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)

Emerging infectious diseases (EIDs) of free-living wild animals can be classified into three major groups on the basis of key epizootiological criteria: (i) EIDs associated with “spill-over” from domestic animals to wildlife populations living in proximity; (ii) EIDs related directly to human intervention, via host or parasite translocations; and (iii) EIDs with no overt human or domestic animal involvement. These phenomena have two major biological implications: first, many wildlife species are reservoirs of pathogens that threaten domestic animal and human health; second, wildlife EIDs pose a substantial threat to the conservation of global biodiversity.

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Emerging Infectious Diseases of Wildlife-- Threats to Biodiversity and Human Health

Evolution of Protein Molecules

Present and future arboviral threats.

We undertook a comprehensive phylogenetic study to establish the genetic relationship among the viruses of the genus Flavivirus and to compare the classification based on molecular phylogeny with the existing serologic method. By using a combination of quantitative definitions (bootstrap support level and the pairwise nucleotide sequence identity), the viruses could be classified into clusters, clades, and species. Our phylogenetic study revealed for the first time that from the putative ancestor two branches, non-vector and vector-borne virus clusters, evolved and from the latter cluster emerged tick-borne and mosquito-borne virus clusters. Provided that the theory of arthropod association being an acquired trait was correct, pairwise nucleotide sequence identity among these three clusters provided supporting data for a possibility that the non-vector cluster evolved first, followed by the separation of tick-borne and mosquito-borne virus clusters in that order. Clades established in our study correlated significantly with existing antigenic complexes. We also resolved many of the past taxonomic problems by establishing phylogenetic relationships of the antigenically unclassified viruses with the well-established viruses and by identifying synonymous viruses.

Phylogeny of the genus Flavivirus.

To evaluate the role of Culex mosquitoes as enzootic and epidemic vectors for WNV, we identified the source of vertebrate blood by polymerase chain reaction amplification and sequencing portions of the cytochrome b gene of mitochondrial DNA. All Cx. restuans and 93% of Cx. pipiens acquired blood from avian hosts; Cx. salinarius fed frequently on both mammals (53%) and birds (36%). Mixed-blood meals were detected in 11% and 4% of Cx. salinarius and Cx. pipiens, respectively. American robin was the most common source of vertebrate blood for Cx. pipiens (38%) and Cx. restuans (37%). American crow represented <1% of the blood meals in Cx. pipiens and none in Cx. restuans. Human-derived blood meals were identified from 2 Cx. salinarius and 1 Cx. pipiens. Results suggest that Cx. salinarius is an important bridge vector to humans, while Cx. pipiens and Cx. restuans are more efficient enzootic vectors in the northeastern United States.

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Host Feeding Patterns of Culex Mosquitoes and West Nile Virus Transmission, Northeastern United States

Dengue is the most common mosquito-borne viral disease in humans. The spread of both mosquito vectors and viruses has led to the resurgence of epidemic dengue fever (a self-limited flu-like syndrome) and the emergence of dengue hemorrhagic fever (severe dengue with bleeding abnormalities) in urban centers of the tropics. There are no animal or laboratory models of dengue disease; indirect evidence suggests that dengue viruses differ in virulence, including their pathogenicities for humans and epidemic potential. We developed two assay systems (using human dendritic cells and Aedes aegypti mosquitoes) for measuring differences in virus replication that correlate with the potential to cause hemorrhagic dengue and increased virus transmission. Infection and growth experiments showed that dengue serotype 2 viruses causing dengue hemorrhagic fever epidemics (Southeast Asian genotype) can outcompete viruses that cause dengue fever only (American genotype). This fact implies that Southeast Asian genotype viruses will continue to displace other viruses, causing more hemorrhagic dengue epidemics.

Selection for Virulent Dengue Viruses Occurs in Humans and Mosquitoes

To determine if eastern North American Ixodes dammini, like related ticks in Eurasia, maintain tick-borne encephalitis group viruses, we analyzed ticks collected from sites where the agent of Lyme disease is zoonotic. Two viral isolates were obtained by inoculating mice with homogenates from tick salivary glands. The virus, which was described by reverse transcriptase polymerase chain reaction and direct sequencing of the amplification products, was similar to, but distinct from, Powassan virus and is provisionally named "deer tick virus." Enzootic tick-borne encephalitis group viruses accompany the agents of Lyme disease, babesiosis, and granulocytic ehrlichiosis in a Holarctic assemblage of emergent deer tick pathogens.

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A new tick-borne encephalitis-like virus infecting New England deer ticks, Ixodes dammini.

The vertebrate hosts of 672 blood-engorged Culex quinquefasciatus Say, collected in Harris County, Texas, during 2005, were identified by nucleotide sequencing PCR products of the cytochrome b gene. Analysis revealed that 39.1% had acquired blood from birds, 52.5% from mammals, and 8.3% were mixed avian and mammalian blood meals. Most frequent vertebrate hosts were dog (41.0%), mourning dove (18.3%), domestic cat (8.8%), white-winged dove (4.3%), house sparrow (3.2%), house finch (3.0%), gray catbird (3.0%), and American robin (2.5%). Results are interpreted in conjunction with concurrent avian and mosquito West Nile virus (WNV) surveillance activities in Harris County. We conclude that Cx. quinquefasciatus is an opportunistic feeder and principal mosquito vector of WNV in this metropolitan area; however, transmission by other mosquito species or by other modes of infection, such as ingestion, must account for the high WNV infection rates among local blue jays and American crows.

Host feeding pattern of Culex quinquefasciatus (Diptera: Culicidae) and its role in transmission of West Nile virus in Harris County, Texas.

Dengue serotype 2 (DEN-2) viruses with the potential to cause dengue hemorrhagic fever have been shown to belong to the Southeast (SE) Asian genotype. These viruses appear to be rapidly displacing the American genotype of DEN-2 in the Western Hemisphere. To determine whether distinct genotypes of DEN-2 virus are better adapted to mosquito transmission, we classified 15 viral strains of DEN-2 phylogenetically and compared their ability to infect and disseminate in different populations of Aedes aegypti mosquitoes. Envelope gene nucleotide sequence analysis confirmed that six strains belonged to the American genotype and nine strains were of the SE Asian genotype. The overall rate of disseminated infection in mosquitoes from Texas was 27% for the SE Asian genotype versus 9% for the American genotype. This pattern of infection was similar in another population of mosquitoes sampled from southern Mexico (30% versus 13%). Together, these findings suggest that Ae. aegypti tends to be more susceptible to infection by DEN-2 viruses of the SE Asian genotype than to those of the American genotype, and this may have epidemiologic implications.

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Philip M. Armstrong

Papers

Host Feeding Patterns of Culex Mosquitoes and West Nile Virus Transmission, Northeastern United States

Selection for Virulent Dengue Viruses Occurs in Humans and Mosquitoes

A new tick-borne encephalitis-like virus infecting New England deer ticks, Ixodes dammini.

Host feeding pattern of Culex quinquefasciatus (Diptera: Culicidae) and its role in transmission of West Nile virus in Harris County, Texas.

Efficiency of dengue serotype 2 virus strains to infect and disseminate in aedes aegypti