Analysis and comparison of the microflora isolated from fresco surface and from surrounding air environment through molecular and biodegradative assays

The aim of this study was to find a correlation among the environmental isolated microflora and the fresco colonizators through the investigation of their biodegradative abilities and DNA characteristics. A molecular technique named RAMP (Random Amplified Microsatellite Polymorphisms) was utilized in order to analyze the DNA diversity of bacterial and fungal species isolated from fresco as well as from air samples. The RAMP-PCR results were combined with the screening of some biodegradative properties obtained through the use of specific agar plate assays detecting the proteolytic, solubilization and biomineralization abilities of the isolated microflora. This comparative analysis showed that only in few cases a direct link among the fresco and airborne isolates of specific microbial group existed. The investigation clearly evidenced that colonization of surface of Ladislav’s fresco occurred in different time and by different strains than those observed at the moment of sampling campaign. Furthermore, the microflora investigation permitted the identification of taxonomically interesting bacteria with particular biodegradative properties, which had been less studied until now.     

Cyanobacteria cause black staining of the National Museum of the American Indian Building, Washington, DC, USA

Microbial deterioration of stone is a widely recognised problem affecting monuments and buildings all over the world. In this paper, dark-coloured staining, putatively attributed to microorganisms, on areas of the National Museum of the American Indian Building, Washington, DC, USA, were studied. Observations by optical and electron microscopy of surfaces and cross sections of limestone indicated that biofilms, which penetrated up to a maximum depth of about 1?mm, were mainly composed of cyanobacteria, with the predominance of Gloeocapsa and Lyngbya. Denaturing gradient gel electrophoresis analysis revealed that the microbial community also included eukaryotic algae (Trebouxiophyceae) and fungi (Ascomycota), along with a consortium of bacteria. Energy-dispersive X-ray spectroscopy analysis showed the same elemental composition in stained and unstained areas of the samples, indicating that the discolouration was not due to abiotic chemical changes within the stone. The dark pigmentation of the stone was correlated with the high content of scytonemin, which was found in all samples.     

Dimeric Smac mimetics/IAP inhibitors as in vivo-active pro-apoptotic agents. Part II: Structural and biological characterization

Novel pro-apoptotic, homodimeric and heterodimeric Smac mimetics/IAPs inhibitors connected through head–head (8), tail–tail (9) or head–tail linkers (10), were biologically and structurally characterized. In vitro characterization (binding to BIR3 and linker-BIR2–BIR3 domains from XIAP and cIAP1, cytotoxicity assays) identified early leads from each dimer family. Computational models and structural studies (crystallography, NMR, gel filtration) partially rationalized the observed properties for each dimer class. Tail–tail dimer 9a was shown to be active in a breast and in an ovary tumor model, highlighting the potential of dimeric Smac mimetics/IAP inhibitors based on the N-AVPI-like 4-substituted 1-aza-2-oxobicyclo[5.3.0]decane scaffold as potential antineoplastic agents.     

Genetic structure and linkage disequilibrium in landrace populations of barley in Sardinia

Multilocus digenic linkage disequilibria (LD) and their population structure were investigated in eleven landrace populations of barley (Hordeum vulgare ssp. vulgare L.) in Sardinia, using 134 dominant simple-sequence amplified polymorphism markers. The analysis of molecular variance for these markers indicated that the populations were partially differentiated (F(ST) = 0.18), and clustered into three geographic areas. Consistent with this population pattern, STRUCTURE analysis allocated individuals from a bulk of all populations into four genetic groups, and these groups also showed geographic patterns. In agreement with other molecular studies in barley, the general level of LD was low (13% of locus pairs, with P < 0.01) in the bulk of 337 lines, and decayed steeply with map distance between markers. The partitioning of multilocus associations into various components indicated that genetic drift and founder effects played a major role in determining the overall genetic makeup of the diversity in these landrace populations, but that epistatic homogenising or diversifying selection was also present. Notably, the variance of the disequilibrium component was relatively high, which implies caution in the pooling of barley lines for association studies. Finally, we compared the analyses of multilocus structure in barley landrace populations with parallel analyses in both composite crosses of barley on the one hand and in natural populations of wild barley on the other. Neither of these serves as suitable mimics of landraces in barley, which require their own study. Overall, the results suggest that these populations can be exploited for LD mapping if population structure is controlled.     

