Journal Marcel's next experimental design - suggestions welcomed

[ElleDee]

@Marcel G, I've been thinking a lot about organic substrates myself as I'm about to start my first new tank in a couple years and am returning to using soil after being disappointed with aquasoil. I've got my mix ready to go and I have high hopes for it, but I don't want to bore anyone with the details unless it is actually successful.

I have previously had good success with a commercial soilless potting mix with a couple of additives thrown in. I know a lot of people have been overwhelmed with algae using similar stuff and I have spent some time thinking about why I only had minor issues* in the early days of the tank and then basically nothing for a few years. I'd love to pretend it was entirely skill on my part, and while I do have a background in horticulture, I don't think that explains it. I didn't know anything about algae being coming to the hobby.

My working theory is that in my area the bulk of the organic material is what we call "pine fines" = very finely shredded and well aged pine bark flecks that are essentially a waste product from the timber industry. They have a lot of lignin, have a high carbon to nitrogen ratio, and are pretty recalcitrant to decomposition, so I'm thinking that might have saved me from the toxic dump of organics people sometimes see. Being slow to decompose could also provide a longer, slower drip of CO2 from the substrate (from microbial respiration), and/or the structure of the pieces may help the substrate resist compaction and aid with the the bulk flow of oxygen and nutrients between the water column and the substrate. I'm not exactly sure which factors are the most important, but that's what has come to mind.

Hopefully I'll have some more to say on this in a few months.

*I did try a no-tech tank that was a disaster of algae for a bit, but once I gave up and got the water flowing I had an immaculately clean tank without having to change out the soil.

 

[Tim Harrison]

I haven't used any organic substrate so far. I plan to use it in my next experiment. I don't know yet which type would be best. Any suggestions?

l’ve used various mixes, from moss peat on its own through to aquatic compost, the type usually used for potting aquatic pond plants. The advantage of the latter is it often provides nutrient levels high enough to aid good plant growth but low enough to prevent excessive release of nitrogenous compounds. And therefore may help mitigate any algae issues. It's well homogenized too, so maybe one less confounding factor to worry about.

However, once mineralised it can turn into heavy clay. To counter that I usually add extra moss peat, in a 1:1 ratio. I also chuck in some horticultural grit, about 10%, for good measure. Both help keep the soil structure open as it mineralises. Obviously this aids water movement, nutrient transference, and gas exchange. The peat also increases the soils CEC making nutrients more readily available to plant roots.

Finally, I cap the soil with 20-30mm of pool filter sand, grade 6/14, or sand with an average particle size of about 3mm. Pool filter sand is composed of inert silicates that will not affect water chemistry. The size and rounded shape of the grains prevents compaction which still allows for water movement, nutrient transference, and gas exchange.

 

[Marcel G]

I apologize for not specifying what I mean by "organic substrate". I simply meant any substrate with an organic component (i.e. both those modern "soil-based" aquarium substrates and various DIY mixes usually made from various garden substrates, up to river or lake sediments). I don't have any criteria set yet to choose by. I'm trying a local river sediment right now, but I don't think I've come across a very suitable type (plants don't seem to grow much in it, even though it had some Cabombas growing in its original habitat). I was inclined to prefer some "Japanese" [granulated] aquarium substrate, which is quite widely used among aquarists these days (and usually with pretty good references). But I'm giving myself some time to think.

 

[Marcel G]

I've been thinking a lot about organic substrates myself as I'm about to start my first new tank in a couple years and am returning to using soil after being disappointed with aquasoil ...

@ElleDee, can you please explain why aquasoil disappointed you?

 

[Marcel G]

l’ve used various mixes, from moss peat on its own through to aquatic compost, the type usually used for potting aquatic pond plants. The advantage of the latter is it often provides nutrient levels high enough to aid good plant growth but low enough to prevent excessive release of nitrogenous compounds. ... However, once mineralised it can turn into heavy clay. To counter that I usually add extra moss peat, in a 1:1 ratio. I also chuck in some horticultural grit, about 10%, for good measure. Both help keep the soil structure open as it mineralises. Obviously this aids water movement, nutrient transference, and gas exchange. The peat also increases the soils CEC making nutrients more readily available to plant roots. Finally, I cap the soil with 20-30mm of pool filter sand, grade 6/14, or sand with an average particle size of about 3mm. Pool filter sand is composed of inert silicates that will not affect water chemistry. The size and rounded shape of the grains prevents compaction which still allows for water movement, nutrient transference, and gas exchange.

