Unanticipated Large-Scale Deletion in Fusarium graminearum Genome Using CRISPR/Cas9 and Its Impact on Growth and Virulence.

Fusarium graminearum, a filamentous fungus, and causal agent of Fusarium head blight (FHB) in wheat and other cereals, leads to significant economic losses globally. This study aimed to investigate the roles of specific genes in F. graminearum virulence using CRISPR/Cas9-mediated gene deletions. Illumina sequencing was used to characterize the genomic changes due to editing. Unexpectedly, a large-scale deletion of 525,223 base pairs on chromosome 2, comprising over 222 genes, occurred in two isolates. Many of the deleted genes were predicted to be involved in essential molecular functions, such as oxidoreductase activity, transmembrane transporter activity, hydrolase activity, as well as biological processes, such as carbohydrate metabolism and transmembrane transport. Despite the substantial loss of genetic material, the mutant isolate exhibited normal growth rates and virulence on wheat under most conditions. However, growth rates were significantly reduced under high temperatures and on some media. Additionally, wheat inoculation assays using clip dipping, seed inoculation, and head point inoculation methods were performed. No significant differences in virulence were observed, suggesting that these genes were not involved in infection or alternative compensatory pathways, and allow the fungi to maintain pathogenicity despite the extensive genomic deletion.

The impact of continuously-variable dose rate VMAT on beam stability, MLC positioning, and overall plan dosimetry.

A recent control system update for Elekta linear accelerators includes the ability to deliver volumetric-modulated arc therapy (VMAT) with continuously variable dose rate (CVDR), rather than a number of fixed binned dose rates (BDR). The capacity to select from a larger range of dose rates allows the linac to maintain higher gantry speeds, resulting in faster, smoother deliveries. The purpose of this study is to investigate two components of CVDR delivery - the increase in average dose rate and gantry speed, and a determination of their effects on beam stability, MLC positioning, and overall plan dosimetry. Initially, ten VMAT plans (5 prostate, 5head and neck) were delivered to a Delta4 dosimetric phantom using both the BDR and CVDR systems. The plans were found to be dosimetrically robust using both delivery methods, although CVDR was observed to give higher gamma pass rates at the 2%/2 mm gamma level for prostates (p < 0.01). For the dual arc head-and-neck plans, CVDR delivery resulted in improved pass rates at all gamma levels (2%/2 mm to 4%/4 mm) for individual arc verifications (p < 0.01), but gave similar results to BDR when both arcs were combined. To investigate the impact of increased gantry speed on MLC positioning, a dynamic leaf-tracking tool was developed using the electronic portal imaging device (EPID). Comparing the detected MLC positions to those expected from the plan, CVDR was observed to result in a larger mean error compared to BDR (0.13 cm and 0.06 cm, respectively, p < 0.01). The EPID images were also used to monitor beam stability during delivery. It was found that the CVDR deliveries had a lower standard deviation of the gun-target (GT) and transverse (AB) profiles (p < 0.01). This study has determined that CVDR may offer a dosimetric advantage for VMAT plans. While the higher gantry speed of CVDR appears to increase deviations in MLC positioning, the relative effect on dosimetry is lower than the positive impact of a flatter and more stable beam profile.

Evaluating the repeatability and set-up sensitivity of a large field of view distortion phantom and software for magnetic resonance-only radiotherapy.

BACKGROUND AND PURPOSE: Magnetic Resonance (MR)-only radiotherapy requires geometrically accurate MR images over the full scanner Field of View (FoV). This study aimed to investigate the repeatability of distortion measurements made using a commercial large FoV phantom and analysis software and the sensitivity of these measurements to small set-up errors. MATERIALS AND METHODS: Geometric distortion was measured using a commercial phantom and software with 2D and 3D acquisition sequences on three different MR scanners. Two sets of repeatability measurements were made: three scans acquired without moving the phantom between scans (single set-up) and five scans acquired with the phantom re-set up in between each scan (repeated set-up). The set-up sensitivity was assessed by scanning the phantom with an intentional 1 mm lateral offset and independently an intentional 1° rotation. RESULTS: The mean standard deviation of distortion for all phantom markers for the repeated set-up scans was < 0.4 mm for all scanners and sequences. For the 1 mm lateral offset scan 90 % of the markers agreed within two standard deviations of the mean of the repeated set-up scan (median of all scanners and sequences, range 78%-93%). For the 1° rotation scan, 80% of markers agreed within two standard deviations of the mean (range 69%-93%). CONCLUSIONS: Geometric distortion measurements using a commercial phantom and associated software appear repeatable, although with some sensitivity to set-up errors. This suggests the phantom and software are appropriate for commissioning a MR-only radiotherapy workflow.

