Biology subjects

Jeunen, G.-J.

Publications and source records attributed to Jeunen, G.-J..

10 recordsLinked to original sources

bioRxiv2026

A streamlined proof of concept CRISPR-based environmental biosurveillance platform for the detection of closely related invasive rats

Environmental biosurveillance requires technologies that can rapidly translate genomic information into sensitive, specific, and deployable detection tools. Here, we present the first proof-of-concept application of SENTINEL (Smart Environmental Nucleic-acid Tracking using Inference from Neural-networks for Early-warning Localization), integrating artificial-intelligence-guided target discovery, isothermal amplification, CRISPR-based detection, and environmental DNA. Using Rattus exulans, R. norvegicus, and R. rattus as a challenging model of closely related invasive vertebrates, SENTINEL achieved species-specific discrimination without detectable cross-reactivity and sensitivities of 10 aM for plasmid DNA and 10 fg/uL for genomic DNA. Selected assays also showed reproducible semi-quantitative performance and successful eDNA validation. This first application of CRISPR-based environmental biosurveillance to terrestrial invasive vertebrates demonstrates a transferable framework for converting sequence data into field-deployable assays, with potential applications across biosecurity, conservation, agriculture, and environmental monitoring.

ecology↗

bioRxiv2025

Recovering historical fish eDNA from museum-preserved Antarctic filter feeders via non-destructive metabarcoding.

O_LIRecent technical advances have significantly enhanced the value of museum specimens for molecular research, with metagenomic and metabarcoding approaches expanding further the utility of museum collections. However, given the finite number of specimens, there is a critical need to move past destructive DNA extraction approaches and to explore non-destructive techniques. C_LIO_LIIn this study, we evaluated the feasibility of extracting historical fish eDNA from the ethanol preservative used to store Antarctic museum specimens. We compared a variety of extraction methods - centrifugation, evaporation, filtration, and precipitation - using ten replicate samples per treatment for statistical analyses. To assess potential differences in preservative-derived eDNA recovery across different filter-feeding taxonomic groups, we included a Bryozoa and two Porifera, i.e., Demospongiae and Hexactinellida. C_LIO_LIComparative analyses with tissue biopsies revealed that the total genomic DNA was significantly reduced for non-destructive approaches. However, 10 ml ethanol filtration performed equal to, or in some instances, outperformed tissue biopsies when investigating the historical eDNA of Antarctic fish using a 16S rRNA metabarcoding approach, both for the number of species detected (-diversity) and community characterisation ({beta}-diversity). C_LIO_LIOur findings demonstrate the potential of ethanol preservative as a valuable, non-destructive source of historical fish eDNA from museum-stored filter-feeding specimens. These results highlight the viability of non-destructive sampling for molecular research on museum collections, preserving specimen integrity while enabling biodiversity assessments. Further refinement of non-destructive eDNA extraction could expand its applicability across taxa, collection types, and preservation methods, ensuring the long-term sustainability of museum-based genomic research. C_LI

ecology↗

bioRxiv2024

CRISPR-based environmental biosurveillance assisted via artificial intelligence design of guide-RNAs

Environmental biosecurity challenges are worsening for aquatic ecosystems as climate change and increased anthropogenic pressures facilitate the spread of invasive species, thereby broadly impacting ecosystem composition, functioning, and services. Environmental DNA (eDNA) has transformed traditional biomonitoring through detection of trace DNA fragments left by organisms in their surroundings, primarily by application of the quantitative polymerase chain reaction (qPCR). However, qPCR presents challenges, including limited portability, reliance on precise thermal cycling, and susceptibility to inhibitors. To address these challenges and enable field-deployable monitoring, isothermal amplification techniques such as Recombinase Polymerase Amplification (RPA) paired with Clustered Regularly Interspaced Short Palindromic Repeats and associated proteins (CRISPR-Cas) have been proposed as alternatives. We report here the development of CORSAIR (CRISPR-based envirOnmental biosuRveillance aSsisted via Artificial Intelligence guide-RNAs), that harnesses the programmability of the CRISPR-Cas technology, RPA and the artificial intelligence (AI)-based tool Activity-informed Design with All-inclusive Patrolling of Targets (ADAPT) to deploy a swift RPA-CRISPR-Cas13a-based method that detects eDNA from two invasive species as proof of concept: Sabella spallanzanii and Undaria pinnatifida. CORSAIR showcased a robust, streamlined method augmented by ADAPT, reaching a high specificity when tested against co-occurring species and a 100% agreement with 12 PCR-benchmarked eDNA samples, reaching a sensitivity of 0.34 copies uL-1 in 1 hour with a cost of 3.5 USD per sample; thus highlighting CORSAIR as a powerful environmental biosurveillance platform for environmental nucleic acid detection. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/627849v1_ufig1.gif" ALT="Figure 1"> View larger version (64K): org.highwire.dtl.DTLVardef@b60942org.highwire.dtl.DTLVardef@119e924org.highwire.dtl.DTLVardef@1956bforg.highwire.dtl.DTLVardef@18e18b9_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical abstractC_FLOATNO C_FIG

molecular biology↗

bioRxiv2024

tombRaider - improved species and haplotype recovery from metabarcoding data through artefact and pseudogene exclusion.

