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Lacroix, V.

Publications and source records attributed to Lacroix, V..

7 recordsLinked to original sources

Extended t-cores for the de novo identification of transposable elements and other inexact repeats from short read RNAseq data

Transcribed repeats represent a major challenge in the de novo assembly of transcriptomes from short RNA-seq reads. Young transposable elements (TEs) and more generally, inexact repeats, create dense and ambiguous regions in the assembly graph, preventing the correct assembly of transcripts. In this paper, we introduce a fully de novo method based on the discovery of dense regions in the compacted De Bruijn graph (DBG) to identify such repeats directly from short read RNA-seq data, without requiring a reference genome or repeat database. Our approach defines the extended t-cores, subgraphs of the DBG that capture the complex topology induced by expressed inexact repeats appearing in RNA-seq reads. Independently of its interest for transcriptome assembly, the proposed method appears to be effective for the de novo identification of repeats in transcriptomes. After classifying cores using sequence-based motifs to distinguish simple repeats from potential TEs, we demonstrate its potential for the de novo discovery of TEs. We validate the approach on a Mus musculus dataset, showing that extended t-cores correspond to known expressed TE families. We also illustrate its de novo discovery potential on a non-model species, Canis lupus familiaris, where the method was also able to recover known TEs.

bioinformatics↗

Metabolic resistance of the tiger mosquito to pyrethroid insecticides in La Reunion Island likely results from local adaptation

The resistance of mosquitoes to insecticides is a valuable model system for studying the genetic bases of xenobiotic adaptation in insects. The spread of the Asian tiger mosquito Aedes albopictus combined to the massive use of pyrethroid insecticides to limit arbovirus transmission resulted in the rise of resistance in various continents. Here, we investigated the genetic mechanisms underlying the recent adaptation of this mosquito to deltamethrin in La Reunion island. Bioassays confirmed the presence of resistance alleles in field populations. The resistance phenotype was further enhanced in the laboratory following a few generations of controlled selection. Combining whole genome Pool-seq and RNA-seq revealed no evidence of target-site resistance mutations but the over-expression and variant selection of detoxification enzymes associated with pyrethroid metabolism including cytochrome P450s, transferases and ABC-transporters. Among over-expressed detoxification genes, only one was linked to a gene duplication while polymorphism data suggest most of them being trans-regulated. Genome-wide selection signatures revealed a 9 Mb inverted superlocus responding to insecticide selection whose phenotypical importance remains uncertain. Altogether, this study indicates that the multigenic metabolic resistance phenotype observed in this insular territory mainly results from local adaptation. From an applied perspective, this study provides a set of markers to track pyrethroid resistance in the tiger mosquito in the South-West Indian Ocean. As this region is subjected to recurrent arbovirus outbreaks, the additive resistance phenotype that may arise from the introduction of Kdr mutations from other territories also calls for improving resistance surveillance at the regional scale. Author summaryWhile novel vector control strategies are being developed, chemical insecticides remain widely used to control mosquitoes transmitting human diseases such as the Asian tiger mosquito. However, the recurrent use of insecticides resulted in the emergence of resistance which can ultimately affect vector control efficacy. Here, we investigate the genetic bases underlying the resistance of the Asian tiger mosquito to the pyrethroid insecticide deltamethrin in La Reunion island. By combining two complementary genomic approaches, we showed that resistance is mainly caused by an increased insecticide detoxification while classical Knock down resistance mutations affecting the target of the insecticide were not detected. We also showed that resistance is underlain by multiple genetic changes spread across the genome, supporting the local selection of resistance rather than the introduction of resistance alleles. Furthermore, we identified a large inverted supergene responding to insecticide selection. This study provides valuable insights into the genetic bases of insecticide resistance, enabling the implementation of molecular makers to improve the tracking of insecticide resistance in this major mosquito vector across the Indian Ocean.

genomics↗

Promoting intestinal healing by exclusive enteral nutrition with TGF-beta in a mouse model of colitis.

