights reserved.)

A large number of cystic fibrosis pathogens such as bacteria of the Burkholderia cepacia complex, Pseudomonas aeruginosa, or Mycobacterium abscessus are associated with complex therapeutic problems due to their inherent resistance to antibiotics. No vaccine is currently available against those pathogens. Vaccines are therefore crucial to combat these multidrug-resistant bacteria in specific clinical situations including cystic fibrosis. Various strategies may be considered to develop these vaccines. Similar virulence factors are expressed during the infection with various pathogens; they could thus be used as antigen to assess cross-protection. Many clinical trials are currently being conducted to try and develop a prophylactic treatment for patients presenting with cystic fibrosis., (Copyright © 2015 Elsevier Masson SAS. All rights reserved.)

Studies of the impact of parasites on host performance have mainly focused on body mass, a phenotypic trait that responds relatively slowly to the presence of parasites, and the expectedly faster response of physiological parameters has been mostly overlooked. We filled the gap by measuring the impact of endoparasites on four hematological/biochemical parameters (hematocrit, albumin, creatinine and fructosamine) in two contrasting free-living populations of roe deer. We generally found negative relationships between parasites and physiological parameters. Our findings also indicate little role of host sex on parasite impact and strongest parasite effects on young and senescent hosts., (Copyright © 2014 Elsevier B.V. All rights reserved.)

The in vitro activity of cefoxitin and imipenem was compared for 43 strains of the Mycobacterium abscessus complex, mostly isolated from cystic fibrosis patients. The MICs of imipenem were lower than those of cefoxitin, although the number of imipenem-resistant strains was higher according to the CLSI breakpoints. Strain comparisons indicated that the MICs of cefoxitin were significantly higher for Mycobacterium bolletii than for M. abscessus. The MICs of both β-lactams were higher for the rough morphotype than for the smooth morphotype. The clinical impact of the in vitro difference between the activity of imipenem and that of cefoxitin remains to be determined., (© 2013 The Authors Clinical Microbiology and Infection © 2013 European Society of Clinical Microbiology and Infectious Diseases.)

The potential role of a patient's resident microbial flora in the risk of acquiring multiresistant bacteria (MRB) during hospitalization is unclear. We investigated this role by cross-sectional study of 103 patients at risk of acquisition of Staphylococcus aureus (SA), resistant (MRSA) or not (MSSA) to methicillin, recruited in four French hospitals. The flora was analysed by an exhaustive culture-based approach combined with molecular and/or mass-spectrometry-based identification, and SA strain typing. Forty-three of the 53 SA-negative patients at entry were followed for up to 52 weeks: 19 (44.2%) remained negative for SA and 24 (55.8%) became positive, including 19 (79%) who acquired an MSSA, four (17%) who acquired an MRSA and one who acquired both (4%). Fifty-one different species were identified among the 103 patients, of which two, Corynebacterium accolens and Staphylococcus haemolyticus (p = 0.02-0.01), were more prevalent in the absence of SA. However, the same number of patients carrying or not these two species acquired an MSSA/MRSA during follow-up, regardless of antibiotic treatment received. Clustering analysis showed that the microbial flora was highly specific to each patient, and not predictive for acquisition of MSSA/MRSA or not. Patient-specific microbial resident flora is not predictive of SA acquisition., (© 2013 The Authors Clinical Microbiology and Infection © 2013 European Society of Clinical Microbiology and Infectious Diseases.)

We report a microbiological process for the documentation of prosthetic joint infection (PJI). Intraoperative periprosthetic tissue samples from 92 consecutive patients undergoing revision surgery for PJI were submitted to mechanized beadmill processing: specimens were aseptically collected in polypropylene vials, filled with sterile water and glass beads and submitted to mechanized agitation with a beadmill. The documentation rate of PJI following culture on solid and liquid media was 83.7% and the contamination rate 8.7%. Final documentation was obtained after overnight culture for 51.9% of cases and with 7 days of broth culture for all documented cases., (© 2010 The Authors. Clinical Microbiology and Infection © 2010 European Society of Clinical Microbiology and Infectious Diseases.)

Clinical-grade human mesenchymal stromal cells (MSCs) have been expanded in vitro for tissue engineering or immunoregulatory purposes without standardized culture conditions or release criteria. Although human MSCs show poor susceptibility for oncogenic transformation, 2 recent studies described their capacity to accumulate chromosomal instability and to give rise to carcinoma in immunocompromised mice after long-term culture. We thus investigated the immunologic and genetic features of MSCs expanded with fetal calf serum and fibroblast growth factor or with platelet lysate in 4 cell-therapy facilities during 2 multicenter clinical trials. Cultured MSCs showed a moderate expression of human leukocyte antigen-DR without alteration of their low immunogenicity or their immunomodulatory capacity. Moreover, some transient and donor-dependent recurring aneuploidy was detected in vitro, independently of the culture process. However, MSCs with or without chromosomal alterations showed progressive growth arrest and entered senescence without evidence of transformation either in vitro or in vivo.

