Cataloguing: visiting e-DNA lab for Kanagawa Prefecture 3

 

I guess you have noticed that for e-DNA primers are very important. Unless there is a suitable primer, no PCR is possible. And we will never be able to have DNA info for all the living creatures on earth, i.e., the catalogue of primers is forever incomplete. In the end, it‘s the realm of the God, Alah, or some deity ... Humans are nonetheless trying to augment the inventory of primers. There is a strong “economic“ incentive. E-DNA is far cheaper option when environmental impact assessment is required before development of some tangible things, like road, port, airport, building, shopping center, etc. Actually Japanese Ministry of Land, Infrastructure, Transport and Tourism will switch to e-DNA completely during project plannings from FY 2028.

You see? The thing we have to do for e-DNA from river is collecting a bottle of water, or straining the liquid. What we need is a bottle or a filter with a syringe, a bucket, and chemicals to stabilize the DNA specimen collected in the container. I first did it alone (; my post on February 17, 2023). Man-power wise, it‘s economical. When we check aquatic creatures from a stream in a good-ol way, we need an army of specimen hunters with, at least, a hand net, a white tray, a bottle of medical alcohol, vial containers, tweezers, and any other gears for humans to wade in the flow of water (; my post on October 28, 2022). Moreover, e-DNA requires less than 30 minutes to collect the specimen from a field, whereas the traditional way is for a one-day expedition. It‘s a simple math for accountants which one is cheaper even when they include the cost of chemicals such as Benzalkonium chloride or RNAlater + postage for courier service. Yup, we need pros for PCR analysis, but the demand for experts is the same for the old way. We need somebody to identify the collected specimen is really from a family of Dipteromimidae, which would guarantee the water quality where it lives is excellent. The labour cost in this regard is the same for new or old way. When it comes to the monetary benefit, there is always somebody who can fund the effort for scientific research. Thus, many endeavours are going to develop at least an operational catalogue of primers.

It’s the chemical to be added for the specimen
before the vial undergo the centrifuge treatment.
The price tag: USD 340 for 10ml.

As of December 2023, Japanese aquatic e-DNA analyzers can use workable primers for fish (MiFish: Miya et al. 2015), crustacean (MiDeca: Komai et al. 2019), mammals (MiMammal: Ushio et al. 2016), birds (Mibird: Ushio eta al. 2018), amphibians (Amphi_16S: Sakata eta al.), eels (MiEel: Takeuchi et al. 2019), salamander (Hynobius: Sakai et al. 2019), and insects (MtInsect16S: Takenaka et al. 2021). Come to think of it, the first e-DNA for French bull flog was made public in 2008, and we have already come this far. This is saying something. Making these more comprehensive is an on-going process. For example, MtInsects, the latest workable addition for primers, is the product of long-hour toil in the lab of Shinshu University where they keep on cataloguing the DNA sequence of insects found in Japan. They hope someday their effort will lead to universal primer that can augment at once the sequence of various taxonomic groups from a bottle.

Someday, the scenery might be more simplified …

There also are several efforts to expand the method for collecting DNA. The lovely one I found is collecting rain droplets accumulated in a dimple of a leaf. It will gather the information of creatures come and go the leaf, won’t it? Romantic … Some are trying to collect them from the air. Sure, hay fever is coming from the airborne pollens constructed by their DNA. In Kobe, researchers are collecting soil for DNA in order to assess the effect of anti-deer measures in their forest. In short, many people in this archipelago try to utilize e-DNA analysis for cataloguing what’s dwelling in our land, besides the homo sapience.

Soon here becomes a safe for DNA data.

Moreover, currently the big project is taking place to make a database that is comprised of DNA data collected from all over Japan. Prof. Michio Kondoh of Tohoku University organized a consortium that establishes and manages a big DNA data gathered from the nation’s landscapes. It’s called ANEMONE. ANEMONE is already operational and anybody in this universe with internet access can search for DNA that is catalogued in Japan. Kanagawa Environmental Research Center is a proud founding member of the consortium (a bragging smile of Mr. Hasebe). This DB is a part of the Japanese effort in 30-by-30 roadmap corresponding to Kunming-Montreal Global Biodiversity Framework adopted at COP15 in 2022. As a national strategy, Japanese government pledges to aim for 30% of domain to be biodiversity sensitive, and there to achieve nature positive way of life. Prof. Kondoh and his gangs make ANEMONE open and to be used for anybody who engages in self-management of our nature that should yield sustainable co-existence of human community and wildlife. (… er, the DB has data from Japan, so I don’t think it is perfectly applicable to, say, Korean Peninsula …) In this regard, what we volunteers enjoy for monitoring water source forest of Yadoriki is in the context of SDGs (a bragging smile of Naomi 😊)!

