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)

The RefSeq project at the National Center for Biotechnology Information (NCBI) maintains and curates a publicly available database of annotated genomic, transcript, and protein sequence records (http://www.ncbi.nlm.nih.gov/refseq/). The RefSeq project leverages the data submitted to the International Nucleotide Sequence Database Collaboration (INSDC) against a combination of computation, manual curation, and collaboration to produce a standard set of stable, non-redundant reference sequences. The RefSeq project augments these reference sequences with current knowledge including publications, functional features and informative nomenclature. The database currently represents sequences from more than 55,000 organisms (>4800 viruses, >40,000 prokaryotes and >10,000 eukaryotes; RefSeq release 71), ranging from a single record to complete genomes. This paper summarizes the current status of the viral, prokaryotic, and eukaryotic branches of the RefSeq project, reports on improvements to data access and details efforts to further expand the taxonomic representation of the collection. We also highlight diverse functional curation initiatives that support multiple uses of RefSeq data including taxonomic validation, genome annotation, comparative genomics, and clinical testing. We summarize our approach to utilizing available RNA-Seq and other data types in our manual curation process for vertebrate, plant, and other species, and describe a new direction for prokaryotic genomes and protein name management.

Reference sequence (RefSeq) database at NCBI: current status, taxonomic expansion, and functional annotation

Rapid, Low-Cost Detection of Zika Virus Using Programmable Biomolecular Components

In April 2008, a nucleotide-sequence-based, complete genome classification system was developed for group A rotaviruses (RVs). This system assigns a specific genotype to each of the 11 genome segments of a particular RV strain according to established nucleotide percent cutoff values. Using this approach, the genome of individual RV strains are given the complete descriptor of Gx-P[x]-Ix-Rx-Cx-Mx-Ax-Nx-Tx-Ex-Hx. The Rotavirus Classification Working Group (RCWG) was formed by scientists in the field to maintain, evaluate and develop the RV genotype classification system, in particular to aid in the designation of new genotypes. Since its conception, the group has ratified 51 new genotypes: as of April 2011, new genotypes for VP7 (G20-G27), VP4 (P[28]-P[35]), VP6 (I12-I16), VP1 (R5-R9), VP2 (C6-C9), VP3 (M7-M8), NSP1 (A15-A16), NSP2 (N6-N9), NSP3 (T8-T12), NSP4 (E12-E14) and NSP5/6 (H7-H11) have been defined for RV strains recovered from humans, cows, pigs, horses, mice, South American camelids (guanaco), chickens, turkeys, pheasants, bats and a sugar glider. With increasing numbers of complete RV genome sequences becoming available, a standardized RV strain nomenclature system is needed, and the RCWG proposes that individual RV strains are named as follows: RV group/species of origin/country of identification/common name/year of identification/G- and P-type. In collaboration with the National Center for Biotechnology Information (NCBI), the RCWG is also working on developing a RV-specific resource for the deposition of nucleotide sequences. This resource will provide useful information regarding RV strains, including, but not limited to, the individual gene genotypes and epidemiological and clinical information. Together, the proposed nomenclature system and the NCBI RV resource will offer highly useful tools for investigators to search for, retrieve, and analyze the ever-growing volume of RV genomic data.

/pdf/uniformity-of-rotavirus-strain-nomenclature-proposed-by-the-4qpjav1aff.pdf

Uniformity of rotavirus strain nomenclature proposed by the Rotavirus Classification Working Group (RCWG).

Rolling circle amplification (RCA) is an isothermal enzymatic process where a short DNA or RNA primer is amplified to form a long single stranded DNA or RNA using a circular DNA template and special DNA or RNA polymerases. The RCA product is a concatemer containing tens to hundreds of tandem repeats that are complementary to the circular template. The power, simplicity, and versatility of the DNA amplification technique have made it an attractive tool for biomedical research and nanobiotechnology. Traditionally, RCA has been used to develop sensitive diagnostic methods for a variety of targets including nucleic acids (DNA, RNA), small molecules, proteins, and cells. RCA has also attracted significant attention in the field of nanotechnology and nanobiotechnology. The RCA-produced long, single-stranded DNA with repeating units has been used as template for the periodic assembly of nanospecies. Moreover, since RCA products can be tailor-designed by manipulating the circular template, RCA has been employed to generate complex DNA nanostructures such as DNA origami, nanotubes, nanoribbons and DNA based metamaterials. These functional RCA based nanotechnologies have been utilized for biodetection, drug delivery, designing bioelectronic circuits and bioseparation. In this review, we introduce the fundamental engineering principles used to design RCA nanotechnologies, discuss recently developed RCA-based diagnostics and bioanalytical tools, and summarize the use of RCA to construct multivalent molecular scaffolds and nanostructures for applications in biology, diagnostics and therapeutics.

Rolling circle amplification: A versatile tool for chemical biology, materials science and medicine

Amyloid plaques, one of the neuropathological hallmarks of Alzheimer's disease, and their main constituent, the amyloid beta-peptide (Abeta), are triggers of the activation of innate inflammatory mechanisms involving the activation of microglia. To dissect the effects of a non-Abeta-specific microglial activation on the Abeta metabolism, we employed a viral infection-based model. Transgenic mice expressing a mutated form of the human amyloid precursor protein (Tg2576) were used. In preceding experiments, 2-week-old transgenic mice and non-transgenic littermates were infected intracerebrally with the neurotropic Borna disease virus and investigated at 2, 4 and 14 weeks post-infection. The Borna disease virus-inoculated mice showed a persisting, subclinical infection of cortical and limbic brain areas characterized by slight T-cell infiltrates, expression of cytokines and a massive microglial activation in the hippocampus and neocortex. Viral-induced effects reached their peak at 4 weeks post-infection. In 14-month-old Tg2576 mice, characterized by the deposition of diffuse and dense-core amyloid plaques in cortical brain regions, Borna disease virus-induced microglial activation in the vicinity of Abeta deposits was used to investigate the influence of a local inflammatory response on these deposits. At 4 weeks post-infection, histometric analyses employing Abeta immunohistochemistry revealed a decrease of the cortical and hippocampal Abeta-immunopositive area. This overall decrease was accompanied by a decrease of parenchymal thioflavin-S-positive amyloid deposits and an increase of such deposits in the walls of cerebral vessels, which indicates that the elicitation of a non-Abeta-specific microglial activation may contribute to a reduction of Abeta in the brain parenchyma.