Role of replication protein A as sensor in activation of the S-phase checkpoint in Xenopus egg extracts

Uncoupling between DNA polymerases and helicase activities at replication forks, induced by diverse DNA lesions or replication inhibitors, generate long stretches of primed single-stranded DNA that is implicated in activation of the S-phase checkpoint. It is currently unclear whether nucleation of the essential replication factor RPA onto this substrate stimulates the ATR-dependent checkpoint response independently of its role in DNA synthesis. Using Xenopus egg extracts to investigate the role of RPA recruitment at uncoupled forks in checkpoint activation we have surprisingly found that in conditions in which DNA synthesis occurs, RPA accumulation at forks stalled by either replication stress or UV irradiation is dispensable for Chk1 phosphorylation. In contrast, when both replication fork uncoupling and RPA hyperloading are suppressed, Chk1 phosphorylation is inhibited. Moreover, we show that extracts containing reduced levels of RPA accumulate ssDNA and induce spontaneous, caffeine-sensitive, Chk1 phosphorylation in S-phase. These results strongly suggest that disturbance of enzymatic activities of replication forks, rather than RPA hyperloading at stalled forks, is a critical determinant of ATR activation.     

Usefulness of Contrast Intracardiac Echocardiography in Performing Pulmonary Vein Balloon Occlusion during Cryo-ablation for Atrial Fibrillation

BackgroundCryoballoon ablation (CBA) has been proven to be very effective for pulmonary vein (PV) isolation (PVI) if complete occlusion is achieved and conventionally assessed by angiographic injection of contrast within PV lumen. The aim of our study was to assess the usefulness of saline contrast intracardiac echocardiography in guiding CBA with respect to PV angiography.MethodsThirty consecutive patients with paroxysmal atrial fibrillation were randomly assigned fluoroscopy plus color-flow Doppler (n = 15; group 1: an iodinated medium as both angiographic and echographic contrast) or contrast intracardiac echocardiography plus color-flow Doppler (n = 15; group 2: saline contrast) for guidance of CBA.ResultsWe evaluated 338 occlusions of 107 PVs. The intracardiac echocontrastography-guided assessment of occlusion, defined as loss of echocontrastographic back-flow to the left atrium after saline injection regardless of the visualization of PV antrum, showed a high level of agreement with the angiographic diagnosis of occlusion. PVI rate was similar in both groups and effectively guided by intracardiac echocontrastography (PVI using ≤ 2 double cryofreezes: 89% of PVs in group 1 vs. 91% in group 2; p=n.s.). Group 2 patients had significantly shorter procedure (127 ± 16 vs. 152 ± 19 minutes; p<0.05) and fluoroscopy times (30 ± 12 vs. 43 ± 9 minutes, p<0.05) and used a lower iodinated contrast (88 ± 26 vs. 190 ± 47 mL, p<0.05).ConclusionsPV occlusion and PVI during cryoablation can be effectively predicted by intracardiac saline echocontrastography. This technique reduces procedural time, radiological exposure and iodinated contrast use.     

In situ hybridization and immunogold localization of vascular endothelial growth factor receptor-2 on the pericytes of the chick chorioallantoic membrane

This paper describes the expression of VEGF and of VEGFR-2 in the vasculature of the chorioallantoic membrane (CAM) as revealed by in situ hybridization and immunoelectron microscopy. Results showed that VEGFR-2 is expressed in both the endothelial cells and the pericytes, while VEGF in the chorionic epithelial cells. VEGF may therefore be released to promote both angiogenesis, by initiating an angiogenic response by endothelial cells expressing VEGFR-2, and the recruitment of pericytes along the capillary wall, playing also a crucial role in maturation and stabilization of the CAM blood vessels.     