The entrance page for ANEMONE DB

… yeah, that all sounds grandiose, but actually, it’s not that so serious. In the end, we forest instructors stroll Yadoriki Water Source Forest alone ourselves, or with visitors, and enjoy lots of findings of natural wonder, from trees, flowers, butterflies, fishes, gadflies, … Sometimes we are greeted by serows or flying squirrels, entertained by beautiful songs of blue-and-white flycathcer, while relaxed watching a flow of clouds in a clear blue sky. We simply record what we have met in the forest, and the result is feeding into the database. Our playground happens to be a part of global effort. Don‘t you think how lucky we are?

For the measures of e-DNA for environmental issues of Kanagawa Prefecture, please make a contact with Kanagawa Environmental Research Center 神奈川県環境科学センター

1-3-39 Shinomiya, Hiratsuka City, 254-0014
〒254-0014平塚市四之宮1-3-39
Phone: 0463-24-3311
FAX: 0463-24-3300

k-center@k-erc.pref.kanagawa.jp 

https://www.pref.kanagawa.jp/docs/b4f/index.html

30-by-30 with Yadoriki Water Source Forest; DNA sampling for water quality monitoring in Kanagawa Prefecture, 3

 

So, Kanagawa Prefecture recruits citizen volunteers to monitor water quality of Sagami 相模川 and Sakawa 酒匂川Rivers both of which provide tap water for the entire Kanagawa. Monitors are collecting biological data from the river system of these two rivers, including the biodiversity condition of land of the estuary. The scheme was started in 2007 when the General Principles for Water Source Environment Conservation and Regeneration Policy かながわ水源環境保全・再生施策大綱 began in effect (; my post for October 28th, 2022). From 2022, DNA sampling is added for monitors to collect data. By analyzing traces of DNA from river water, we can estimate the number of species and the quantity of them which live in that particular water. According to Mr. Yuta Hasebe, from roughly 1km upstream from the point of water sampling, the snippets of DNA can be collected in a DNA filter (; my post on February 17th) or in a 1L bottle (; last week’s post on March 3rd). The merits for employing DNA sampling for water quality monitoring are 5-holds.

1. It’s far easier to collect biological data in a field, compared with landing nets and vats to make specimen. For DNA, we just collect water in a research container, and done. Landing nets and vats require us to go very near or to enter the water, catch specimen, throw them in a bottle of alcohol, then do microscopic study. DNA is easier and safer, you see?

2. All the stages of studying samples can be divided into a simpler segment. If that is with a bottle-method, we collect water, the bottle is sent to the lab, the scientists filter the water to catch DNA, the DNA sample is checked with database, then the result is analyzed by researchers. Each stage can be done by a different person and/or team, and so collaborative work would improve productivity for environmental analysis. That’s VERY important as the budget of the Office is always tight.

3. Extracted DNA samples can be stored for 10-20 years in a freezer at -20 °C, which gives chances for future researchers to meta-monitor long-term changes in water quality.

4. Especially for DNA of fishes, Japanese database is the world largest. We can utilize it fully to analyze the environment fishes indicate for their living condition.

5. It is possible to find DNA for a creature that is not caught so far by landing nets. Say, no one has caught a fish ABC in that part of Sagami River, but the water from the point can contain their DNA. That’s news!

The total evaluation of water quality is done by counting the existence of indicator species. Indicators are not only for fishes, but also insects and the other water creatures. Though, the comprehensive DNA database is yet only for fishes. Scientists are not sitting idly to leave the matter as such. At this moment, the labs in Shinshu University and Kanagawa Institute of Technology are collaborating to expand the registered number of species of zoobenthos in the DNA database. Mr. Hasebe asked us, “Please send your specimen to our lab. We check it for identification, then do DNA sampling. The result will be reported to the team for creating zoobenthos database. We REALLY appreciate your help!!” As of February 2023, the database for non-fish species is nearing the start of operation. So, in FY 2023, the Prefecture is planning to analyze and report DNA samples of non-fish creatures collected by monitors. Such citizen mobilization for environmental DNA sampling is the first in Japan. Ahem! From this April Kanagawa Forest Instructors Association starts to collaborate with Kanagawa Environmental Research Center to monitor Yadoriki Stream. We have about 30 years of experience observing water creatures in Yadoriki Water Source Forest やどりき水源林 so that the partnership is a natural fit. DNA sampling last year (; my posts for February 17th and March 3rd) was our trial. We have had mixed results for the 3 spots in the estuary of Yadoriki Stream. By continuing the systematic monitoring with a help of professional researchers with the advanced technology we would be able to know more about our Water Source Forest … And there is more.