Environmental DNA metabarcoding has revolutionized ecological surveys of natural systems. By amplifying and sequencing small gene fragments from environmental samples containing complex DNA mixtures, scientists are now capable of exploring biodiversity patterns across the tree of life in a time-efficient and cost-effective manner. However, the accuracy of species and haplotype identification can be compromised by sequence artefacts and pseudogenes. Despite various strategies developed over the years, effective removal of artefacts remains challenging and inconsistent data reporting standards hinder reproducibility in eDNA metabarcoding experiments. To address these issues, we introduce tombRaider, an open-source command line software program (https://github.com/gjeunen/tombRaider) and R package (https://github.com/gjeunen/tombRaider_R) to remove artefacts and pseudogenes from metabarcoding data post clustering and denoising. tombRaider features a modular algorithm capable of evaluating multiple criteria, including sequence similarity, co-occurrence patterns, taxonomic assignment, and the presence of stop codons. We validated tombRaider using various published data sets, including mock invertebrate communities, air eDNA from a zoo, and salmon haplotypes from aquatic eDNA. Our results demonstrate that tombRaider effectively removed a higher proportion of artefacts while retaining authentic sequences, thus enhancing the accuracy and reliability of eDNA-derived diversity metrics. This user-friendly software program not only improves data quality in eDNA metabarcoding studies, but also contributes to standardised reporting practices, an aspect currently lacking in this emerging research field.

bioinformatics↗

bioRxiv2024

Unlocking Antarctic molecular time-capsules - recovering historical environmental DNA from museum-preserved sponges

Marine sponges have recently emerged as efficient natural environmental DNA (eDNA) samplers. The ability of sponges to accumulate eDNA provides an exciting opportunity to reconstruct contemporary communities and ecosystems with high temporal and spatial precision. However, the use of historical eDNA (heDNA), trapped within the vast number of specimens stored in scientific collections, opens up the opportunity to begin to reconstruct the communities and ecosystems of the past. Here, using a variety of Antarctic sponge specimens stored in an extensive marine invertebrate collection, we were able to recover information on Antarctic fish biodiversity from specimens up to 20 years old. We successfully recovered 64 fish heDNA signals from 27 sponge specimens. Alpha diversity measures did not differ among preservation methods, but sponges stored frozen had a significantly different fish community composition compared to those stored dry or in ethanol. Our results show that we were consistently and reliably able to extract the heDNA trapped within marine sponge specimens, thereby enabling the reconstruction and investigation of communities and ecosystems of the recent past with a spatial and temporal resolution previously unattainable. Future research into heDNA extraction from other preservation methods, as well as the impact of specimen age and collection method will strengthen and expand the opportunities for this novel resource to access new knowledge on ecological change during the last century.

molecular biology↗

bioRxiv2022

Non-invasive real-time genomic monitoring of the critically endangered kakapo

We used non-invasive real-time genomic approaches to monitor one of the last surviving populations of the critically endangered k[a]k[a]p[o] (Strigops habroptilus). We first established an environmental DNA metabarcoding protocol to identify the distribution of k[a]k[a]p[o] and other vertebrate species in a highly localized manner using soil samples. Harnessing real-time nanopore sequencing and the high-quality k[a]k[a]p[o] reference genome, we then extracted species-specific DNA from soil. We combined long read-based haplotype phasing with known individual genomic variation in the k[a]k[a]p[o] population to identify the presence of individuals, and confirmed these genomically informed predictions through detailed metadata on k[a]k[a]p[o] distributions. This study shows that individual identification is feasible through nanopore sequencing of environmental DNA, with important implications for future efforts in the application of genomics to the conservation of rare species, potentially expanding the application of real-time environmental DNA research from monitoring species distribution to inferring fitness parameters such as genomic diversity and inbreeding.

genomics↗

bioRxiv2022

Environmental DNA reflects common haplotypic variation

Analysis of environmental DNA (eDNA) has gained widespread usage for taxonomically based biodiversity assessment. While interest in applying non-invasive eDNA monitoring for population genetic assessments has grown, its usage in this sphere remains limited. One barrier to uptake is that the effectiveness of eDNA detection below the species level remains to be determined for multiple species and environments. Here, we test the utility of this emergent technology in a population genetic framework using eDNA samples derived from water along New Zealands South Island (Otago Coast: n=9; Kaik[o]ura: n=7) and DNA obtained from tissue samples (n=76) of individual blackfoot p[a]ua (Haliotis iris) sampled from New Zealands Otago coast. We recovered four mitochondrial haplotypes from eDNA versus six from the tissue samples collected. Three common haplotypes were recovered with both eDNA and tissue samples, while only one out of three rare haplotypes - represented in tissue samples by one individual each - was recovered with our eDNA methods. We demonstrate that eDNA monitoring is an effective tool for recovering common genetic diversity from p[a]ua, although rare (< 5%) haplotypes are seldom recovered. Our results show the potential of eDNA to identify population-level haplotypes for gastropods in the marine environment identification below the species level and for studying the population genetic diversity of gastropods. This work supports eDNA methods as effective, non-invasive tools for genetic monitoring. Non-invasive eDNA sampling could minimize target organism stress and human interaction enabling population genetic research for hard-to-sample, delicate, or sensitive species.

