It's Still in the Water!

[saltshop]

I think you guys just need to spend a day in each others shoes! I have a father who is a chemistry professor, while I am rather interested in biology and ecology when it comes to my glass box (dare I say I was a business/law major ). From the chemist point of view, one usually knows what is going to happen beforehand and then you perform simple experiments to prove what you had already formulted on paper and from what is already known. In this case organics+life+metals=chelators and detoxification of various elements. From the biology point of view, you usually have an effect and then set about determining the cause through experimentation, observation, and previous works. In this case we have the mysterious "old tank syndrome" combined with what is known about heavy metals in "real life" ecosystems.

I am really not sure why the two fields have clashed so much here. FWIW, from someone who is "rather familiar" with you both through postings I am actually quite surprised! I know Randy prefers to keep things, if at all possible, as close to NSW be it good or bad. Iodine for instance is added to keep NSW values even if there is no conclusive evidence of what it may do...but there really is no arguing with what is the natural environment, correct? Now if metals are measured through ICP in the wild and shown to be a problem then, even if our "organic soup" tanks most likely are somewhat different, why not try to keep NSW values if at all possible just in case. Now for Doc, I know you are not real fond of someone making conclusions without thoroughly investigating all other possibilties and are quick to point it out when someone else does it. To be honest I was quite surprised when I read the last column stating that LR and sand should be discarded every four years or so. I saw nothing to support that LR and sand is releasing large amounts of these metals and that there is a demonstrated chain of events leading to death of organisms...heck it could be some build-up of some type of organic that cause old tanks to crumble or even a combination of the two You have mentioned here that it may be looked in to as far as what the levels are in the sand. I am really unsure of how you came to the conclusion you did already when there is still more to be explored???

Now, you both can banish me from your forums or whatever you want to do (I can still hang out with Eric ), but as an outsider this is how it looks to me. You are arguing about the forest but can't see through the trees. If you were to put your heads together you could probably come up with some really good "experiments/trials" that would lead to a much better understanding than either of you could provide by yourselves. Peace....

 

[jayo]

What bothers me about the toxicity argument is that people that use natural seawater (with "normal" metal levels) don't seem to have systems that are any healthier than those that use artificial "toxic metal levels" salt. I've never heard anyone claim that they could keep delicate animals better with natural sea water (unless it was an open system, and then it is more likely plankton that is making the difference). Even Ron is working hard to get more "natural" water (with a salt mix lower in metals) - why bother when using water with ideal concentrations (natural sea water) doesn't seem to work any better than artificial for any reef tank I've ever seen?

jayo

 

[saltshop]

LOL...hey Scott at least us business guys see things clearly! What are the odds of tax law coming up in a reef forum? I have mentioned this elsewhere, but in some cases the sample size was less than N=23, for the case of Arsenic N=1, Iodide N=14, etc. as the results that were below detection limits were thrown out and considered non-consequencial. I am sure that would cause much distress among any statisticians out there.

 

[Habib]

Ron,

I will be running some sea urchin bioassays on tank water and sea water mixes within the next month. Provided that I can complete the tests in time for MACNA, I will report on them there.

I assume you will be counting the number of fertilized eggs.
Is this correct?

If so which percentage range of fertilized eggs would you consider to be acceptable?

And if the results are within the acceptable range what would your conclusion be?

And what would your conclusion be if it is not within the acceptable range?

TIA

 

[Randy Holmes-Farley]

Ah, yes... but they are scheduled.

Wonderful When you finish testing on corals and find them suffering, I'll help you promote the concern far and wide that we are harming corals .

 

[wade]

Sorry to keep this going, but I'm still a bit confused...

If I get around to doing this I will look at the sediment proper, basically to find out the metals levels are. I don't want to get into sampling pore water, etc. Simply put, at this time it isn't worth the money for me to do this, as all of this work has been either self-funded or funded with the help of some aquarists, money really is a driving force.

When you say sediment proper, what do you mean? Typically, sediment is tested by pouring off any residual water and then dessicating it, which excludes any water soluble (non-bound) chemicals. Perhaps, the misunderstanding is mine about ICP methodology. Can you run a sample that is equal parts water and araganite/sediment?

