De novo Genome Assembly of the Fungal Plant Pathogen Pyrenophora semeniperda

Pyrenophora semeniperda (anamorph Drechslera campulata) is a necrotrophic fungal seed pathogen that has a wide host range within the Poaceae. One of its hosts is cheatgrass (Bromus tectorum), a species exotic to the United States that has invaded natural ecosystems of the Intermountain West. As a natural pathogen of cheatgrass, P. semeniperda has potential as a biocontrol agent due to its effectiveness at killing seeds within the seed bank; however, few genetic resources exist for the fungus. Here, the genome of P. semeniperda isolate assembled from sequence reads of 454 pyrosequencing is presented. The total assembly is 32.5 Mb and includes 11,453 gene models encoding putative proteins larger than 24 amino acids. The models represent a variety of putative genes that are involved in pathogenic pathways typically found in necrotrophic fungi. In addition, extensive rearrangements, including inter- and intrachromosomal rearrangements, were found when the P. semeniperda genome was compared to P. tritici-repentis, a related fungal species.     

De novo assembly of human genome at single-cell levels

Genome assembly has been benefited from long-read sequencing technologies with higher accuracy and higher continuity. However, most human genome assembly require large amount of DNAs from homogeneous cell lines without keeping cell heterogeneities, since cell heterogeneity could profoundly affect haplotype assembly results. Herein, using single-cell genome long-read sequencing technology (SMOOTH-seq), we have sequenced K562 and HG002 cells on PacBio HiFi and Oxford Nanopore Technologies (ONT) platforms and conducted de novo genome assembly. For the first time, we have completed the human genome assembly with high continuity (with NG50 of ∼2 Mb using 95 individual K562 cells) at single-cell levels, and explored the impact of different assemblers and sequencing strategies on genome assembly. With sequencing data from 30 diploid individual HG002 cells of relatively high genome coverage (average coverage ∼41.7%) on ONT platform, the NG50 can reach over 1.3 Mb. Furthermore, with the assembled genome from K562 single-cell dataset, more complete and accurate set of insertion events and complex structural variations could be identified. This study opened a new chapter on the practice of single-cell genome de novo assembly.     

De novo GTP Biosynthesis Is Critical for Virulence of the Fungal Pathogen Cryptococcus neoformans

We have investigated the potential of the GTP synthesis pathways as chemotherapeutic targets in the human pathogen Cryptococcus neoformans, a common cause of fatal fungal meningoencephalitis. We find that de novo GTP biosynthesis, but not the alternate salvage pathway, is critical to cryptococcal dissemination and survival in vivo. Loss of inosine monophosphate dehydrogenase (IMPDH) in the de novo pathway results in slow growth and virulence factor defects, while loss of the cognate phosphoribosyltransferase in the salvage pathway yielded no phenotypes. Further, the Cryptococcus species complex displays variable sensitivity to the IMPDH inhibitor mycophenolic acid, and we uncover a rare drug-resistant subtype of C. gattii that suggests an adaptive response to microbial IMPDH inhibitors in its environmental niche. We report the structural and functional characterization of IMPDH from Cryptococcus, revealing insights into the basis for drug resistance and suggesting strategies for the development of fungal-specific inhibitors. The crystal structure reveals the position of the IMPDH moveable flap and catalytic arginine in the open conformation for the first time, plus unique, exploitable differences in the highly conserved active site. Treatment with mycophenolic acid led to significantly increased survival times in a nematode model, validating de novo GTP biosynthesis as an antifungal target in Cryptococcus.     

De novo reconstruction of DNA origami structures through atomistic molecular dynamics simulation

The DNA origami method has brought nanometer-precision fabrication to molecular biology labs, offering myriads of potential applications in the fields of synthetic biology, medicine, molecular computation, etc. Advancing the method further requires controlling self-assembly down to the atomic scale. Here we demonstrate a computational method that allows the equilibrium structure of a large, complex DNA origami object to be determined to atomic resolution. Through direct comparison with the results of cryo-electron microscopy, we demonstrate de novo reconstruction of a 4.7 megadalton pointer structure by means of fully atomistic molecular dynamics simulations. Furthermore, we show that elastic network-guided simulations performed without solvent can yield similar accuracy at a fraction of the computational cost, making this method an attractive approach for prototyping and validation of self-assembled DNA nanostructures.     

De novo Cancers Following Liver Transplantation: A Single Center Experience in China

Background

De novo cancers are a growing problem that has become one of the leading causes of late mortality after liver transplantation. The incidences and risk factors varied among literatures and fewer concerned the Eastern population.

Aims

The aim of this study was to examine the incidence and clinical features of de novo cancers after liver transplantation in a single Chinese center.

Methods

569 patients who received liver transplantation and survived for more than 3 months in a single Chinese center were retrospectively reviewed.

Results

A total of 18 de novo cancers were diagnosed in 17 recipients (13 male and 4 female) after a mean of 41±26 months, with an overall incidence of 3.2%, which was lower than that in Western people. Of these, 8 (3.32%) cases were from 241 recipients with malignant liver diseases before transplant, while 10 (3.05%) cases were from 328 recipients with benign diseases. The incidence rates were comparable, p = 0.86. Furthermore, 2 cases developed in 1 year, 5 cases in 3 years and 11 cases over 3 years. The most frequent cancers developed after liver transplantation were similar to those in the general Chinese population but had much higher incidence rates.

Conclusions

Liver transplant recipients were at increased risk for developing de novo cancers. The incidence rates and pattern of de novo cancers in Chinese population are different from Western people due to racial and social factors. Pre-transplant malignant condition had no relationship to de novo cancer. Exact risk factors need further studies.