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At least 433 records · Page 24

High‐quality genome‐scale metabolic modelling of Pseudomonas putida highlights its broad metabolic capabilities

Summary Genome‐scale reconstructions of metabolism are computational species‐specific knowledge bases able to compute systemic metabolic properties. We present a comprehensive and validated reconstruction of the biotechnologically relevant bacterium Pseudomonas putida KT2440 that greatly expands computable predictions of its metabolic states. The reconstruction represents a significant reactome expansion over available reconstructed bacterial metabolic networks. Specifically, i JN1462 (i) incorporates several hundred additional genes and associated reactions resulting in new predictive capabilities, including new nutrients supporting growth; (ii) was validated by in vivo growth screens that included previously untested carbon (48) and nitrogen (41) sources; (iii) yielded gene essentiality predictions showing large accuracy when compared with a knock‐out library and Bar‐seq data; and (iv) allowed mapping of its network to 82 P . putida sequenced strains revealing functional core that reflect the large metabolic versatility of this species, including aromatic compounds derived from lignin. Thus, this study provides a thoroughly updated metabolic reconstruction and new computable phenotypes for P . putida , which can be leveraged as a first step toward understanding the pan metabolic capabilities of Pseudomonas .

Microbiology↗

A cell wall reference profile for Miscanthus bioenergy crops highlights compositional and structural variations associated with development and organ origin

Summary Miscanthus spp. are promising lignocellulosic energy crops, but cell wall recalcitrance to deconstruction still hinders their widespread use as bioenergy and biomaterial feedstocks. Identification of cell wall characteristics desirable for biorefining applications is crucial for lignocellulosic biomass improvement. However, the task of scoring biomass quality is often complicated by the lack of a reference for a given feedstock. A multidimensional cell wall analysis was performed to generate a reference profile for leaf and stem biomass from several miscanthus genotypes harvested at three developmentally distinct time points. A comprehensive suite of 155 monoclonal antibodies was used to monitor changes in distribution, structure and extractability of noncellulosic cell wall matrix glycans. Glycan microarrays complemented with immunohistochemistry elucidated the nature of compositional variation, and in situ distribution of carbohydrate epitopes. Key observations demonstrated that there are crucial differences in miscanthus cell wall glycomes, which may impact biomass amenability to deconstruction. For the first time, variations in miscanthus cell wall glycan components were comprehensively characterized across different harvests, organs and genotypes, to generate a representative reference profile for miscanthus cell wall biomass. Ultimately, this portrait of the miscanthus cell wall will help to steer breeding and genetic engineering strategies for the development of superior energy crops.

Plant Sciences↗

Comparative genomics of Rhizophagus irregularis , R. cerebriforme , R. diaphanus and Gigaspora rosea highlights specific genetic features in Glomeromycotina

Glomeromycotina is a lineage of early diverging fungi that establish arbuscular mycorrhizal (AM) symbiosis with land plants. Despite their major ecological role, the genetic basis of their obligate mutualism remains largely unknown, hindering our understanding of their evolution and biology. Here, we compared the genomes of Glomerales (Rhizophagus irregularis, Rhizophagus diaphanus, Rhizophagus cerebriforme) and Diversisporales (Gigaspora rosea) species, together with those of saprotrophic Mucoromycota, to identify gene families and processes associated with these lineages and to understand the molecular underpinning of their symbiotic lifestyle. Genomic features in Glomeromycotina appear to be very similar with a very high content in transposons and protein-coding genes, extensive duplications of protein kinase genes, and loss of genes coding for lignocellulose degradation, thiamin biosynthesis and cytosolic fatty acid synthase. Most symbiosis-related genes in R. irregularis and G. rosea are specific to Glomeromycotina. We also confirmed that the present species have a homokaryotic genome organisation. The high interspecific diversity of Glomeromycotina gene repertoires, affecting all known protein domains, as well as symbiosis-related orphan genes, may explain the known adaptation of Glomeromycotina to a wide range of environmental settings. Our findings contribute to an increasingly detailed portrait of genomic features defining the biology of AM fungi.