Also, from a scientific standpoint/methodology issue, I would like to see your water column tests performed on samples that have had a bit more in the way of control/explanation. I realize that you had to draw from aquarists who don't know too much about science (generally speaking, no offense meant to the readers), but were things such as PTFE solvent rinsed bottles used? were samples microwaved or microfiltered prior to use? Questions like that make me nervous about the data you presented. The reason being, as an example, what if the water from a given tank had bacteria/algae counts on the order of 10^3 per ml? This would indicate that many of the elements you analyzed for were likely a part of that biological system and then we get into issues of coral/fish feeding and selectivity.

In all, this is a very complex issue and a few simple tests will tell us little. True tox testing (urchin bioassay, et al ) will give us an indicator of overall health, but only if statistically sigificant sampling is used (with every effort made to control and get random samples). I guess my point to this is that it would be nice to have the data you are striving for, but I question its worth in a standalone fashion. And as you say, money drives everything, especially science, so the cost of doing this 'right' would be noticable.

That make sense?

Wade

 

[Habib]

Wade,

but were things such as PTFE solvent rinsed bottles used? were samples microwaved or microfiltered prior to use? Questions like that make me nervous about the data you presented. The reason being, as an example, what if the water from a given tank had bacteria/algae counts on the order of 10^3 per ml?

I also had concerns regarding this and also some other related matters.

A discussion with Ron about this can be found here:

What we test in the water

 

[rshimek]

Originally posted by SDBDRZ

Hi,


IIs a statistical sample size of 23 adequate to draw conclusions about the population as a whole?

No, probably not. And I hope that other studies will be done. I think reef tanks are relatively diverse. However, that having been said....

This series of tanks was taken from across America (NY to AZ to TX to MT, etc). The data are surprisingly consistent.

As I have said, there is no compelling reason to regard these few tanks as representative. However, there is likewise no compelling reason not to either.

Presently, they are all we have. I would love to see more studies done testing for these materials in a comparable manner. Such tests are unlikely. I advertise on both Reef Central and Reefs.Org for volunteers (who had to help pay for the the tests) and this is the best I got. The tests are spendy - the base costs are over $200 per tank (although we got a volume discount )

There that should keep me from getting flamed

:thumbsup:

 

[rshimek]

Originally posted by Habib

Habib,

I assume you will be counting the number of fertilized eggs.
Is this correct?

No, this will be a developmental test, running from fertilization up to the end of the prism stage.

If so which percentage range of fertilized eggs would you consider to be acceptable?

And if the results are within the acceptable range what would your conclusion be?

And what would your conclusion be if it is not within the acceptable range?

The test will be a modified EPA protocol (I can send it to you if you wish). The modification will primarily be the use of a tropical urchin, as the test was designed for a temperate species.

Anyway the endpoint will be a comparison between the number of normal embryos versus the number of deformities.

I will be testing development in:

Catalina Salt Water (NSW) being shipped to me;
Several salt mixes newly made up;
Several tank waters.

The working hypothesis is that that there is no difference in the waters from all the tanks.

If salt mixes contain toxic materials, these will have an effect on the tests.

If tank wates are modified (by any means - unfortunately I will not be able to determine how they are modified) so that the embryos survive that will show up as well in the tank waters.

I have not yet decided whether I wish to do other modifications - such as dosing the NSW with various levels of metals to mimic salt levels, etc.

 

[rshimek]

Originally posted by Randy Holmes-Farley

Wonderful When you finish testing on corals and find them suffering, I'll help you promote the concern far and wide that we are harming corals .

There is a voluminous literature on the basis of using bioassays as a way of determining toxicity of waters. If you are unfamiliar with such methods, I can provide you with some literature citations to get you started.

It is a standard methodology, and has been used in coral reef environments as well as elsewhere. The results are accepted by effectively every govenmental agency examining environmental degradation in the world, as well as being considered definitive by environmental scientists world wide.

The presumption is that injury to the the bioassay organism reflects injury to all organisms in the community. Coral bioassays exist, and they are cited in my recent article, and that metals concentrations resulting in coral mortality are lower than those found in our tanks. I would love to use these tests, but unfortunately, I cannot afford to purchase the corals to run them. Fortunately, the urchin developmental test, which I can afford to do, is widely accepted throughout the environmental assessment scientific community.

 

[rshimek]

Originally posted by wade

When you say sediment proper, what do you mean? Typically, sediment is tested by pouring off any residual water and then dessicating it, which excludes any water soluble (non-bound) chemicals.

Yes. And this is what would be done.

, but were things such as PTFE solvent rinsed bottles used?

