Insights into the representativeness of biodiversity assessment in large reservoir through eDNA metabarcoding

Cortez, T.; Torres, A. L.; Guimaraes, M.; Pinheiro, H. B.; Cabral, M.; Zielinsky, G.; Pereira, C. M.; de Castro, G. M.; Guerreiro, L. T.; Americo, J. A.; do Amaral, D. L.; Rebelo, M. F.

doi:10.1101/2024.05.02.592166

bioRxiv · 10.1101/2024.05.02.592166

Insights into the representativeness of biodiversity assessment in large reservoir through eDNA metabarcoding

Cortez, T.·Torres, A. L.·Guimaraes, M.·Pinheiro, H. B.·Cabral, M.·Zielinsky, G.·Pereira, C. M.·de Castro, G. M.·Guerreiro, L. T.·Americo, J. A.·do Amaral, D. L.·Rebelo, M. F.

View original source ↗Export metadata ↓

Abstract

Monitoring biodiversity on a large scale, such as in hydropower reservoirs, poses scientific challenges. Conventional methods such as passive fishing gear are prone to various biases, while the utilization of environmental DNA (eDNA) metabarcoding has been restricted. Most eDNA studies have primarily focused on replicating results from traditional methods, which themselves have limitations regarding representativeness and bias. In our study, we employed eDNA metabarcoding with three markers (12SrRNA, COI, and 16SrRNA) to evaluate the biodiversity of an 800 km{superscript 2} reservoir. We utilized hydrodynamic modeling to determine water flow velocity and the water renewal ratio throughout the study area. Additionally, we conducted statistical comparisons - rarefaction curves and multivariate methods - among samples as an alternative approach to assess biodiversity representation. The eDNA identified taxa previously documented in the reservoir by traditional monitoring methods, as well as revealed 29 - nine fishes and 20 non-fish - previously unreported species. These results highlight the robustness of eDNA as a biodiversity monitoring technique. Our findings also indicated that by randomly sampling 30% of the original number of samples, we could effectively capture the same biodiversity. This approach enabled us to comprehend the reservoirs biodiversity profile and propose a straightforward, cost-effective monitoring protocol for the future based on eDNA.

Source connections

Published article ↗

Explore related subjects

molecular biology Preprint

Keep this discovery

Explore connections, maps & timelines

BibTeX RIS

Cortez, T., Torres, A. L., Guimaraes, M., Pinheiro, H. B., Cabral, M., Zielinsky, G., Pereira, C. M., de Castro, G. M., Guerreiro, L. T., Americo, J. A., do Amaral, D. L., Rebelo, M. F.. 2024-05-02. Insights into the representativeness of biodiversity assessment in large reservoir through eDNA metabarcoding. https://doi.org/10.1101/2024.05.02.592166

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related preprints

bioRxiv2026

Plasmid architecture determines the stability of inverted terminal repeats in adeno-associated virus vectors

Recombinant vectors derived from adeno-associated viruses (rAAVs) are a mainstay of human gene therapy. rAAVs are produced from plasmids containing transgene cassettes flanked by inverted terminal repeats (ITRs), which form structured DNA elements that stabilize the ends of the single-stranded viral genome and are the only viral sequences required in cis for genome packaging. For decades, it has been recognized that propagation of ITR-containing plasmids can result in deletions and other mutations, prompting the use of specialized bacterial strains, modified growth conditions, and truncated or altered ITRs. Despite these practices, ITR instability remains a persistent source of plasmid heterogeneity. To identify determinants of ITR stability, we evaluated ITR integrity in one of the original cloned AAV2 genome isolates, a reconstructed AAV2 plasmid, and a synthetic rAAV vector containing full-length native AAV2 ITRs. We established a quantitative bioinformatic workflow for analyzing ITR-containing plasmids and virus preparations from raw Oxford Nanopore sequencing data. These experiments showed that ITRs were highly stable during short-term culture, whereas prolonged culture revealed strong positional effects, with preferential loss or mutation of the ITR nearest the plasmid origin of replication. Consistent with this model, a survey of 7,041 sequence-verifiable AAV plasmids from the Addgene repository identified a widely disseminated 11-bp ITR deletion in 4,773 plasmids; among analyzable two-ITR plasmids, this deletion was located in the origin-proximal ITR in 95.3% of cases. Guided by these findings, we constructed a novel rAAV entry vector with stable full-length native AAV2 ITRs that enabled efficient packaging of a 4,750-bp all-in-one CRISPR-Cas9 cassette. Finally, we developed a cell-based strategy to compare the effects of ITR mutations on rAAV genome integration, providing preliminary evidence that ITR sequence variation can influence integration outcomes. Together, these findings show that ITR instability is a preventable, position-dependent property of plasmid architecture and identify ITR integrity as an important variable in rAAV vector design and quality control.

molecular biology↗

bioRxiv2026

Single-point mutation alters odorant receptor sensitivity associated with host plant specialization in Spodoptera moths

Host specialization in herbivorous insects is often associated with divergence in chemosensory abilities. Here, we investigated the possible contribution of odorant receptors (ORs) in host plant restriction in the lily moth Spodoptera picta, a species specialized on Amaryllidaceae. Manual annotation of S. picta ORs in its genome revealed a repertoire similar in size and composition to those of its polyphagous sister species, S. littoralis and S. litura, suggesting that specialization did not involve major gene loss or expansion in the lily moth. To assess functional divergence beyond gene number, we applied a large scaled structure-based virtual screening approach to the entire OR repertoires of these three Spodoptera species, generating ligand-binding profiles for 120,591 volatile compounds. Among 69 1:1:1 OR orthologs, 24 exhibited divergent predicted binding spectra. We pinpointed OR29 that we also found to be highly expressed in both male and female antennae of S. picta through a RNAseq approach. Functional assays demonstrated that S. picta OR29 acquired heightened sensitivity to limonene enantiomers, volatiles emitted by host Amaryllidaceae inflorescences. Site-directed mutagenesis revealed that a single amino acid substitution within the predicted binding region underlies this shift in sensitivity. These results show that host specialization in S. picta has not been accompanied by significant OR repertoire remodeling, but rather by subtle molecular changes that fine-tune receptor sensitivity to host-derived volatiles.

molecular biology↗

bioRxiv2026

Arc represses gene expression in IS605-family transposons

Bacterial insertion sequences (IS) are compact transposable elements that encode proteins required for their mobility and maintenance, yet many also encode accessory proteins with poorly understood functions. For example, IS605-family elements often encode a transposase called TnpA and an RNA-guided nuclease called TnpB that supports transposon maintenance, alongside an additional ribbon-helix-helix protein named Arc. Though the roles of TnpA and TnpB have been extensively studied in recent years, the enigmatic function of Arc has not been investigated. Here, we show that Arc acts as a transcriptional repressor to directly bind the transposon's native promoter sequence regulating TnpA and TnpB gene expression. By systematically testing Arc-containing IS605 elements, we identified a conserved binding pattern at intergenic transposon sequences neighboring protein-coding genes through chromatin immunoprecipitation and sequencing analyses. We then used fluorescence reporter assays and demonstrated that these intergenic sequences function as strong promoters, and that the presence of Arc dramatically reduces their gene expression. Together, these findings identify Arc as a transposon-encoded transcriptional repressor, revealing a regulatory layer that may promote long-term persistence of IS605-family elements by keeping their activity in check. The widespread association of Arc homologs with diverse mobile elements and cellular genes suggests that these compact regulators may more broadly restrain the expression of neighboring genetic machinery across varied genomic contexts. neighboring genetic machinery across varied genomic contexts.

molecular biology↗