I also can buy "substrate for aquatic plants" here, which I have used in my aquarium a few times. However, I never did any methodical comparison with other substrates, so I can't say how the substrate stood up to other "soil-based" substrates. I've also considered some sort of DIY version, but I see the problem with that one being the difficulty of replication. My goal is to find recipes that would be relatively easy to replicate. Therefore I would like to stick to some "standard" substrates ... if there is such a thing. However, it is possible that due to large differences in the properties of different substrates, it may not be possible to replicate this factor in the end. When I once had lab analysis done on three different samples of ADA Aqua Soil Amazonia, it turned out that each contained a different ratio of sand:silt:clay, which would indicate that the manufacturer probably used different soil types to make them. In other words, even if I use a substrate from a particular manufacturer, I probably can't guarantee that if other aquarists use the same substrate, they will get the same results as I did ... which is sad. But what can be done about it?

 

[ElleDee]

I apologize for not specifying what I mean by "organic substrate". I simply meant any substrate with an organic component (i.e. both those modern "soil-based" aquarium substrates and various DIY mixes usually made from various garden substrates, up to river or lake sediments).

This is what I assumed you meant!

@ElleDee, can you please explain why aquasoil disappointed you?

This is largely just a personal thing. I have found that my dirted tanks (the soilless mix with cap as described above) have been more productive over a longer time span, but obviously aquasoil is more easily topped up or replaced, and I may like a richer aquasoil like the original Amazonia more than Controsoil.

But I also don't like how it looks, and I don't like how lightweight it is. I keep disturbing the top layer as I clean the tank, and I feel like that is interfering with root growth and muddying the barrier between water and substrate. It's like, when I clean my tank and I have a cap, I can get all the surface detritus up without disturbing anything, but with aquasoil I keep stirring stuff up from the deeper layers and the surface is never really clean. I do want some detritus to work its way back into the substrate to feed the microbes/plants, but I want the top clean for algae control; I can achieve that with a cap and can't with aquasoil. I have never heard anyone have this issue, so I guess that makes it a personal problem!

 

[Tim Harrison]

When I once had lab analysis done on three different samples of ADA Aqua Soil Amazonia, it turned out that each contained a different ratio of sand:silt:clay, which would indicate that the manufacturer probably used different soil types to make them.

That's an interesting discovery Marcel, and may well account for some problems folk have experienced. I like AS type Gucci substrate for it's convenience with regards scaping. However, it's expensive and now you've highlighted it's not necessarily the same bag to bag, I'd suggest a compost based alternative capped with sand might be better.

I've not had aquatic compost analysed but I'm guessing it's pretty similar bag to bag. Either way, regardless it's also dirt cheap, and I should imagine a large bag will last several rounds of experiments across time so they will be repeatalbe without having to worry about substrate becoming another confounding variable.

 

[Yugang]

@Marcel G I have been reading all this thread, including your detailed posts, and I LOVE it! Your hands are still dry, your tanks empty, but just discussing the design of experiment brings up so many good thoughts.

I like to check with you how you feel about moving this thread forward. I am a believer that every minute spent on design of experiment is worth it, because the costs of the experiment (especially time, and cost) is such that any idea that gives a better final result it worth it. But, I am just cautious that the exercise may feel a bit like criticism. So do you like us to suggest minor changes so that you can go ahead and just do the experiment, or are you enjoying this "dry run" exercise and feel that any suggestion is worth it - for this or otherwise future experiments?

 

[Yugang]

The experiment aims amongst others to differentiate between various uptake mechanisms for inorganic carbon - CO2 from water, soil, or HCO3. From literature we learn that there is a variety of strategies that plants use, here is a statistical analysis of 100 aquatic plants




The lower median capability of freshwater macrophytes to exploit the reserves of inorganic carbon is consistent with their alternative strategies for acquiring inorganic carbon described above. Thus, only 50 species of the 110 tested (45%) are able to use HCO3− (Fig. 5A). Four species perform C4 photosynthesis and nine species perform CAM based on diel acidity changes. All the species with C4 also use HCO3−, while only three species, Ottelia alismoides (Zhang et al., 2014) and possibly Scirpus subterminalis (Beer and Wetzel, 1981) and Vallisneria spiralis (Yin et al., 2017), combine both CAM and HCO3− use. Ottelia alismoides appears, uniquely, to combine three different types of CCM, HCO3− use, C4, and CAM (Shao et al., 2017). Fifty-two species have no apparent CCM. Of these, 69% have access to CO2 from the atmosphere or sediment, or grow in environments where CO2 is likely to be high locally (Fig. 5B). In contrast, only ~24% of species with an ability to use HCO3− have alternative strategies, and these largely comprise C4 metabolism and access to atmospheric CO2.

