The rapid evolution of 454 GS-FLX sequencing technology has not been accompanied by a reassessment of the quality and accuracy of the sequences obtained. Current strategies for decision-making and ...error-correction are based on an initial analysis by Huse et al. in 2007, for the older GS20 system based on experimental sequences. We analyze here the quality of 454 sequencing data and identify factors playing a role in sequencing error, through the use of an extensive dataset for Roche control DNA fragments.
We obtained a mean error rate for 454 sequences of 1.07%. More importantly, the error rate is not randomly distributed; it occasionally rose to more than 50% in certain positions, and its distribution was linked to several experimental variables. The main factors related to error are the presence of homopolymers, position in the sequence, size of the sequence and spatial localization in PT plates for insertion and deletion errors. These factors can be described by considering seven variables. No single variable can account for the error rate distribution, but most of the variation is explained by the combination of all seven variables.
The pattern identified here calls for the use of internal controls and error-correcting base callers, to correct for errors, when available (e.g. when sequencing amplicons). For shotgun libraries, the use of both sequencing primers and deep coverage, combined with the use of random sequencing primer sites should partly compensate for even high error rates, although it may prove more difficult than previous thought to distinguish between low-frequency alleles and errors.
QDD is an open access program providing a user-friendly tool for microsatellite detection and primer design from large sets of DNA sequences. The program is designed to deal with all steps of ...treatment of raw sequences obtained from pyrosequencing of enriched DNA libraries, but it is also applicable to data obtained through other sequencing methods, using FASTA files as input. The following tasks are completed by QDD: tag sorting, adapter/vector removal, elimination of redundant sequences, detection of possible genomic multicopies (duplicated loci or transposable elements), stringent selection of target microsatellites and customizable primer design. It can treat up to one million sequences of a few hundred base pairs in the tag-sorting step, and up to 50 000 sequences in a single input file for the steps involving estimation of sequence similarity. Availability: QDD is freely available under the GPL licence for Windows and Linux from the following web site: http://www.univ-provence.fr/gsite/Local/egee/dir/meglecz/QDD.html Contact: emese.meglecz@univ-provence.fr Supplementary information: Supplementary data are available at Bioinformatics online.
Amphibians are rapidly vanishing. At the same time, it is most likely that the number of amphibian species is highly underestimated. Recent DNA barcoding work has attempted to define a threshold ...between intra- and inter-specific genetic distances to help identify candidate species. In groups with high extinction rates and poorly known species boundaries, like amphibians, such tools may provide a way to rapidly evaluate species richness.
Here we analyse published and new 16S rDNA sequences from 60 frog species of Amazonia-Guianas to obtain a minimum estimate of the number of undescribed species in this region. We combined isolation by distance, phylogenetic analyses, and comparison of molecular distances to evaluate threshold values for the identification of candidate species among these frogs.
In most cases, geographically distant populations belong to genetically highly distinct lineages that could be considered as candidate new species. This was not universal among the taxa studied and thus widespread species of Neotropical frogs really do exist, contrary to previous assumptions. Moreover, the many instances of paraphyly and the wide overlap between distributions of inter- and intra-specific distances reinforce the hypothesis that many cryptic species remain to be described. In our data set, pairwise genetic distances below 0.02 are strongly correlated with geographical distances. This correlation remains statistically significant until genetic distance is 0.05, with no such relation thereafter. This suggests that for higher distances allopatric and sympatric cryptic species prevail. Based on our analyses, we propose a more inclusive pairwise genetic distance of 0.03 between taxa to target lineages that could correspond to candidate species.
Using this approach, we identify 129 candidate species, two-fold greater than the 60 species included in the current study. This leads to estimates of around 170 to 460 frog taxa unrecognized in Amazonia-Guianas.
As a consequence the global amphibian decline detected especially in the Neotropics may be worse than realised.
Microsatellite marker development has been greatly simplified by the use of high‐throughput sequencing followed by in silico microsatellite detection and primer design. However, the selection of ...markers designed by the existing pipelines depends either on arbitrary criteria, or older studies on PCR success. Based on wet laboratory experiments, we have identified the following factors that are most likely to influence genotyping success rate: alignment score between the primers and the amplicon; the distance between primers and microsatellites; the length of the PCR product; target region complexity and the number of reads underlying the sequence. The QDD pipeline has been modified to include these most pertinent factors in the output to help the selection of markers. Furthermore, new features are also included in the present version: (i) not only raw sequencing reads are accepted as input, but also contigs, allowing the analysis of assembled high‐coverage data; (ii) input data can be both in fasta and fastq format to facilitate the use of Illumina and IonTorrent reads; (iii) A comparison to known transposable elements allows their detection; (iv) A contamination check can be carried out by BLASTing potential markers against the nucleotide (nt) database of NCBI; (v) QDD3 is now also available imbedded into a virtual machine making installation easier and operating system independent. It can be used both on command‐line version as well as integrated into a Galaxy server, providing a user‐friendly interface, as well as the possibility to utilize a large variety of NGS tools.