Structural Basis for Group B Streptococcus Pilus 1 Sortases C Regulation and Specificity

Gram-positive bacteria assemble pili through class C sortase enzymes specialized in polymerizing pilin subunits into covalently linked, high-molecular-weight, elongated structures. Here we report the crystal structures of two class C sortases (SrtC1 and SrtC2) from Group B Streptococcus (GBS) Pilus Island 1. The structures show that both sortases are comprised of two domains: an 8-stranded β-barrel catalytic core conserved among all sortase family members and a flexible N-terminal region made of two α-helices followed by a loop, known as the lid, which acts as a pseudo-substrate. In vitro experiments performed with recombinant SrtC enzymes lacking the N-terminal portion demonstrate that this region of the enzyme is dispensable for catalysis but may have key roles in substrate specificity and regulation. Moreover, in vitro FRET-based assays show that the LPXTG motif common to many sortase substrates is not the sole determinant of sortase C specificity during pilin protein recognition.     

Waking and Sleeping following Water Deprivation in the Rat

Wake-sleep (W-S) states are affected by thermoregulation. In particular, REM sleep (REMS) is reduced in homeotherms under a thermal load, due to an impairment of hypothalamic regulation of body temperature. The aim of this work was to assess whether osmoregulation, which is regulated at a hypothalamic level, but, unlike thermoregulation, is maintained across the different W-S states, could influence W-S occurrence. Sprague-Dawley rats, kept at an ambient temperature of 24°C and under a 12 h∶12 h light-dark cycle, were exposed to a prolonged osmotic challenge of three days of water deprivation (WD) and two days of recovery in which free access to water was restored. Two sets of parameters were determined in order to assess: i) the maintenance of osmotic homeostasis (water and food consumption; changes in body weight and fluid composition); ii) the effects of the osmotic challenge on behavioral states (hypothalamic temperature (Thy), motor activity, and W-S states). The first set of parameters changed in WD as expected and control levels were restored on the second day of recovery, with the exception of urinary Ca⁺⁺ that almost disappeared in WD, and increased to a high level in recovery. As far as the second set is concerned, WD was characterized by the maintenance of the daily oscillation of Thy and by a decrease in activity during the dark periods. Changes in W-S states were small and mainly confined to the dark period: i) REMS slightly decreased at the end of WD and increased in recovery; ii) non-REM sleep (NREMS) increased in both WD and recovery, but EEG delta power, a sign of NREMS intensity, decreased in WD and increased in recovery. Our data suggest that osmoregulation interferes with the regulation of W-S states to a much lesser extent than thermoregulation.     

Combination training in aging individuals modifies functional connectivity and cognition, and is potentially affected by dopamine-related genes

Background

Aging is a major co-risk factor in many neurodegenerative diseases. Cognitive enrichment positively affects the structural plasticity of the aging brain. In this study, we evaluated effects of a set of structured multimodal activities (Combination Training; CT) on cognitive performances, functional connectivity, and cortical thickness of a group of healthy elderly individuals. CT lasted six months.

Methodology

Neuropsychological and occupational performances were evaluated before and at the end of the training period. fMRI was used to assess effects of training on resting state network (RSN) functional connectivity using Independent Component Analysis (ICA). Effects on cortical thickness were also studied. Finally, we evaluated whether specific dopamine-related genes can affect the response to training.

Principal Findings

Results of the study indicate that CT improves cognitive/occupational performances and reorganizes functional connectivity. Intriguingly, individuals responding to CT showed specific dopamine-related genotypes. Indeed, analysis of dopamine-related genes revealed that carriers of DRD3 ser9gly and COMT Val158Met polymorphisms had the greatest benefits from exposure to CT.

Conclusions and Significance

Overall, our findings support the idea that exposure to a set of structured multimodal activities can be an effective strategy to counteract aging-related cognitive decline and also indicate that significant capability of functional and structural changes are maintained in the elderly.