Viral-induced inflammation is accompanied by β-amyloid plaque reduction in brains of amyloid precursor protein transgenic Tg2576 mice

Rotaviruses infect humans and animals and are a main cause of diarrhea. They are non-enveloped viruses with a genome of 11 double-stranded RNA segments. Based on genome analysis and amino acid sequence identities of the capsid protein VP6, the rotavirus species A to J (RVA-RVJ) have been defined so far. In addition, rotaviruses putatively assigned to the novel rotavirus species K (RVK) and L (RVL) have been recently identified in common shrews (Sorex araneus), based on partial genome sequences. Here, the complete genome sequence of strain KS14/0241, a prototype strain of RVL, is presented. The deduced amino acid sequence for VP6 of this strain shows only up to 47% identity to that of RVA to RVJ reference strains. Phylogenetic analyses indicate a clustering separated from the established rotavirus species for all 11 genome segments of RVL, with the closest relationship to RVH and RVJ within the phylogenetic RVB-like clade. The non-coding genome segment termini of RVL showed conserved sequences at the 5′-end (positive-sense RNA strand), which are common to all rotaviruses, and those conserved among the RVB-like clade at the 3′-end. The results are consistent with a classification of the virus into a novel rotavirus species L.

/pdf/whole-genome-sequence-analysis-of-a-prototype-strain-of-the-1zaabqsr.pdf

Whole Genome Sequence Analysis of a Prototype Strain of the Novel Putative Rotavirus Species L

Between 2000 and 2004 a disease occurred in an aviary in Germany affecting various bird species belonging to the order Passeriformes including Collared Grosbeaks (Mycerobas affinis), Eurasian Bullfinches (Pyrrhula pyrrhula griseiventris), Brown Bullfinches (Pyrrhula nipalensis), Grey-headed bullfinches (Pyrrhula erythaca) and Yellow-bellied Tits (Periparus venustulus). The major clinical signs included increased mortality of fledglings and young birds, as well as feather disorders and feather loss in adult birds. In addition, adult Eurasian Bullfinches showed in one year a disease course, in which the major symptom was inflammation of the skin beginning on the basis of the beak and spreading over the head occurring a few days before death. Bacteriological and parasitological investigations did not reveal any consistent findings. Using a newly developed polymerase chain reaction protocol, DNA of the recently discovered finch polyomavirus (FPyV) was demonstrated in several affected birds. Because of the consistent detection of FPyV-DNA and the similarity of the symptoms with those observed during infection with the closely related avian polyomavirus in other bird species, an etiological role of FPyV in the observed disease is assumed

Nachweis von DNA des Finken-Polyomavirus bei Erkrankungen verschiedener Vogelarten der Ordnung Passeriformes

Predictive models for thermal inactivation of human norovirus and surrogates in strawberry puree

Hepatitis E virus (HEV) infection can cause acute and chronic hepatitis in humans. The zoonotic HEV genotype 3, which is highly prevalent in Europe, is mainly transmitted by consumption of raw meat and raw meat products produced from infected pigs or wild boars. High salt concentrations represent an important measure to preserve meat products and to inactivate foodborne pathogens. Here, an HEV preparation in phosphate-buffered saline (PBS) was subjected to different salt concentrations and the remaining infectivity was measured in a cell culture assay. Treatments with up to 20% sodium chloride for 24 h at 23 °C, with and without addition of 0.015% sodium nitrite or 0.03% sodium nitrate, did not lead to virus inactivation as compared to PBS only. Conditions usually applied for short-term and long-term fermented raw sausages were simulated by incubation at 22 °C for up to 6 days and at 16 °C for up to 8 weeks, respectively. Only 2% sodium chloride with 0.015% sodium nitrite showed a weak (< 1 log10), but significant, infectivity reduction after 2 and 4 days as compared to PBS only. Addition of 2% sodium chloride and 0.03% sodium nitrate showed a slight, but not significant, decrease in infectivity after 2 and 8 weeks as compared to PBS only. In conclusion, HEV is highly stable at high salt concentrations and at salt conditions usually applied to preserve raw meat products.

/pdf/effect-of-sodium-chloride-sodium-nitrite-and-sodium-nitrate-2u34ohhxl6.pdf

Reimar Johne

Papers

Viral-induced inflammation is accompanied by β-amyloid plaque reduction in brains of amyloid precursor protein transgenic Tg2576 mice

Whole Genome Sequence Analysis of a Prototype Strain of the Novel Putative Rotavirus Species L

Nachweis von DNA des Finken-Polyomavirus bei Erkrankungen verschiedener Vogelarten der Ordnung Passeriformes

Predictive models for thermal inactivation of human norovirus and surrogates in strawberry puree

Effect of Sodium Chloride, Sodium Nitrite and Sodium Nitrate on the Infectivity of Hepatitis E Virus