The lion's share of bacteria in various environments cannot be cloned in the laboratory and thus cannot be sequenced using existing technologies. A major goal of single-cell genomics is to complement gene-centric metagenomic data with whole-genome assemblies of uncultivated organisms. Assembly of single-cell data is challenging because of highly non-uniform read coverage as well as elevated levels of sequencing errors and chimeric reads. We describe SPAdes, a new assembler for both single-cell and standard (multicell) assembly, and demonstrate that it improves on the recently released E+V-SC assembler (specialized for single-cell data) and on popular assemblers Velvet and SoapDeNovo (for multicell data). SPAdes generates single-cell assemblies, providing information about genomes of uncultivatable bacteria that vastly exceeds what may be obtained via traditional metagenomics studies. SPAdes is available online ( http://bioinf.spbau.ru/spades ). It is distributed as open source software.

SPAdes, a new genome assembly algorithm and its applications to single-cell sequencing ( 7th Annual SFAF Meeting, 2012)

Rhabdoviruses infect a large number of plant species and cause significant crop diseases. They have a negative-sense, single-stranded unsegmented or bisegmented RNA genome. The number of plant-associated rhabdovirid sequences has grown in the last few years in concert with the extensive use of high-throughput sequencing platforms. Here, we report the discovery of 27 novel rhabdovirus genomes associated with 25 different host plant species and one insect, which were hidden in public databases. These viral sequences were identified through homology searches in more than 3000 plant and insect transcriptomes from the National Center for Biotechnology Information (NCBI) Sequence Read Archive (SRA) using known plant rhabdovirus sequences as the query. The identification, assembly and curation of raw SRA reads resulted in sixteen viral genome sequences with full-length coding regions and ten partial genomes. Highlights of the obtained sequences include viruses with unique and novel genome organizations among known plant rhabdoviruses. Phylogenetic analysis showed that thirteen of the novel viruses were related to cytorhabdoviruses, one to alphanucleorhabdoviruses, five to betanucleorhabdoviruses, one to dichorhaviruses and seven to varicosaviruses. These findings resulted in the most complete phylogeny of plant rhabdoviruses to date and shed new light on the phylogenetic relationships and evolutionary landscape of this group of plant viruses. Furthermore, this study provided additional evidence for the complexity and diversity of plant rhabdovirus genomes and demonstrated that analyzing SRA public data provides an invaluable tool to accelerate virus discovery, gain evolutionary insights and refine virus taxonomy.

Illuminating the Plant Rhabdovirus Landscape through Metatranscriptomics Data

MYB transcription factors are important regulators of the plant response to abiotic stress. Their participation in the salinity stress of the key forage legume species alfalfa (Medicago sativa) was investigated here by comparing the transcriptomes of the two cultivars Dryland (DL) and Sundory (SD), which differed with respect to their ability to tolerate salinity stress. When challenged by the stress, DL plants were better able than SD ones to scavenge reactive oxygen species. A large number of genes encoding transcription regulators, signal transducers and proteins involved in both primary and secondary metabolism were differentially transcribed in the two cultivars, especially when plants were subjected to salinity stress. The set of induced genes included 17 MYB family of transcription factors, all of which were subsequently isolated. The effect of constitutively expressing these genes on the salinity tolerance expressed by Arabidopsis thaliana was investigated. The introduction of MsMYB4 significantly increased the plants’ salinity tolerance in an abscisic acid-dependent manner. A sub-cellular localization experiment and a transactivation assay indicated that MsMYB4 was deposited in the nucleus and was able to activate transcription in yeast. Based on this information, we propose that the MsMYB4 products is likely directly involved in alfalfa’s response to salinity stress.

/pdf/transcriptional-profiling-reveals-that-a-myb-transcription-1ckjeonrk1.pdf

Transcriptional profiling reveals that a MYB transcription factor MsMYB4 contributes to the salinity stress response of alfalfa.

Compendium of Alfalfa Diseases and Pests, Third Edition

Alfalfa (Medicago sativa L.), also known as lucerne, is a major forage crop worldwide. In the United States, it has recently become the third most valuable field crop, with an estimated value of over $9.3 billion. Alfalfa is naturally infected by many different pathogens, including viruses, obligate parasites that reproduce only inside living host cells. Traditionally, viral infections of alfalfa have been considered by breeders, growers, producers and researchers to be diseases of limited importance, although they are widespread in all major cultivation areas. However, over the past few years, due to the rapid development of high-throughput sequencing (HTS), viral metagenomics, bioinformatics tools for interpreting massive amounts of HTS data and the increasing accessibility of public data repositories for transcriptomic discoveries, several emerging viruses of alfalfa with the potential to cause serious yield losses have been described. They include alfalfa leaf curl virus (family Geminiviridae), alfalfa dwarf virus (family Rhabdoviridae), alfalfa enamovirus 1 (family Luteoviridae), alfalfa virus S (family Alphaflexiviridae) and others. These discoveries have called into question the assumed low economic impact of viral diseases in alfalfa and further suggested their possible contribution to the severity of complex infections involving multiple pathogens. In this review, we will focus on viruses of alfalfa recently described in different laboratories on the basis of the above research methodologies.

/pdf/high-throughput-sequencing-for-deciphering-the-virome-of-2ezphepdlm.pdf

High-Throughput Sequencing for Deciphering the Virome of Alfalfa (Medicago sativa L.)

Publicly available transcriptomic datasets have become a valuable tool for the discovery of new pathogens, particularly viruses. In this study, several coding-complete viral genomes previously not found or experimentally confirmed in alfalfa were identified in the plant datasets retrieved from the NCBI Sequence Read Archive. Publicly available Medicago spp. transcriptomic datasets were retrieved from the NCBI SRA database. The raw reads were first mapped to the reference genomes of Medicago sativa and Medigago truncatula followed by the alignment of the unmapped reads to the NCBI viral genome database and de novo assembly using the SPAdes tool. When possible, assemblies were experimentally confirmed using 5′/3′ RACE and RT-PCRs. Twenty three different viruses were identified in the analyzed datasets, of which several represented emerging viruses not reported in alfalfa prior to this study. Among them were two strains of cnidium vein yellowing virus, lychnis mottle virus and Cactus virus X, for which coding-complete genomic sequences were obtained by a de novo assembly. The results improve our knowledge of the diversity and host range of viruses infecting alfalfa, provide essential tools for their diagnostics and characterization and demonstrate the utility of transcriptomic datasets for the discovery of new pathogens.

/pdf/identification-of-emerging-viral-genomes-in-transcriptomic-23jqugx4cf.pdf

Identification of emerging viral genomes in transcriptomic datasets of alfalfa (Medicago sativa L.).

We present evidence here that alfalfa (Medicago sativa L.) can be a natural host species for a new strain of Cycas necrotic stunt virus (CNSV), for which the name CNSV-A (alfalfa) is proposed. Prior to this report, the virus has not been identified in alfalfa.

Identification of the Coding-Complete Genome of Cycas Necrotic Stunt Virus in Transcriptomic Data Sets of Alfalfa (Medicago sativa).

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Identification of emerging viral genomes in transcriptomic datasets of alfalfa (Medicago sativa L.).

Identification of the Coding-Complete Genome of Cycas Necrotic Stunt Virus in Transcriptomic Data Sets of Alfalfa (Medicago sativa).