Influence of xenobiotic perturberation on soil mcrobial community

The effects of xenobiotics application on the composition and function of soil microbial community were investigated in mountain grassland (Slovakia) and agricultural (UK) soils. Slovak soil was Cambisol, sampled from the mountain grassland regions of Greater Fatra, Lesser Fatra, Lower Tatras and Slovak Ore Moutains. UK soil was sampled from the Cottenham, Faulkborne and Denchworth series located in an agriculture farm at Silsoe. Soils thereby differed in climate conditions, soil type, soil texture and land use. Initially soils were characterized by texture, moisture, pH, total carbon, oxidizable carbon, total nitrogen, microbial biomass, hydrolytic enzyme activity, soil respiration and PLFA composition. Results distinctly showed that the microbial community structure, especially abundance of G+ , G- bacteria and fungi, varied between different soil types. An experiment was established using the UK soils. The effects of the xenobiotics polyvinylalcohol, a fungicide (Fundazol) and a herbicide (Gesagard) on soil microbial community and activity were investigated one day and forty-two days following xenobiotic application. The functional stability in the terms of resistance and resilience using the method described by Orwin and Wardle (2004) was calculated from the soil respiration rate data. The experimental treatments caused a significant difference in the PCA profile of PLFA data. Soil type and textural classification affected the altered microbial profile. Treatments also altered microbial activity and microbial biomass. The arable soils were more resistant to xenobiotic perturbation than grassland soils. The reduction of functional stability was associated with the altered soil microbial community composition. Thus, soil type had a greater role than treatment type in determining microbial community composition whereas the treatment type was more determining factor of catabolic profile

Calcium and ROS: A mutual interplay

AbstractCalcium is an important second messenger involved in intra- and extracellular signaling cascades and plays an essential role in cell life and death decisions. The Ca2+ signaling network works in many different ways to regulate cellular processes that function over a wide dynamic range due to the action of buffers, pumps and exchangers on the plasma membrane as well as in internal stores. Calcium signaling pathways interact with other cellular signaling systems such as reactive oxygen species (ROS). Although initially considered to be potentially detrimental byproducts of aerobic metabolism, it is now clear that ROS generated in sub-toxic levels by different intracellular systems act as signaling molecules involved in various cellular processes including growth and cell death. Increasing evidence suggests a mutual interplay between calcium and ROS signaling systems which seems to have important implications for fine tuning cellular signaling networks. However, dysfunction in either of the systems might affect the other system thus potentiating harmful effects which might contribute to the pathogenesis of various disorders

Combination of Resonance and Non-Resonance Chiral Raman Scattering in a Cobalt(III) Complex

Resonance Raman optical activity (RROA) spectra with high sensitivity reveal details on molecular structure, chirality, and excited electronic properties. Despite the difficulty of the measurements, the recorded data for the Co(III) complex with S,S-N,N-ethylenediaminedisuccinic acid are of exceptional quality and, coupled with the theory, spectacularly document the molecular behavior in resonance. This includes a huge enhancement of the chiral scattering, contribution of the antisymmetric polarizabilities to the signal, and the Herzberg-Teller effect significantly shaping the spectra. The chiral component is by about one order of magnitude bigger than for an analogous aluminum complex. The band assignment and intensity profile were confirmed by simulations based on density functional and vibronic theories. The resonance was attributed to the S0→S3 transition, with the strongest signal enhancement of Raman and ROA spectral bands below about 800 cm−1. For higher wavenumbers, other excited electronic states contribute to the scattering in a less resonant way. RROA spectroscopy thus appears as a unique tool to study the structure and electronic states of absorbing molecules in analytical chemistry, biology, and material science

Concept of a Wearable Temperature Sensor for Intelligent Textile

This paper proposes the intelligent textile for human body temperature measurement. The main concept of the textile is based on two commercially available LM35DM temperature sensors, knitted electrically conductivity yarns and five areas for press-stud. Three different measurements methods were used in our experiment. Three measurements were performed in the first step, digital thermometer in person’s armpit, contact-less thermometer in the places close to our LM35DM sensors and contact-less thermometer on the forehead. Next, two LM35DM in SOIC8 package were used for temperature measurement within intelligent textile. Finally, the first step was repeated. All measurements were done on ten subjects with their permission to collect body temperature data for scientific purposes. The results show that the average measurement error for first sensor is 0.844°C and 0.278°C for second sensor