Obinutuzumab (GA101), a type II, glycoengineered, humanized anti-CD20 monoclonal antibody, was superior to rituximab in human diffuse large B-cell lymphoma (DLBCL) and mantle-cell lymphoma (MCL) ...xenograft models. In phase I of our study, obinutuzumab (GA101) exhibited encouraging activity but no clear dose-response relationship, and few patients had aggressive histologies. The efficacy and safety of two doses of obinutuzumab (GA101) were explored in our randomized phase II trial in patients with heavily pretreated DBLCL and MCL.
Patients were randomly assigned to receive eight cycles of obinutuzumab (GA101) either as a flat dose of 400 mg for all infusions (days 1 and 8 of cycle 1; day 1 of cycles 2 to 8) or 1,600 mg on days 1 and 8 of cycle 1 and 800 mg on day 1 of cycles 2 to 8.
Forty patients were enrolled: 21 patients in the 400/400-mg treatment arm (DLBCL, n = 10; MCL, n = 11) and 19 patients in the 1,600/800-mg arm (DLBCL, n = 15; MCL, n = 4). End-of-treatment response was 28% (32% and 24% in the 1,600/800-mg and 400/400-mg study arms, respectively). Best overall response rates were 37% in the 1,600/800-mg arm and 24% in the 400/400-mg study arm (DLBCL, eight 32% of 25 patients; MCL, four 27% of 15 patients). Five (20%) of 25 rituximab-refractory patients exhibited treatment response, including four of 12 in the 1,600/800-mg group. The most common adverse events were infusion-related reactions (IRRs), which were manageable. Three patients had grade 3/4 IRRs. Grade 3/4 neutropenia was seen in only one patient.
Obinutuzumab (GA101) 1,600/800 mg achieves early steady-state concentration and clinical activity with an acceptable safety profile in relapsed/refractory DLBCL and MCL, supporting further exploration.
The close relationships between hosts and their microbiota underlie the current concept of holobionts. This biological entity enlarges our perception of the genomic diversity supported by ...individuals/species and the potential abilities of hosts to respond to the environment.
We investigated a total of 54 specimens of the sofie fish Parachrondrostoma toxostoma (Pt) for the taxonomic and functional composition of microbiota associated with the skin, gills, and gut tissues. Five wild populations from the Ardèche and Durance river basins in the Mediterranean part of the large catchment of the Rhône (southern France) were compared.
We found that the skin microbiota was more diverse and variable in bacterial community composition than the gut microbiota. These differences suggest that different deterministic factors underlie the composition of these bacterial communities, resulting in tissue‐specific variation of the microbiota composition between wild fish populations inhabiting two rivers. Microbiota diversity and composition differed between wild Pt populations at both regional and local spatial scales.
Using the Picrust tool, the taxonomic variation of the microbiota composition in the river fish populations could be associated with the enrichment of predicted metabolic pathways. In particular, carbohydrate metabolism was more represented and xenobiotic biodegradation pathways less represented in the gut microbiota of Pt populations from the Ardèche than of those from the Durance. This result suggests variability in the ability to acclimate to environmental features through the assimilation of food resources from plants and the detoxification of xenobiotic components.
This study opens perspectives in the detection of subtle shifts in environmental features through the description of the microbiota composition, and in the potential prediction of the functional capacities of species to acclimate.
Estimating the consequences of environmental changes, specifically in a global change context, is essential for conservation issues. In the case of pollutants, the interest in using an evolutionary ...approach to investigate their consequences has been emphasized since the 2000s, but these studies remain rare compared to the characterization of direct effects on individual features. We focused on the study case of anthropogenic ionizing radiation because, despite its potential strong impact on evolution, the scarcity of evolutionary approaches to study the biological consequences of this stressor is particularly true. In this study, by investigating some particular features of the biological effects of this stressor, and by reviewing existing studies on evolution under ionizing radiation, we suggest that evolutionary approach may help provide an integrative view on the biological consequences of ionizing radiation. We focused on three topics: (i) the mutagenic properties of ionizing radiation and its disruption of evolutionary processes, (ii) exposures at different time scales, leading to an interaction between past and contemporary evolution, and (iii) the special features of contaminated areas called exclusion zones and how evolution could match field and laboratory observed effects. This approach can contribute to answering several key issues in radioecology: to explain species differences in the sensitivity to ionizing radiation, to improve our estimation of the impacts of ionizing radiation on populations, and to help identify the environmental features impacting organisms (e.g., interaction with other pollution, migration of populations, anthropogenic environmental changes). Evolutionary approach would benefit from being integrated to the ecological risk assessment process.
