肥満糖尿病ラットにおける胃バイパス後の選択的腸管刺激による糖代謝への影響

研究科: 医学薬学府学位:千大院医薬博甲第医1000号要約博士(医学)千葉大

Transport Distance of Invertebrate Environmental DNA in a Natural River

<div><p>Environmental DNA (eDNA) monitoring is a novel molecular technique to detect species in natural habitats. Many eDNA studies in aquatic systems have focused on lake or ponds, and/or on large vertebrate species, but applications to invertebrates in river systems are emerging. A challenge in applying eDNA monitoring in flowing waters is that a species' DNA can be transported downstream. Whether and how far eDNA can be detected due to downstream transport remains largely unknown. In this study we tested for downstream detection of eDNA for two invertebrate species, <i>Daphnia longispina</i> and <i>Unio tumidus</i>, which are lake dwelling species in our study area. The goal was to determine how far away from the source population in a lake their eDNA could be detected in an outflowing river. We sampled water from eleven river sites in regular intervals up to 12.3 km downstream of the lake, developed new eDNA probes for both species, and used a standard PCR and Sanger sequencing detection method to confirm presence of each species' eDNA in the river. We detected <i>D. longispina</i> at all locations and across two time points (July and October); whereas with <i>U. tumidus,</i> we observed a decreased detection rate and did not detect its eDNA after 9.1 km. We also observed a difference in detection for this species at different times of year. The observed movement of eDNA from the source amounting to nearly 10 km for these species indicates that the resolution of an eDNA sample can be large in river systems. Our results indicate that there may be species' specific transport distances for eDNA and demonstrate for the first time that invertebrate eDNA can persist over relatively large distances in a natural river system.</p></div

Depicted is the geographic area of study and river system where transport of eDNA was measured.

<p>Study area within Switzerland (A) and sampling locations (B) in the outflowing (direction indicated by red arrow) River Glatt. Red dot is the sampling location in the Lake Greifensee where the source populations for both species, <i>Daphnia longispina</i> and <i>Unio tumidus</i> are found. Chimlibach (yellow dot) is a small tributary feeding into the Glatt river system (direction indicated by yellow arrow) and served as a negative control. Black dots are sampling locations tested for presence of eDNA form the two species. Tributaries to the Glatt indicated in blue lines and additional dilution sources from wastewater treatment plant release points are indicated with black arrows. Numbers are the distance (in km) of the sampling sites away from the lake (measured as along-stream distance).</p

Primer pair sequences and specifications used for detection of environmental DNA of targeted species.

<p>* Mismatches in primer region come from comparisons with closely related co-occurring species used in primer design (<i>Unio crassus, Daphnia galeata</i>)</p><p>** Including target region and primers</p

Estimated pairwise differentiation (Фst) from the COI gene among vernal pool regions.

<p>Estimated pairwise differentiation (Фst) from the COI gene among vernal pool regions.</p

Map depicting in the extent of vernal pool regions throughout California (gray) and Southern Oregon and sampling localities (black dots) from which genetic data was generated for an analysis of phylogeographic structure and diversity for the species of <i>Branchinecta lynchi</i>.

<p>Vernal pool region abbreviations are as follows: Carrizo (CZVP), Central Coast (CCVP), Klamath Mountains (KMVP), Livermore (LMVP), Northeastern Sacramento Valley (NESV), Northwestern Sacramento Valley (NWVP), San Joaquin Valley (SJVP), Santa Barbara (SBVP), Solano-Colusa (SCVP), Southeastern Sacramento Valley (SESV), Southern Sierra Foothills (SSFH), and Western Riverside (WRVP). <i>B</i>. <i>lynchi</i> are not known from the Santa Rosa (SRVP), Lake-Napa (LNVP), Modoc Plateau (MPVP) or San Diego (SDVP) vernal pool regions in historical or contemporary samples and thus no sample locations were evaluated from these locations for this study.</p

Depicts the negative relationship between the age of the <i>Branchinecta lynchi</i> specimen and its success for amplification and sequencing for two mitochondrial genes (COI and 16S).

<p>Depicts the negative relationship between the age of the <i>Branchinecta lynchi</i> specimen and its success for amplification and sequencing for two mitochondrial genes (COI and 16S).</p

Depicts the patterns of isolation by distance within each vernal pool region between geographic distance in kilometers and genetic distance estimated with corrected K2P.

<p>CZVP, SJVP and WRVP regions were not analyzed due to low sample size.</p

Estimated molecular diversity indexes for <i>Branchinecta lynchi</i> for vernal pool regions.

<p>Estimated molecular diversity indexes for <i>Branchinecta lynchi</i> for vernal pool regions.</p

Range-wide phylogeographic structure of the vernal pool fairy shrimp (<i>Branchinecta lynchi</i>)

<div><p>Wetland habitats across the world are experiencing rapid modification and loss due to accelerating habitat conversion. Impacts to wetland habitats are particularly acute in California where up to 90% of wetland habitats have been modified or lost. Vernal pool ecosystems have therefore undergone a dramatic loss in habitat and along with them an entire endemic fauna is under threat of extinction. Recent efforts to conserve vernal pool habitat and associated species have involved restoration and creation of vernal pools as well as translocations of threatened species. The vernal pool fairy shrimp, <i>Branchinecta lynchi</i>, is one of several endemic and federally listed species being targeted for translocations. To guide reintroduction and conservation, detailed information on range-wide population structure and diversity is needed. We collected genetic data from two mitochondrial genes throughout the known extant range of <i>B</i>. <i>lynchi</i> to elucidate population structure and diversity of the species. We found support for phylogeographic structure throughout the range of <i>B</i>. <i>lynch</i> associated with isolated watersheds and vernal pool regions previously identified in the recovery plan for the species. The underlying mechanisms responsible for this broad pattern of genetic structure have yet to be identified. However, the evidence of only a few haplotypes being shared across the species range and patterns of isolation by distance within vernal pool regions suggests dispersal limitation may play a role. These results stress that conservation programs, at a minimum, should consider using individuals from regional populations as sources for reintroductions to maintain historical patterns of genetic differentiation. Additionally, because genetic structure is associated with vernal pool regions which are based on local hydrology and geology, translocations should proceed considering the distance between donor and recipient sites.</p></div