Noninvasive detection of F8 int22h-related inversions and sequence variants in maternal plasma of hemophilia carriers

Direct detection of F8 and F9 sequence variants in maternal plasma of hemophilia carriers has been demonstrated by microfluidics digital PCR. Noninvasive prenatal assessment of the most clinically relevant group of sequence variants among patients with hemophilia, namely, those involving int22h-related inversions disrupting the F8 gene, poses additional challenges because of its molecular complexity. We investigated the use of droplet digital PCR (ddPCR) and targeted massively parallel sequencing (MPS) for maternal plasma DNA analysis to noninvasively determine fetal mutational status in pregnancies at risk for hemophilia. We designed family-specific ddPCR assays to detect causative sequence variants scattered across the F8 and F9 genes. A haplotype-based approach coupled with targeted MPS was applied to deduce fetal genotype by capturing a 7.6-Mb region spanning the F8 gene in carriers with int22h-related inversions. The ddPCR analysis correctly determined fetal hemophilia status in 15 at-risk pregnancies in samples obtained from 8 to 42 weeks of gestation. There were 3 unclassified samples, but no misclassification. Detailed fetal haplotype maps of the F8 gene region involving int22h-related inversions obtained through targeted MPS enabled correct diagnoses of fetal mutational status in 3 hemophilia families. Our data suggest it is feasible to apply targeted MPS to interrogate maternally inherited F8 int22h-related inversions, whereas ddPCR represents an affordable approach for the identification of F8 and F9 sequence variants in maternal plasma. These advancements may bring benefits for the pregnancy management for carriers of hemophilia sequence variants; in particular, the common F8 int22h-related inversions, associated with the most severe clinical phenotype

Detection of ctDNA from dried blood spots after DNA size selection

Background: Recent advances in the study and clinical applications of circulating tumor DNA (ctDNA) are limited by practical considerations of sample collection. Whole genome sequencing (WGS) is increasingly used for analysis of ctDNA, identifying copy-number alterations and fragmentation patterns. We hypothesized that low-depth/shallow WGS (sWGS) data may be generated from minute amounts of cell-free DNA, and that fragment-size selection may remove contaminating genomic DNA from small blood volumes. Dried blood spots have practical advantages for sample collection, may facilitate serial sampling, and could support novel study designs in humans and animal models. Methods: We developed a protocol for the isolation and analysis of cell-free DNA from dried blood spots using filter paper cards and bead-based size selection. DNA extracted and size-selected from dried spots was analyzed using sWGS and PCR. Results: Analyzing a 50L dried blood spot from frozen whole blood of a patient with melanoma, we identified ctDNA based on the presence of tumor-specific somatic copy-number alterations, and found a fragment size profile similar to that observed in plasma DNA. We found alterations in different chromosomes in blood-spots from two patients with high-grade serous ovarian carcinoma. Extending this approach to serial dried blood spots from mouse xenograft models, we detect tumor-derived cell-free DNA and identified ctDNA from the originally grafted ascites. Conclusion: Our data suggest that ctDNA can be detected and monitored in dried blood spots from archived and fresh blood samples, enabling new approaches for sample collection and novel study/trial designs for both patients and in vivo models.The University of Cambridge and Cancer Research UK (grant numbers A20240 and A29580). The research leading to these results has received funding from the European Research Council under the European Union’s Seventh Framework Programme (FP/2007-2013)/ERC Grant Agreement n.337905. Healthy volunteer samples were provided by the Cambridge Blood and Stem Cell Biobank, which is supported by the Cambridge NIHR Biomedical Research Centre, Wellcome Trust - MRC Stem Cell Institute and the Cambridge Experimental Cancer Medicine Centre, UK

Conformational effects on the Circular Dichroism of Human Carbonic Anhydrase II: a multilevel computational study

Circular Dichroism (CD) spectroscopy is a powerful method for investigating conformational changes in proteins and therefore has numerous applications in structural and molecular biology. Here a computational investigation of the CD spectrum of the Human Carbonic Anhydrase II (HCAII), with main focus on the near-UV CD spectra of the wild-type enzyme and it seven tryptophan mutant forms, is presented and compared to experimental studies. Multilevel computational methods (Molecular Dynamics, Semiempirical Quantum Mechanics, Time-Dependent Density Functional Theory) were applied in order to gain insight into the mechanisms of interaction between the aromatic chromophores within the protein environment and understand how the conformational flexibility of the protein influences these mechanisms. The analysis suggests that combining CD semi empirical calculations, crystal structures and molecular dynamics (MD) could help in achieving a better agreement between the computed and experimental protein spectra and provide some unique insight into the dynamic nature of the mechanisms of chromophore interactions

Fragmentation patterns and personalized sequencing of cell-free DNA in urine and plasma of glioma patients.