Background and aimsExclusive Enteral Nutrition (EEN) is the first line of treatment for pediatric Crohns disease (CD), but its mechanisms of action remain poorly understood. We studied EEN nutritional composition and TGF-{beta} effect in a mouse model of colitis, as well as the role of intestinal microbiota. MethodsMice were treated with Dextran Sulfate Sodium (DSS) during 5 days to induce colitis, until the inflammatory peak (day 7) or gut restitution (day 14). After DSS treatment, some of them received EEN formula such as Modulen IBD(R) (DM mice) or Infatrini Peptisorb(R) (DINF mice), with TGF-{beta} supplementation or neutralization, and clinical inflammation was evaluated. After sacrifice, macroscopic and microscopic inflammation were analyzed, as well as intestinal permeability (IP). The composition of mucosal colonic microbiota was analyzed and fecal microbiota transplantation was performed to evaluate its capacity to mediate anti-inflammatory and pro-regenerative effect. Colonic crypts from DSS and EEN mice were cultured as 3D organoids and cellular properties were analyzed. ResultsDSS mice developed colitis, as evidenced by the weight loss and clinical inflammation. It was accompanied by macroscopic inflammation such as colon thickness and edemas, and an elevated IP. In contrast, EEN mice with TGF-{beta} formula present faster weight recovery and decreased inflammatory parameters, with a normalized IP, suggesting gut restitution and functionality. These functional improvements were not obtained for EEN mice without TGF-{beta} formula. Moreover, EEN with Modulen IBD(R) (DM mice) modified the microbiota in comparison to DSS condition and attenuated inflammation. In addition, the organoids from DM mice colonic crypts treated had an enhanced survival, and re-epithelialization capacity. ConclusionsBoth EEN formula have anti-inflammatory properties, certainly by the nutritional composition. However, TGF-{beta} plays a significant role in intestinal restitution and restoring barrier function. These beneficial effects are partly mediated by the microbiota to maintain gut homeostasis.

physiology↗

Detection and identification of transposable element transcripts using Long Read RNA-seq in Drosophila germline tissues

Transposable elements (TEs) are repeated DNA sequences potentially able to move throughout the genome. In addition to their inherent mutagenic effects, TEs can disrupt nearby genes by donating their intrinsic regulatory sequences, for instance, promoting the ectopic expression of a cellular gene. TE transcription is therefore not only necessary for TE transposition per se but can also be associated with TE-gene fusion transcripts, and in some cases, be the product of pervasive transcription. Hence, correctly determining the transcription state of a TE copy is essential to apprehend the impact of the TE in the host genome. Methods to identify and quantify TE transcription have mostly relied on short RNA-seq reads to estimate TE expression at the family level while using specific algorithms to discriminate copy-specific transcription. However, assigning short reads to their correct genomic location, and genomic feature is not trivial. Here we retrieved full-length cDNA (TeloPrime, Lexogen) of Drosophila melanogaster gonads and sequenced them using Oxford Nanopore Technologies. We show that long-read RNA-seq can be used to identify and quantify transcribed TEs at the copy level. In particular, TE insertions overlapping annotated genes are better estimated using long reads than short reads. Nevertheless, long TE transcripts (> 4.5 kb) are not well captured. Most expressed TE insertions correspond to copies that have lost their ability to transpose, and within a family, only a few copies are indeed expressed. Long-read sequencing also allowed the identification of spliced transcripts for around 107 TE copies. Overall, this first comparison of TEs between testes and ovaries uncovers differences in their transcriptional landscape, at the subclass and insertion level.

bioinformatics↗

Transcriptome profiling of mouse samples using nanopore sequencing of cDNA and RNA molecules

