Publications by authors named "Céline Lopez-Roques"

20 Publications

  • Page 1 of 1

Characterisation of hydrocarbon degradation, biosurfactant production, and biofilm formation in Serratia sp. Tan611: a new strain isolated from industrially contaminated environment in Algeria.

Antonie Van Leeuwenhoek 2021 Apr 15;114(4):411-424. Epub 2021 Feb 15.

Génétique Moléculaire, Génomique et Microbiologie (GMGM), UMR 7156, Université de Strasbourg, Strasbourg, France.

A novel bacterial strain was isolated from industrially contaminated waste water. In the presence of crude oil, this strain was shown to reduce the rate of total petroleum hydrocarbons (TPH) up to 97.10% in 24 h. This bacterium was subsequently identified by 16S rRNA gene sequence analysis and affiliated to the Serratia genus by the RDP classifier. Its genome was sequenced and annotated, and genes coding for catechol 1,2 dioxygenase and naphthalene 1,2-dioxygenase system involved in aromatic hydrocarbon catabolism, and LadA-type monooxygenases involved in alkane degradation, were identified. Gas Chromatography-Mass Spectrometry (GC-MS) analysis of crude oil after biological treatment showed that Serratia sp. Tan611 strain was able to degrade n-alkanes (from C to C). This bacterium was also shown to produce a biosurfactant, the emulsification index (E24) reaching 43.47% and 65.22%, against vegetable and crude oil, respectively. Finally, the formation of a biofilm was increased in the presence of crude oil. These observations make Serratia sp. Tan611 a good candidate for hydrocarbon bioremediation.
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http://dx.doi.org/10.1007/s10482-021-01527-5DOI Listing
April 2021

DNAModAnnot: a R toolbox for DNA modification filtering and annotation.

Bioinformatics 2021 Jan 20. Epub 2021 Jan 20.

Université Libre de Bruxelles, Interuniversity Institute of Bioinformatics in Brussels (IB2), Brussels, 1050, Belgium.

Motivation: Long-read sequencing technologies can be employed to detect and map DNA modifications at the nucleotide resolution on a genome-wide scale. However, published software packages neglect the integration of genomic annotation and comprehensive filtering when analyzing patterns of modified bases detected using Pacific Biosciences (PacBio) or Oxford Nanopore Technologies (ONT) data. Here, we present DNAModAnnot, a R package designed for the global analysis of DNA modification patterns using adapted filtering and visualization tools.

Results: We tested our package using PacBio sequencing data to analyze patterns of the 6-methyladenine (6 mA) in the ciliate Paramecium tetraurelia, in which high 6 mA amounts were previously reported. We found Paramecium tetraurelia 6 mA genome-wide distribution to be similar to other ciliates. We also performed 5-methylcytosine (5mC) analysis in human lymphoblastoid cells using ONT data and confirmed previously known patterns of 5mC. DNAModAnnot provides a toolbox for the genome-wide analysis of different DNA modifications using PacBio and ONT long-read sequencing data.

Availability: DNAModAnnot is distributed as a R package available via GitHub (https://github.com/AlexisHardy/DNAModAnnot).

Supplementary Information: Supplementary data are available at Bioinformatics online.
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http://dx.doi.org/10.1093/bioinformatics/btab032DOI Listing
January 2021

Convergent rewiring of the virulence regulatory network promotes adaptation of Ralstonia solanacearum on resistant tomato.

Mol Biol Evol 2020 Dec 11. Epub 2020 Dec 11.

LIPM, Université de Toulouse, INRAE, CNRS, Castanet-Tolosan, France.