The bottles were provided by the laboratory.

were samples microwaved or microfiltered prior to use?

No.

Questions like that make me nervous about the data you presented.

Yes, they should. On the other hand, these data are all we have. So... Accept them or reject them, unless someone else wants to do something else, they are all we have.

In all, this is a very complex issue and a few simple tests will tell us little.

With the amazingly high concentations found, they certainly give some indications.

but I question its worth in a standalone fashion.

I certainly welcome any effort to duplicate or replicate the sample analysis in any other form, so go to it...

And as you say, money drives everything, especially science, so the cost of doing this 'right' would be noticable.

Well, I don't think there is any evidence that the results are "wrong."

To invalidate these data would require additional testing and analysis, and I would love to see the results of such tests.

 

[saltshop]

IIs a statistical sample size of 23 adequate to draw conclusions about the population as a whole?

No, probably not. And I hope that other studies will be done. I think reef tanks are relatively diverse. However, that having been said....

I have a few questions along these lines, but first I have to say "holy cow, don't get on the computer after coming home from a Nascar party!" My apologies for the rambling post!!

I was looking through the Feb. article that lists the tank conc. v. IO and for the most part most of the elements seem to be held in tow relative to the starting point. Are you at any point going to put up the data from each tank in some sort of spreadsheet format? The reason I ask, is that in some cases the values fluctuate by significant amounts resulting in too large of a standard deviation to have it make any sort of sense, partly because of the limited sampling. An example of this would be for Lithium which has a standard deviation which is larger than the mean itself. Some of the others that would fall in line as being too random would include Cobalt, Iodide, Molybdenum, Phosphorus, and Silicon. I also have a question about Vanadium which has a Min. of 0.030, a Max of 0.037, and a mean of .023 +/- .047? I think there has to be something wrong with that one to get a mean larger than the minimum and a crazy SSTD. I am not sure if it was corrected in the later issues? I think in these cases it would be really nice to see what the data sets actually were to get an idea of the representation both above and below mean. If done, it would also be nice to note what salt mix was used to begin with. In some cases a single tank or 2 that uses Sea Chem products might throw off the whole curve for something like Boron, likewise certain salts have really high Lithium levels.

Lastly, have you done (or will do) any statistical runs on tank age versus the elements listed? If the premise is that these elements are building up over time there should be a statistically meaningful measure showing the older the tank the higher the build-up. I could not find such a thing in any of the articles.

From just looking at the means while taking into account the large SSTD and range it appears that we start with a crappy mix to begin with and it doesn't get any better over time and in a few cases a little worse. In some cases people probably started with an even worse mix ... I am hopeful that someone will PLEASE find a good source for the raw materials to make adequate salt at home to start with and if the elements "hold the line" like they did against IO most of this thread would become moot. I would have no problem paying a dollar plus per gallon to make up water that was even close to NSW to start with, as I am pretty sure water changes could be held to something like a semi-annual event. Heck we might even end up saving money in the long run. Thanks!

 

[El-ahrairah]

Ron,

Just thought I'd chime in on this interesting discusson. I have a couple of questions regarding your urchin experiment. What tropical urchin species are you planning to use? Will it be in season when you perform your experiments?

Actually, I can pretty much tell you what the results of your urchin experiment are going to be, since urchins don't like ASW. The tropical urchin will most likely give pretty much the same results as temperate species.

Your cultures using the Catalina sea water should have 90% or survivourship and high synchronicity or your doing something wrong. Whereas your cultures using ASW regardless of the brand will show much higher mortality and non-sychronous embryos. The water from your sample tanks will likely match the ASW samples, especially if the tanks use ASW.

The embryos and larvae of many other species will show the same results as well. This is not entirely due to the higher amounts of various metals (though it does play a large part) in the ASW but also the lack of many other compounds mostly organic that are found in NSW. Larvae are very intolerant to ASW this has been known for the past thirty years. I've actually tested this with L. pictus, S. purpuratus, S. neumayeri (antarctic species), C. gigas, and R. pachyptila (hydrothermal vent worm)myself.

