Biology subjects

Boyer, F.

Publications and source records attributed to Boyer, F..

14 recordsLinked to original sources

bioRxiv2026

Workflow for multiplex microsatellite panel development and sample preparation for robust amplicon sequencing of low-template and degraded DNA: validation for non-invasive genotyping in three large carnivore species

High-throughput amplicon sequencing has transformed microsatellite (STR) genotyping by overcoming many of the limitations of fragment-length analysis, enabling more accurate, cost-effective, and standardized genotyping. Yet, protocols specifically designed for high-throughput sequencing (HTS)-based STR genotyping from low-template and degraded DNA remain scarce, despite the prevalence of these challenging sample types in ecological and conservation contexts. We present a methodology for the de novo development of robust STR multiplex panels together with a laboratory protocol for efficient and reliable STR genotyping by sequencing with low quantity and quality DNA samples. The protocol comprises (i) an automated bioinformatic pipeline to design large sets of short tetranucleotide markers optimized for multiplex amplicon sequencing of degraded and low-template DNA; (ii) guidelines for efficient in vitro optimization of multiplex amplification using directly low quantity/quality template DNA; and (iii) a library preparation procedure that improves detection of low-level allele signal while enabling quality assessment of STR amplicon sequencing under limiting DNA conditions. We demonstrate the approach by developing and validating STR panels for non-invasive genotyping of three large carnivore species: a 44-plex for the grey wolf (Canis lupus), a 41-plex for the Eurasian lynx (Lynx lynx), and a 30-plex for the brown bear (Ursus arctos). Multiplex performance was high, with [≥]91% of samples successfully genotyped at [≥]50% of loci (allele size range 28-110 bp across panels) and correctly assigned to known individuals, negligible levels of noise in the controls, and high discriminatory power (PIDsibs [≤]2.4 x 1e-12), also owing to sequence variation among same-length alleles at 15-50% of loci. The approach is broadly applicable to animal and plant species, a wide range of sample types, and large-scale analysis such as genetic monitoring. Our study reinforces the value of STR amplicon sequencing for ecological and conservation applications while highlighting the importance of marker design and laboratory workflows tailored to HTS-based genotyping for accurate and efficient implementation.

ecology↗

bioRxiv2025

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↗

bioRxiv2025

Rainwash eDNA to monitor tropical rainforest biodiversity

Tropical rainforest canopies shelter an under-explored reservoir of biodiversity. In recent years, the amplification and sequencing of taxonomically informative DNA fragments from environmental samples (i.e. eDNA) has revolutionized biomonitoring. Here, we explore the potential of DNA contained in canopy throughfall water to sample the biological diversity of rainforest canopies. By sampling rainwash eDNA in two 1ha-plots, one mature Amazonian forest and a nearby tree plantation, we were able to detect 170 plant taxa, 72 vertebrate taxa including mammals, birds, and amphibians, and 313 insect taxa including e.g. mosquitoes, ants, beetles. The taxonomic composition retrieved in these two plots reflected their different disturbance status, and the plantation exhibited a much lower diversity. We also found that rainwash eDNA persists over ten days in passive eDNA collectors while providing a local picture of the diversity. This suggests that this method can be scaled up for a cost-effective environmental management of rainforest, and more generally of all forests canopies. TeaserRainwash DNA reveals the identity of plants, animals, and insects diversity, offering a practical tool for monitoring canopy biodiversity.

ecology↗

bioRxiv2024

Untangling the contribution of adaptive versus non-adaptive processes in the evolution of reproductive isolation between Coenonympha butterflies.