Background Aims: Advances in bone tissue engineering with mesenchymal stromal cells (MSC) as an alternative to conventional orthopedic procedures has opened new horizons for the treatment of large bone defects. Bone marrow (BM) and trabecular bone are both sources of MSC. Regarding clinical use, we tested the potency of MSC from different sources., Methods: We obtained MSC from 17 donors (mean age 64.6 years) by extensive washing of trabecular bone from the femoral head and trochanter, as well as BM aspirates of the iliac crest and trochanter. The starting material was evaluated by histologic analysis and assessment of colony-forming unit-fibroblasts (CFU-F). The MSC populations were compared for proliferation and differentiation potential, at RNA and morphologic levels., Results: MSC proliferation potential and immunophenotype (expression of CD49a, CD73, CD90, CD105, CD146 and Stro-1) were similar whatever the starting material. However, the differentiation potential of MSC obtained by bone washing was impaired compared with aspiration; culture-amplified cells showed few Oil Red O-positive adipocytes and few mineralized areas and formed inconsistent Alcian blue-positive high-density micropellets after growth under adipogenic, osteogenic and chondrogenic conditions, respectively. MSC cultured with 1 ng/mL fibroblast growth factor 2 (FGF-2) showed better differentiation potential., Conclusions: Trabecular bone MSC from elderly patients is not good starting material for use in cell therapy for bone repair and regeneration, unless cultured in the presence of FGF-2.

Objective: To study the beta-lactamases content of Stenotrophomonas maltophilia strains and to evaluate the virulence potential of these strains with the in vivo Caenorhabditis elegans model., Methodology: From 1st January 2006 to 31st December 2006, a monitoring programme to study multidrug resistant Gram-negative bacteria including extended-spectrum beta-lactamases (ESBL)-producing S. maltophilia was conducted at Nîmes University Hospital and Perpignan Hospital. The ESBL production was confirmed by the double-disk synergy test using ceftazidime, cefotaxime and cefepime disks associated with clavulanic acid disk. The strains were characterized phenotypically (beta-lactamase[s] identification) and genotypically (pulsed-field gel electrophoresis, plasmid analysis) and evaluated for their virulence with the in vivo nematode C. elegans model (establishment of survival curves [LT50])., Results: Twelve ESBL-producing S. maltophilia strains were isolated in eight patients (median age: 65 years+/-19) mainly during skin infections (41.7%). The ESBL content revealed the presence of four CTX-M-15-producing strains at the same patient. The analysis by ECP confirmed that the four strains were identical. The plasmid analysis demonstrated that the plasmid carrying CTX-M-15 in the worldwide clonal Escherichia coli O25-ST131 strain and S. maltophilia were different. The C. elegans model confirmed that S. maltophilia strains presented a low virulence potential (LT50=4.5days+/-0.5 according to the strains and nematode death in 10days+/-1) whatever their resistance., Conclusion: For the first time in France, a CTX-M-15-producing S. maltophilia strain has been identified. The in vivo model confirmed that these bacteria have a low potential virulence. However, these strains were isolated from "immunocompromised" and multihospital patients demonstrating the necessary monitoring of these patients. The CTX-M after diffusing in hospitals and community in E. coli strains seem to spread in other Gram-negative bacteria.

The knockdown of a [FeFe]-hydrogenase-like gene in the model plants Medicago truncatula and Arabidopsis thaliana resulted in a mutant with a dwarf phenotype. Surprisingly, the phenotype is undistinguishable from wild type under hypoxic conditions. The heterologous expression of the plant gene in Escherichia coli indicates that the resulting protein probably coordinates two [Fe-S] clusters with different magnetic properties. Sequence alignment analysis indicates that these two clusters would be topologically equivalent to the mesial and proximal [Fe-S] centers of [FeFe]-hydrogenases. A possible role of the gene product in oxygen signaling pathways is discussed.

