Biology subjects

Sado, T.

Publications and source records attributed to Sado, T..

8 recordsLinked to original sources

bioRxiv2025

μCeta: a set of cetacean-specific primers for environmental DNA metabarcoding with minimal amplification of non-target vertebrates

Biodiversity monitoring is crucial for understanding ecosystem dynamics and species distributions, particularly in the context of anthropogenic impacts and climate change. Cetaceans, as key indicator species of marine ecosystems, face increasing threats from human activities, highlighting the need for effective, non-invasive monitoring methods. In the present study, we developed novel Cetacea-specific primer sets to enhance the detection efficiency of cetacean species through environmental DNA (eDNA) metabarcoding, while minimizing the amplification of non-target vertebrates, such as fish and humans. We retrieved mitochondrial genomes of 71 cetacean species from a public database and designed 20 candidate primer sets, which were assessed in silico for their specificity and capacity to differentiate cetacean sequences. Four primer sets with the best in silico performance were selected for empirical validation using DNA from tissue samples and eDNA from seawater collected from aquarium pools and Hong Kong coastal waters. All four primer sets effectively amplified cetacean DNA from tissue samples. However, in the aquarium pool tests, three primer sets failed to accurately identify one or more cetacean species due to a lack of interspecific variation within the amplified region. From these, we selected one primer set targeting a 267 bp region of the mitochondrial 12S rRNA gene, named {micro}Ceta, and applied it to water samples collected from Hong Kong coastal waters, where the iconic Indo-Pacific humpback dolphin (Sousa chinensis) was observed. {micro}Ceta successfully detected S. chinensis eDNA while avoiding amplification of non-target species such as fish or humans. Our results demonstrate that {micro}Ceta is a reliable tool for cetacean eDNA detection in Hong Kong waters, contributing to cetacean conservation and enhancing our understanding of marine biodiversity.

ecology↗

bioRxiv2024

A deletion at the X-linked ARHGAP36 gene locus is associated with the orange coloration of tortoiseshell and calico cats

The X-linked orange (O) locus in domestic cats controls an unknown molecular mechanism that causes the suppression of black-brownish pigmentation in favor of orange coloration. The alternating black-brownish and orange patches seen in tortoiseshell and calico cats are considered as classic examples of the phenotypic expression of random X-chromosome inactivation (XCI) occurring in female mammals. However, the O gene in the cat genome has not been identified, and the genetic variation responsible for the orange coloration remains unknown. We report here that a 5.1-kilobase (kb) deletion within an intron of the X-linked ARHGAP36 gene, encoding a Rho GTPase activating protein, is closely and exclusively associated with orange coloration. The deleted region contains a highly conserved putative regulatory element, whose removal presumably cause altered ARHGAP36 expression. Notably, ARHGAP36 expression in cat skin tissues is linked to the suppression of many melanogenesis genes, potentially shifting pigment synthesis from eumelanin to pheomelanin. Furthermore, we find evidence that the gene undergoes XCI in female human and mouse cells, and XCI-dependent CpG island methylation consistent with random XCI in female domestic cats. The 5.1-kb deletion seems widespread in domestic cats with orange coat coloration, suggesting a single origin of this coat color phenotype.

genetics↗

bioRxiv2022

Temperature sensitivity of the interspecific interaction strength of coastal marine fish communities