The evolutionary and adaptive potential of a pathogen is a key determinant for successful host colonization and proliferation, but remains poorly known for most of the pathogens. Here, we used experimental evolution combined with phenotyping, genomics and transcriptomics to estimate the adaptive potential of the bacterial plant pathogen Ralstonia solanacearum to overcome the quantitative resistance of the tomato cultivar Hawaii 7996. After serial passaging over 300 generations, we observed pathogen adaptation to within-plant environment of the resistant cultivar but no plant resistance breakdown. Genomic sequence analysis of the adapted clones revealed few genetic alterations but we provide evidence that all but one were gain of function mutations. Transcriptomic analyses revealed that even if different adaptive events occurred in independently evolved clones, there is convergence towards a global rewiring of the virulence regulatory network as evidenced by largely overlapping gene expression profiles. A subset of four transcription regulators, including HrpB, the activator of the type 3 secretion system regulon and EfpR, a global regulator of virulence and metabolic functions, emerged as key nodes of this regulatory network that are frequently targeted to redirect the pathogen's physiology and improve its fitness in adverse conditions. Significant transcriptomic variations were also detected in evolved clones showing no genomic polymorphism, suggesting that epigenetic modifications regulate expression of some of the virulence network components and play a major role in adaptation as well.
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http://dx.doi.org/10.1093/molbev/msaa320DOI Listing
December 2020

Complete Genome Sequence of sp. Strain Nx66, Isolated from Waters Contaminated with Petrochemicals in El Saf-Saf Valley, Algeria.

Microbiol Resour Announc 2020 Nov 19;9(47). Epub 2020 Nov 19.

Génétique Moléculaire, Génomique et Microbiologie, Université de Strasbourg, Strasbourg, France

sp. strain Nx66 was isolated from waters contaminated by petrochemical effluents collected in Algeria. Its genome was sequenced using Illumina MiSeq (2 × 150-bp read pairs) and Oxford Nanopore (long reads) technologies and was assembled using Unicycler. It is composed of one chromosome of 3.42 Mb and one plasmid of 34.22 kb.
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http://dx.doi.org/10.1128/MRA.01130-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7679099PMC
November 2020

Genome structure and content of the rice root-knot nematode ().

Ecol Evol 2020 Oct 13;10(20):11006-11021. Epub 2020 Sep 13.

IRD-CIRAD-University of Montpellier UMR Interactions Plantes Microorganismes Environnement (IPME) Montpellier France.

Discovered in the 1960s, is a root-knot nematode species considered as a major threat to rice production. Yet, its origin, genomic structure, and intraspecific diversity are poorly understood. So far, such studies have been limited by the unavailability of a sufficiently complete and well-assembled genome. In this study, using a combination of Oxford Nanopore Technologies and Illumina sequencing data, we generated a highly contiguous reference genome (283 scaffolds with an N50 length of 294 kb, totaling 41.5 Mb). The completeness scores of our assembly are among the highest currently published for genomes. We predicted 10,284 protein-coding genes spanning 75.5% of the genome. Among them, 67 are identified as possibly originating from horizontal gene transfers (mostly from bacteria), which supposedly contribute to nematode infection, nutrient processing, and plant defense manipulation. Besides, we detected 575 canonical transposable elements (TEs) belonging to seven orders and spanning 2.61% of the genome. These TEs might promote genomic plasticity putatively related to the evolution of parasitism. This high-quality genome assembly constitutes a major improvement regarding previously available versions and represents a valuable molecular resource for future phylogenomic studies of species. In particular, this will foster comparative genomic studies to trace back the evolutionary history of .  and its closest relatives.
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http://dx.doi.org/10.1002/ece3.6680DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7593179PMC
October 2020

Complete Genome Sequence of the Type Strain Pectobacterium punjabense SS95, Isolated from a Potato Plant with Blackleg Symptoms.

Microbiol Resour Announc 2020 Aug 6;9(32). Epub 2020 Aug 6.

