The interrogation of genetic markers in environmental meta-barcoding studies is currently seriously hindered by the lack of taxonomically curated reference data sets for the targeted genes. The Protist Ribosomal Reference database (PR2, http://ssu-rrna.org/) provides a unique access to eukaryotic small sub-unit (SSU) ribosomal RNA and DNA sequences, with curated taxonomy. The database mainly consists of nuclear-encoded protistan sequences. However, metazoans, land plants, macrosporic fungi and eukaryotic organelles (mitochondrion, plastid and others) are also included because they are useful for the analysis of high-troughput sequencing data sets. Introns and putative chimeric sequences have been also carefully checked. Taxonomic assignation of sequences consists of eight unique taxonomic fields. In total, 136 866 sequences are nuclear encoded, 45 708 (36 501 mitochondrial and 9657 chloroplastic) are from organelles, the remaining being putative chimeric sequences. The website allows the users to download sequences from the entire and partial databases (including representative sequences after clustering at a given level of similarity). Different web tools also allow searches by sequence similarity. The presence of both rRNA and rDNA sequences, taking into account introns (crucial for eukaryotic sequences), a normalized eight terms ranked-taxonomy and updates of new GenBank releases were made possible by a long-term collaboration between experts in taxonomy and computer scientists.

Sequencing of ribosomal DNA clone libraries amplified from environmental DNA has revolutionized our understanding of microbial eukaryote diversity and ecology. The results of these analyses have shown that protist groups are far more genetically heterogeneous than their morphological diversity suggests. However, the clone library approach is labour-intensive, relatively expensive, and methodologically biased. Therefore, even the most intensive rDNA library analyses have recovered only small samples of much larger assemblages, indicating that global environments harbour a vast array of unexplored biodiversity. High-throughput parallel tag 454 sequencing offers an unprecedented scale of sampling for molecular detection of microbial diversity. Here, we report a 454 protocol for sampling and characterizing assemblages of eukaryote microbes. We use this approach to sequence two SSU rDNA diversity markers-the variable V4 and V9 regions-from 10 L of anoxic Norwegian fjord water. We identified 38 116 V4 and 15 156 V9 unique sequences. Both markers detect a wide range of taxonomic groups but in both cases the diversity detected was dominated by dinoflagellates and close relatives. Long-tailed rank abundance curves suggest that the 454 sequencing approach provides improved access to rare genotypes. Most tags detected represent genotypes not currently in GenBank, although many are similar to database sequences. We suggest that current understanding of the ecological complexity of protist communities, genetic diversity, and global species richness are severely limited by the sequence data hitherto available, and we discuss the biological significance of this high amplicon diversity.

Background: Analyses of biomolecules for biodiversity, phylogeny or structure/function studies often use graphical tree representations. Many powerful tree editors are now available, but existing tree visualization tools make little use of meta-information related to the entities under study such as taxonomic descriptions or gene functions that can hardly be encoded within the tree itself (if using popular tree formats). Consequently, a tedious manual analysis and post-processing of the tree graphics are required if one needs to use external information for displaying or investigating trees.

The susceptibility of Anopheles mosquitoes to Plasmodium infections relies on complex interactions between the insect vector and the malaria parasite. A number of studies have shown that the mosquito innate immune responses play an important role in controlling the malaria infection and that the strength of parasite clearance is under genetic control, but little is known about the influence of environmental factors on the transmission success. We present here evidence that the composition of the vector gut microbiota is one of the major components that determine the outcome of mosquito infections. A. gambiae mosquitoes collected in natural breeding sites from Cameroon were experimentally challenged with a wild P. falciparum isolate, and their gut bacterial content was submitted for pyrosequencing analysis. The meta-taxogenomic approach revealed a broader richness of the midgut bacterial flora than previously described. Unexpectedly, the majority of bacterial species were found in only a small proportion of mosquitoes, and only 20 genera were shared by 80% of individuals. We show that observed differences in gut bacterial flora of adult mosquitoes is a result of breeding in distinct sites, suggesting that the native aquatic source where larvae were grown determines the composition of the midgut microbiota. Importantly, the abundance of Enterobacteriaceae in the mosquito midgut correlates significantly with the Plasmodium infection status. This striking relationship highlights the role of natural gut environment in parasite transmission. Deciphering microbe-pathogen interactions offers new perspectives to control disease transmission.