Speciation is a key evolutionary process which has been studied in numerous organisms and at multiple scales, from lineage radiation to gene expression. However, the factors explaining the rise of new species are not yet fully understood, and the relative contribution of neutral versus selective evolutionary processes in triggering and maintaining reproductive isolation between lineages is still debated. To explore this question, we study the divergence of two butterfly species, Coenonympha arcania and C. gardetta (Nymphalidae), which diverged relatively recently but show strong ecological differences. Whole genome sequence data reveal high overall differentiation between the two lineages, best explained by a long period of isolation at the early stage of their divergence. Demographically explicit approaches identify that 6.6% of the genome (32.7 Mbp) is impermeable to gene flow between the two species. Lots of these barrier loci are located on the Z chromosome, potentially spanning 75% of its length which would indicate that a large Z effect is at play in this speciation. Moreover, only a small proportion of barriers showed signatures of selection, suggesting that non-adaptive processes largely contributed to the build-up of reproductive isolation. Therefore, although genes involved in stress response and response to hypoxia are interesting candidates under selection, the adaptation of C. gardetta to alpine conditions may not to be the main driver of speciation. Our study brings an original example of intertwined adaptive and non-adaptive processes leading to reproductive isolation in a speciation with secondary contact and improves our understanding of the genomic underpinnings of species divergence.

evolutionary biology↗

bioRxiv2024

From single species to communities: microsatellite amplicon sequencing to monitor felids using Feliplex

O_LIThe current biodiversity crisis demands a shift from single-species to multispecies approaches in conservation, particularly for rare and endangered species. However, this transition requires tools optimised for multispecies research, which are currently limited. C_LIO_LIRecent advances in high-throughput sequencing (HTS) technologies and bioinformatics have enabled efficient and robust acquisition of genetic data. Amplicon sequencing approaches, in particular, have demonstrated potential for enhancing non-invasive genetic studies of endangered species, but their application has been mostly limited to single species. C_LIO_LITo enable multispecies genetic research, we introduce a cost-effective and robust HTS-based amplicon sequencing approach for genotyping multiple species simultaneously, designed for population monitoring, including individual identification and ascertaining patterns of population structure. C_LIO_LIWe developed Feliplex, a multiplex panel of 85 co-amplifying tetranucleotide microsatellite markers for cross-genotyping Felidae species, to demonstrate the utility of our approach. Feliplex was validated on known samples from nine Indian felid species across the genera Panthera, Prionailurus, and Felis. We applied it to invasive (blood and tissue) and non-invasive (hair and faeces) DNA extracts from 173 wild individuals obtaining respectively 70% and 56% multilocus genotyping success rates. The panel accurately identified known population clusters in tigers (Panthera tigris, n=19) and revealed hitherto unknown genetic structure in fishing cats (Prionailurus viverrinus, n=40). C_LIO_LIFeliplexs wide applicability across Felidae allows reliable multispecies genotyping from low-quality/quantity samples, while supporting cost-effective genetic studies and conservation monitoring of lesser-known species like small cats. Our approach has a broad applicability and can be adapted to develop similar multispecies panels for closely related species groups. C_LI

ecology↗

bioRxiv2024

A genomic duplication spanning multiple P450s contributes to insecticide resistance in the dengue mosquito Aedes aegypti

Resistance of mosquitoes to insecticides is one example of rapid adaptation to anthropogenic selection pressures having a strong impact on human health and activities. Target-site modification and increased insecticide detoxification are the two main mechanisms underlying insecticide resistance in mosquitoes. While target-sites mutations are well characterised and often used to track resistance in the field, the genomic events associated with insecticide detoxification remain partially characterised. Recent studies evidenced the key role of gene duplications in the over-expression of detoxification enzymes and their potential use to track metabolic resistance alleles in the field. However, such genomic events remain difficult to characterise due to their complex genomic architecture and their co-occurrence with other resistance alleles. In this concern, the present work investigated the role of a large genomic duplication affecting a cluster of detoxification enzymes in conferring resistance to the pyrethroid insecticide deltamethrin in the mosquito Aedes aegypti. Two isofemale lines originating from French Guiana and being deprived from major target-site mutations showed distinct insecticide resistance levels. Combining RNA-seq and whole genome pool-seq identified a 220 Kb genomic duplication enhancing the expression of multiple contiguous cytochrome P450s in the resistant line. The genomic architecture of the duplicated loci was elucidated through long read sequencing, evidencing its transposon-mediated evolutionary origin. The involvement of this P450 duplication in deltamethrin survival was supported by a significant phenotypic response to the P450 inhibitor piperonyl butoxide together with genotype-phenotype association and RNA interference. Experimental evolution suggested that this P450 duplication is associated with a significant fitness cost, potentially affecting its adaptive value in presence of other resistance alleles. Overall, this study supports the importance of genomic duplications affecting detoxification enzymes in the rapid adaptation of mosquitoes to insecticides. Deciphering their genomic architecture provides new insights into the evolutionary processes underlying such rapid adaptation. Such findings provide new tools for the surveillance and management of resistance in the field.