59 BASIC BIOLOGICAL SCIENCES↗

Promiscuous terpene synthases from Prunella vulgaris highlight the importance of substrate and compartment switching in terpene synthase evolution

The mint family (Lamiaceae) is well documented as a rich source of terpene natural products. More than 200 diterpene skeletons have been reported from mints, but biosynthetic pathways are known for just a few of these. We crossreferenced chemotaxonomic data with publicly available transcriptomes to select common selfheal (Prunella vulgaris) and its highly unusual vulgarisin diterpenoids as a case study for exploring the origins of diterpene skeletal diversity in Lamiaceae. Four terpene synthases (TPS) from the TPS-a subfamily, including two localised to the plastid, were cloned and functionally characterised. Previous examples of TPS-a enzymes from Lamiaceae were cytosolic and reported to act on the 15-carbon farnesyl diphosphate. Plastidial TPS-a enzymes using the 20-carbon geranylgeranyl diphosphate are known from other plant families, having apparently arisen independently in each family. All four new enzymes were found to be active on multiple prenyl-diphosphate substrates with different chain lengths and stereochemistries. One of the new enzymes catalysed the cyclisation of geranylgeranyl diphosphate into 11-hydroxy vulgarisane, the likely biosynthetic precursor of the vulgarisins. We uncovered the pathway to a rare diterpene skeleton. Our results support an emerging paradigm of substrate and compartment switching as important aspects of TPS evolution and diversification.

59 BASIC BIOLOGICAL SCIENCES↗

Genome-wide scans of selection highlight the impact of biotic and abiotic constraints in natural populations of the model grass Brachypodium distachyon

Grasses are essential plants for ecosystem functioning. Quantifying the selective pressures that act on natural variation in grass species is therefore essential regarding biodiversity maintenance. Here, in this study, we investigate the selection pressures that act on two distinct populations of the grass model Brachypodium distachyon without prior knowledge about the traits under selection. We took advantage of whole-genome sequencing data produced for 44 natural accessions of B. distachyon and used complementary genome-wide selection scans (GWSS) methods to detect genomic regions under balancing and positive selection. We show that selection is shaping genetic diversity at multiple temporal and spatial scales in this species, and affects different genomic regions across the two populations. Gene ontology annotation of candidate genes reveals that pathogens may constitute important factors of positive and balancing selection in B. distachyon. We eventually cross-validated our results with quantitative trait locus data available for leaf-rust resistance in this species and demonstrate that, when paired with classical trait mapping, GWSS can help pinpointing candidate genes for further molecular validation. Thanks to a near base-perfect reference genome and the large collection of freely available natural accessions collected across its natural range, B. distachyon appears as a prime system for studies in ecology, population genomics and evolutionary biology.

60 APPLIED LIFE SCIENCES↗

MO-E-BRF-01: Research Opportunities in Technology for Innovation in Radiation Oncology (Highlight of ASTRO NCI 2013 Workshop)

Radiotherapy is one of the most effective treatments for solid tumors, in large part due to significant technological advances associated with, for instance, the ability to target tumors to very high levels of accuracy (within millimeters). Technological advances have played a central role in the success of radiation therapy as an oncologic treatment option for patients. ASTRO, AAPM and NCI sponsored a workshop “Technology for Innovation in Radiation Oncology” at the NCI campus in Bethesda, MD on June 13–14, 2013. The purpose of this workshop was to bring together expert clinicians and scientists to discuss the role of disruptive technologies in radiation oncology, in particular with regard to how they are being developed and translated to clinical practice in the face of current and future challenges and opportunities. The technologies discussed encompassed imaging and delivery aspects, along with methods to enable/facilitate application of them in the clinic. Measures for assessment of the performance of these technologies, such as techniques to validate quantitative imaging, were reviewed. Novel delivery technologies, incorporating efficient and safe delivery mechanisms enabled by development of tools for process automation and the associated field of oncology informatics formed one of the central themes of the workshop. The discussion on disruptive technologies was grounded in the need for evidence of efficacy. Scientists in the areas of technology assessment and bioinformatics provided expert views on different approaches toward evaluation of technology efficacy. Clinicians well versed in clinical trials incorporating disruptive technologies (e.g. SBRT for early stage lung cancer) discussed the important role of these technologies in significantly improving local tumor control and survival for these cohorts of patients. Recommendations summary focused on the opportunities associated with translating the technologies into the clinic and assessing their efficacy, and provided a glimpse into the future. Learning Objectives: To understand the impact of technology on the field of radiation therapy To learn about the trends of technology development for the field of radiation oncology To understand the opportunities for in innovative technology research.