Glioma-derived cell-free DNA (cfDNA) is challenging to detect using liquid biopsy because quantities in body fluids are low. We determined the glioma-derived DNA fraction in cerebrospinal fluid (CSF), plasma, and urine samples from patients using sequencing of personalized capture panels guided by analysis of matched tumor biopsies. By sequencing cfDNA across thousands of mutations, identified individually in each patient's tumor, we detected tumor-derived DNA in the majority of CSF (7/8), plasma (10/12), and urine samples (10/16), with a median tumor fraction of 6.4 × 10-3 , 3.1 × 10-5 , and 4.7 × 10-5 , respectively. We identified a shift in the size distribution of tumor-derived cfDNA fragments in these body fluids. We further analyzed cfDNA fragment sizes using whole-genome sequencing, in urine samples from 35 glioma patients, 27 individuals with non-malignant brain disorders, and 26 healthy individuals. cfDNA in urine of glioma patients was significantly more fragmented compared to urine from patients with non-malignant brain disorders (P = 1.7 × 10-2 ) and healthy individuals (P = 5.2 × 10-9 ). Machine learning models integrating fragment length could differentiate urine samples from glioma patients (AUC = 0.80-0.91) suggesting possibilities for truly non-invasive cancer detection

Prenatal maternal plasma DNA screening for cystic fibrosis: A computer modelling study of screening performance.

Background: Prenatal cystic fibrosis (CF) screening is currently based on determining the carrier status of both parents. We propose a new method based only on the analysis of DNA in maternal plasma. Methods: The method relies on the quantitative amplification of the CF gene to determine the percentage of DNA fragments in maternal plasma at targeted CF mutation sites that carry a CF mutation. Computer modelling was carried out to estimate the distributions of these percentages in pregnancies with and without a fetus affected with CF. This was done according to the number of DNA fragments counted and fetal fraction, using the 23 CF mutations recommended by the American College of Medical Genetics for parental carrier testing. Results: The estimated detection rate (sensitivity) is 70% (100% of those detected using the 23 mutations), the false-positive rate 0.002%, and the odds of being affected given a positive screening result 14:1, compared with 70%, 0.12%, and 1:3, respectively, with current prenatal screening based on parental carrier testing. Conclusions: Compared with current screening practice based on parental carrier testing, the proposed method would substantially reduce the number of invasive diagnostic procedures (amniocentesis or chorionic villus sampling) without reducing the CF detection rate. The expected advantages of the proposed method justify carrying out the necessary test development for use in a clinical validation study.The author(s) declared that no grants were involved in supporting this work

CXCR4 inhibition in human pancreatic and colorectal cancers induces an integrated immune response.

Inhibition of the chemokine receptor CXCR4 in combination with blockade of the PD-1/PD-L1 T cell checkpoint induces T cell infiltration and anticancer responses in murine and human pancreatic cancer. Here we elucidate the mechanism by which CXCR4 inhibition affects the tumor immune microenvironment. In human immune cell-based chemotaxis assays, we find that CXCL12-stimulated CXCR4 inhibits the directed migration mediated by CXCR1, CXCR3, CXCR5, CXCR6, and CCR2, respectively, chemokine receptors expressed by all of the immune cell types that participate in an integrated immune response. Inhibiting CXCR4 in an experimental cancer medicine study by 1-wk continuous infusion of the small-molecule inhibitor AMD3100 (plerixafor) induces an integrated immune response that is detected by transcriptional analysis of paired biopsies of metastases from patients with microsatellite stable colorectal and pancreatic cancer. This integrated immune response occurs in three other examples of immune-mediated damage to noninfected tissues: Rejecting renal allografts, melanomas clinically responding to anti-PD1 antibody therapy, and microsatellite instable colorectal cancers. Thus, signaling by CXCR4 causes immune suppression in human pancreatic ductal adenocarcinoma and colorectal cancer by impairing the function of the chemokine receptors that mediate the intratumoral accumulation of immune cells.Stand Up 2 Cancer, Lustgarten Foundation, NIH