Need to be registered for their DNA …

Such water monitoring can be done only by continuous and patient efforts of local monitors cooperating with professional scientists. It should be followed by the next level endeavor to sustain and improve the biodiversity of the environment where the locals live. To this end, Prof. Michio Kondoh for Tohoku University has become a lead for creating a platform of citizen-scientists collaboration and for making the DNA database open source, usable for better nature conservation and management. The pals of Prof. Kondoh created a consortium named ANEMONE that is the portal for open database of fishes and zoobenthos of Japan. Citizen monitors and scientists feed the monitoring results to enrich the contents of this nationwide database. Kanagawa Prefecture is helping ANEMONE to have more data. Yadoriki’s last year’s result will become a part of ANEMONE’s database. But, alas, the problem is always money. For the help, the Ministry of Environment of Japan has bought Prof. Kondoh’s idea and now prepares preferential national tax treatment for private donation and cooperate sponsorship supporting activities for ANEMONE. At the moment, ANEMONE is welcoming registration of anybody who’s interested in it as a partner for the scheme. Especially corporate partners can expect benefit from the coming new tax system.

The reason why the MOE of Japan comes in here is the existence of international pledge Japan has made for 30-by-30 Conservation Goal at COP 15. The diplomats and national delegation brought back the homework to materialize what the international convention agreed, and give it to locals, like us in Kanagawa Prefecture, to meet the deadline of 2030. The tricky part for registering 30% of our land for 30-by-30 is, how we can prove that the area is environmentally protected with sufficient biodiversity possible to be counted for 30-by-30. Data stored in ANEMONE will provide evidence for that. Moreover, it can be a convenient tool for corporate accounting. When corporations help the idea of open database of environmental DNA, ANEMONE will give them certificate for helping improvement of biodiversity. Based on this paper, the national government will provide preferential tax treatment. “Donation to ANEMONE is nice money for your bottom line and CSR, isn’t it?” With more moneyed help, Japanese promise for international community will approach to the goal faster. Gotcha.

Actually, many corporations already financially support
 the management of Yadoriki Water Source Forest.
 This part of the Forest is generously funded by NGK.
 Hopefully they will be able to utilize coming new tax system
 with our monitoring data …

Within the scheme of General Principles for Water Source Environment Conservation and Regeneration Policy, Kanagawa Prefecture has Committee for Regenerating Nature of Tanzawa-Oyama Area. The panel for the Committee includes lots of names of professors, NPO chairpersons, Corporations, etc. They are people for mountains, and its Secretariat is at Kanagawa Natural Environment Conservation Center 神奈川県自然環境保全センター. Inevitably, they are now discussing how to verify Tanzawa-Oyama Area fit for 30-by-30 which requires properly scientific endorsement with data for biodiversity. Mr. Hasebe of Kanagawa Environmental Research Center 神奈川県環境科学センター says, “Well, at least for estuaries of Sagami and Sakawa Rivers, we have the system of citizen monitoring using professional resources and being connected to national database of ANEMONE. If corporations and large landlords help our monitoring with ANEMONE, we can provide sufficient certificate for 30-by-30 enrolment, and they have tax benefit. Not a bad deal, huh?”

This part of Yadoriki Water Source Forest is
funded by private donations …
Will they receive preferential tax treatment
as corporations under the new tax system?

In any case, nature monitoring using environmental DNA in Kanagawa Prefecture is just 1 year or so old. Let us see how we can proceed from here … Meanwhile, Yadoriki Water Source Forest is welcoming sweet smells of Oriental Paperbush. Yellow flowers are dotting here and there in the estuary of Yadoriki Stream. We are entering a nice season …

Now in Yadoriki Stream, tadpoles are having their form
 in eggs of Montane Brown Frog …

If you find environmental issues in waters of Kanagawa Prefecture, please make a contact with Kanagawa Environmental Research Center 神奈川県環境科学センター

1-3-39 Shinomiya, Hiratsuka City, 254-0014
〒254-0014平塚市四之宮1-3-39
Phone: 0463-24-3311
FAX: 0463-24-3300

k-center@k-erc.pref.kanagawa.jp
https://www.pref.kanagawa.jp/docs/b4f/index.html