evolutionary biology↗

bioRxiv2021

Multiple adaptive solutions to face climatic constraints: novel insights in the debate over the role of convergence in local adaptation

The extent to which genomic convergence shapes locally adapted phenotypes in different species remains a fundamental question in evolutionary biology. To bring new insights to this debate we set up a framework which aimed to compare the adaptive trajectories of two domesticated mammal species co-distributed in diversified landscapes. We sequenced the genomes of 160 sheep and 161 goats extensively managed along environmental gradients, including temperature, rainfall, seasonality and altitude, to identify genes and biological processes shaping local adaptation. Allele frequencies at adaptive loci were rarely found to vary gradually along environmental gradients, but rather displayed a discontinuous shift at the extremities of environmental clines. Of the more than 430 adaptive genes identified, only 6 were orthologous between sheep and goats and those responded differently to environmental pressures, suggesting different adaptive mechanisms in these two closely related species. Such diversity of adaptive pathways may result from a high number of biological functions involved in adaptation to multiple eco-climatic gradients, and provides more arguments for the role of contingency and stochasticity in adaptation rather than repeatability.

evolutionary biology↗

bioRxiv2020

Splice switching oligonucleotide mediated gene knockdown in B cells and plasma cells

The need to identify new therapeutic approaches to the treatment of cancers of the B lymphoid lineage is crucial. Unlike CRISPR/Cas technology, antisense strategies result in transient modifications of gene expression and lack mutagenic effects at the DNA level. Here, we provide evidence for efficient knockdown of c-REL and RELA expression after treatment with splice switching antisense oligonucleotides (SSO) inducing exon skipping and reading frameshifts. We also developed a tool to facilitate the choice of exons for on purpose inhibition of mouse and human gene expression. Interestingly, treatments with morpholino SSO targeting the c-REL exon 2 donor splice site or RELA exon 5 acceptor splice site elicited very efficient knockdown in diffuse large B cell lymphoma (DLBCL) cell lines and antibody-secreting cells derived from primary human B cells. Consistent with the clinical relevance of c-REL activation in DLBCLs, treatment with c-REL SSO induced major alterations in NF-{kappa}B and TNF signalling pathways and strongly decreased cell viability. Altogether, SSO-mediated knockdown is a powerful approach to transiently inhibit the expression of given genes in B-lineage cells that should pave the way for cancer treatments, provided optimized ligand-conjugations for in vivo delivery. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=169 SRC="FIGDIR/small/302984v1_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@170de4forg.highwire.dtl.DTLVardef@113229corg.highwire.dtl.DTLVardef@145377forg.highwire.dtl.DTLVardef@13e8c1e_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2020

metabaR : an R package for the evaluation and improvement of DNA metabarcoding data quality

O_LIDNA metabarcoding is becoming the tool of choice for biodiversity studies across taxa and large-scale environmental gradients. Yet, the artefacts present in metabarcoding datasets often preclude a proper interpretation of ecological patterns. Bioinformatic pipelines removing experimental noise have been designed to address this issue. However, these often only partially target produced artefacts, or are marker specific. In addition, assessments of data curation quality and the appropriateness of filtering thresholds are seldom available in existing pipelines, partly due to the lack of appropriate visualisation tools. C_LIO_LIHere, we present metabaR, an R package that provides a comprehensive suite of tools to effectively curate DNA metabarcoding data after basic bioinformatic analyses. In particular, metabaR uses experimental negative or positive controls to identify different types of artefactual sequences, i.e. reagent contaminants and tag-jumps. It also flags potentially dysfunctional PCRs based on PCR replicate similarities when those are available. Finally, metabaR provides tools to visualise DNA metabarcoding data characteristics in their experimental context as well as their distribution, and facilitate assessment of the appropriateness of data curation filtering thresholds. C_LIO_LImetabaR is applicable to any DNA metabarcoding experimental design but is most powerful when the design includes experimental controls and replicates. More generally, the simplicity and flexibility of the package makes it applicable any DNA marker, and data generated with any sequencing platform, and pre-analysed with any bioinformatic pipeline. Its outputs are easily usable for downstream analyses with any ecological R package. C_LIO_LImetabaR complements existing bioinformatics pipelines by providing scientists with a variety of functions with customisable methods that will allow the user to effectively clean DNA metabarcoding data and avoid serious misinterpretations. It thus offers a promising platform for automatised data quality assessments of DNA metabarcoding data for environmental research and biomonitoring. C_LI