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•Evolutionary processes are discussed but still poorly studied in radioecology.•Ionizing radiation is particularly prone to disrupt evolutionary processes.•Evolution studies provide new light on wildlife status from nuclear exclusion zones.•An evolutionary approach could be beneficial for radioecological risk assessment.•A wide range of environmental changes would require such evolutionary approach.
Analyses of high-throughput transcriptome sequences of non-model organisms are based on two main approaches: de novo assembly and genome-guided assembly using mapping to assign reads prior to ...assembly. Given the limits of mapping reads to a reference when it is highly divergent, as is frequently the case for non-model species, we evaluate whether using blastn would outperform mapping methods for read assignment in such situations (>15% divergence). We demonstrate its high performance by using simulated reads of lengths corresponding to those generated by the most common sequencing platforms, and over a realistic range of genetic divergence (0% to 30% divergence). Here we focus on gene identification and not on resolving the whole set of transcripts (i.e. the complete transcriptome). For simulated datasets, the transcriptome-guided assembly based on blastn recovers 94.8% of genes irrespective of read length at 0% divergence; however, assignment rate of reads is negatively correlated with both increasing divergence level and reducing read lengths. Nevertheless, we still observe 92.6% of recovered genes at 30% divergence irrespective of read length. This analysis also produces a categorization of genes relative to their assignment, and suggests guidelines for data processing prior to analyses of comparative transcriptomics and gene expression to minimize potential inferential bias associated with incorrect transcript assignment. We also compare the performances of de novo assembly alone vs in combination with a transcriptome-guided assembly based on blastn both via simulation and empirically, using data from a cyprinid fish species and from an oak species. For any simulated scenario, the transcriptome-guided assembly using blastn outperforms the de novo approach alone, including when the divergence level is beyond the reach of traditional mapping methods. Combining de novo assembly and a related reference transcriptome for read assignment also addresses the bias/error in contigs caused by the dependence on a related reference alone. Empirical data corroborate these findings when assembling transcriptomes from the two non-model organisms: Parachondrostoma toxostoma (fish) and Quercus pubescens (plant). For the fish species, out of the 31,944 genes known from D. rerio, the guided and de novo assemblies recover respectively 20,605 and 20,032 genes but the performance of the guided assembly approach is much higher for both the contiguity and completeness metrics. For the oak, out of the 29,971 genes known from Vitis vinifera, the transcriptome-guided and de novo assemblies display similar performance, but the new guided approach detects 16,326 genes where the de novo assembly only detects 9,385 genes.
This paper is based on the interdisciplinary research conducted in the south of France that analyses the different economic, social and environmental roles played by agricultural irrigation canals. ...We argue that beyond their productive role, which is to supply farmers with water, they fulfil other environmental services and play an important role in the context of future climate change to face challenges of adaptation. We point up several ecosystem services provided by such canals, e.g. replenishing the groundwater table, the development of riparian vegetation and wet areas in the Mediterranean zone, tools for regulating flooding and drought, the bases for new cultural approaches to nature. Moreover, they play an important role in the maintenance of an ichthyological biodiversity that is indispensable for the persistence of natural ecosystem. Functioning as an ecological corridor, they display interesting capacities as refuges for certain fish species under stress. Indeed, they can potentially connect upstream and downstream zones over a continuum of more than 300 km and thus covering very contrasted climatic zones (alpine versus Mediterranean). For now, most of these services remain largely unknown and underestimated. However, they serve as assets for territorial development since they combine economic, ecological and social factors whose remodelling is becoming increasingly necessary in the face of climate change.
The group of large five-spotted
Hemichromis
species from West Africa and Lower Guinea was revised using an integrative approach combining morphometry (measurements, meristics and colour patterns) and ...genetics (cytochrome b gene). Specimens from 100 populations were sorted into three main groups on the basis of morphological criteria. Samples from Lake Bosumtwi clustered together with others from West Africa in a large well-supported clade (bootstrap values and posterior probabilities: 98/96/100), both in morphometric and genetic analyses, raising again the hypothesis of synonymy of
H. frempongi
with
H. fasciatus
. Samples from southern Lower Guinea, including the type locality (Gabon) of
H. elongatus
, constituted a second weakly resolved cluster (64/38/51), resulting from a potential admixture of populations of diverse origins which share common morphological traits. Populations from northern Lower Guinea (Atlantic coastal basins) and the upper Chad basin in Cameroon formed a third strongly supported clade (99/100/100), which corresponds to a new species,
Hemichromis camerounensis
sp. nov., herein described.
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