Systematic Review of Synthetic Computed Tomography Generation Methodologies for Use in Magnetic Resonance Imaging-Only Radiation Therapy.

Magnetic resonance imaging (MRI) offers superior soft-tissue contrast as compared with computed tomography (CT), which is conventionally used for radiation therapy treatment planning (RTP) and patient positioning verification, resulting in improved target definition. The 2 modalities are co-registered for RTP; however, this introduces a systematic error. Implementing an MRI-only radiation therapy workflow would be advantageous because this error would be eliminated, the patient pathway simplified, and patient dose reduced. Unlike CT, in MRI there is no direct relationship between signal intensity and electron density; however, various methodologies for MRI-only RTP have been reported. A systematic review of these methods was undertaken. The PRISMA guidelines were followed. Embase and Medline databases were searched (1996 to March, 2017) for studies that generated synthetic CT scans (sCT)s for MRI-only radiation therapy. Sixty-one articles met the inclusion criteria. This review showed that MRI-only RTP techniques could be grouped into 3 categories: (1) bulk density override; (2) atlas-based; and (3) voxel-based techniques, which all produce an sCT scan from MR images. Bulk density override techniques either used a single homogeneous or multiple tissue override. The former produced large dosimetric errors (>2%) in some cases and the latter frequently required manual bone contouring. Atlas-based techniques used both single and multiple atlases and included methods incorporating pattern recognition techniques. Clinically acceptable sCTs were reported, but atypical anatomy led to erroneous results in some cases. Voxel-based techniques included methods using routine and specialized MRI sequences, namely ultra-short echo time imaging. High-quality sCTs were produced; however, use of multiple sequences led to long scanning times increasing the chances of patient movement. Using nonroutine sequences would currently be problematic in most radiation therapy centers. Atlas-based and voxel-based techniques were found to be the most clinically useful methods, with some studies reporting dosimetric differences of <1% between planning on the sCT and CT and <1-mm deviations when using sCTs for positional verification.

Investigating the generalisation of an atlas-based synthetic-CT algorithm to another centre and MR scanner for prostate MR-only radiotherapy.

There is increasing interest in MR-only radiotherapy planning since it provides superb soft-tissue contrast without the registration uncertainties inherent in a CT-MR registration. However, MR images cannot readily provide the electron density information necessary for radiotherapy dose calculation. An algorithm which generates synthetic CTs for dose calculations from MR images of the prostate using an atlas of 3 T MR images has been previously reported by two of the authors. This paper aimed to evaluate this algorithm using MR data acquired at a different field strength and a different centre to the algorithm atlas. Twenty-one prostate patients received planning 1.5 T MR and CT scans with routine immobilisation devices on a flat-top couch set-up using external lasers. The MR receive coils were supported by a coil bridge. Synthetic CTs were generated from the planning MR images with ([Formula: see text]) and without (sCT) a one voxel body contour expansion included in the algorithm. This was to test whether this expansion was required for 1.5 T images. Both synthetic CTs were rigidly registered to the planning CT (pCT). A 6 MV volumetric modulated arc therapy plan was created on the pCT and recalculated on the sCT and [Formula: see text]. The synthetic CTs' dose distributions were compared to the dose distribution calculated on the pCT. The percentage dose difference at isocentre without the body contour expansion (sCT-pCT) was [Formula: see text] and with ([Formula: see text]-pCT) was [Formula: see text] (mean ± one standard deviation). The [Formula: see text] result was within one standard deviation of zero and agreed with the result reported previously using 3 T MR data. The sCT dose difference only agreed within two standard deviations. The mean ± one standard deviation gamma pass rate was [Formula: see text] for the sCT and [Formula: see text] for the [Formula: see text] (with [Formula: see text] global dose difference and [Formula: see text] distance to agreement gamma criteria). The one voxel body contour expansion improves the synthetic CT accuracy for MR images acquired at 1.5 T but requires the MR voxel size to be similar to the atlas MR voxel size. This study suggests that the atlas-based algorithm can be generalised to MR data acquired using a different field strength at a different centre.