Concept and performance evaluation of two 3 GHz buncher units optimizing the dose rate of a novel preclinical proton minibeam irradiation facility.

To demonstrate the large potential of proton minibeam radiotherapy (pMBRT) as a new method to treat tumor diseases, a preclinical proton minibeam radiation facility was designed. It is based on a tandem Van-de-Graaff accelerator providing a 16 MeV proton beam and a 3 GHz linac post-accelerator (designs: AVO-ADAM S.A, Geneva, Switzerland and ENEA, Frascati, Italy). To enhance the transmission of the tandem beam through the post-accelerator by a factor of 3, two drift tube buncher units were designed and constructed: A brazed 5-gap structure (adapted SCDTL tank of the TOP-IMPLART project (ENEA)) and a non-brazed low budget 4-gap structure. Both are made of copper. The performance of the two differently manufactured units was evaluated using a 16 MeV tandem accelerator beam and a Q3D magnetic spectrograph. Both buncher units achieve the required summed voltage amplitude of 42 kV and amplitude stability at a power feed of less than 800 W.

Magnetically focused 70 MeV proton minibeams for preclinical experiments combining a tandem accelerator and a 3 GHz linear post-accelerator.

PURPOSE: Radiotherapy plays an important role for the treatment of tumor diseases in two-thirds of all cases, but it is limited by side effects in the surrounding healthy tissue. Proton minibeam radiotherapy (pMBRT) is a promising option to widen the therapeutic window for tumor control at reduced side effects. An accelerator concept based on an existing tandem Van de Graaff accelerator and a linac enables the focusing of 70 MeV protons to form minibeams with a size of only 0.1 mm for a preclinical small animal irradiation facility, while avoiding the cost of an RFQ injector. METHODS: The tandem accelerator provides a 16 MeV proton beam with a beam brightness of B = 4 nA mm 2 mrad 2 as averaged from 5 µs long pulses with a flat top current of 17 µA at 200 Hz repetition rate. Subsequently, the protons are accelerated to 70 MeV by a 3 GHz linear post-accelerator consisting of two Side Coupled Drift Tube Linac (SCDTL) structures and four Coupled Cavity Linac (CCL) structures [design: AVO-ADAM S.A (Geneva, Switzerland)]. A 3 GHz buncher and four magnetic quadrupole lenses are placed between the tandem and the post-accelerator to maximize the transmission through the linac. A quadrupole triplet situated downstream of the linac structure focuses the protons into an area of (0.1 × 0.1) mm2 . The beam dynamics of the facility is optimized using the particle optics code TRACE three-dimensional (3D). Proton transmission through the facility is elaborated using the particle tracking code TRAVEL. RESULTS: A study about buncher amplitude and phase shift between buncher and linac is showing that 49% of all protons available from the tandem can be transported through the post-accelerator. A mean beam current up to 19 nA is expected within an area of (0.1 × 0.1) mm2 at the beam focus. CONCLUSION: An extension of existing tandem accelerators by commercially available 3 GHz structures is able to deliver a proton minibeam that serves all requirements to obtain proton minibeams to perform preclinical minibeam irradiations as it would be the case for a complete commercial 3 GHz injector-RFQ-linac combination. Due to the modularity of the linac structure, the irradiation facility can be extended to clinically relevant proton energies up to or above 200 MeV.

Aspartic Protease Zymography Case Study: Detection of Fungal Acid Proteases by Zymography.

This chapter describes a method for the production and characterization of fungal acid proteases. Protease production is induced by growth on BSA media over a pH gradient and protein levels are monitored over time with the Bradford assay. Once protein is depleted, the media is purified and proteases are characterized by gelatin zymography using acrylamide and buffers at near-neutral pH. Maintaining pH levels below those found in traditional zymographic systems avoids the potential loss of activity that may occur in aspartic proteases under alkaline conditions.

Fungal endophytes of wild and hybrid Vitis leaves and their potential for vineyard biocontrol.

Plants are colonized by diverse assemblages of fungal endophytes that have potential as biocontrol agents for a variety of crops, including grapevine. Although the diversity of symbionts can be very high in wild plants, the fungal endophytes of wild Vitis plants have not yet been investigated. We surveyed the fungal endophytes of 6 wild populations of Vitis riparia, as well as a cold-tolerant, hybrid grapevine in 5 vineyards (1 certified organic), using 454 pyrosequencing. We detected between 43 and 235 operational taxonomic units per sample, with the highest richness and diversity in the wild, the lowest in conventional vineyards, and intermediate levels in the organic vineyard. Wild plants supported a range of taxa not seen in the conventional vineyards, and vineyards were dominated by relatively few taxa. We also isolated fungi from the wild plants and tested them for their ability to inhibit pathogens of grapevine. Several wild isolates (e.g., Ramularia spp.) were strongly inhibitory to grapevine pathogens. We show that wild Vitis supports a distinct and highly diverse community of fungal endophytes and may represent a rich repository of potential vineyard biocontrol agents.

Acid protease production in fungal root endophytes.

Fungal endophytes are ubiquitous in healthy root tissue, but little is known about their ecosystem functions, including their ability to utilize organic nutrient sources such as proteins. Root-associated fungi may secrete proteases to access the carbon and mineral nutrients within proteins in the soil or in the cells of their plant host. We compared the protein utilization patterns of multiple isolates of the root endophytes Phialocephala fortinii s.l., Meliniomyces variabilis and Umbelopsis isabellina with those of two ectomycorrhizal (ECM) fungi, Hebeloma incarnatulum and Laccaria bicolor, and the wood-decay fungus Irpex lacteus at pH values of 2-9 on liquid BSA media. We also assessed protease activity using a fluorescently labeled casein assay and gelatin zymography and characterized proteases using specific protease inhibitors. I. lacteus and U. isabellina utilized protein efficiently, while the ECM fungi exhibited poor protein utilization. ECM fungi secreted metallo-proteases and had pH optima above 4, while other fungi produced aspartic proteases with lower pH optima. The ascomycetous root endophytes M. variabilis and P. fortinii exhibited intermediate levels of protein utilization and M. variabilis exhibited a very low pH optimum. Comparing proteolytic profiles between fungal root endophytes and fungi with well defined ecological roles provides insight into the ecology of these cryptic root associates.

The effects of fungal root endophytes on plant growth: a meta-analysis.

Fungal root endophytes are plant associates that colonize root tissue internally without causing any obvious harm to their host. Although ubiquitous, this relationship is not well understood. Our objectives were to determine the effects of fungal root endophyte inoculation on plant biomass and nitrogen concentration by conducting an extensive meta-analysis. We also explored the effects of experimental conditions on the host-endophyte relationship. We performed analyses weighted with non-parametric variance on plant response to root endophytes from the Ascomycetes (excluding the Clavacipitaceae), including categorical analyses of 21 experimental factors, ranging from the identity of the host and the endophyte, to the composition of the growing medium. The response of total biomass to endophyte inoculation was 18% lower than non-inoculated controls, while individually, root biomass, shoot biomass, and nitrogen concentration responses to endophyte inoculation were neutral. The identities of both the host and the endophyte had an influence, as did the original source of the endophyte (whether or not the isolate used originated from the same host species). Experimental conditions also influenced the plant-endophyte relationship, with the most important being the availability and sources of carbon and organic nitrogen, particularly peat moss. Although our analysis demonstrates that overall plant biomass and nitrogen concentration responses to ascomycetous root endophyte inoculation is neutral to negative, these results are somewhat confounded by among-study differences in experimental conditions, which undoubtedly contribute to the high levels of variability in plant response seen in the literature.