So statistically we expect about 50% of our tested aquarium plants to do well in a tank with access to sufficient HCO3 (Maybe less, my personal opinion, as the tropics have waterways with more CO2 than we would see in cooler climates and plants may have evolved accordingly with a focus on less costly CO2 capture). They will only use HCO3 if CO2 supply is really low, otherwise prefer and be more efficient using CO2. Plants that have the capability to use HCO3 are generally a bit less efficient at CO2 capture than plants specialised in CO2 only.

Obviously sorting out which commonly used aquarium plants can use HCO3 is a substantial project, but may not be necessary as we may assume that they will do well when given sufficient CO2 either via injection or microbial respiration in the substrate.

Above data are from the paper attached to Resource - Fresh water biology - useful articles and science references

 

[Marcel G]

I like to check with you how you feel about moving this thread forward. ... I am just cautious that the exercise may feel a bit like criticism. So (1) do you like us to suggest minor changes so that you can go ahead and just do the experiment, or (2) are you enjoying this "dry run" exercise and feel that any suggestion is worth it - for this or otherwise future experiments?

I'm afraid I don't see much difference in the options. To me, a discussion is a conversation in which one side formulates an opinion and attempts to explain how it arrived at that opinion (ideally using logical arguments). The other side then asks follow-up questions if they don't understand the opinion presented. And if he/she thinks he understands the opinion but disagrees with it, he will present a counter-argument, again with appropriate (logical) reasons. Both debaters are respectful of each other and welcome any counter-arguments, because their goal is to achieve the truth (i.e., the correct view of the matter or the actual resolution of the problem). Criticism is seen here as a "healing tool" that helps the person to "correct" an incorrect view that does not lead to knowledge of the truth. Such a discussion is called "fruitful" (useful). If it is just presenting different opinions without proper [rational] justification, and the aim is not to arrive at the truth but to advance one's own opinion, then it is not in the true sense of the word discussion, but just idle chatter or shouting.

In light of the above, I can safely say that any criticism is warmly welcomed, because my goal is to know what parameters are crucial for good growth of different groups of aquarium plants. My "opinion" is just my [admittedly imperfect] attempt to identify these parameters. However, I am under no illusion that it is correct. On the contrary, I sincerely fear that it is not correct, and that by using it in practice I will waste another three months (and money) without finding what I am looking for. So I will be very grateful for any other "opinion" that convinces me [with better arguments] of the greater weight of other parameters/factors. That is, after all, the reason why I am here (i.e. why I registered here and raised my question) ... it is not to make myself visible and achieve fame, but to solve a problem that is bothering me.

 

[Marcel G]

The experiment aims amongst others to differentiate between various uptake mechanisms for inorganic carbon (DIC) - CO2 from water, soil, or HCO3. From literature we learn that there is a variety of strategies that plants use ... Sorting out which commonly used aquarium plants can use HCO3 is a substantial project, but may not be necessary as we may assume that they will do well when given sufficient CO2 either via injection or microbial respiration in the substrate.

I'll take the liberty of modifying it a bit: The goal of the experiment is not to distinguish between different mechanisms of DIC uptake, but to identify key parameters without which different plant species cannot grow well. So only if one of these parameters is DIC, we will need to devise a procedure to distinguish between the different mechanisms of its uptake. I, of course, believe that DIC is on of the key parameter (and various results of scientific studies and our practical experience suggest this), but I could be wrong.

Thus, we can formulate the hypothesis that while some plant species (i.e. "strict CO2 users") require a minimum concentration of CO2 to grow well because they cannot obtain carbon in other ways, other species can and thus are not dependent on CO2 (if they have various alternative carbon sources available).

The problem is that we don't know how high that "minimum concentration of CO2" should be, and we also don't know what all the "alternative carbon sources" might be in play. This would be a good thing to estimate somehow, because I have to use some specific values or sources in the experiment. The table I posted in the "useful sources" thread suggests that we should have at least 5-8 ppm CO2 in an aquarium recipe that targets "strict CO2 users" (with no other carbon sources). In the opposite aquarium with a recipe that will be aimed at "HCO3 users", then logically we should have a lower CO2 concentration (ideally corresponding to natural equilibrium values) and we should use HCO3 as a carbon source there (again, the question is at what concentration). You already pointed this out to me and I suggested using 52 ppm HCO3 (2.4°dKH). Next, we could try to test the ability of plants to use CO2 from sediment. We could test this in a third aquarium, where we could again have a naturally low CO2 concentration in the water + organic sediment to serve as a potential CO2 source. So with these three options we could have the parameter "DIC" (dissolved inorganic carbon) covered. Do you agree with this or did I miss something and do you have any comments?