Institute for Integrative Biology of the Cell (I2BC), CEA CNRS University Paris-Saclay, Gif-sur-Yvette, France

is a newly described species causing blackleg disease in potato plants. Therefore, by the combination of long (Oxford Nanopore Technologies, MinION) and short (Illumina MiSeq) reads, we sequenced the complete genome of SS95, which contains a circular chromosome of 4.793 Mb with a GC content of 50.7%.
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http://dx.doi.org/10.1128/MRA.00420-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7409842PMC
August 2020

Diversity of Pectobacteriaceae Species in Potato Growing Regions in Northern Morocco.

Microorganisms 2020 Jun 13;8(6). Epub 2020 Jun 13.

Institute for Integrative Biology of the Cell (I2BC), Université Paris-Saclay, CEA, CNRS, 91198 Gif-sur-Yvette, France.

Dickeya and Pectobacterium pathogens are causative agents of several diseases that affect many crops worldwide. This work investigated the species diversity of these pathogens in Morocco, where Dickeya pathogens have only been isolated from potato fields recently. To this end, samplings were conducted in three major potato growing areas over a three-year period (2015-2017). Pathogens were characterized by sequence determination of both the gene marker and genomes using Illumina and Oxford Nanopore technologies. We isolated 119 pathogens belonging to (19%), (3%), (5%), (56%) and (17%). Their taxonomic assignation was confirmed by draft genome analyses of 10 representative strains of the collected species. were isolated from a unique area where a wide species diversity of pectinolytic pathogens was observed. In tuber rotting assays, isolates were more aggressive than Pectobacterium isolates. The complete genome sequence of LAR.16.03.LID was obtained and compared with other genomes from public databases. Overall, this study highlighted the ecological context from which some Dickeya and Pectobacterium species emerged in Morocco, and reported the first complete genome of a strain isolated in Morocco that will be suitable for further epidemiological studies.
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http://dx.doi.org/10.3390/microorganisms8060895DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7356628PMC
June 2020

A Chromosome-Scale Genome Assembly Resource for Infecting Sedge Grass ( sp.).

Mol Plant Microbe Interact 2020 Jul 1;33(7):880-883. Epub 2020 Jun 1.

Laboratoire des Interactions Plantes-Microorganismes (LIPM), INRA, CNRS, 24 Chemin de Borde Rouge - Auzeville, CS52627, F31326 Castanet Tolosan Cedex, France.

The fungus is a close relative of the notorious polyphagous plant pathogens and but exhibits a host range restricted to plants from the genus ( family). To date, there are no genomic resources available for fungi in the genus. Here, we present a chromosome-scale reference genome assembly for . The assembly contains 24 contigs with a total length of 43.53 Mbp, with scaffold N of 2,649.7 kbp and N of 1,133.1 kbp. BRAKER-predicted gene models were manually curated using WebApollo, resulting in 11,275 protein-coding genes that we functionally annotated. We provide a high-quality reference genome assembly and annotation for as a resource for studying evolution and pathogenicity in fungi from the family.
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http://dx.doi.org/10.1094/MPMI-03-20-0060-ADOI Listing
July 2020

Long-Read Genome Sequence of the Sugar Beet Rhizosphere Mycoparasite .

G3 (Bethesda) 2020 02 6;10(2):431-436. Epub 2020 Feb 6.

Laboratoire de Recherche en Sciences Végétales, Université de Toulouse, CNRS, UPS, 24 Chemin de Borde Rouge, Auzeville, BP42617, 31326 Castanet-Tolosan, France,

is a soil born free living oomycete able to parasitize fungi and oomycetes prey, including important plant and animals pathogens. can colonize endophytically the root tissues of diverse plants where it induces plant defenses. Here we report the first long-read genome sequencing of a strain sequenced by PacBio technology. Sequencing of genomic DNA loaded onto six SMRT cells permitted the acquisition of 913,728 total reads resulting in 112X genome coverage. The assembly and polishing of the genome sequence yielded180 contigs (N50 = 1.3 Mb; L50 = 12). The size of the genome assembly is 41.9 Mb with a longest contig of 2.7 Mb and 15,007 predicted protein-coding genes among which 95.25% were supported by RNAseq data, thus constituting a new genome reference. This data will facilitate genomic comparisons of species that are commensal, beneficial or pathogenic on plant, or parasitic on fungi and oomycete to identify key genetic determinants underpinning their diverse lifestyles. In addition comparison with plant pathogenic or zoopathogenic species will illuminate genomic adaptations for pathogenesis toward widely diverse hosts.
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http://dx.doi.org/10.1534/g3.119.400746DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7003069PMC
February 2020

