Evaluation of Marine Biodiversity Using Environmental DNA and Understanding the Amount of Resources of Aquatic Products
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The study of the diversity of marine organisms and ecosystems provides fundamental knowledge for protecting the richness of the ocean. In addition, the environmental DNA method is useful for the protection of biological resources because it does not involve the capture or destruction of organisms. This research method is friendly to rare species, such as endangered species and endemic species.
The United Nations has designated the decade starting in 2021 as the "Decade of Ocean Science," with the aim of contributing to the SDGs. Ocean science, as defined by the UN, includes the field of fisheries.
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Biodiversity is very high around Japan, with more than 4,000 fish species. Of these, hundreds of fish species are sold in the market. Japan’s diet and food culture are supported by a wide variety of marine life, including not only fish but also shellfish, crustaceans, and algae. This diversity is the most important point of the fishing industry that is not found in the agriculture and livestock industries.
For the sustainable use of these diverse and abundant marine resources in the future, it is important to properly manage the abundance. Because any marine organism is a member of the ecosystem, the management requires not only resource information, such as the amount, distribution, age, and size composition of the single resource species but also information on the environments and the organisms surrounding them. However, it is very difficult for us to know where and how many organisms live in the water. Current marine product surveys often use nets to capture target organisms and estimate the amount (concentration) based on the amount obtained, the hauling speed, and the size of the net mouth. When investigating marine organisms in coastal areas, we sometimes dive in the water and visually count the number of target organisms. Fishermen, on the other hand, use a fish sonar to confirm the presence of the fish. However, research using these methods requires not only a great deal of time and energy but also specialized knowledge for fish classification, as well as special equipment and techniques for capturing the fish. Thus, survey opportunities and the data obtained from the surveys are limited. In addition, the results of the surveys may vary depending on the investigator, investigation equipment, and investigation method, and the results often lack credibility. Environmental DNA analyses have attracted attention in recent years as a new research method to overcome these problems.
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All environments such as water, soil, and air, contain DNA derived from the individual organism. This is called environmental DNA (eDNA). Every organism contains DNA as genes. The eDNA analysis is a technology that was originally developed as a means of analyzing microorganisms such as bacteria, as any environment naturally contains their DNA. Molecular biology techniques were developed to directly examine the DNA of microorganisms in the environment, replacing traditional culture analysis in the 1990s. The development of the eDNA analysis has elucidated the existence of various microorganisms in different environments and play important roles in their respective ecosystem.
Water also contains DNA released from macro-organisms in the form of excrements, such as feces and secretions. Unlike microorganisms, the eDNA of macroorganisms is DNA isolated from living organisms. It was discovered in 2014 that such a small amount of DNA outside of the living organism can be detected. Because each species has specific DNA, the eDNA analysis allows us to obtain accurate information on the biological communities in an ecosystem. For example, you can find species of organisms that inhabit a certain area. Biomass and the number of organisms might be estimated from the concentration of DNA if it correlates with biomass. This method is particularly useful for surveying the distribution of species with small populations, such as endangered species and endemic species, as you do not capture or kill the organisms. Moreover, it is possible to identify the species in marine protected areas, where you cannot conduct direct biological surveys.
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The eDNA analysis starts with collecting water. Then the eDNA is collected by filtering the water. The DNA is extracted and purified. The purified DNA is amplified and analyzed. There are currently two main methods involved in the analysis process. One is the metabarcoding method, which detects many species within a particular taxon. The other is the species-specific detection method, which detects single species. The two methods have different advantages. The former can detect many species simultaneously in one analysis, while the latter can obtain data that can be used as an index of the biomass of the target.
You need universal primers that can simultaneously amplify many types of DNA. By searching relatively short sequences that contain two conserved regions with an intervening hypervariable region, a large number of species can be discriminated at once. The primers are used to amplify fragments of DNA by PCR (Polymerase Chain Reaction), which are then sequenced by a next-generation sequencer. If the obtained nucleotide sequence matches with the reference, it can be inferred that the species with the sequence exists in the environment. Universal primers that are effective for various taxonomic groups, including fish, amphibians, and mammals have been developed.
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You can analyze the concentration of the eDNA of a specific organism using species-specific primers. It leads to the estimation of biomass without collecting and/or killing organisms. However, the flow is complicated in the sea, and the DNA released from organisms is transported and diffused widely by the flow. In addition, the DNA is gradually decomposed due to the influence of bacteria, ultraviolet rays, and so on. Therefore, we first developed a numerical model that can reproduce the detailed physical process in the water. Applying the model to Maizuru Bay, we simulated the behavior of eDNA released from Japanese horse mackerel. Their distribution was estimated by a fish finder. As a result, we succeeded in reproducing the observed distribution of the eDNA concentration.
Even in the sea, which is not a closed but an open system, the biomass can be estimated from the simulation of the eDNA. We divided the target sea area into fine grids and used the model to simulate how much DNA released from each grid will reach all the grids. Then, we statistically estimated the fish biomass for each grid by calculating the combination to best match the distribution of the actual eDNA concentration. As a result, we obtained similar biomass to that estimated by the fish finder
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