And what about other key parameters (besides DIC)?

 

[Yugang]

In light of the above, I can safely say that any criticism is warmly welcomed, because my goal is to know what parameters are crucial for good growth of different groups of aquarium plants.

Thank you @Marcel G . I am struggling a bit what insights are expected to be gained from the experiment. My struggle does not necessarily mean that the design of experiment is suboptimal, but it is more likely that I mistunder stood, or still misunderstand, what information you hope to extract.

The main goal is to find (= identify) a specific set of parameters that will ensure a good plant growth and can be easily replicated.

Without doing any experiment, my answer here is that plants need water, a source of inorganic carbon, macros and micros, light. This in itself is obvious, and an experiment to verify this would probably be trivial, so I am assuming that you want to dig one lever deeper.

Looking at your 8 tanks, each of them with a bunch of plant species the focus seems to be

• Organic substrate, versus no substrate at all
• micros and micros originate from the water column versus from a substrate, or both.
• Source of inorganic carbon is CO2, or bicarbonates, or both

It seems that at least to some extent all essentials are in place in tanks 1-7, where only tank 8 seems to lack any sufficient source of organic carbon. You do not mention dosing micros in the tanks that have no substrate, but I am assuming that this is not intentional.

Now the question is if the goal of the experiment is met when we see some plants successful in al tanks, except tank 8 perhaps.
I am assuming that your aim is to learn more than this, you want to dig deeper and have more valuable information from the experiment, but then I struggle to precisely understand what it is, what will be the observables and how would you analyse the observations into some insights?

Besides, I would have a few other thoughts
- Especially for the tanks with organic substrate would a 2-3 weeks experiment be sufficient for microorganisms to build and support CO2 through respiration? Also, could plants still harvest from their pre-test reservoir and have not fully acclimatised to the test conditions?
- We expect approximately half of the aquatic plants to have the ability to use bicarbonates, the other half needs at least some CO2. All plants prefer CO2 if it is available, over bicarbonates As you aim for general recipes for tanks, can't we dismiss the recipes with no inorganic carbon or only bicarbonates right away?
- We know that even the most experienced aquarists need their tank to cycle, then often see diatoms, cyano and several algae before the tank matures and the plants show their best. Is there a risk that a short duration experiment will not differentiate between good and suboptimal test parameters?

So in summary @Marcel G I am still struggling a bit understanding the objectives (assuming these are not trivial), and how would observations be evaluated to valuable new insights. It could be a nice exercise trying to make a prediction what will happen in the experiment, and work out how to analyse if the experiments shows what is expected, or rather some unexpected observations. When I do that for this experiment, I have the feeling that it makes sense to narrow it down to a more focussed objective, limit the scope.

 

[GreggZ]

I know this is probably a touchy subject here, but I think your question is misleading. In fact, its meaning is similar to asking if plants can be "caused" by low/high nutrient levels. Plants will grow whenever they have adequate (1) light, (2) heat, (3) water and (4) nutrients, and an (5) environment without an excessive amount of disturbance/toxicity (simplistically speaking). Similarly, algae will grow whenever adequate light, heat, water and nutrients are available, and an environment without an excessive amount of disturbance/toxicity. Most of my experiments were designed with different conditions in each aquarium to see which plants would prefer which conditions. It is therefore logical in principle that if a plant would thrive in one set of conditions, it would probably not thrive in the others (where it will naturally suffer). Poor plant growth is therefore hardly avoidable in experiments designed in this way. The algae will simply grow there because they have everything they need. The only way to limit (or prevent) this is to use those "disturbing/toxic influences" => limiting the intensity of lighting, applying downright toxic substances, moving some nutrients from the water to the substrate, introducing enough algae eaters, changing pH, regular maintenance etc.

Bear with me as I am trying to understand your point of view. In your comments above have you concluded that plant health has no relation to algae? That is given enough light, heat, water, and nutrients that algae is unavoidable? Regardless of plant health? Forgive me if I misunderstood your comments.

