Development of a real-time PCR for the specific detection of Waddlia chondrophila in clinical samples.

Waddlia chondrophila is considered as an emerging human pathogen likely involved in miscarriage and lower respiratory tract infections. Given the low sensitivity of cell culture to recover such an obligate intracellular bacteria, molecular-based diagnostic approaches are warranted. We thus developed a real-time PCR that amplifies Waddlia chondrophila DNA. Specific primers and probe were selected to target the 16S rRNA gene. The PCR specifically amplified W. chondrophila but did not amplify other related-bacteria such as Parachlamydia acanthamoebae, Simkania negevensis and Chlamydia pneumoniae. The PCR exhibited a good intra-run and inter-run reproducibility and a sensitivity of less than ten copies of the positive control. This real-time PCR was then applied to 32 nasopharyngeal aspirates taken from children with bronchiolitis not due to respiratory syncytial virus (RSV). Three samples revealed to be Waddlia positive, suggesting a possible role of this Chlamydia-related bacteria in this setting.

Marine and transitional Middle/Upper Eocenen units of the Southeastern Pyrenean Foreland Basin (NE Spain)

The stratigraphic basis of this work has allowed the use of larger foraminifers in the biostratigraphic characterisation of the new Shallow Benthic Zones (SBZ). This part of the volume presents a description of the sedimentary cycles formed by the transgressive-regressive systems of the Lutetian and Bartonian in the southeastern sector of the Ebro Foreland Basin. Concerning the Lutetian deposits studied in the Amer-Vic and Empordà areas, four sedimentary cycles have been characterised. The first and second are found within the Tavertet/Girona Limestone Formation (Reguant, 1967; Pallí, 1972), while the third and fourth cycles cover the Coll de Malla Marl Formation (Clavell et al., 1970), the Bracons Formation (Gich, 1969, 1972), the Banyoles Marl Formation (Almela and Ríos, 1943), and the Bellmunt Formation (Gich, 1969, 1972). In the Bartonian deposits studied in the Igualada area, two transgressive-regressive sedimentary cycles have been characterised in the Collbàs Formation (Ferrer, 1971), the Igualada Formation (Ferrer, 1971), and the Tossa Formation (Ferrer, 1971). The Shallow Benthic Zones (SBZs) recognised within the Lutetian are the following: SBZ 13, from the Early Lutetian, in the transgressive system of the first cycle; SBZ 14, from the Middle Lutetian, in the second cycle and the lower part of the transgressive system of the third cycle; SBZ 15, from the Middle Lutetian, in the remaining parts of the third system; SBZ 16, from the Late Lutetian, throughout the fourth cycle. The association of larger foraminifers in the first and second cycles of the Bartonian in the Igualada area has been used as the basis for the definition of SBZs 17 and 18 recognised in the Bartonian of the western Tethys.

An inventory of the marine and transitional Middle/Upper Eocene deposits of the Southeastern Pyrenean Foreland Basin (NE Spain)

In the southeastern Ebro Foreland Basin, the marine deposits of Lutetian and Bartonian age show excellent outcrop conditions, with a great lateral and horizontal continuity of lithostratigraphic units. In addition, the rich fossil record -mainly larger foraminifers-, provides biostratigraphic data of regional relevance for the whole Paleogene Pyrenean Basin, that can be used for the Middle Eocene biocorrelation of the western Tethys. This contribution is a sedimentary and biostratigraphic synthesis of the basic outcrops and sections of the Lutetian andBartonian marine and transitional deposits in the southeastern sector of the Ebro Foreland Basin.

The Upper Oligocene of Montgat (Catalan Coastal Ranges, Spain): new age constraints to the western Mediterranean Basin opening

The Oligocene deposits of Montgat are integrated in a small outcrop made up of Cenozoic and Mesozoic rocks located in the Garraf-Montnegre horst, close to the major Barcelona fault. The Oligocene of Montgat consists of detrital sediments of continental origin mainly deposited in alluvial fan environments; these deposits are folded and affected by thrusts and strike-slip faults. They can be divided in two lithostratigraphic units separated by a minor southwest-directed thrust: (i) the Turó de Montgat Unit composed of litharenites and lithorudites with high contents of quartz, feldspar, plutonic and limestone rock fragments; and (ii) the Pla de la Concòrdia Unit composed of calcilitharenites and calcilithorudites with high contents of dolosparite and dolomicrite rock fragments. The petrological composition of both units indicates that sediments were derived from the erosion of Triassic (Buntsandstein, Muschelkalk and Keuper facies), Jurassic and Lower Cretaceous rocks (Barremian to Aptian in age). Stratigraphic and petrological data suggest that these units correspond to two coalescent alluvial fans with a source area located northwestwards in the adjoining Collserola and Montnegre inner areas. Micromammal fossils (Archaeomys sp.) found in a mudstone layer of the Pla de la Concòrdia Unit assign a Chattian age (Late Oligocene) to the studied materials. Thus, the Montgat deposits are the youngest dated deposits affected by the contractional deformation that led to the development of the Catalan Intraplate Chain. Taking into account that the oldest syn-rift deposits in the Catalan Coastal Ranges are Aquitanian in age, this allows to precise that the change from a compressive to an extensional regime in this area took place during latest Oligocene-earliest Aquitanian times. This age indicates that the onset of crustal extension related to the opening of the western Mediterranean Basin started in southern France during latest Eocene-early Oligocene and propagated southwestward, affecting the Catalan Coastal Ranges and the northeastern part of the Valencia trough during the latest Chattian-earliest Aquitanian times.