genetics↗

bioRxiv2021

Assessing the utility of marine filter feeders for environmental DNA (eDNA) biodiversity monitoring

Aquatic environmental DNA (eDNA) surveys are transforming how we monitor marine ecosystems. The time-consuming pre-processing step of active filtration, however, remains a bottleneck. Hence, new approaches omitting active filtration are in great demand. One exciting prospect is to use the filtering power of invertebrates to collect eDNA. While proof-of-concept has been achieved, comparative studies between aquatic and filter feeder eDNA signals are lacking. Here, we investigated the differences among four eDNA sources (water; bivalves; sponges; and ethanol in which filter-feeding organisms were stored) along a vertical transect in Doubtful Sound, New Zealand using three metabarcoding primers (fish (16S); MiFish-E/U). While concurrent SCUBA diver observations validated eDNA results, laboratory trials corroborated in-field bivalve eDNA detection results. Combined, eDNA sources detected 59 vertebrates, while divers observed eight fish species. There were no significant differences in alpha and beta diversity between water and sponge eDNA and both sources were highly correlated. Vertebrate eDNA was detected in ethanol, although only a reduced number of species were detected. Bivalves failed to reliably detect eDNA in both field and mesocosm experiments. While additional research into filter feeder eDNA accumulation efficiency is essential, our results provide strong evidence for the potential of incorporating sponges into eDNA surveys.

molecular biology↗

bioRxiv2021

Environmental DNA (eDNA) metabarcoding surveys extend the range of invasion for non-indigenous freshwater species in Eastern Europe.

Active environmental DNA (eDNA) surveillance through species-specific amplification has shown increased sensitivity in the detection of non-indigenous species (NIS) compared to traditional approaches. When many NIS are of interest, however, active surveillance decreases in cost- and time-efficiency. Passive surveillance through eDNA metabarcoding takes advantage of the complex DNA signal in environmental samples and facilitates the simultaneous detection of multiple species. While passive eDNA surveillance has previously detected NIS, comparative studies are essential to determine the ability of eDNA metabarcoding to accurately describe the range of invasion for multiple NIS versus alternative approaches. Here, we surveyed twelve sites, covering nine rivers across Belarus for NIS with three different techniques, i.e., an ichthyological, hydrobiological, and eDNA survey, whereby DNA was extracted from 500 mL surface water samples and amplified with two 16S rRNA primer assays targeting the fish and macro-invertebrate biodiversity. Nine non-indigenous fish and ten non-indigenous sediment-living macro-invertebrates were detected by traditional surveys, while seven NIS eDNA signals were picked up, including four fish, one aquatic and two sediment-living macro-invertebrates. Passive eDNA surveillance extended the range of invasion further north for two invasive fish and identified a new NIS for Belarus, the freshwater jellyfish Craspedacusta sowerbii. False-negative detections for the eDNA survey could be attributed to (i) preferential amplification of aquatic over sediment-living macro-invertebrates from surface water samples and (ii) an incomplete reference database. The evidence provided in this study recommends the implementation of both molecular-based and traditional approaches to maximize the probability of early detection of non-native organisms.

ecology↗

bioRxiv2018

Bio-On-Magnetic-Beads (BOMB): Open platform for high-throughput nucleic acid extraction and manipulation

Current molecular biology laboratories rely heavily on the purification and manipulation of nucleic acids. Yet, commonly used centrifuge-and column-based protocols require specialised equipment, often use toxic reagents and are not economically scalable or practical to use in a high-throughput manner. Although it has been known for some time that magnetic beads can provide an elegant answer to these issues, the development of open-source protocols based on beads has been limited. In this article, we provide step-by-step instructions for an easy synthesis of functionalised magnetic beads, and detailed protocols for their use in the high-throughput purification of plasmids, genomic DNA and total RNA from different sources, as well as environmental TNA and PCR amplicons. We also provide a bead-based protocol for bisulfite conversion, and size selection of DNA and RNA fragments. Comparison to other methods highlights the capability, versatility and extreme cost-effectiveness of using magnetic beads. These open source protocols and the associated webpage (https://bomb.bio) can serve as a platform for further protocol customisation and community engagement.

molecular biology↗