We propose that marine planktonic microeukaryote assemblages incorporate dynamic and metabolically active abundant and rare subcommunities, with contrasting structuring patterns but fairly regular proportions, across space and time.

Growth conditions. M. hydrocarbonoclasticus was aerobically cultivated in synthetic medium (SM) containing 1.23% (wtivol) Tris, 0.37% (wtivol) NH,Cl, 0.62% (wtkol)

A collection of 75 strains of Pectobacterium chrysanthemi (including all biovars and pathovars) and the type strains of Brenneria paradisiaca (CFBP 4178T) and Pectobacterium cypripedii (CFBP 3613T) were studied by DNA–DNA hybridization, numerical taxonomy of 121 phenotypic characteristics, serology and 16S rRNA gene-based phylogenetic analyses. From analysis of 16S rRNA gene sequences, it was deduced that P. chrysanthemi strains and B. paradisiaca CFBP 4178T formed a clade distinct from the genera Pectobacterium and Brenneria; therefore, it is proposed to transfer all the strains to a novel genus, Dickeya gen. nov. By DNA–DNA hybridization, the strains of P. chrysanthemi were distributed among six genomic species: genomospecies 1 harbouring 16 strains of biovar 3 and four strains of biovar 8, genomospecies 2 harbouring 16 strains of biovar 3, genomospecies 3 harbouring two strains of biovar 6 and five strains of biovar 5, genomospecies 4 harbouring five strains of biovar 2, genomospecies 5 harbouring six strains of biovar 1, four strains of biovar 7 and five strains of biovar 9 and genomospecies 6 harbouring five strains of biovar 4 and B. paradisiaca CFBP 4178T. Two strains of biovar 3 remained unclustered. Biochemical criteria, deduced from a numerical taxonomic study of phenotypic characteristics, and serological reactions allowed discrimination of the strains belonging to the six genomic species. Thus, it is proposed that the strains clustered in these six genomic species be assigned to the species Dickeya zeae sp. nov. (type strain CFBP 2052T=NCPPB 2538T), Dickeya dadantii sp. nov. (type strain CFBP 1269T=NCPPB 898T), Dickeya chrysanthemi comb. nov. (subdivided into two biovars, bv. chrysanthemi and bv. parthenii), Dickeya dieffenbachiae sp. nov. (type strain CFBP 2051T=NCPPB 2976T), Dickeya dianthicola sp. nov. (type strain CFBP 1200T=NCPPB 453T) and Dickeya paradisiaca comb. nov., respectively.

Summary Although protists are critical components of marine ecosystems, they are still poorly characterized. Here we analysed the taxonomic diversity of planktonic and benthic protist communities collected in six distant European coastal sites. Environmental deoxyribonucleic acid (DNA) and ribonucleic acid (RNA) from three size fractions (pico‐, nano‐ and micro/mesoplankton), as well as from dissolved DNA and surface sediments were used as templates for tag pyrosequencing of the V4 region of the 18S ribosomal DNA. Beta‐diversity analyses split the protist community structure into three main clusters: picoplankton‐nanoplankton‐dissolved DNA, micro/mesoplankton and sediments. Within each cluster, protist communities from the same site and time clustered together, while communities from the same site but different seasons were unrelated. Both DNA and RNA‐based surveys provided similar relative abundances for most class‐level taxonomic groups. Yet, particular groups were overrepresented in one of the two templates, such as marine alveolates (MALV)‐I and MALV‐II that were much more abundant in DNA surveys. Overall, the groups displaying the highest relative contribution were Dinophyceae, Diatomea, Ciliophora and Acantharia. Also, well represented were Mamiellophyceae, Cryptomonadales, marine alveolates and marine stramenopiles in the picoplankton, and Monadofilosa and basal Fungi in sediments. Our extensive and systematic sequencing of geographically separated sites provides the most comprehensive molecular description of coastal marine protist diversity to date.