bioinformatics↗

bioRxiv2020

BAdabouM: a genomic structural variations discovery tool for polymorphism analyses

Genomic Structural Variations (SVs) are known to impact the evolution of genomes and to have consequences on individuals fitness. Nevertheless, they remain challenging to detect in whole genome re-sequencing (WGS) data. Lots of methods detecting SVs are described in the literature but they might be hard to install, have non-trivial settings, do not detect all SVs categories and have generally high levels of false positive. Here we introduce BAdabouM, a fast (C written) and easy to install SVs discovery tool. BAdabouM auto evaluates read length, library size and mean coverage to set thresholds specific to each experiment. BAdabouM interprets multiple SVs signatures (reads aligned with a split, non-concordant mapped pairs or uneven coverage) to detect insertions, deletions, copy number variations, inversions, and translocations at single-nucleotide resolution. When compared with two widely used methods on simulated and real datasets, BAdabouM was faster, exhibited a similar accuracy with a good concordance on SVs detected, and detected significantly more insertions. BAdabouM was more reproducible to detect independently SVs across individuals, which is a clear advantage when characterizing population polymorphism. Furthermore, BAdabouM demonstrated a superior ability to detect breakpoints with a base pair resolution. BAdabouM proved to be efficient, fast and accurate to detect SVs, and handle. BAdabouM is a complementary method to be used for a more comprehensive detection of SVs, and is especially suited for studying polymorphism for all types of SVs with a high accuracy.

bioinformatics↗

bioRxiv2019

Glomerulosclerosis and kidney failure in a mouse model of monoclonal immunoglobulin light-chain deposition disease

Light chain deposition disease (LCDD) is a rare disorder characterized by glomerular and peritubular amorphous deposits of a monoclonal immunoglobulin (Ig) light chain (LC), leading to nodular glomerulosclerosis and nephrotic syndrome. We developed a transgenic model using site-directed insertion of the variable domain of a pathogenic human LC gene into the mouse Ig kappa locus, ensuring its production by all plasma cells. High free LC levels were achieved after backcrossing with mice presenting increased plasma cell differentiation and no Ig heavy chain (HC) production. Our mouse model recapitulates the characteristic features of LCDD, including progressive glomerulosclerosis, nephrotic-range proteinuria and finally, kidney failure. The variable domain of the LC bears alone the structural properties involved in its pathogenicity. RNA sequencing conducted on plasma cells demonstrated that LCDD LC induces endoplasmic reticulum stress, likely accounting for the high efficiency of proteasome inhibitor-based therapy. Accordingly, reduction of circulating pathogenic LC was efficiently achieved and not only preserved renal function, but partially reversed kidney lesions. Finally, transcriptome analysis of pre-sclerotic glomeruli revealed that proliferation and extracellular matrix remodelling represented the first steps of glomerulosclerosis, paving the way for future therapeutic strategies in LCDD and other kidney diseases featuring diffuse glomerulosclerosis, particularly diabetic nephropathy.

pathology↗

bioRxiv2018

Reproducing Transformation of Indolent B-cell Lymphoma by T-cell Immunosuppression of L.CD40 Mice.