Independent Origin of XY and ZW Sex Determination Mechanisms in Mosquitofish Sister Species.

Genetics 2020 01 8;214(1):193-209. Epub 2019 Nov 8.

Physiological Chemistry, Biocenter, University of Wuerzburg, 97074, Germany

Fish are known for the outstanding variety of their sex determination mechanisms and sex chromosome systems. The western () and eastern mosquitofish () are sister species for which different sex determination mechanisms have been described: ZZ/ZW for and XX/XY for Here, we carried out restriction-site associated DNA (RAD-) and pool sequencing (Pool-seq) to characterize the sex chromosomes of both species. We found that the ZW chromosomes of females and the XY chromosomes of males correspond to different linkage groups, and thus evolved independently from separate autosomes. In interspecific hybrids, the Y chromosome is dominant over the W chromosome, and X is dominant over Z. In , we identified a candidate region for the Y-linked melanic pigmentation locus, a rare male phenotype that constitutes a potentially sexually antagonistic trait and is associated with other such characteristics, , large body size and aggressive behavior. We developed a SNP-based marker in the Y-linked allele of (), which was linked to melanism in all tested populations. This locus represents an example for a color locus that is located in close proximity to a putative sex determiner, and most likely substantially contributed to the evolution of the Y.
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http://dx.doi.org/10.1534/genetics.119.302698DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6944411PMC
January 2020

Author Correction: De novo phased assembly of the Vitis riparia grape genome.

Sci Data 2019 Oct 30;6(1):249. Epub 2019 Oct 30.

EGFV, Bordeaux Sciences Agro - INRA - Université de Bordeaux, ISVV, 210 chemin de Leysotte, 33882, Villenave d'Ornon, France.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.
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http://dx.doi.org/10.1038/s41597-019-0268-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6820792PMC
October 2019

Structure of the intergenic spacers in chicken ribosomal DNA.

Genet Sel Evol 2019 Oct 26;51(1):59. Epub 2019 Oct 26.

Saint Petersburg State University, Universitetskaya emb. 7/9, Saint Petersburg, 199034, Russian Federation.

Background: Ribosomal DNA (rDNA) repeats are situated in the nucleolus organizer regions (NOR) of chromosomes and transcribed into rRNA for ribosome biogenesis. Thus, they are an essential component of eukaryotic genomes. rDNA repeat units consist of rRNA gene clusters that are transcribed into single pre-rRNA molecules, each separated by intergenic spacers (IGS) that contain regulatory elements for rRNA gene cluster transcription. Because of their high repeat content, rDNA sequences are usually absent from genome assemblies. In this work, we used the long-read sequencing technology to describe the chicken IGS and fill the knowledge gap on rDNA sequences of one of the key domesticated animals.

Methods: We used the long-read PacBio RSII technique to sequence the BAC clone WAG137G04 (Wageningen BAC library) known to contain chicken NOR elements and the HGAP workflow software suit to assemble the PacBio RSII reads. Whole-genome sequence contigs homologous to the chicken rDNA repetitive unit were identified based on the Gallus_gallus-5.0 assembly with BLAST. We used the Geneious 9.0.5 and Mega software, maximum likelihood method and Chickspress project for sequence evolution analysis, phylogenetic tree construction and analysis of the raw transcriptome data.