Polarity and specific sequence requirements of peroxisome proliferator-activated receptor (PPAR)/retinoid X receptor heterodimer binding to DNA. A functional analysis of the malic enzyme gene PPAR response element.

The malic enzyme (ME) gene is a target for both thyroid hormone receptors and peroxisome proliferator-activated receptors (PPAR). Within the ME promoter, two direct repeat (DR)-1-like elements, MEp and MEd, have been identified as putative PPAR response elements (PPRE). We demonstrate that only MEp and not MEd is able to bind PPAR/retinoid X receptor (RXR) heterodimers and mediate peroxisome proliferator signaling. Taking advantage of the close sequence resemblance of MEp and MEd, we have identified crucial determinants of a PPRE. Using reciprocal mutation analyses of these two elements, we show the preference for adenine as the spacing nucleotide between the two half-sites of the PPRE and demonstrate the importance of the two first bases flanking the core DR1 in 5'. This latter feature of the PPRE lead us to consider the polarity of the PPAR/RXR heterodimer bound to its cognate element. We demonstrate that, in contrast to the polarity of RXR/TR and RXR/RAR bound to DR4 and DR5 elements respectively, PPAR binds to the 5' extended half-site of the response element, while RXR occupies the 3' half-site. Consistent with this polarity is our finding that formation and binding of the PPAR/RXR heterodimer requires an intact hinge T region in RXR while its integrity is not required for binding of the RXR/TR heterodimer to a DR4.

Evaluation of the specific IgG- and IgA-antibody response against immundominant Aspergillus fumigatus antigens as diagnostic markers of invasive aspergillosis

An improvement in the serological diagnostic toolbox of invasive aspergillosis (IA) is necessary. So far, most laboratories do not perform antibody detection assays at all to diagnose IA, as commercial test systems are based on crude and undefined antigen mixtures of A. fumigatus. Utilizing the A. fumigatus protein mitogillin, we could demonstrate that the use of selected characterized immunodominant antigens can improve the serodiagnosis of Aspergillus-related diseases. In an animal model we were able to identify additional 36 immunodominant antigens of a cDNA library of A. fumigatus germlings. Five selected antigens were expressed recombinantly in E. coli, purified and used for Westernblot und ELISA analyses to study the kinetics of the specific antibody response in rabbits that were infected systemically with A. fumigatus. Subsequently, the specific IgG- and IgA-antibody responses against these antigens were studied in patients suffering from proven IA and compared to healthy blood donors and patients with other forms of pneumonia. Furthermore, we examined how total IgG- and IgA-levels influence the diagnostic value of antibody detection in IA patients.

Assessment of vascular invasion by bone and soft tissue tumours of the limbs: usefulness of MDCT angiography.

OBJECTIVE: To evaluate the accuracy of computed tomography angiography (CTA) in predicting arterial encasement by limb tumours, by comparing CTA with surgical findings (gold standard). METHODS: Preoperative CTA images of 55 arteries in 48 patients were assessed for arterial status: cross-sectional CTA images were scored as showing a fat plane between artery and tumour (score 0), slight contact between artery and tumour (score 1), partial arterial encasement (score 2) or total arterial encasement (score 3). Reformatted CTA images were assessed for arterial displacement, rigid wall, stenosis or occlusion. At surgery, arteries were classified as free or surgically encased; 45 arteries were free and 10 were surgically encased. RESULTS: Multivariate logistic regression identified the axial CTA score as a relevant predictor for arterial encasement and subsequent vascular intervention during surgery. All sites where CTA showed a fat plane between the tumour and the artery were classified as free at surgery (n = 28/28). The sensitivity of total arterial encasement on CTA (score 3) was 90%, specificity 93%, accuracy 93% and positive likelihood ratio 13.5. CONCLUSION: CTA evidence of total arterial encasement is a highly specific indication of arterial encasement. The presence of fat between the tumour and the artery on CTA rules out arterial involvement at surgery.

A downstream mediator in the growth repression limb of the jasmonate pathway.