Transformation of an indolent B-cell lymphoma is associated with a more aggressive clinical course and poor survival. The role of immune surveillance in the transformation of a B-cell indolent lymphoma towards a more aggressive form is poorly documented. To experimentally address this question, we used the L.CD40 mouse model, which is characterized by B-cell specific continuous CD40 signaling, responsible for spleen indolent clonal or oligoclonal B-cell lymphoma after one year in 60% cases. Immunosuppression was obtained either by T/NK cell depletion or by treatment with the T-cell immunosuppressive drug cyclosporin A. Immunosuppressed L.CD40 mice had larger splenomegaly with increased numbers of B-cells in both spleen and peripheral blood. High-throughput sequencing of immunoglobulin variable segments revealed that clonal expansion was increased in immunosuppressed L.CD40 mice. Tumor B cells of immunosuppressed mice were larger with an immunoblastic aspect, both on blood smears and spleen tissue sections, with increased proliferation rate and increased numbers of activated B-cells. Collectively, these features suggest that immune suppression induced a shift from indolent lymphomas into aggressive ones. Thus, as a preclinical model, immunosuppressed L.CD40 mice reproduce aggressive transformation of an indolent B-cell tumor and highlight the role of the immune surveillance in its clinical course, opening new perspective for immune restoration therapies.\n\nSummary statementHighlighting the role of immune surveillance, transformation of indolent B-cell lymphoma into an aggressive malignancy is experimentally reproduced after T-cell immune suppression in the L.CD40 preclinical mouse model.

cancer biology↗

bioRxiv2018

Double-digest RAD-sequencing: do wet and dry protocol parameters impact biological results?

O_LINext-generation sequencing technologies have opened a new era of research in genomics. Among these, restriction enzyme-based techniques such as restriction-site associated DNA sequencing (RADseq) or double-digest RAD-sequencing (ddRADseq) are now widely used in many population genomics fields. From DNA sampling to SNP calling, both wet and dry protocols have been discussed in the literature to identify key parameters for an optimal loci reconstruction.\nC_LIO_LIThe impact of these parameters on downstream analyses and biological results drawn from RADseq or ddRADseq data has however not been fully explored yet. In this study, we tackled this issue by investigating the effects of ddRADseq laboratory (i.e. wet protocol) and bioinformatics (i.e. dry protocol) settings on loci reconstruction and inferred biological signal at two evolutionary scale using two systems: a complex of butterfly species (Coenonympha sp.) and populations of Common beech (Fagus sylvatica).\nC_LIO_LIResults suggest an impact of wet protocol parameters (DNA quantity, number of PCR cycles during library preparation) on the number of recovered reads and SNPs, the number of unique alleles and individual heterozygosity. We also found that bioinformatic settings (i.e. clustering and minimum coverage thresholds) impact loci reconstruction (e.g. number of loci, mean coverage) and SNP calling (e.g. number of SNPs, heterozygosity). We however do not detect an impact of parameter settings on three types of analysis performed with ddRADseq data: measure of genetic differentiation, estimation of individual admixture, and demographic inferences. In addition, our work demonstrates the high reproducibility and low rate of genotyping inconsistencies of the ddRADseq protocol.\nC_LIO_LIThus, our study highlights the impact of wet parameters on ddRADseq protocol with strong consequences on experimental success and biological conclusions. Dry parameters affects loci reconstruction and descriptive statistics but not biological conclusion for the two studied systems. Overall, this study illustrates, with others, the relevance of ddRADseq for population and evolutionary genomics at the inter- or intraspecific scales.\nC_LI

molecular biology↗

bioRxiv2017

Soil community assembly varies across body sizes in a tropical forest

The relative influence of deterministic niche-based (i.e. abiotic conditions, biotic interactions) and stochastic-distance dependent neutral processes (i.e. demography, dispersal) in shaping communities has been extensively studied for various organisms, but is far less explored jointly across the tree of life, in particular in soil environments. Here, using a thorough DNA-based census of the whole soil biota in a large tropical forest plot, we show that soil aluminium, topography, and plant species identity are all important drivers of soil richness and community composition. Body size emerges as an important feature of the comparative ecology of the different taxa at the studied spatial scale, with microorganisms being more importantly controlled by environmental factors, while soil mesofauna rather display random spatial distribution. We infer that niche-based processes contribute differently to community assembly across trophic levels due to spatial scaling. Body size could hence help better quantifying important properties of multitrophic assemblages.

ecology↗