Results: Three complete IGS sequences in the White Leghorn chicken genome and one IGS sequence in the red junglefowl contig AADN04001305.1 (Gallus_gallus-5.0) were detected. They had various lengths and contained three groups of tandem repeats (some of them being very GC rich) that form highly organized arrays. Initiation and termination sites of rDNA transcription were located within small and large unique regions (SUR and LUR), respectively. No functionally significant sites were detected within the tandem repeat sequences.

Conclusions: Due to the highly organized GC-rich repeats, the structure of the chicken IGS differs from that of IGS in human, apes, Xenopus or fish rDNA. However, the chicken IGS shares some molecular organization features with that of the turtles, which are other representatives of the Sauropsida clade that includes birds and reptiles. Our current results on the structure of chicken IGS together with the previously reported ribosomal gene cluster sequence provide sufficient data to consider that the complete chicken rDNA sequence is assembled with confidence in terms of molecular DNA organization.
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http://dx.doi.org/10.1186/s12711-019-0501-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6815422PMC
October 2019

De novo phased assembly of the Vitis riparia grape genome.

Sci Data 2019 07 19;6(1):127. Epub 2019 Jul 19.

EGFV, Bordeaux Sciences Agro - INRA - Université de Bordeaux, ISVV, 210 chemin de Leysotte, 33882, Villenave d'Ornon, France.

Grapevine is one of the most important fruit species in the world. In order to better understand genetic basis of traits variation and facilitate the breeding of new genotypes, we sequenced, assembled, and annotated the genome of the American native Vitis riparia, one of the main species used worldwide for rootstock and scion breeding. A total of 164 Gb raw DNA reads were obtained from Vitis riparia resulting in a 225X depth of coverage. We generated a genome assembly of the V. riparia grape de novo using the PacBio long-reads that was phased with the 10x Genomics Chromium linked-reads. At the chromosome level, a 500 Mb genome was generated with a scaffold N50 size of 1 Mb. More than 34% of the whole genome were identified as repeat sequences, and 37,207 protein-coding genes were predicted. This genome assembly sets the stage for comparative genomic analysis of the diversification and adaptation of grapevine and will provide a solid resource for further genetic analysis and breeding of this economically important species.
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http://dx.doi.org/10.1038/s41597-019-0133-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6642119PMC
July 2019

Whole-genome landscape of Medicago truncatula symbiotic genes.

Nat Plants 2018 12 5;4(12):1017-1025. Epub 2018 Nov 5.

IPS2, CNRS, INRA, Universities of Paris Diderot and Sorbonne Paris Cité, Gif sur Yvette, France.

Advances in deciphering the functional architecture of eukaryotic genomes have been facilitated by recent breakthroughs in sequencing technologies, enabling a more comprehensive representation of genes and repeat elements in genome sequence assemblies, as well as more sensitive and tissue-specific analyses of gene expression. Here we show that PacBio sequencing has led to a substantially improved genome assembly of Medicago truncatula A17, a legume model species notable for endosymbiosis studies, and has enabled the identification of genome rearrangements between genotypes at a near-base-pair resolution. Annotation of the new M. truncatula genome sequence has allowed for a thorough analysis of transposable elements and their dynamics, as well as the identification of new players involved in symbiotic nodule development, in particular 1,037 upregulated long non-coding RNAs (lncRNAs). We have also discovered that a substantial proportion (~35% and 38%, respectively) of the genes upregulated in nodules or expressed in the nodule differentiation zone colocalize in genomic clusters (270 and 211, respectively), here termed symbiotic islands. These islands contain numerous expressed lncRNA genes and display differentially both DNA methylation and histone marks. Epigenetic regulations and lncRNAs are therefore attractive candidate elements for the orchestration of symbiotic gene expression in the M. truncatula genome.
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http://dx.doi.org/10.1038/s41477-018-0286-7DOI Listing
December 2018

Author Correction: Parallels between experimental and natural evolution of legume symbionts.