Guillain-Barre syndrome (GBS) is a rare disease triggered by postinfectious mechanisms. The disease concerns all ages, and is widely distributed around the world. The principal risks are respiratory failure, especially during the initial phase of the disease, and persisting deficit at long term. Among the infectious known agents, Campylobacter jejuni and CMV represent more than 40% of GBS causes. The clinical presentation, and the long-term prognosis of GBS related to these two etiologies are different. The physiopathological mechanisms of the nervous attack are probably also different. There is no proof, at this time, that anti-infectious treatment can improve the prognosis. The treatment is based on the early use of immunomodulatory treatments like intravenous immunoglobulins or plasma exchanges.

Aim of the Study: We aimed to study prevalence and features of Campylobacter jejuni and cytomegalovirus (CMV)-associated Guillain-Barré syndromes (GBS) in a French care unit., Patients and Methods: We studied 264 patients with GBS admitted at Raymond Poincaré hospital (Garches) between 1996 and 2001. Clinical data were obtained prospectively. Sera were collected at patients entry and tested retrospectively for anti-C. jejuni, anti-CMV and antigangliosides GM1 et GM2 antibodies., Results: GBS associated with a serological evidence for a recent C. jejuni infection were the more frequent (58/264, 22%); they affected predominantly men of mature years (mean age: 51.3 years; sex-ratio M/F: 1.76), mostly after a gastrointestinal illness (52%); they were often pure motor forms (57%), were severe (mechanical ventilation: 40%) and associated to an anti-GM1 IgG and/or IgM response (44%). GBS cases involving a primary CMV infection were less frequent (40/264, 15%), but were severe too (mechanical ventilation: 37.5%); they occurred preferentially in young women (mean age: 35.9 years; sex-ratio MF: 0.82), often after respiratory tract symptoms (28%) or an influenza-like syndrome (15%) and were frequently associated with sensory loss (73%) and with an anti-GM2 IgM response (47%)., Conclusion: C. jejuni and CMV proved to be major triggering agents of GBS in France. They are associated with distinct presentations, which are both severe.

Patients with cystic fibrosis are particularly at risk of infection with non-tuberculous mycobacteria (NTM). Prevalence of these infections increases with age to around 15 %. The main species involved are M. abscessus and M. avium, the latter not found in children under 15. Diagnosis relies on clinical, radiological and above all bacteriological criteria defined by the ATS. Identification of the causal species of NTM is essential and requires genetic techniques, some of which are currently evaluated. Treatment depends on the mycobacterial species. For M. avium, combined therapy with rifampicin, clarithromycin and ethambutol must be extended 12 months after negativation. M. abscessus infection is particularly resistant to therapy. Usual treatment is a one month course of intravenous imipenem or cefoxitin plus amikacin followed by oral clarithromycin plus ethambutol for at least 12 months after negativation. In case of local lesions, surgery is an option.

Objectives: The objective of this study was to evaluate the epidemiology of antibiotic-resistant bacteria among motor impaired patients admitted to an acute rehabilitation unit., Methods: From January 2000 to December 2002, the acute rehabilitation units of R. Poincare Hospital have screened patients for methicillin-resistant Staphylococcus aureus (MRSA) and extended-spectrum beta-lactamase enterobacteria (ESBL-EB) carriage by nasal and rectal swab at admission, every month and exit., Results: Finally, MRSA was isolated form screening or diagnosis samples of 360 patients and ESBL-EB from screening or diagnosis samples of 170 patients, corresponding respectively to an incidence of 3.6 for 1000 days of hospitalization (DH) and 1.7 for 1000 DH. 66% (236/360) of MRSA carriers and 58% of ESBL-EB carriers were identified only by screening samples. Carriage origin was identified for year 2002: Cases were imported for 40% (26/65) of MRSA carriers and 43% (18/42) of ESBL-EB carriers. The median acquisition delays were of 31 days [3-154] for MRSA and 19 days [3-317] for ESBL-EB., Conclusion: This allowed to set up contact precautions for more than 2 fold patients that would have allowed diagnosis samples alone.

This study evaluated the possible advantages provided by a genotypic method over commercially available biochemical systems for the identification of clinical isolates of coagulase-negative staphylococci (CNS). Partial sequencing of the sodA gene was performed for 168 coagulase-negative clinical isolates of staphylococci identified previously with the ID32 STAPH system. Of these, 101 (60.1%) were identified to the species level with ID32 STAPH, while 67 (39.9%) were misidentified or not identified with certainty. Sequencing of sodA proved useful for resolving all ambiguities or inconclusive identifications generated by the commercially available biochemical identification system.