I have to agree with you that the majority of tanks contain concentrations of metals that far exceed that of NSW, and that for most invertebrates these are poisons and that many(but not all) of the unexplained deaths that hobbyists see can be attributed to this. However how can you honestly say that the thousands of people with tanks that are capable of keeping healthy corals and other sensitive invertebrates is simply conjecture and that it is in itself not sufficient evidence that elevated levels of various elements will always poison the tank. Please do not argue with "those specimens that thrive are pollutant tolerant" I've been force to sit through enough toxicity seminars that that argument just doesn't wash (since even the tolerant species can't handle the levels of pollutants you say are in tank water).
I don't intend to come off harsh and apologis if I do.
-Michael

 

[Randy Holmes-Farley]

For those thinking about sediment toxicity, here's an article where truly polluted sediments are being tested for toxicity, with clean sediments in Instant Ocean Salt mix used as the nontoxic control sample:

http://www.wes.army.mil/el/dots/pdfs/eedp03-1.pdf

 

[Habib]

Ron,

No, this will be a developmental test, running from fertilization up to the end of the prism stage.

I don't know the EPA procedure for this test. I however know that the EPA procedure to say something about toxicity by determination of the percentage of fertilised eggs allows the use of NSW and synthetic seasalts such as Forty Fantoms, IO, HW-marinemix,...

The control should have a value of above 70% (IIRC).

The results of the controls that I have seen for some saltmixes which we use in our hobby are typically 80 - 90%!!!-----> Non-Toxic?!?

Spiking artificial seawater with IIRC 0.0025 ppm (2.5 ug/L) Cu --> 70%, 0.02 ppm Cu --> 40% (IIRC).


I also wonder what the EPA procedure allows to be used in the the test you intend to do? NSW and also artificial seawater??

In your last article you mention various results obtained for various coral stages. Abstracts which I have read for some of such studies mention the use of artificial seawater as control!

The studies you mention in your last article, has any of these used artificial seawater as control or was NSW used??

TIA

 

[El-ahrairah]

I don't know the EPA procedure for this test. I however know that the EPA procedure to say something about toxicity by determination of the percentage of fertilised eggs allows the use of NSW and synthetic seasalts such as Forty Fantoms, IO, HW-marinemix,...

The control should have a value of above 70% (IIRC).

The results of the controls that I have seen for some saltmixes which we use in our hobby are typically 80 - 90%!!!-----> Non-Toxic?!?

Ferilization tests are basically of no value. Sea urchins are basically foolproof when it comes to fertilizing them. If you don't get better than 90% fertilization then there is something wrong with the eggs. The real test for toxicity is if they can make it past gastrulation. In our lab we can get around 90% fertilization everytime, even in conditions that kill them by the time they hit blastula. I'd like to see ASW mixes that can regularly get urchins to settle. ASW is a poor substitute when trying to culture invertebrate larvae.
-Michael

 

[rshimek]

Originally posted by El-ahrairah

Michael,

ASW is a poor substitute when trying to culture invertebrate larvae.

As you have indicated this has been the case for some time, but it may not necessarily be so with all ASW. I have now recieved a mix that claims to have the trace element concentrations of NSW, and it is apparently marketed to labs working with EPA standardized bioassays. I will test this during the tests. I am interested to see if the larvae fare better in it than in other ASW mixes.

Actually, I can pretty much tell you what the results of your urchin experiment are going to be, since urchins don't like ASW

Maybe so, and maybe no. The last time I cultured urchins in Instant Ocean was for an invert class I taught 3 years ago, and we got in an open class room situation, larvae that lived for 5 weeks and had good juvenile rudiments in them prior to the termination of the class.

The urchins I will use are Lytechinus and they will be obtained from Carolina Biological Supply, and generally some individuals are ripe all year round.

As far as raising urchins through to competancy, I have raised:
Strongylocentrotus purpuratus, S. franciscanus, S. droebachiensis, S. pallidus, S pallidus x S. droebachiensis (I was the first to do this), and D. excentricus. Other animals I have spawned and raised include:numerous echinoderms of all classes except concentricycloideans, Serpula vermicularis, several anemones (no-brainers, they don't feed), sea pens (Ptilosarcus gurneyi) numerous gastropods with feeding planktotropic larvae, but specifically turrid gastropods such as Oenopota levidensis, and really a whole slug of others.

The water from your sample tanks will likely match the ASW samples, especially if the tanks use ASW.

I doubt this very much. After short periods in reef tanks, the salt water often seems quite fine for many larvae (I have collected gastropod, polychaete and flatworm larvae from my tanks and others have done the same). As much as I have been giving Randy a hard time about him not having any data to support his chelation myth, I actually think it or something like it is occurring, and that ASW may be effectively detoxified in a tank. However, the price that is paid for such detoxification is that the chelated or bound toxins end up in the sediments or live rock and will come back to haunt us.