60 APPLIED LIFE SCIENCES↗

Sci-Thur PM – Colourful Interactions: Highlights 01: Design to delivery of spatially fractionated mini-beam canine radiotherapy

Spatial fractionation of radiation using arrays of narrow parallel micro-planar beams (less than 1 mm), is a relatively new concept with many unknowns specifically within the underlying biology of cell death. A tungsten collimator has been designed to produce mini-beams with a Varian linear accelerator for translational animal research into the effectiveness of spatial fractionation mini-beam radiotherapy (MBRT). This work presents the treatment planning process and workflow for the application of MBRT treatments within a clinical study. For patient dose calculations, the MBRT collimator was incorporated into a Monte Carlo based treatment planning system called MMCTP. Treatment planning was split between Eclipse and MMCTP, as the field apertures were determined within Eclipse prior to being sent to MMCTP for dose calculations. The calculated plan was transferred back into Aria with updated MUs per field for patient treatment. Patients were positioned within a vac-lock bag lying prone with a bite block and a thermoplastic mask to immobilize the head. Prior to treatment, a delivery verification plan was created within MMCTP. DQA output measurements of the treatment fields agreed with the calculated dose to within 1.5%. We have presented a workflow for MBRT treatments that include the planning technique, dose calculation method, DQA process and data integration into a record and verify system. The clinical study following this workflow represent the first series of linac based MBRT patients and depending on the clinical outcome of the study, our technique could be applied to human MBRT treatments.

60 APPLIED LIFE SCIENCES↗

Sci-Thur PM – Colourful Interactions: Highlights 03: Radiation induced glycogen accumulation in non-small cell lung cancer xenografts detected using Raman spectroscopy

Purpose: This study presents the novel application of Raman spectroscopy (RS) to identify biochemical signatures of radiation response in human non-small cell lung cancer (NSCLC) xenografts, irradiated in vivo. Methods: Human NSCLC cells (H460) were subcutaneously injected into the flanks of 12 mice. Tumours were treated with single fraction radiation doses (0, 5 or 15 Gy) and harvested at 3 days post irradiation. A Renishaw inVia Raman microscope coupled to a 785 nm laser was used to collect Raman spectral maps for each tumour. Immunohistochemistry (IHC) staining for CAIX was used to visualize hypoxia, and co-registration between IHC fluorescence and Raman images was carried out. Results: Principal component analysis revealed radiation induced spectral signatures linked to changes in protein, nucleic acid, lipid and carbohydrates. In particular, a marked increase in glycogen for irradiated tumours was observed. Spatial mapping revealed intra-tumoural heterogeneity in the distribution of glycogen within the tumour, suggesting tumour response to radiation is not globally uniform. Furthermore, co-registration of Raman glycogen maps with CAIX IHC staining showed a correlation between glycogen rich and hypoxic regions of the tissue. Conclusions: We identify glycogen as a unique radiation induced response in NSCLC tumour xenografts, which may reflect inherent metabolic changes associated with radiation response in tissue. This study provides unique insight into the biochemical response of tumours, irradiated in vivo, and demonstrates the potential of RS for detecting radiobiological responses in tumours.

60 APPLIED LIFE SCIENCES↗

Sci-Thur PM - Colourful Interactions: Highlights 04: A Fast Quantitative MRI Acquisition and Processing Pipeline for Radiation Treatment Planning and Simulation