We used partial 16S rRNA gene (16S DNA) sequencing for the prospective identification of nonfermenting Gram-negative bacilli recovered from patients attending our cystic fibrosis center (hôpital Necker-Enfants malades), which gave problematic results with conventional phenotypic tests. During 1999, we recovered 1093 isolates of nonfermenting Gram-negative bacilli from 702 sputum sampled from 148 patients. Forty-six of these isolates (27 patients) were not identified satisfactorily in routine laboratory tests. These isolates were identified by 16S DNA sequencing as Pseudomonas aeruginosa (19 isolates, 12 patients), Achromobacter xylosoxidans (10 isolates, 8 patients), Stenotrophomonas maltophilia (9 isolates, 9 patients), Burkholderia cepacia genomovar I/III (3 isolates, 3 patients), Burkholderia vietnamiensis (1 isolate), Burkholderia gladioli (1 isolate) and Ralstonia mannitolilytica (3 isolates, 2 patients). Fifteen isolates (33%) were resistant to all antibiotics in routine testing. Sixteen isolates (39%) resistant to colistin were recovered on B. cepacia-selective medium: 2 P. aeruginosa, 3 A. xylosoxidans, 3 S. maltophilia and the 8 Burkholderia--Ralstonia isolates. The API 20NE system gave no identification for 35 isolates and misidentified 11 isolates (2 P. aeruginosa, 2 A. xylosoxidans and 1 S. maltophilia classified as B. cepacia ). Control measures and/or treatment were clearly improved as a result of 16S DNA sequencing in three of these cases. This study confirms the weakness of phenotypic methods for identification of atypical nonfermenting Gram-negative bacilli recovered from cystic fibrosis patients. The genotypic methods, such as 16S DNA sequencing which allows identification of strains in routine practice, appears to have a small, but significant impact on the clinical management of CF patients.

Objective: Improvement of neurosurgical techniques with a more detailed description of brain tumors and their functional environment., Methods: We performed: (1) anatomical magnetic resonance imaging (MRI) for reference, (2) functional sequences dedicated to the adjacent cortical structures (sensorimotor, visual, language paradigms), and (3) thallium 201 cerebral tomoscintigraphy to visualize active tumor invasion. Data were transferred to a workstation for automatic registration., Results: All data were combined into one synthetic image showing the foci of high proliferative activity, which have to be completely resected, and the peritumoral functional structures, which have to be spared in order to minimize postoperative sequelae. This trimodal image is entered into a surgical neuronavigation computer for preoperative planning in order to outline tumoral target and functional risk areas. All this information is displayed in the operative microscope (Zeiss MKM) optically linked to MR images. This multimodality technique diminishes operative time by reducing electrocorticography and improves the operative short-term outcome., Conclusion: Multimodal imaging is useful for optimization of neurosurgical tumor resection.

18 FDG positon emission tomography provides metabolic images and allows better local and metastatic staging than radiologic methods. A best cartography of node involvement and a best delineation of the tumor zone should allow an optimal radiotherapy. Lung cancer is a good example of the interest of this new method.

The optimisation of the field of irradiation is the aim of the radiotherapist. Pet-Scan provides information revealing a better cartography of node involvement and allowing a better delineation of the tumour zone. In the follow-up of irradiated patients, PET-Scan provides information on the nature of residual lesions and a possible recurrence.

The green fluorescent protein (GFP) of the jellyfish Aequorea victoria is a useful reporter molecule for monitoring in vivo gene expression in eukaryotic and prokaryotic cells. We constructed a series of GFP vectors for in situ detection of the intracellular pathogen Listeria monocytogenes. The gfp-mutl gene, which encodes a red-shifted GFP, was transcriptionally fused to a strong L. monocytogenes promoter and inserted into various Escherichia coli-Listeria shuttle vectors: i) the integrative monocopy plasmid pAT113; ii) the low copy number plasmid pTCV-Exl; iii) the high copy number plasmid pAT18. Listeria cells harboring pNF6 and pNF7, constructed from pAT113 and pTCV-Exl, respectively, gave low fluorescence intensities, and were optically detected in cultured macrophages, but not in tissue sections. The fluorescence of Listeria with the pAT18 derivative pNF8 was about 40 times greater than that with pNF6 and 15 times greater than that with pNF7. Listeria cells harboring pNF8 were readily detected in both cultured macrophages and tissue sections. Constructed GFP vectors did not affect the virulence of L. monocytogenes in a murine model of infection.