Nat Commun 2018 11 2;9(1):4641. Epub 2018 Nov 2.

Microbial Evolutionary Genomics, Institut Pasteur, 28 Rue Dr. Roux, 75015, Paris, France.

Clémence Genthon and Céline Lopez-Roques, who performed sequencing, were inadvertently omitted from the author list. This has now been corrected in the PDF and HTML versions of the Article.
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http://dx.doi.org/10.1038/s41467-018-07205-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6214916PMC
November 2018

The Genomic Basis of Color Pattern Polymorphism in the Harlequin Ladybird.

Curr Biol 2018 10 23;28(20):3296-3302.e7. Epub 2018 Aug 23.

Aix Marseille Université, CNRS, IBDM, Marseille, France. Electronic address:

Many animal species comprise discrete phenotypic forms. A common example in natural populations of insects is the occurrence of different color patterns, which has motivated a rich body of ecological and genetic research [1-6]. The occurrence of dark, i.e., melanic, forms displaying discrete color patterns is found across multiple taxa, but the underlying genomic basis remains poorly characterized. In numerous ladybird species (Coccinellidae), the spatial arrangement of black and red patches on adult elytra varies wildly within species, forming strikingly different complex color patterns [7, 8]. In the harlequin ladybird, Harmonia axyridis, more than 200 distinct color forms have been described, which classic genetic studies suggest result from allelic variation at a single, unknown, locus [9, 10]. Here, we combined whole-genome sequencing, population-based genome-wide association studies, gene expression, and functional analyses to establish that the transcription factor Pannier controls melanic pattern polymorphism in H. axyridis. We show that pannier is necessary for the formation of melanic elements on the elytra. Allelic variation in pannier leads to protein expression in distinct domains on the elytra and thus determines the distinct color patterns in H. axyridis. Recombination between pannier alleles may be reduced by a highly divergent sequence of ∼170 kb in the cis-regulatory regions of pannier, with a 50 kb inversion between color forms. This most likely helps maintain the distinct alleles found in natural populations. Thus, we propose that highly variable discrete color forms can arise in natural populations through cis-regulatory allelic variation of a single gene.
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http://dx.doi.org/10.1016/j.cub.2018.08.023DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6203698PMC
October 2018

Parallels between experimental and natural evolution of legume symbionts.

Nat Commun 2018 06 11;9(1):2264. Epub 2018 Jun 11.

Microbial Evolutionary Genomics, Institut Pasteur, 28 rue Dr. Roux, 75015, Paris, France.

The emergence of symbiotic interactions has been studied using population genomics in nature and experimental evolution in the laboratory, but the parallels between these processes remain unknown. Here we compare the emergence of rhizobia after the horizontal transfer of a symbiotic plasmid in natural populations of Cupriavidus taiwanensis, over 10 MY ago, with the experimental evolution of symbiotic Ralstonia solanacearum for a few hundred generations. In spite of major differences in terms of time span, environment, genetic background, and phenotypic achievement, both processes resulted in rapid genetic diversification dominated by purifying selection. We observe no adaptation in the plasmid carrying the genes responsible for the ecological transition. Instead, adaptation was associated with positive selection in a set of genes that led to the co-option of the same quorum-sensing system in both processes. Our results provide evidence for similarities in experimental and natural evolutionary transitions and highlight the potential of comparisons between both processes to understand symbiogenesis.
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http://dx.doi.org/10.1038/s41467-018-04778-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5995829PMC
June 2018

The Rosa genome provides new insights into the domestication of modern roses.

Nat Genet 2018 06 30;50(6):772-777. Epub 2018 Apr 30.

Laboratoire Reproduction et Développement des Plantes, Univ Lyon, ENS de Lyon, UCB Lyon 1, CNRS, INRA, Lyon, France.

