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Estimating the genetic diversity of Pacific salmon and trout using multigene eDNA metabarcoding.

ABSTRACT: Genetic diversity underpins species conservation and management goals, and ultimately determines a species' ability to adapt. Using freshwater environmental DNA (eDNA) samples, we examined mitochondrial genetic diversity using multigene metabarcode sequence data from four Oncorhynchus species across 16 sites in Oregon and northern California. Our multigene metabarcode panel included targets commonly used in population genetic NADH dehydrogenase 2 (ND2), phylogenetic cytochrome c oxidase subunit 1 (COI) and eDNA (12S ribosomal DNA) screening. The ND2 locus showed the greatest within-species haplotype diversity for all species, followed by COI and then 12S rDNA for all species except Oncorhynchus kisutch. Sequences recovered for O. clarkii clarkii were either identical to, or one mutation different from, previously characterized haplotypes (95.3% and 4.5% of reads, respectively). The greatest diversity in O. c. clarkii was among coastal watersheds, and subsets of this diversity were shared with fish in inland watersheds. However, coastal streams and the Umpqua River watershed appear to harbour unique haplotypes. Sequences from O. mykiss revealed a disjunction between the Willamette watershed and southern watersheds suggesting divergent histories. We also identified similarities between populations in the northern Deschutes and southern Klamath watersheds, consistent with previously hypothesized connections between the two via inland basins. Oncorhynchus kisutch was only identified in coastal streams and the Klamath River watershed, with most diversity concentrated in the coastal Coquille watershed. Oncorhynchus tshawytscha was only observed at one site, but contained multiple haplotypes at each locus. The characterization of genetic diversity at multiple loci expands the knowledge gained from eDNA sampling and provides crucial information for conservation actions and genetic management.

SUBMITTER: Weitemier K

PROVIDER: S-EPMC8597136 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:DNA metabarcoding is used to generate species composition data for entire communities. However, sequencing errors in high-throughput sequencing instruments are fairly common, usually requiring reads to be clustered into operational taxonomic units (OTUs), losing information on intraspecific diversity in the process. While Cytochrome c oxidase subunit I (COI) haplotype information is limited in resolving intraspecific diversity it is nevertheless often useful e.g. in a phylogeographic context, helping to formulate hypotheses on taxon distribution and dispersal.This study combines sequence denoising strategies, normally applied in microbial research, with additional abundance-based filtering to extract haplotype information from freshwater macroinvertebrate metabarcoding datasets. This novel approach was added to the R package "JAMP" and can be applied to COI amplicon datasets. We tested our haplotyping method by sequencing (i) a single-species mock community composed of 31 individuals with 15 different haplotypes spanning three orders of magnitude in biomass and (ii) 18 monitoring samples each amplified with four different primer sets and two PCR replicates.We detected all 15 haplotypes of the single specimens in the mock community with relaxed filtering and denoising settings. However, up to 480 additional unexpected haplotypes remained in both replicates. Rigorous filtering removes most unexpected haplotypes, but also can discard expected haplotypes mainly from the small specimens. In the monitoring samples, the different primer sets detected 177-200 OTUs, each containing an average of 2.40-3.30 haplotypes per OTU. The derived intraspecific diversity data showed population structures that were consistent between replicates and similar between primer pairs but resolution depended on the primer length. A closer look at abundant taxa in the dataset revealed various population genetic patterns, e.g. the stonefly Taeniopteryx nebulosa and the caddisfly Hydropsyche pellucidula showed a distinct north-south cline with respect to haplotype distribution, while the beetle Oulimnius tuberculatus and the isopod Asellus aquaticus displayed no clear population pattern but differed in genetic diversity.We developed a strategy to infer intraspecific genetic diversity from bulk invertebrate metabarcoding data. It needs to be stressed that at this point this metabarcoding-informed haplotyping is not capable of capturing the full diversity present in such samples, due to variation in specimen size, primer bias and loss of sequence variants with low abundance. Nevertheless, for a high number of species intraspecific diversity was recovered, identifying potentially isolated populations and taxa for further more detailed phylogeographic investigation. While we are currently lacking large-scale metabarcoding datasets to fully take advantage of our new approach, metabarcoding-informed haplotyping holds great promise for biomonitoring efforts that not only seek information about species diversity but also underlying genetic diversity.

Project description:Environments characterized by physical extremes harbor unique species diversity with particular adaptations. Cliffs are harsh environments for organisms but host a great diversity of specialized plants with many endemics, rare and even endangered species. It is, however, less known which fungal diversity the cliff habitats contain and whether it differs among different cliff locations. We thus sampled soil from three separate cliff locations in the North, Centre, and South of Spain and used eDNA metabarcoding to determine fungal diversity. To better understand whether cliff specialist plants may promote particular fungal communities, we have sampled soil from crevices with cliff specialist plants and no apparent plants as controls. Major lifestyles found in cliff soils were saprotrophs, and major fungal orders were Dothideomycetes, Sordariomycetes, and Eurotiomycetes, while the amount of symbiotrophic fungi was relatively low. We found no significant differences in fungal amplicon sequence variant (ASV) richness among the three sampled locations, but the sites were significantly different in their community composition and their main indicator species. Overall, there were no significant differences in fungal ASV richness or composition between soils from cliff specialist plants and soils without plants, suggesting a unique fungal diversity in cliff soils independent from specialized plants. However, preliminary findings on soils of the specialist cliff plant Sedum dasyphyllum against control soils suggest that the presence of a specialist plant may be a relevant factor affecting the specificity of the fungal community in cliff soils. Our results indicate the existence of particular cliff fungal communities in each location, and that, despite limited and poorly developed soils and harsh conditions, cliffs can harbor a great diversity of fungal species, comparable to other ecosystems of Spain. This study points out that some fungi may be cliff-specific, shaping particular communities that mediate plant adaptations to cliffs' extreme conditions.

Dataset Information

Estimating the genetic diversity of Pacific salmon and trout using multigene eDNA metabarcoding.

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