Antibiotherapy is one of the main treatments of cystic fibrosis, contributing to a better nutritional and respiratory status and a prolonged survival. The choice of antibiotics depends on quantitative and qualitative analysis of sputum, bacteria resistance phenotypes and severity of infection. Haemophilus influenzae infection can be treated orally with the association of amoxicillin-clavulanic acid or a cephalosporin. Staphylococcus aureus generally remains sensitive to usual antibiotics; in case of a methicillin-resistant strain, an oral bitherapy or a parenteral cure can be proposed. Treatment of Pseudomonas aeruginosa is different in case of first colonization or chronic infection: in first colonization, parenteral antibiotherapy (beta-lactams-aminoglycosids) followed by inhaled antibiotherapy may eradicate the bacteria; in chronic infections, exacerbations require parenteral bi-antibiotherapy (beta-lactams or quinolons and aminoglycosids) for 15 to 21 days, inhaled antibiotics between the cures being useful to decrease the number of exacerbation. A careful monitoring of antibiotherapy is necessary because of possible induction of bacterial resistance, nephrotoxicity and ototoxicity of aminosids and allergy to beta-lactams.

Antibiotherapy is one of the main treatment in cystic fibrosis. Antibiotic administration schedules are different from normal patients because of pharmacokinetic and pharmacodynamic particularities. In moderate disease, the digestive resorption of antibiotics is delayed and their half-life is reduced due to an increase in total clearance. In severe disease, the volume of distribution of antibiotics is increased due to the higher proportion of lean mass in these malnourished patients. Other particularities limit the action of antibiotics such as thick sputum, which limits drug penetration; the property of Pseudomonas aeruginosa to be surrounded by a biofilm; alteration of local antibacterial defense; and inhibition of antibiotics by local factors. Systematic prescription of a biotherapy beta-lactam-aminoglycoside and obtaining high antibiotic concentration in situ might limit this antagonism. In spite of particular therapeutic schedules such as single daily dose for aminoglycoside and continuous infusion for beta-lactams, the intervals between administrations must be narrowed for time-dependent antibiotics, and the total daily dose increased by 20 to 30% for concentration-dependent antibiotics.

The details of most electron transfer reactions involving iron-sulfur proteins have remained undisclosed because of the lack of experimental methods suitable to measure precisely the relevant rates. Nuclear magnetic resonance (NMR) provides a powerful means to overcome these problems, at least with selected proteins. A combination of NMR studies and site-directed mutagenesis experiments has been instrumental in defining both the site of interaction and the main trends of the intracomplex electron transfer in the case of rubredoxin electron self-exchange. Analysis of the NMR data obtained for mixtures of different redox levels of several 2[4Fe-4S] ferredoxins provided both first-order, for intramolecular, and second-order, for intermolecular, rate constants. Their dependence as a function of structural changes gave insight into the mechanism of electron transfer in this type of protein. Contrary to some expectations, the high-spin [4Fe-4Se]+ clusters assembled in isopotential ferredoxins do not change the intramolecular electron transfer rate as compared to low-spin [4Fe-4S]+ homologs. In combination with activity measurements, the kinetic data have been used to model the electron transfer competent complexes between Clostridium pasteurianum ferredoxin and the main enzymes acting as redox partners in vivo.

Overexpression in Escherichia coli of the fdx4 gene from Aquifex aeolicus has allowed isolation and characterization of the first hyperthermophilic [2Fe-2S](Scys)(4) protein, a homodimer of M = 2 x 12.4 kDa with one [2Fe-2S] cluster per subunit. This protein is undamaged by heating to 100 degrees C for at least three hours. The primary structure, in particular the characteristic distribution of the four cysteine ligands of the metal site, and the spectroscopic properties of the A. aeolicus protein relate it to well characterized [2Fe-2S] proteins from Clostridium pasteurianum and Azotobacter vinelandii. These proteins are also homologous to subunits or domains of hydrogenases and NADH-ubiquinone oxidoreductase (Complex I) of respiratory chains. The A. aeolicus [2Fe-2S] protein is thus representative of a presumably novel protein fold involved in a variety of functions in very diverse cellular backgrounds., (Copyright 1999 Academic Press.)

Human phagocytes (polymorphonuclear neutrophils and monocytes) play a critical role in host defense against invading microorganisms. Recent studies reported that circulating phagocytes undergo a final maturation process, in particular in terms of oxidative burst, during extravasation and migration to local sites of inflammation. This process is known as priming. We report here on a nine-year-old boy with successive disseminated infections due to intracellular microorganisms (Mycobacterium bovis, BCG, and Salmonella typhimurium). No T- or B-cell quantitative or qualitative defects were found. Polymorphonuclear neutrophil (PMN) migration and NADPH oxidase in PMNs and monocytes stimulated with various agents at optimal concentrations were normal, ruling out a leukocyte adhesion deficiency syndrome, a Chediak Higashi syndrome, and a chronic granulomatous disease. Nevertheless, the patient's PMNs and monocytes showed defective priming capacity, as measured by H(2)O(2) production after pretreatment with LPS (5 microg/mL for 30 min), TNFalpha (100 units/mL for 30 min), or IL-8 (50 ng/mL for 30 min) in response to bacterial N-formyl peptides (fMLP 10(-6) M for 5 min). In these conditions, H(2)O(2) production of PMNs and monocytes from the patient did not exceed that of the samples treated with fMLP or LPS alone, while the controls strongly produced H(2)O(2). Moreover, monocytes from the patient showed an impaired capacity to kill S. typhimurium in vitro. Such an impairment could be related at least in part to the priming deficiency of phagocyte oxidative burst. This case suggests, for the first time, that in vivo priming processes are critical in host defence against intracellular pathogens.