MRI-only Radiation Treatment Planning (RTP) is becoming increasingly popular because of a simplified work-flow, and less inconvenience to the patient who avoids multiple scans. The advantages of MRI-based RTP over traditional CT-based RTP lie in its superior soft-tissue contrast, and absence of ionizing radiation dose. The lack of electron-density information in MRI can be addressed by automatic tissue classification. To distinguish bone from air, which both appear dark in MRI, an ultra-short echo time (UTE) pulse sequence may be used. Quantitative MRI parametric maps can provide improved tissue segmentation/classification and better sensitivity in monitoring disease progression and treatment outcome than standard weighted images. Superior tumor contrast can be achieved on pure T{sub 1} images compared to conventional T{sub 1}-weighted images acquired in the same scan duration and voxel resolution. In this study, we have developed a robust and fast quantitative MRI acquisition and post-processing work-flow that integrates these latest advances into the MRI-based RTP of brain lesions. Using 3D multi-echo FLASH images at two different optimized flip angles (both acquired in under 9 min, and 1mm isotropic resolution), parametric maps of T{sub 1}, proton-density (M{sub 0}), and T{sub 2}{sup *} are obtained with high contrast-to-noise ratio, and negligible geometrical distortions, water-fat shifts and susceptibility effects. An additional 3D UTE MRI dataset is acquired (in under 4 min) and post-processed to classify tissues for dose simulation. The pipeline was tested on four healthy volunteers and a clinical trial on brain cancer patients is underway.

60 APPLIED LIFE SCIENCES↗

Sci-Thur PM – Colourful Interactions: Highlights 05: Opal–the Oncology Patient Application

We describe Opal (Oncology portal and application), the mobile phone app and patient portal that we have developed and are deploying for Radiation Oncology patients at our cancer centre. Opal is a novel tool to empower patients with their own personal medical data, including appointment schedules, consultation notes, test results, radiotherapy treatment planning information and wait time management. Furthermore, due to its integration with our electronic medical record and treatment planning database, Opal will allow us to collect patient reported outcomes from consenting patients and link them directly with dose volume histograms and other treatment data.

60 APPLIED LIFE SCIENCES↗

Sci-Thur PM - Colourful Interactions: Highlights 06: Patient-specific calibration of cone-beam computed tomographic (CBCT) image sets for on-line dose tracking and plan assessment

Purpose: To develop a radiotherapy dose tracking and plan evaluation technique using cone-beam computed tomography (CBCT) images. Methods: We developed a patient-specific method of calibrating CBCT image sets for dose calculation. The planning CT was first registered with the CBCT using deformable image registration (DIR). A scatter plot was generated between the CT numbers of the planning CT and CBCT for each slice. The CBCT calibration curve was obtained by least-square fitting of the data, and applied to each CBCT slice. The calibrated CBCT was then merged with original planning CT to extend the small field of view of CBCT. Finally, the treatment plan was copied to the merged CT for dose tracking and plan evaluation. The proposed patient-specific calibration method was also compared to two methods proposed in literature. To evaluate the accuracy of each technique, 15 head-and-neck patients requiring plan adaptation were arbitrarily selected from our institution. The original plan was calculated on each method’s data set, including a second planning CT acquired within 48 hours of the CBCT (serving as gold standard). Clinically relevant dose metrics and 3D gamma analysis of dose distributions were compared between the different techniques. Results: Compared to the gold standard of using planning CTs, the patient-specific CBCT calibration method was shown to provide promising results with gamma pass rates above 95% and average dose metric agreement within 2.5%. Conclusions: The patient-specific CBCT calibration method could potentially be used for on-line dose tracking and plan evaluation, without requiring a re-planning CT session.

60 APPLIED LIFE SCIENCES↗

Sci-Thur PM – Colourful Interactions: Highlights 07: Canadian Computed Tomography Survey: National Diagnostic Reference Levels

Purpose: The Canadian Computed (CT) Tomography Survey sought to collect CT technology and dose index data (CTDI and DLP) at the national level in order to establish national diagnostic reference levels (DRLs) for seven common CT examinations of standard-sized adults and pediatric patients. Methods: A single survey booklet (consisting of four sections) was mailed to and completed for each participating CT scanner. Survey sections collected data on (i) General facility and scanner information, (ii) routine protocols (as available), (iii) individual patient data (as applied) and (iv) manual CTDI measurements. Results: Dose index (CTDIvol and DLP) and associated patient data from 24 280 individual patient exam sequences was analyzed for seven common CT examinations performed in Canada: Adult Head, Chest, Abdomen/Pelvis, and Chest/Abdomen/Pelvis, and Pediatric Head, Chest, and Abdomen. Pediatric examination data was sub-divided into three age ranges: 0–3, 3–7 and 7–13 years. DRLs (75th percentile of dose index distributions) were found for all thirteen groups. Further analysis also permitted segmentation of examination data into 8 sub-groups, whose dose index data was displayed along with group histograms – showing relative contribution of axial vs. helical, contrast use (C+ vs. C-), and application of fixed current vs. dose reduction (DR) – 75th percentiles of DR sub-groups were, in almost all cases, lower than whole group (examination) DRLs. Conclusions: The analysis and summaries presented in the pending survey report can serve to aid local CT imaging optimization efforts within Canada and also contribute further to international efforts in radiation protection of patients.