Roses have high cultural and economic importance as ornamental plants and in the perfume industry. We report the rose whole-genome sequencing and assembly and resequencing of major genotypes that contributed to rose domestication. We generated a homozygous genotype from a heterozygous diploid modern rose progenitor, Rosa chinensis 'Old Blush'. Using single-molecule real-time sequencing and a meta-assembly approach, we obtained one of the most comprehensive plant genomes to date. Diversity analyses highlighted the mosaic origin of 'La France', one of the first hybrids combining the growth vigor of European species and the recurrent blooming of Chinese species. Genomic segments of Chinese ancestry identified new candidate genes for recurrent blooming. Reconstructing regulatory and secondary metabolism pathways allowed us to propose a model of interconnected regulation of scent and flower color. This genome provides a foundation for understanding the mechanisms governing rose traits and should accelerate improvement in roses, Rosaceae and ornamentals.
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http://dx.doi.org/10.1038/s41588-018-0110-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5984618PMC
June 2018

The Complete Genome Sequence of the Fish Pathogen Provides Insights into Virulence Mechanisms.

Front Microbiol 2017 16;8:1542. Epub 2017 Aug 16.

Virologie et Immunologie Moléculaires, Institut National de la Recherche Agronomique, Université Paris-SaclayJouy-en-Josas, France.

is a devastating bacterial pathogen of wild and farmed marine fish with a broad host range and a worldwide distribution. We report here the complete genome sequence of the type strain NCIMB 2154. The genome consists of a 3,435,971-base pair circular chromosome with 2,866 predicted protein-coding genes. Genes encoding the biosynthesis of exopolysaccharides, the type IX secretion system, iron uptake systems, adhesins, hemolysins, proteases, and glycoside hydrolases were identified. They are likely involved in the virulence process including immune escape, invasion, colonization, destruction of host tissues, and nutrient scavenging. Among the predicted virulence factors, type IX secretion-mediated and cell-surface exposed proteins were identified including an atypical sialidase, a sphingomyelinase and a chondroitin AC lyase which activities were demonstrated .
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http://dx.doi.org/10.3389/fmicb.2017.01542DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5561996PMC
August 2017

Accurate Identification and Quantification of DNA Species by Next-Generation Sequencing in Adeno-Associated Viral Vectors Produced in Insect Cells.

Hum Gene Ther Methods 2017 06 21;28(3):148-162. Epub 2017 Apr 21.

1 INSERM UMR1089, University of Nantes, Centre Hospitalier Universitaire, Nantes, France.

Recombinant adeno-associated viral (rAAV) vectors have proven excellent tools for the treatment of many genetic diseases and other complex diseases. However, the illegitimate encapsidation of DNA contaminants within viral particles constitutes a major safety concern for rAAV-based therapies. Moreover, the development of rAAV vectors for early-phase clinical trials has revealed the limited accuracy of the analytical tools used to characterize these new and complex drugs. Although most published data concerning residual DNA in rAAV preparations have been generated by quantitative PCR, we have developed a novel single-strand virus sequencing (SSV-Seq) method for quantification of DNA contaminants in AAV vectors produced in mammalian cells by next-generation sequencing (NGS). Here, we describe the adaptation of SSV-Seq for the accurate identification and quantification of DNA species in rAAV stocks produced in insect cells. We found that baculoviral DNA was the most abundant contaminant, representing less than 2.1% of NGS reads regardless of serotype (2, 8, or rh10). Sf9 producer cell DNA was detected at low frequency (≤0.03%) in rAAV lots. Advanced computational analyses revealed that (1) baculoviral sequences close to the inverted terminal repeats preferentially underwent illegitimate encapsidation, and (2) single-nucleotide variants were absent from the rAAV genome. The high-throughput sequencing protocol described here enables effective DNA quality control of rAAV vectors produced in insect cells, and is adapted to conform with regulatory agency safety requirements.
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http://dx.doi.org/10.1089/hgtb.2016.185DOI Listing
June 2017