 

[Marcel G]

I am struggling a bit what insights are expected to be gained from the experiment. My struggle does not necessarily mean that the design of experiment is suboptimal, but it is more likely that I mistunder stood, or still misunderstand, what information you hope to extract. Without doing any experiment, my answer here is that plants need water, a source of inorganic carbon, macros and micros, light. This in itself is obvious, and an experiment to verify this would probably be trivial ...

My understanding is that some plants just don't grow well even if we provide them with all the "basic/obvious things" you mentioned: water, light, heat and all the essential nutrients (= carbon + mineral macro and micro elements). So I would like to use my experiments to find out why this is so, and what we are missing in the list of those "basic things", or what other influences can cause the bad result. That's it.

CONTENT vs FORM

I think of it as follows (and I try to take into account our experience and previous scientific discoveries): For plants it is obviously not only the CONTENT (= water, light, heat and nutrients) that is important, but also the FORM we provide them with that content, and also the various physico-chemical PROCESSES or PROPERTIES of the environment we often overlook.

To give you an example: Iron (Fe) can be supplied to plants either in the form of inorganic salts (e.g. FeSO4 or FeCl3), or in the form of organic/synthetic chelates (Fe-citrate, Fe-gluconate, Fe-EDTA, Fe-DTPA, etc.), or in the form of organic sediment, so to speak. Our experience and some scientific experiments show that at higher pH and higher redox iron oxidises and precipitates, making it unavailable to plants. This can be solved by using suitable chelates. Furthermore, scientific experiments show that there is a strongly reducing environment in the sediment due to the low redox, which reduces and dissolves many of the oxidized substances, so that plants can be threatened in turn by their toxicity (this applies most often to iron and manganese). From what I've read so far, I'm definitely convinced that this also applies to our aquarium substrates (including granular ones like 'ADA Amazonia' and the like). The idea that oxygenated water flows in our substrates is, in my opinion, a misconception (contradicting the laws of physics). At most, the first 5 mm of the substrate can be oxygenated. Soon after the substrate is flooded (within a few hours at the latest), the bacteria will have drained all the oxygen from the substrate. But now I'm getting sidetracked again.

CONTENT SHAPERS

So suddenly we have other factors (such as pH and redox) that can modify or SHAPE the CONTENT quite dramatically. I've given these factors the working title of "content shapers", and in addition to the aforementioned pH and redox, I include the microflora here (which affects a whole host of properties [not just] of the sediment).

Could any of these be the reason why, for example, Ammannia pedicellata 'Gold' is not growing well? I have no idea. But it is possible. So, from my point of view, this is one of the relevant possibilities (= factors that could have a decisive influence on the success or failure in growing some 'sensitive/challenging' plants).

BASIC THINGS + the X factors

So, on the one hand we have the "basic things" (= water, light, heat and nutrients), on the other hand we have various "other factors" (= pH, redox, microflora ...) that can make life really difficult for plants [especially with nutrients] ... if they are somehow out of the right shape. But what is this "right shape" from the point of view of various "sensitive/problematic" plants?

I don't have much clarity on this yet. I don't know which of those factors play a key role and which play a marginal role. I don't know whether pH or bicarbonate content (the amount of which determines pH in natural waters), or both, is important for plants. I don't know if a strongly reducing environment in the sediment can be a problem for our plants (because of the threat of heavy metal toxicity), or if they can always cope with it without problems. I don't know if we can provide plants with a perfect biodiversity of microorganisms in the sediment, or a well-functioning redox cascade ... or if any of that matters at all. I'm just thinking about it [out loud] and gathering input so that I can design some experiment based on it that might help us move a little closer to successfully growing ALL plants (even the "problematic" ones). Plants that we don't usually have any problems with probably don't have problems with any of this. But we all know that there are plants that have problems with at least some of it. But we still don't know EXACTLY what bothers them. So that's the goal of my experiments => to try to find out, or identify those factors that we MUST have there on the one hand (that plants need), but also factors that we MUST NOT have there on the other hand (that bother some plants extremely).

But FIRST, however, we must try to identify these factors. Only then, can we discuss the most appropriate way to test them. If you would be willing to help me with finding these potential factors (with your ideas and thoughts), I would be very happy.

I'm not good at expressing my thoughts "succinctly and clearly", so I apologize for my often wordy posts. Hopefully I have managed to express myself clearly enough this time.

 

[Marcel G]

In your comments above have you concluded that plant health has no relation to algae? That is given enough light, heat, water, and nutrients that algae is unavoidable? Regardless of plant health? ...