60 APPLIED LIFE SCIENCES↗

Sci-Thur PM - Colourful Interactions: Highlights 08: ARC TBI using Single-Step Optimized VMAT Fields

Purpose: This work outlines a new TBI delivery technique to replace a lateral POP full bolus technique. The new technique is done with VMAT arc delivery, without bolus, treating the patient prone and supine. The benefits of the arc technique include: increased patient experience and safety, better dose conformity, better organ at risk sparing, decreased therapist time and reduction of therapist injuries. Methods: In this work we build on a technique developed by Jahnke et al. We use standard arc fields with gantry speeds corrected for varying distance to the patient followed by a single step VMAT optimization on a patient CT to increase dose inhomogeneity and to reduce dose to the lungs (vs. blocks). To compare the arc TBI technique to our full bolus technique, we produced plans on patient CTs for both techniques and evaluated several dosimetric parameters using an ANOVA test. Results and Conclusions: The arc technique is able reduce both the hot areas to the body (D2% reduced from 122.2% to 111.8% p<0.01) and the lungs (mean lung dose reduced from 107.5% to 99.1%, p<0.01), both statistically significant, while maintaining coverage (D98% = 97.8% vs. 94.6%, p=0.313, not statistically significant). We developed a more patient and therapist-friendly TBI treatment technique that utilizes single-step optimized VMAT plans. It was found that this technique was dosimetrically equivalent to our previous lateral technique in terms of coverage and statistically superior in terms of reduced lung dose.

60 APPLIED LIFE SCIENCES↗

Metabolomics Highlights Different Life History Strategies of White and Brown Rot Wood-Degrading Fungi

Despite the ecological and economic importance of wood-degrading fungi, little is known about the array of metabolites that fungi produce during wood decomposition. This study provides an in-depth insight into the wood decomposition process by analyzing and comparing the changes of >100 compounds produced by fungi with metabolic distinct nutritional modes (white and brown rot fungi) at different decay stages.

59 BASIC BIOLOGICAL SCIENCES↗

Hantavirus is Associated With Open Developed Areas and Arid Climates, Highlighting Increased Risk in the Western United States

In the United States, hantaviruses can cause hantavirus pulmonary syndrome (HPS) in humans, an acute respiratory illness with a high mortality rate. Most people contract HPS from exposure to infected rodent excrement. The interannual dynamics of hantavirus transmission are tied to both environmental and human-related factors, including changes in annual climate conditions, rodent populations, and the built environment in which humans are more likely to be exposed. Similar environmental conditions and socioeconomic factors also likely determine the long-term risk of hantavirus exposure. Here, we use ecological niche models and human cases of HPS in the U.S. from 1993 to 2022 to assess hantavirus risk using four socioeconomic variables, 17 land use variables, one variable of rodent richness, and seven climate variables to determine both the geographical locations of highest exposure risk and leading environmental predictors. We found that areas with higher relative risk tend to be where it is drier, higher social vulnerability, increased rodent richness, and more open to low levels of development—this largely mapped to the western U.S. We found evidence that fringe ecosystems may be important areas of hantavirus transmission, similar to other emerging diseases. Increased rodent richness was associated with increased hantavirus risk, warranting further investigation into how the abundance and community composition of rodents could impact long-term risk. These risk maps can help public health officials develop plans for mitigating hantavirus, especially for the most susceptible populations. They can also be used to further investigate regions estimated to be at high risk for hantavirus where disease cases have not been as common but may be underreported.

54 ENVIRONMENTAL SCIENCES↗