The effects of temperature on interaction strengths are important for understanding and forecasting how global climate change impacts marine ecosystems; however, tracking and quantifying interactions of marine fish species is practically difficult especially under field conditions, and thus, how temperature influences their interaction strengths under field conditions remains poorly understood. We herein performed quantitative fish environmental DNA (eDNA) metabarcoding on 550 seawater samples that were collected twice a month from 11 coastal sites for two years in the Boso Peninsula, Japan, and analyzed eDNA monitoring data using nonlinear time-series analytical tools. We detected fish-fish interactions as information flow between eDNA time series, reconstructed interaction networks for the top 50 frequently detected species, and quantified pairwise, fluctuating interaction strengths. Although there was a large variation, water temperature influenced fish-fish interaction strengths. The impact of water temperature on interspecific interaction strengths varied among fish species, suggesting that fish species identity influences the temperature effects on interactions. For example, interaction strengths that Halichoeres tenuispinis and Microcanthus stringatus received strongly increased with water temperature, while those of Engraulis japonicus and Girella punctata decreased with water temperature. An increase in water temperature induced by global climate change may change fish interactions in a complex way, which consequently influences marine community dynamics and stability. Our research demonstrates a practical research framework to study the effects of environmental variables on interaction strengths of marine communities in nature, which would contribute to understanding and predicting natural marine ecosystem dynamics.

ecology↗

bioRxiv2021

SmcHD1 underlies the formation of H3K9me3 blocks on the inactive X chromosome in mice

Stable silencing of the inactive X chromosome (Xi) in female mammals is critical for the development of embryos and their postnatal health. SmcHD1 is essential for stable silencing of the Xi, and its functional deficiency results in derepression of many X-inactivated genes. Although SmcHD1 has been suggested to play an important role in the formation of higher order chromatin structure of the Xi, the underlying mechanism is largely obscure. Here we explore the epigenetic state of the Xi in SmcHD1-deficient epiblast stem cells (EpiSCs) and mouse embryonic fibroblasts (MEFs) in comparison with their wild-type counterparts. The results suggest that SmcHD1 underlies the formation of H3K9me3-enriched blocks on the Xi, which, although the importance of H3K9me3 has been largely overlooked in mice, play a critical role in the establishment of the stably silenced state. We propose that the H3K9me3 blocks formed on the Xi facilitate robust heterochromatin formation in combination with H3K27me3, and the substantial loss of H3K9me3 caused by SmcHD1 deficiency leads to aberrant distribution of H3K27me3 on the Xi and derepression of X-inactivated genes.

genetics↗

bioRxiv2021

The critical role of natural history museums in advancing eDNA for biodiversity studies: a case study with Amazonian fishes

Ichthyological surveys have traditionally been conducted using whole-specimen, capture-based sampling with varied, but conventional fishing gear. Recently, environmental DNA (eDNA) metabarcoding has emerged as a complementary, and possible alternative, approach to whole-specimen methodologies. In the tropics, where much of the diversity remains undescribed, vast reaches continue unexplored, and anthropogenic activities are constant threats; there have been few eDNA attempts for ichthyological inventories. We tested the discriminatory power of eDNA using MiFish primers with existing public reference libraries and compared this with capture-based methods in two distinct ecosystems in the megadiverse Amazon basin. In our study, eDNA provided an accurate snapshot of the fishes at higher taxonomic levels and corroborated its effectiveness to detect specialized fish assemblages. Some flaws in fish metabarcoding studies are routine issues addressed in natural history museums. Thus, by expanding their archives to include eDNA and adopting a series of initiatives linking collection-based research, training and outreach, natural history museums can enable the effective use of eDNA to survey Earths hotspots of biodiversity before taxa go extinct. Our project surveying poorly explored rivers and using DNA vouchered archives to build metabarcoding libraries for Neotropical fishes can serve as a model of this protocol.

ecology↗

bioRxiv2020

Environmental DNA metabarcoding for biodiversity monitoring of a highly-diverse tropical fish community in a coral-reef lagoon: Estimation of species richness and detection of habitat segregation