Yeah, sort of. Please try to find some recipes (culture media) on the internet for growing algae. You will find that scientists routinely grow and study algae in laboratories. There is absolutely nothing mysterious about it. This fact alone implies that if you give algae what it needs (while making sure that no negative influences interfere with its growth), then the result will be beautifully overgrown algae, plain and simple. This is of course also true in our aquariums. But there, as I have already mentioned, there are a number of those "negative influences" that inhibit their growth and reset their environment. Of course, one of those "negative influences" for the algae are the plants (which deprive them of light and nutrients, maybe some of them also release some allelochemicals into the water, plus healthy and fast enough growing plants don't provide a very good place for the algae to attach). Even though the plants are a "positive factor" for us, they are a "negative factor" for the algae. Of course, more algae will take hold on poorly growing plants than on well-grown ones. But I don't think you can draw an equation between "good growing plants" and "no algae". Such a view is highly simplistic and misleading, because it does not take into account the other "negative influences" at all. I had plenty of healthy and fast-growing plants in my previous experiments, and yet the aquarium was full of algae by the end of the experiment (even though the plants themselves were sometimes not so affected). Do you find this argument convincing or do you object to it? Feel free to make a counter-argument.

 

[Marcel G]

Looking at your 8 tanks, each of them with a bunch of plant species the focus seems to be

• Organic substrate, versus no substrate at all
• micros and micros originate from the water column versus from a substrate, or both.
• Source of inorganic carbon is CO2, or bicarbonates, or both

It seems that at least to some extent all essentials are in place in tanks 1-7, where only tank 8 seems to lack any sufficient source of organic carbon. You do not mention dosing micros in the tanks that have no substrate, but I am assuming that this is not intentional.

Now the question is if the goal of the experiment is met when we see some plants successful in al tanks, except tank 8 perhaps.
I am assuming that your aim is to learn more than this, you want to dig deeper and have more valuable information from the experiment, but then I struggle to precisely understand what it is, what will be the observables and how would you analyse the observations into some insights?

Try to think of "organic substrate" or "water column" as the FORM and the nutrients themselves as the CONTENT. Plants need the CONTENT. That is not my concern, because we all know that (so there is no need to test it). What I am interested in is which of those plants can take up that CONTENT in the FORM of "nutrients dissolved in the water column" and which, on the other hand, require the "substrate form" of nutrients. So I'm not testing whether plants grow better with nutrients in water or in a substrate, but what those plants NEED or REQUIRE to grow well. For example, if I find that Rotala wallichii will grow well in water without any substrate, then I will know that this plant does NOT NEED substrate to grow well (although it may grow a little better in substrate). On the other hand, Ammannia pedicellata 'Gold' has never grown well [in my tanks] in just water without organic substrate. So maybe this plant NEEDS/REQUIRES organic substrate to grow well. (PS: I'm just speculating now.) Got it? Then it may be similar with carbon (and other factors that still need to be identified). I know that Rotala wallichii does NOT NEED extra CO2 to grow well (it can easily get by with a naturally low concentration of CO2). On the other hand, all the other plants I have grown recently without CO2 have not grown well. So maybe these plants NEED some extra CO2 to grow well. Maybe the CO2 in the organic substrate is enough for them, but what they have available in naturally low amounts in the water column is probably not enough (unless the problem was something else => some other factor, like too low nutrient concentration, or inappropriate form of microelements, or wrong ratio of some nutrients, etc.).

PS: The goal of my experiment will be fulfilled if at least in one aquarium the test plant grows well. Then I have figured out what this plant NEEDS. It needs what it has in that aquarium. If it grows well in multiple aquariums, then that means it NEEDS what it has in one or the other aquarium (in other words, it can be successfully grown in multiple conditions). If it grows well everywhere, then that means that NONE of those parameters are important to it, and that it has NO requirements for a particular FORM (i.e., you can just supply it with the essential content in any form).

Besides, I would have a few other thoughts
- Especially for the tanks with organic substrate would a 2-3 weeks experiment be sufficient for microorganisms to build and support CO2 through respiration? Also, could plants still harvest from their pre-test reservoir and have not fully acclimatised to the test conditions?
- We expect approximately half of the aquatic plants to have the ability to use bicarbonates, the other half needs at least some CO2. All plants prefer CO2 if it is available, over bicarbonates As you aim for general recipes for tanks, can't we dismiss the recipes with no inorganic carbon or only bicarbonates right away?
- We know that even the most experienced aquarists need their tank to cycle, then often see diatoms, cyano and several algae before the tank matures and the plants show their best. Is there a risk that a short duration experiment will not differentiate between good and suboptimal test parameters?