We studied the prognostic significance of plasmablastic (PB) multiple myeloma (MM) in Eastern Cooperative Oncology Group Phase III trial E9486. Two reviewers independently reviewed 453 cases. They agreed on 37 PB (8.2%) cases and 416 non-PB cases, achieving an 85% concordance (P < .0001). These PB cases had significantly lower hemoglobin and serum albumin levels, higher calcium and beta 2-microglobuin levels, and higher percentage BM plasma cells (PC) by immunofluorescence. They had higher bone marrow PC labeling indices, higher serum soluble interleukin-6 receptor (sIL-6R) levels, and a higher probability of ras mutations. Three treatment regimens were used: vincristine, bis-chloro-ethyl nitrosourea (BCNU) melphalan, cyclophosphamide, and prednisone (VBMCP) alone; VBMCP with added cyclophosphamide (HiCy); or recombinant interferon alpha 2 (rIFNalpha2). Although the numbers are low, patients with PB had a significantly lower response rate versus non-PB MM when treated with VBMCP (treated, 47.1% v nontreated, 66.5% [P = .015]). Patients with nonresponding PB had a significantly higher progression rate than non-PB cases (30.6% v 11.8% [P < .0001]), especially with VBMCP alone (35.3% v 15.8% [P = .002]), and with added HiCy (37.5% v 9.8% [P < .0001]), but not with added rIFNalpha2. Event-free and overall survival of PB MM was shorter (median years, 1.1 v 2.7 and 1.9 v 3.7, respectively [P < .0001 for both]). In multivariate analysis, PB classification was also highly prognostic. There is no survival difference between the patients who were classified as PB by both reviewers versus patients classified as PB by only one reviewer. We conclude that PB MM is a discrete entity associated with more aggressive disease and shortened survival. Tumor cell ras mutations and increased sIL-6R may contribute to a higher proliferation rate and reduced survival. There were significant improvements in response and progression with the addition of HiCy and rIFNalpha2 to VBMCP, but the numbers were small and improved survival could not be shown.

Background: Since a significant proportion of Streptococcus pneumoniae strains is now resistant to penicillin and sometimes to third-generation cephalosporin, it is necessary to reevaluate the initial therapy of bacterial meningitis proposed before identification of the organism and its susceptibility pattern., Population: From 1 January 1992 to 31 March 1994, nine children with acute S pneumoniae meningitis were treated with ceftriaxone plus aminoglycoside as conventional initial therapy. Eight children were less than 1 year-old (five from 3 to 6 months). Five S pneumoniae strains were penicillin-resistant; four had a ceftriaxone minimal inhibitory concentration (MIC) of 0.047 to 0.094 mg/L and one of 1.5 mg/L. Ceftriaxone was given intravenously at doses of 50 mg/kg twice a day to patients less than 12 months old and 100 mg/kg once a day to patients older than 12 months. Intravenous amikacin (7.5 mg/kg twice daily) or netilmicin (3 mg/kg twice daily) were administered in combination. Dexomethasone was given to all children as adjunctive therapy. Follow-up lumbar puncture was performed after 24 to 36 hours of treatment., Results: For each of the nine patients, cerebrospinal fluid was sterile with normal glucose level. After 2 or 4 days, initial therapy had been modified according to antibiogram and MIC. Monotherapy with ceftriaxone was continued in five children. Rifampicin was associated with initial bitherapy in one case. In two other patients, initial empiric therapy was stopped and changed to chloramphenicol., Conclusion: No case of bacteriological failure was noted in our patients but evolution of epidemiology and emergence of decreased penicillin sensibility in S pneumoniae strains (55% in our study) suggests that a third antibiotic (vancocin or rifampicin) should be associated with the standard first-line drug when S pneumoniae is suspected.