An environmental DNA (eDNA) metabarcoding approach has been widely used for biodiversity monitoring of fishes, although it has rarely been applied to tropical and subtropical aquatic ecosystems, where species diversity is remarkably high. This study examined the extent to which species richness can be estimated in a small coral reef lagoon (1500 x 900 m) near Okinawa Island, southern Japan, where the surrounding waters are likely to harbor more than 1500 species of fish. During 2015-2017, a total of 16 capture-based surveys were conducted to create a faunal list of fish species, followed by eDNA metabarcoding based on seawater samples taken from 11 sites in the lagoon on a day in May 2019. We also tested whether eDNA metabarcoding could detect differences between adjacent fish communities inhabiting the offshore reef edge and shore-side seagrass beds within the lagoon. A total of 217 fish species were confirmed by the capture-based samplings, while 291 fish species were detected by eDNA metabarcoding, identifying a total of 410 species distributed across 119 families and 193 genera. Of these 410 species, only 96 (24% of the total) were commonly identified by both methods, indicating that capture-based surveys failed to collect a number of species detected by eDNA metabarcoding. Interestingly, two different approaches to estimate species richness based on eDNA data yielded values close to the 410 species, including one that suggested an additional three or more eDNA surveys from 11 sites (36 samples) would detect 90% of the 410 species. In addition, non-metric multi-dimensional scaling for fish assemblages clearly distinguished between the fish communities of the offshore reef edge and those of the shore-side seagrass beds.

ecology↗

bioRxiv2017

Demonstration of the potential of environmental DNA as a tool for the detection of avian species

Birds play unique functional roles in the maintenance of ecosystems, such as pollina-tion and seed dispersal, and thus monitoring bird communities (e.g., monitoring bird species diversity) is a first step towards avoiding undesirable consequences of anthro-pogenic impacts on bird communities. In the present study, we hypothesized that birds, regardless of their main habitats, must have frequent contact with water and that tissues that contain their DNA that persists in the environment (environmental DNA; eDNA) could be used to detect the presence of avian species. To this end, we applied a set of universal PCR primers (MiBird, a modified version of fish/mammal universal primers) for metabarcoding avian eDNA. We confirmed the versatility of MiBird primers by performing in silico analyses and by amplifying DNAs extracted from bird tissues. Analyses of water samples from zoo cages of birds with known species composition suggested that the use of MiBird primers combined with the Il-lumina MiSeq platform could successfully detect avian species from water samples. Additionally, analysis of water samples collected from a natural pond detected five avian species common to the sampling areas. The present findings suggest that avian eDNA metabarcoding would be a complementary detection/identification tool in cases where visual detection and identification of bird species is difficult.

ecology↗

bioRxiv2017

Quantitative monitoring of multispecies fish environmental DNA using high-throughput sequencing

Effective ecosystem conservation and resource management require quantitative monitoring of biodiversity, including accurate descriptions of species composition and temporal variations of species abundance. Therefore, quantitative monitoring of biodiversity has been performed for many ecosystems, but it is often time- and effort-consuming and costly. Recent studies have shown that environmental DNA (eDNA), which is released to the environment from macro-organisms living in a habitat, contains information about species identity and abundance. Thus, analyzing eDNA would be a promising approach for more efficient biodiversity monitoring. In the present study, we added internal standard DNAs (i.e., known amounts of short DNA fragments from fish species that have never been observed in a sampling area) to eDNA samples, which were collected weekly from a coastal marine ecosystem in Maizuru-Bay, Kyoto, Japan (from April 2015 to March 2016), and performed metabarcoding analysis using Illumina MiSeq to simultaneously identify fish species and quantify fish eDNA copy numbers. A correction equation was obtained for each sample using the relationship between the number of sequence reads and the added amount of the standard DNAs, and this equation was used to estimate the copy numbers from the sequence reads of non-standard fish eDNA. The calculated copy numbers showed significant positive correlation with those determined by quantitative PCR, suggesting that eDNA metabarcoding with standard DNA enabled useful quantification of eDNA. Furthermore, for samples that show a high level of PCR inhibition, our method might allow more accurate quantification than qPCR because the correction equations generated using internal standard DNAs would include the effect of PCR inhibition. A single run of Illumina MiSeq produced > 70 quantitative fish eDNA time series in our study, showing that our method could contribute to more efficient and quantitative monitoring of biodiversity.

ecology↗