So in summary @Marcel G I am still struggling a bit understanding the objectives (assuming these are not trivial), and how would observations be evaluated to valuable new insights. It could be a nice exercise trying to make a prediction what will happen in the experiment, and work out how to analyse if the experiments shows what is expected, or rather some unexpected observations. When I do that for this experiment, I have the feeling that it makes sense to narrow it down to a more focussed objective, limit the scope.

(1) If I mentioned somewhere that the length of the experiment will be 2-3 weeks, I was wrong. It will be 2-3 months. That will hopefully answer your other question about the nutrient reserve ("nutritional status") they bring with them. I think that within a month or so, all the plants will have exhausted their supply (in extra CO2 tanks this should happen even earlier). And in a month, I'm sure enough bacteria will have multiplied.

(2) For example, Rotala wallichii grows well for me in aquariums without extra CO2 and without any bicarbonate (KH/HCO3=0). If I exclude this option from the experiment, then I won't find out what plants can get by with just the natural concentration of carbon (CO2) in the water column without needing any "extra" from the organic substrate. Moreover, this can also distinguish plants that need some non-zero bicarbonate in the water to grow well. As I mentioned in another thread, "HCO3 users" probably require about 20 ppm of HCO3 if they are growing in an environment where only naturally low concentrations of CO2 are available. Plain and simple: Plant "A" may be accustomed to growing in CO2-poor water and nutrient-poor sandy substrate, and these conditions are sufficient for it to grow well (e.g. low-demand "strict CO2 users" like Rotala wallichii). Plant "B", however, may need slightly higher CO2 concentrations, while not caring about bicarbonate content (e.g. high-demand "strict CO2 users"). Plant "C" may be used to bicarbonate, but if we give it carbon in the form of extra CO2, it will be fine (e.g. "HCO3 users"); but if we don't give it any extra carbon, it will either make do with what little CO2 it naturally has (e.g. "low-demand HCO3 users"), or we will find that it can't make do and that it needs at least some bicarbonate (e.g. "high-demand HCO3 users"). And so on. [Don't take these options as exhaustive or necessarily correct; I wrote haphazardly as they occurred to me.] So we should account for all the possibilities that might tell us something important about our plants.

(3) Yes, the risk is there. But on the other hand, scientists do this quite routinely and the plants grow well for them (if they provide the right content in the right form). But they [unlike us] know what optimal parameters their plants need, because they test plants that grow under their noses even in a nearby lake or river. So they always have one aquarium as a "control" in which they simulate the same conditions as the plants have in nature. But we don't know what optimal parameters our plants need. We are still looking for these optimum parameters. So I would like to design the experiment so that there are as many different combinations of the key factors as possible... in the hope that at least one of those combinations will turn out to be the right one (for a particular plant species).

But maybe that goal is too ambitious. Still, you can cover quite a few combinations with eight aquariums:

 

[Yugang]

But maybe that goal is too ambitious.

I am happy to follow your experiment, but worry that this may be the case. Also I do not see yet how the experiment design leads to your goals, but look forward seeing the progresses over the next months. There may be a trade off between level of ambition and chances of success.

I believe the best chances of learning and success is by following some of the best scapers who keep multiple species successfully, and scientific papers that have been peer reviewed. Besides the nutrients and inorganic carbon already mentioned they would mention KH, GH, pH, stability, low organic waste as well as tank maturity. Stability applies to all parameters in the tank, as the assumption is that plants can adapt, but the less variation there is the lower the cost of adapting the plants machinery. Low organic waste seems especially relevant to disadvantage algae in the power balance with plants in the ecosystem. And then tank maturity is only partially understood, but refers probably to the huge field of micro-organisms and competition by chemicals and biological agents.

You mention R. Wallichii, I can grow that quite well and use it as an indicator plant for CO2 stability. It seems to adapt to multiple environments, I play a lot with different CO2 levels, but it hates change and will show that by stunting.

If I had 8 tanks for experiments I would use them to verify, and perhaps debunk, some of the assumptions that we seem to take for granted in the hobby.