In order to test a possible depression-associated defect in signal transduction, platelet alpha 2-adrenergic-mediated phosphoinositide (PI) hydrolysis was measured, both in drug-free major depressed patients and in control healthy subjects. Results that express phospholipase C activity have shown significant increase in the metabolites of epinephrine-stimulated tritiated phosphatidyl-4,5-biphosphate (3H-PIP2) with respect to basal activity (saline-stimulated). Thrombin (2 units) and 10 mM sodium fluoride (NaF) also induced an increase in 3H-PIP2 metabolites. These increases were potentiated in drug-free depressed patients both in epinephrine-and thrombin-stimulated platelets. In contrast, sodium fluoride, which directly stimulates G protein without receptor interaction, did not differentiate between patients and controls with respect to PI hydrolysis. This result suggests a possible depression-associated defect in heterologous receptor-G protein interaction.

The effects of RO 41-9067, a D2 dopamine receptors agonist, on different sleep parameters were studied in the rat. RO 41-9067 dose dependently decreased paradoxical sleep, and only at the higher dose increased waking during the light period. In contrast, the higher dose of RO 41-9067 increased paradoxical sleep and decreased waking during the dark period. Finally, the combination of RO 41-9067 and clonidine significantly prevent the decrease of total sleep time and paradoxical sleep found after clonidine alone. These results, compared with those of classical D2 dopamine receptors agonists, suggest an action for RO 41-9067 on D2 dopamine receptors depending on the cerebral structure, a different action particularly on the striatum and/or on the structures responsible for paradoxical sleep. An active role for D2 dopamine receptors and an interaction between noradrenergic and dopaminergic systems in the regulation of sleep is proposed.

This study examined whether the pharmacologic manipulation of catecholaminergic systems would affect the hypnotic action of flunitrazepam in rats. Flunitrazepam, a potent hypnotic, was used alone or in combination with alpha-methyl paratyrosine (alpha MPT), an inhibitor of the synthesis of catecholamines, and clonidine (CLN), an alpha 2-adrenoceptor agonist. Flunitrazepam significantly increased the amount of slow-wave sleep and the latency of paradoxical sleep (PS) and decreased the amount of PS. Administration of flunitrazepam to alpha MPT-treated rats significantly increased the number of sleep cycles and PS episodes as compared with flunitrazepam alone. Clonidine decreased total sleep time and significantly decreased PS. The association of flunitrazepam with CLN induced a decrease in PS and waking as compared with flunitrazepam alone. The possible involvement of noradrenergic mechanisms in modulating the effect of flunitrazepam on the rat sleep-waking cycle is proposed.

A newly synthesized agent S-12024-2 was shown to improve some aspects of cognitive processes such as memory consolidation. The relationships between sleep and memory lead us to investigate the effects of the intraperitoneal administration of three different doses (1, 3, and 10 mg/kg) of S-12024-2 on sleep variables in the rat. The results showed that S-12024-2 (10 mg/kg) increased slow wave sleep (SWS) and decreased wakefulness during the light period of the first 24 h of sleep recording. During day 1 of sleep recording, S-12024-2 tended to increase paradoxical sleep (PS) with a maximal effect observed with 3 mg/kg. Four days after administration of S-12024-2 (3 mg/kg), PS remained significantly high. These data suggest an active role for S-12024-2 on SWS and PS, compatible with its favourable effects on memory.

The active site of rubredoxins consists of a single iron tetrahedrally coordinated to four cysteinate sulfurs. One of the iron ligands, cysteine 42, has been mutated into serine in Clostridium pasteurianum rubredoxin. This mutation resulted in a shift to higher energy of the 320-800 nm region of the UV-visible absorption spectrum. Resonance Raman spectra showed that the nu 1 breathing mode of the iron chromophore was upshifted as a result of the C42S mutation. The spectral pattern, however, was not largely disturbed by the mutation. The EPR spectra of both the wild type and the C42S mutated protein displayed the characteristic features, at g = 4.3 and g = 9.5, of the "3/2" and "1/2" Kramers' doublets, respectively, of a S = 5/2 multiplet. These combined data afford strong evidence that in the C42S mutated rubredoxin serine has replaced cysteine 42 as a ligand of the iron, while maintaining the tetrahedral coordination of the metal. The most spectacular effect of the C42S mutation was a ca. 200 mV downshift of the redox potential of rubredoxin.