(since even the tolerant species can't handle the levels of pollutants you say are in tank water).

Well, Michael, the pollutants are in the tank. And I have certainly seen pollution-tolerant species that handle far greater loads. See the comments below regarding bioassay labs.

As you indicate the problems with ASW have been know for 30 years or more - and actually I was part of the group (as a grad student at the UW Friday Harbor Labs) that first did some experimentation with such mixes.

Our presumption at the time was the the mixes lacked something. After the tests I have run, I suspect instead that they have too much of the trace metals.

Additionally, Michael, may I add....

Do you have Meg Strathmann's book on "Invert Reproduction and Development?" If so, you will find I am chapter advisor for the asteroid and prosobranch chapters. I have had significant experience with invertebrate embryology, development, and larval ecology.

One of the larger engineer-consulting firms in the Pacific NW is Parametrix, Inc. From 1988 until 1993 I was the head of their natural resources section, and set up and was the head of their bioassay lab from 1988 until 1991. During that time we ran literally thousands of urchin bioassays on Superfund site work (given the potential for litigation, these tests were very closely monitored for procedural compliance). I think I know how do the procedure .

Additionally, most of these tests concerned excess heavy metals concentrations (although quite a number also dealt with excess toxic organic compounds - so Randy, some folks can figure something to test for. ).

Habib

Yes, you will find tests done using several ASW mixes as controls. I think these tests are flawed, but if they show increased mortality above and beyond what is caused by the salt mix due to metals concentrations, it must mean the metals are REALLY nasty.

I will try to find the URL for the test protocol and you can down load it if you want.

 

[El-ahrairah]

Ron,

First off I am not challenging your ability to raise larvae or your knowledge on them. I posted the species that I had worked with to back up my claim that I had some experience with invertebrate larvae (that I didn't just walk in off the street - since you don't know me). Also the larval biology world isn't that big so naturally I know the Strathmann book quite well. You must have noticed then the fact that larvae - while they may reach settlement the numbers are not as high as when using NSW. When I first came into my lab I had already had a decades experience with marine aquaria and was quite shocked when my advisor told me ASW was a no-no, so naturally I had to check for myself. Consistantly the mortality with ASW is much greater than with NSW.

Our presumption at the time was the the mixes lacked something. After the tests I have run, I suspect instead that they have too much of the trace metals.

I still believe they are missing something. I agree that the heavy metals that are present are highly toxic to urchins (again this has already been done to death), but even if you make up seawater without the metals, I would say they don't do as well as with NSW (by "do" I mean their physiology not just thier mortality - ie respiration, protein synthesis, lipid metabolism...). I would argue that the seawater needs the natural DOC (animo acids and the like) for the urchins to survive as in NSW. I agree that the "toxins" present are part of the problem but only part of the problem with ASW - if you are trying to raise larvae.

As much as I have been giving Randy a hard time about him not having any data to support his chelation myth, I actually think it or something like it is occurring, and that ASW may be effectively detoxified in a tank. However, the price that is paid for such detoxification is that the chelated or bound toxins end up in the sediments or live rock and will come back to haunt us.

Okay then, from reading the earlier posts it wasn't apparent to me that you even considered accepting the idea. As far as coming back to haunt us, it might not be the case. The diversity of a reef tank will greatly differ from that in the wild - most likely from the bacterial point of view. Bacteria have a wonderful way of using everything and anything as an energy source. It may be (yes I know it's just specualtion) that the microbial community of a closed reef tank is able to utilize and "permenantly" detoxify many of the toxins in the water, thus the need for better toxin analysis of the sediments and rock of aged tanks. Naturally a monetary and timely undertaking for someone. Since I have never had it come back to haunt me personally and I have had tanks run (and still running) for a lot longer than four years, I am not yet convinced that the elevated levels of compounds in the ASW are that problematic.
-Michael

 

[El-ahrairah]

Ron,

One other thing, you didn't quite answer my first question as to which species you plan to use, Lytechinus... ??

Since you covered an entire genus I'm not surprised their are some ripe all year round.
-Michael

 

[Jabrams]

The urchins I will use are Lytechinus and they will be obtained from Carolina Biological Supply, and generally some individuals are ripe all year round.