Wounding plant tissues initiates large-scale changes in transcription coupled to growth arrest, allowing resource diversion for defense. These processes are mediated in large part by the potent lipid regulator jasmonic acid (JA). Genes selected from a list of wound-inducible transcripts regulated by the jasmonate pathway were overexpressed in Arabidopsis thaliana, and the transgenic plants were then assayed for sensitivity to methyl jasmonate (MeJA). When grown in the presence of MeJA, the roots of plants overexpressing a gene of unknown function were longer than those of wild-type plants. When transcript levels for this gene, which we named JASMONATE-ASSOCIATED1 (JAS1), were reduced by RNA interference, the plants showed increased sensitivity to MeJA and growth was inhibited. These gain- and loss-of-function assays suggest that this gene acts as a repressor of JA-inhibited growth. An alternative transcript from the gene encoding a second protein isoform with a longer C terminus failed to repress jasmonate sensitivity. This identified a conserved C-terminal sequence in JAS1 and related genes, all of which also contain Zim motifs and many of which are jasmonate-regulated. Both forms of JAS1 were found to localize to the nucleus in transient expression assays. Physiological tests of growth responses after wounding were consistent with the fact that JAS1 is a repressor of JA-regulated growth retardation.

CD8(+) T cells specific for tumor antigens can be rendered dysfunctional by the tumor microenvironment through upregulation of the inhibitory receptors BTLA and PD-1.

Cytotoxic T cells that are present in tumors and capable of recognizing tumor epitopes are nevertheless generally impotent in eliciting tumor rejection. Thus, identifying the immune escape mechanisms responsible for inducing tumor-specific CD8(+) T-cell dysfunction may reveal effective strategies for immune therapy. The inhibitory receptors PD-1 and Tim-3 are known to negatively regulate CD8(+) T-cell responses directed against the well-characterized tumor antigen NY-ESO-1. Here, we report that the upregulation of the inhibitory molecule BTLA also plays a critical role in restricting NY-ESO-1-specific CD8(+) T-cell expansion and function in melanoma. BTLA-expressing PD-1(+)Tim-3(-) CD8(+) T cells represented the largest subset of NY-ESO-1-specific CD8(+) T cells in patients with melanoma. These cells were partially dysfunctional, producing less IFN-γ than BTLA(-) T cells but more IFN-γ, TNF, and interleukin-2 than the highly dysfunctional subset expressing all three receptors. Expression of BTLA did not increase with higher T-cell dysfunction or upon cognate antigen stimulation, as it does with PD-1, suggesting that BTLA upregulation occurs independently of functional exhaustion driven by high antigen load. Added with PD-1 and Tim-3 blockades, BTLA blockade enhanced the expansion, proliferation, and cytokine production of NY-ESO-1-specific CD8(+) T cells. Collectively, our findings indicate that targeting BTLA along with the PD-1 and Tim-3 pathways is critical to reverse an important mechanism of immune escape in patients with advanced melanoma.

Measurement of the specific heat and determination of the thermodynamic functions of relaxed amorphous silicon

The specific heat, cp, of two amorphous silicon (a-Si) samples has been measured by differential scanning calorimetry in the 100–900K temperature range. When the hydrogen content is reduced by thermal annealing, cp approaches the value of crystalline Si (c-Si). Within experimental accuracy, we conclude that cp of relaxed pure a-Si coincides with that of c-Si. This result is used to determine the enthalpy, entropy, and Gibbs free energy of defect-free relaxed a-Si. Finally, the contribution of structural defects on these quantities is calculated and the melting point of several states of a-Si is predicted

Critical role for the chemokine receptor CXCR6 in NK cell-mediated antigen-specific memory of haptens and viruses.

Hepatic natural killer (NK) cells mediate antigen-specific contact hypersensitivity (CHS) in mice deficient in T cells and B cells. We report here that hepatic NK cells, but not splenic or naive NK cells, also developed specific memory of vaccines containing antigens from influenza, vesicular stomatitis virus (VSV) or human immunodeficiency virus type 1 (HIV-1). Adoptive transfer of virus-sensitized NK cells into naive recipient mice enhanced the survival of the mice after lethal challenge with the sensitizing virus but not after lethal challenge with a different virus. NK cell memory of haptens and viruses depended on CXCR6, a chemokine receptor on hepatic NK cells that was required for the persistence of memory NK cells but not for antigen recognition. Thus, hepatic NK cells can develop adaptive immunity to structurally diverse antigens, an activity that requires NK cell-expressed CXCR6.

Absence of association between specific common variants of the obesity-related FTO gene and psychological and behavioral eating disorder phenotypes.

Extensive population-based genome-wide association studies have identified an association between the FTO gene and BMI; however, the mechanism of action is still unknown. To determine whether FTO may influence weight regulation through psychological and behavioral factors, seven single-nucleotide polymorphisms (SNPs) of the FTO gene were genotyped in 1,085 individuals with anorexia nervosa (AN) and 677 healthy weight controls from the international Price Foundation Genetic Studies of Eating Disorders. Each SNP was tested in association with eating disorder phenotypes and measures that have previously been associated with eating behavior pathology: trait anxiety, harm-avoidance, novelty seeking, impulsivity, obsessionality, compulsivity, and concern over mistakes. After appropriate correction for multiple comparisons, no significant associations between individual FTO gene SNPs and eating disorder phenotypes or related eating behavior pathology were identified in cases or controls. Thus, this study found no evidence that FTO gene variants associated with weight regulation in the general population are associated with eating disorder phenotypes in AN participants or matched controls. © 2011 Wiley-Liss, Inc.