Ganoderma Fusions with High Yield of Ergothioneine and Comparative Analysis of Its Genomics.

Ergothioneine (EGT), an exceptional antioxidant found ubiquitously across diverse living organisms, plays a pivotal role in various vital physiological regulatory functions. Its principal natural sources are mushrooms and animal liver tissues. Ganoderma spp., a traditional Chinese food and medicinal mushroom, boasts high concentrations of EGT. To advance the development of novel Ganoderma spp. strains with enhanced EGT yields, we employed an efficient Ganoderma spp. protoplasmic fusion system. Through molecular and biological characterization, we successfully generated seven novel fusion strains. Notably, fusion strain RS7 demonstrated a remarkable increase in mycelial EGT production (12.70 ± 1.85 mg/L), surpassing the parental strains FQ16 and FQ23 by 34.23% and 39.10%, respectively. Furthermore, in the context of the fruiting body, fusion strain RS11 displayed a notable 53.58% enhancement in EGT production (11.24 ± 1.96 mg/L) compared to its parental strains. Genomic analysis of the RS7, the strain with the highest levels of mycelial EGT production, revealed mutations in the gene EVM0005141 associated with EGT metabolism. These mutations led to a reduction in non-productive shunts, subsequently redirecting more substrate towards the EGT synthesis pathway. This redirection significantly boosted EGT production in the RS7 strain. The insights gained from this study provide valuable guidance for the commercial-scale production of EGT and the selective breeding of Ganoderma spp. strains.

Multiplex gene precise editing and large DNA fragment deletion by the CRISPR-Cas9-TRAMA system in edible mushroom Cordyceps militaris.

The medicinal mushroom Cordyceps militaris contains abundant valuable bioactive ingredients that have attracted a great deal of attention in the pharmaceutical and cosmetic industries. However, the development of this valuable mushroom faces the obstacle of lacking powerful genomic engineering tools. Here, by excavating the endogenous tRNA-processed element, introducing the extrachromosomal plasmid and alongside with homologous template, we develop a marker-free CRISPR-Cas9-TRAMA genomic editing system to achieve the multiplex gene precise editing and large synthetic cluster deletion in C. militaris. We further operated editing in the synthetases of cordycepin and ergothioneine to demonstrate the application of Cas9-TRAMA system in protein modification, promoter strength evaluation and 10 kb metabolic synthetic cluster deletion. The Cas9-TRAMA system provides a scalable method for excavating the valuable metabolic resource of medicinal mushrooms and constructing a mystical cellular pathway to elucidate the complex cell behaviours of the edible mushroom.

Hemodynamic Effect of Flow Diverter and Coils in Treatment of Large and Giant Intracranial Aneurysms.

BACKGROUND: This study aimed to investigate the hemodynamic changes induced by a flow diverter (FD) and coils in the treatment of internal carotid artery aneurysms, as well as to evaluate the effect of this treatment by using angiographic follow-up data. METHODS: Six large and giant aneurysms were treated by the Tubridge FD and loose packing coils between June 2013 and May 2015. Patient-specific models were constructed and analyzed using a computational fluid dynamics (CFD) method. The virtual FD deployment method was used to implant the Tubridge stent into a 3-dimensional digital subtraction angiographic image of the aneurysms, and the coils were simulated by a porous medium model. RESULTS: Tubridge FD alone can significantly reduce the intra-aneurysmal flow velocity (0.17 ± 0.05 m/s-0.11 ± 0.06 m/s, P < 0.001) and wall shear stress (WSS, 1.39 ± 0.29 Pa-0.77 ± 0.34 Pa, P = 0.001) and increase the low wall shear area (LSA, 6.38% ± 1.49%-34.60% ± 28.90%, P = 0.047). Coils, as a supplementary measure, further reduced the velocity (0.11 ± 0.06 m/s-0.08 ± 0.05 m/s, P = 0.03) and WSS (0.77 ± 0.34 Pa-0.47 ± 0.35 Pa, P = 0.04) and increased the LSA (34.60% ± 28.90%-63.33% ± 34.82%, P = 0.044). Aneurysm with sustained strong inflow after treatment (case 3, 25% reduction in velocity, 12% reduction in WSS, and 16% increment in LSA) showed partial patency, whereas others with a weaker inflow jet (mean 56% reduction in velocity, 74% reduction in WSS, and 1081% increment in LSA) showed complete occlusion at follow-up. CONCLUSIONS: On the basis of using the CFD method, adjunctive coiling with the Tubridge FD placement may significantly reduce intra-aneurysmal flow velocity and WSS, promoting thrombosis formation and occlusion of aneurysms.

Fast Virtual Stenting with Active Contour Models in Intracranical Aneurysm.

Intracranial stents are becoming increasingly a useful option in the treatment of intracranial aneurysms (IAs). Image simulation of the releasing stent configuration together with computational fluid dynamics (CFD) simulation prior to intervention will help surgeons optimize intervention scheme. This paper proposed a fast virtual stenting of IAs based on active contour model (ACM) which was able to virtually release stents within any patient-specific shaped vessel and aneurysm models built on real medical image data. In this method, an initial stent mesh was generated along the centerline of the parent artery without the need for registration between the stent contour and the vessel. Additionally, the diameter of the initial stent volumetric mesh was set to the maximum inscribed sphere diameter of the parent artery to improve the stenting accuracy and save computational cost. At last, a novel criterion for terminating virtual stent expanding that was based on the collision detection of the axis aligned bounding boxes was applied, making the stent expansion free of edge effect. The experiment results of the virtual stenting and the corresponding CFD simulations exhibited the efficacy and accuracy of the ACM based method, which are valuable to intervention scheme selection and therapy plan confirmation.

A scaling aneurysm model-based approach to assessing the role of flow pattern and energy loss in aneurysm rupture prediction.

BACKGROUND AND PURPOSE: Energy loss (EL) was regarded to be one of the key parameters in predicting the rupture risk of IA. In this paper, we took varied aspect ratio (AR) as a scaling law to create a series of longitudinal models to investigate the longitudinal changes of flow pattern and EL as the AR varies, in order to explore the relationship between the longitudinal characteristic EL parameters with aneurysm rupture risk. METHODS: Seven original intracranial aneurysms (IA) models with similar locations were reconstructed from patient 3D rotational angiography (3DRA) images. Based on these models, a series of scaling aneurysm models with different ARs were created with our proposed scaling algorithms. Fluid-solid interaction (FSI) simulations were performed on every model to obtain hemodynamics flow pattern and EL. RESULTS: With AR increasing, flow pattern became more complex, with vortices appearing gradually in the aneurysms (AR > 1.5). Furthermore, the velocity significantly decreased in aneurysms with high ARs (>1.5). Meanwhile, the aneurysm EL increased with increasing AR. Once AR exceeded 1.5, EL changed drastically. CONCLUSION: EL was a potential parameter predicting future rupture of unruptured aneurysms. If the EL during the growth of the unruptured aneurysms increased sharply, we strongly recommend an intervention.