Our vision of DNA transcription and splicing has changed dramatically with the introduction of short-read sequencing. These high-throughput sequencing technologies promised to unravel the complexity of any transcriptome. Generally gene expression levels are well-captured using these technologies, but there are still remaining caveats due to the limited read length and the fact that RNA molecules had to be reverse transcribed before sequencing. Oxford Nanopore Technologies has recently launched a portable sequencer which offers the possibility of sequencing long reads and most importantly RNA molecules. Here we generated a full mouse transcriptome from brain and liver using the Oxford Nanopore device. As a comparison, we sequenced RNA (RNA-Seq) and cDNA (cDNA-Seq) molecules using both long and short reads technologies and tested the TeloPrime preparation kit, dedicated to the enrichment of full-length transcripts. Using spike-in data, we confirmed that expression levels are efficiently captured by cDNA-Seq using short reads. More importantly, Oxford Nanopore RNA-Seq tends to be more efficient, while cDNA-Seq appears to be more biased. We further show that the cDNA library preparation of the Nanopore protocol induces read truncation for transcripts containing internal runs of Ts. This bias is marked for runs of at least 15 Ts, but is already detectable for runs of at least 9 Ts and therefore concerns more than 20% of expressed transcripts in mouse brain and liver. Finally, we outline that bioinformatics challenges remain ahead for quantifying at the transcript level, especially when reads are not full-length. Accurate quantification of repeat-associated genes such as processed pseudogenes also remains difficult, and we show that current mapping protocols which map reads to the genome largely over-estimate their expression, at the expense of their parent gene. The entire dataset is available from http://www.genoscope.cns.fr/externe/ONT_mouse_RNA.

bioinformatics↗

A fast and agnostic method for bacterial genome-wide association studies: bridging the gap between kmers and genetic events

MotivationGenome-wide association study (GWAS) methods applied to bacterial genomes have shown promising results for genetic marker discovery or fine-assessment of marker effect. Recently, alignment-free methods based on kmer composition have proven their ability to explore the accessory genome. However, they lead to redundant descriptions and results which are hard to interpret.\n\nMethodsHere, we introduce DBGWAS, an extended kmer-based GWAS method producing interpretable genetic variants associated with pheno-types. Relying on compacted De Bruijn graphs (cDBG), our method gathers cDBG nodes identified by the association model into subgraphs defined from their neighbourhood in the initial cDBG. DBGWAS is fast, alignment-free and only requires a set of contigs and phenotypes. It produces annotated subgraphs representing local polymorphisms as well as mobile genetic elements (MGE) and offers a graphical framework to interpret GWAS results.\n\nResultsWe validated our method using antibiotic resistance phenotypes for three bacterial species. DBGWAS recovered known resistance determinants such as mutations in core genes in Mycobacterium tuberculosis and genes acquired by horizontal transfer in Staphylococcus aureus and Pseudomonas aeruginosa - along with their MGE context. It also enabled us to formulate new hypotheses involving genetic variants not yet described in the antibiotic resistance literature.\n\nConclusionOur novel method proved its efficiency to retrieve any type of phenotype-associated genetic variant without prior knowledge. All experiments were computed in less than two hours and produced a compact set of meaningful subgraphs, thereby outperforming other GWAS approaches and facilitating the interpretation of the results.\n\nAvailabilityOpen-source tool available at https://gitlab.com/leoisl/dbgwas

bioinformatics↗

Representing Genetic Determinants in BacterialGWAS with Compacted De Bruijn Graphs

MotivationAntimicrobial resistance has become a major worldwide public health concern, calling for a better characterization of existing and novel resistance mechanisms. GWAS methods applied to bacterial genomes have shown encouraging results for new genetic marker discovery. Most existing approaches either look at SNPs obtained by sequence alignment or consider sets of kmers, whose presence in the genome is associated with the phenotype of interest. While the former approach can only be performed when genomes are similar enough for an alignment to make sense, the latter can lead to redundant descriptions and to results which are hard to interpret.\n\nResultsWe propose an alignment-free GWAS method detecting haplotypes of variable length associated to resistance, using compacted De Bruijn graphs. Our representation is flexible enough to deal with very plastic genomes subject to gene transfers while drastically reducing the number of features to explore compared to kmers, without loss of information. It accomodates polymorphisms in core genes, accessory genes and noncoding regions. Using our representation in a GWAS leads to the selection of a small number of entities which are easier to visualize and interpret than fixed-length kmers. We illustrate the benefit of our approach by describing known as well as potential novel determinants of antimicrobial resistance in P. aeruginosa, a pathogenic bacteria with a highly plastic genome.\n\nAvailability and implementationThe code and data used in the experiments will be made available upon acceptance of this manuscript.\n\nContactmagali.dancette@biomerieux.com

bioinformatics↗