De novo Sequencing of Novel Mycoviruses From Fusarium sambucinum: An Attempt on Direct RNA Sequencing of Viral dsRNAs.

An increasing number of viruses are continuously being found in a wide range of organisms, including fungi. Recent studies have revealed a wide viral diversity in microbes and a potential importance of these viruses in the natural environment. Although virus exploration has been accelerated by short-read, high-throughput sequencing (HTS), and viral de novo sequencing is still challenging because of several biological/molecular features such as micro-diversity and secondary structure of RNA genomes. This study conducted de novo sequencing of multiple double-stranded (ds) RNA (dsRNA) elements that were obtained from fungal viruses infecting two Fusarium sambucinum strains, FA1837 and FA2242, using conventional HTS and long-read direct RNA sequencing (DRS). De novo assembly of the read data from both technologies generated near-entire genomic sequence of the viruses, and the sequence homology search and phylogenetic analysis suggested that these represented novel species of the Hypoviridae, Totiviridae, and Mitoviridae families. However, the DRS-based consensus sequences contained numerous indel errors that differed from the HTS consensus sequences, and these errors hampered accurate open reading frame (ORF) prediction. Although with its present performance, the use of DRS is premature to determine viral genome sequences, the DRS-mediated sequencing shows great potential as a user-friendly platform for a one-shot, whole-genome sequencing of RNA viruses due to its long-reading ability and relative structure-tolerant nature.

Geographic Distribution, Genetic Variability and Biological Properties of Rice Orange Leaf Phytoplasma in Southeast Asia.

Rice orange leaf phytoplasma (ROLP) causes clear orange to yellowish leaf discoloration and severe stunting in rice seedlings. The ecological and biological characteristics of ROLP are largely unknown because the disease has not widely caused serious problems in rice cultivated areas, thereby leading to the low accumulation of research data. However, in the past decade, the disease became a threat to rice production, particularly in South China and India; it has also been recognised in other Asian countries, such as Vietnam, Thailand and the Philippines. Here, we observed the occurrence of ROLP in paddies of the Southeast Asian counties (Cambodia, Vietnam and the Philippines) and found that the isolates in the Philippines and Vietnam were monophyletic, while those in India, Thailand and Cambodia were more diverse, suggesting their potential origins. In Cambodia, it was revealed that following polymerase chain reaction (PCR) detection, the known ROLP-insect vectors, N. virescens Distant and Recilia dorsalis Motchulsky, were ROLP-positive, indicating their roles in pathogen dispersal. Moreover, fluorescent and scanning electron microscopy revealed the intensive accumulation of the phytoplasma in phloem tissues and massive accumulation of storage starch in vascular bundle sheath and parenchyma. Altogether, this study illustrated the genetic variability of global ROLP isolates and the pathogen's biological impact on rice tissue.

Cauliflower Mosaic Virus TAV, a Plant Virus Protein That Functions like Ribonuclease H1 and is Cytotoxic to Glioma Cells.

Recent comparisons between plant and animal viruses reveal many common principles that underlie how all viruses express their genetic material, amplify their genomes, and link virion assembly with replication. Cauliflower mosaic virus (CaMV) is not infectious for human beings. Here, we show that CaMV transactivator/viroplasmin protein (TAV) shares sequence similarity with and behaves like the human ribonuclease H1 (RNase H1) in reducing DNA/RNA hybrids detected with S9.6 antibody in HEK293T cells. We showed that TAV is clearly expressed in the cytosol and in the nuclei of transiently transfected human cells, similar to its distribution in plants. TAV also showed remarkable cytotoxic effects in U251 human glioma cells in vitro. These characteristics pave the way for future analysis on the use of the plant virus protein TAV, as an alternative to human RNAse H1 during gene therapy in human cells.

Microscopic ossicle analyses and the complete mitochondrial genome sequence of Holothuria (Roweothuria) polii (Echinodermata; Holothuroidea) provide new information to support the phylogenetic positioning of this sea cucumber species.

Sea cucumbers (Holothuroidea) are ecologically important organisms for their bioturbation and alkalinization activities of the seabed. These species are extensively fished as they are considered luxury food. Sea cucumbers are also relevant for biomedical studies and the production of bioactive compounds. A few initiatives are recently evaluating sea cucumbers as novel aquaculture species. The aim of this study was to provide morphological and genetic information useful for the identification of Holothuria polii, the white spot sea cucumber (a common species of the Mediterranean Sea). We generated the complete sequence of the mitochondrial DNA (mtDNA) genome of this species and combined it with a detailed ossicle characterization of the sequenced specimen by scanning electron microscopic analysis. Ossicles (known also as sclerites) are anatomical features that can discriminate Holothuroidea species, including the closely related ones of the genus Holothuria. The complete mitochondrial genome was assembled, functionally annotated and then used to evaluate the phylogenetic relationship of H. polii against the other few Holothuroidea species for which the whole mtDNA was available. The 15,907 bp H. polii mtDNA sequence has the same gene order already reported for H. scabra, H. forskali and other species of the same class. Cox1 and 16S gene sequences were informative for species identification across the genus and could be used for the authentication of commercialized Holothuria spp. The mitochondrial genome sequence presented here provides the basis to a future analysis of the variability of H. polii populations in the Mediterranean region.