• I am currently mostly interested to explore when lowering CO2 levels (like 15 ppm and below) start to show deterioration as compared to the more popular 30 ppm for high tech. So setting 80 tanks from 5 ppm up to 40 ppm in steps of 5 and testing a bunch of plants would be great.
• There is also the eternal debate on lean dosing versus EI dosing, so also for that a controlled test environment could have a huge benefit for our hobby. Is toxicity real, and can we demonstrate it?
• I am also very interested if there is any benefit of root dosing, versus providing all micros and macros via the water column. Such experiment could be combined with comparing organic substrate, high CEC substrate, as well as inert substrates.
• A lot of good experimental work could be done on understanding algae, and how plant health and algae interact in the ecosystem. Some claim they understand BBA, so we could set up controlled experiments to turn BBA on and off, following their understanding of the relevant levers. Same for several other types of algae, and how they depend on plant health and/or waste organics.

 

[Marcel G]

If I had 8 tanks for experiments I would use them to verify, and perhaps debunk, some of the assumptions that we seem to take for granted in the hobby.

• I am currently mostly interested to explore when lowering CO2 levels (like 15 ppm and below) start to show deterioration as compared to the more popular 30 ppm for high tech. So setting 80 tanks from 5 ppm up to 40 ppm in steps of 5 and testing a bunch of plants would be great.
• There is also the eternal debate on lean dosing versus EI dosing, so also for that a controlled test environment could have a huge benefit for our hobby. Is toxicity real, and can we demonstrate it?
• I am also very interested if there is any benefit of root dosing, versus providing all micros and macros via the water column. Such experiment could be combined with comparing organic substrate, high CEC substrate, as well as inert substrates.
• A lot of good experimental work could be done on understanding algae, and how plant health and algae interact in the ecosystem. Some claim they understand BBA, so we could set up controlled experiments to turn BBA on and off, following their understanding of the relevant levers. Same for several other types of algae, and how they depend on plant health and/or waste organics.

These all seem like good suggestions, but from my point of view they are hampered by one huge obstacle: the absence of knowledge of the optimal parameters of the plants being tested.

I will give again an illustrative example:

I used to follow a so-called growth curve where I tested different nutrient concentrations in five different aquariums. For example, I had 2 ppm NO3 in the first aquarium, 4 ppm in the second, 8 ppm in the third, 16 ppm in the fourth and 32 ppm in the fifth. For some plants the curve came out perfectly...


...while for others it didn't come out at all:

The reason is simple:

I did not use optimal water/sediment parameters in these experiments. Thus, while the chosen water parameters were problem-free for Pogostemon, Rotala was severely underperforming, resulting in poor growth ... regardless of the chosen nutrient concentrations. In other words, the fact that I was growing this plant in sub-optimal conditions meant that the experiment to model its growth curve was doomed to failure.

My point is that if we can't grow each plant species in optimal (or at least suitable) conditions, then there is no point in testing anything. How can I test for optimal CO2 concentrations if I prepare water with carbonates, for example, which Rotala wallichii doesn't like (or doesn't like under some conditions)? Or, conversely, if I prepare water with no carbonates and Hygrophila corymbosa, which needs them, dies? Do you understand? Unless I know what each plant needs (let's call it "vital parameters"), I'm not likely to prepare water (or "testing environment") that's suitable for any further experiments.

I have already experienced so many botched experiments (due to not knowing the vital environmental parameters) that I don't want to go into any more [without knowing what these vital parameters for different groups of plants are]. If you help me find these vital parameters, then you can come up with any test you want, and I'll run it for you (assuming my current equipment is up to the task). PS: This offer applies to everyone.

 

[Yugang]

My point is that if we can't grow each plant species in optimal (or at least suitable) conditions, then there is no point in testing anything.

So let me ask, from the 100 most common plant species in our hobby, how many would not grow well in reverse osmosis water, 4 GH, 2 dKH, EI water column dosing, 30 ppm CO2, 24 C, and 100 PAR light? I believe less than 5%, if any. This is not to say that this is the only correct solution for the planted tank, but surely it is a safe bet, wouldn't you agree?
From here there can be fine-tuning, but I am not sure if the argument holds that we don't know well enough how to grow plants sucessfully as a starting point for further exploration.

 

[Marcel G]

So let me ask, from the 100 most common plant species in our hobby, how many would not grow well in reverse osmosis water, 4 GH, 2 dKH, EI water column dosing, 30 ppm CO2, 24 C, and 100 PAR light? I believe less than 5%, if any. This is not to say that this is the only correct solution for the planted tank, but surely it is a safe bet, wouldn't you agree?
From here there can be fine-tuning, but I am not sure if the argument holds that we don't know well enough how to grow plants sucessfully as a starting point for further exploration.

I apologize, but I give up further attempts to clarify my intent. With my limited powers of expression, I don't think I'm capable of it.