Increased interleukin-6 (IL-6) production and expression by malignant cells of the IL-6 receptor has been evidenced in a subgroup of non-Hodgkin's lymphomas, suggesting that this cytokine plays a role in lymphoma growth and in B clinical symptoms. In this study, the effect of the administration of an anti-IL-6 monoclonal antibody (MoAb) was analyzed in 11 patients seropositive for human immunodeficiency virus-1 and suffering from an immunoblastic or a polymorphic large-cell lymphoma. The antibody (BE-8, 10 to 40 mg/day) was administered for 21 days. Neutralization of in vivo IL-6 effect was assessed by monitoring C-reactive protein levels in the serum. In 5 patients, the lymphoma progressed during treatment. Among them were the 2 patients in whom endogenous IL-6 effect was not neutralized. Five patients experienced a stabilization, and 1 a partial remission. This effect on lymphoma growth lasted for 8 to 28 weeks. The anti-IL-6 MoAb had a clear effect on lymphoma-associated fever and cachexia. The mean body weight increase was 1.4 +/- 0.5 kg between day 1 and day 21, and reached 12 kg in 120 days in 1 patient who received three courses of treatment. Side effects were a consistent but moderate thrombocytopenia, and an occasional and moderate decrease of neutrophil counts. Immunization against the MoAb was observed in only 2 patients. These results indicate that in some cases of lymphomas growth of malignant cells may be partially IL-6-dependent and that neutralizing endogenous effect of IL-6 completely abrogates B clinical symptoms.

We report a novel, reproducible methodology which enabled 10 human myeloma cell lines (HMCL) to be obtained from each of 10 tumor samples harvested from 9 patients with extramedullary proliferation. Fresh samples were cultured with interleukin 6 (IL-6) and granulocyte macrophage-colony stimulating factor (GM-CSF) at a high cell density and resulting HMCL growth became progressively dependent on IL-6 alone, no longer requiring GM-CSF. These HMCL, which had the same immunoglobulin gene rearrangements as the patients' original myeloma cells, were designated XG-1 to XG-9. XG HMCL had a plasma cell morphology, expressed plasma cell antigen (Ag), namely cytoplasmic immunoglobulins, CD38, B-B4 Ag, and CD77, and lacked the usual B-cell Ag. They also expressed activation antigens such as CD28 with coexpression of CD28 and its ligand, B7 Ag, in four HMCL. Six HMCL expressed CD40, 4 CD23, and 5 its ligand, CD21. The XG HMCL bore adhesion molecules VLA-4 and CD44 (all 10 HMCL), VLA-5 (7 HMCL), and CD56 (4 HMCL). Finally, cytogenetic study of 8 HMCL indicated a 14q+ chromosome, and t(11,14) translocation was found in 6 of 8 and 5 of 8 HMCL, respectively. The possibility of obtaining malignant plasma cell lines reproducibly from each patient with extramedullary proliferation offers a unique tool for studying the phenotype and abnormalities of the still unidentified tumor stem cell in this disease.

Acetylcholine promotes paradoxical sleep (PS), but the role of noradrenaline in this stimulation is controversial. The relationship between cholinergic and noradrenergic systems in the production of PS was investigated in the rat implanted on a continuous basis for sleep recordings. Stimulation of PS was obtained with microinjections of carbachol (1 microgram) into the pontine reticular formation. In the presence of the alpha 2-agonist clonidine (5 micrograms/kg, IP), the carbachol activation of PS was abolished. This stimulation also disappeared when the animals were pretreated with alpha-methyl-paratyrosine (150 mg/kg, IP), an inhibitor of catecholamine synthesis. Thus, carbachol stimulation appeared inefficient when brain noradrenergic activation was decreased. This observation supports the view that the realization of PS by the cholinergic system requires a certain level of noradrenergic activity.

Soluble forms of most cytokine receptors, able to bind effectively to their respective ligands, have now been described. A soluble interleukin-6-binding molecule derived from the gp80 component of the multichain IL-6 receptor can be detected in biological fluids, and can act as an agonist of IL-6 activity. The clinical significance of the soluble receptor levels still remains to be explored. We took advantage of the characterization of an anti-IL-6 monoclonal antibody and of an anti-IL-6R monoclonal antibody that both bound to IL-6/IL-6R complexes to design an immunometric assay for the measurement of soluble IL-6R complexed to IL-6. This reaction scheme was designated as ELIA (enzyme-ligand immunoassay). When exogeneous IL-6 was added in excess to an sIL-6R containing sample, all sIL-6R was present in a complexed form. Thus, the reaction scheme could also be used to determine total sIL-6R concentrations. A recombinant sIL-6R standard was prepared from the supernatant of murine thymoma cells transfected with a gene coding for an extracellular portion of the IL-6 receptor. The assay permitted the precise and reproducible measurement of sIL-6R in serum or plasma. This approach is of general relevance for the determination of soluble cytokine receptors in biological fluids, provided that adequate anti-cytokine and anti-receptor antibodies are available.