Coral Coloration & Trace Element Experiment

[taricha]

Let's go a little deeper on coral color and how the fluorescent pigments I'm measuring relate to the appearance of these corals.

I'll go from the easiest to measure to the messiest.
First, Sarcophyton. This pigment is a booming yellow-green. Top is under normal tank lighting, and bottom is under royal blue ~450nm LED shot through a 90% yellow filter.

And here's what that pigment profile looks spectrally. The Blue data is using a fiber optic cable placed at the coral to measure the light "in situ" from the coral surface when lit by ~450nm royal blue. And the Red data is from my pigment extraction in water showing that the fluorescent peaks I'm measuring in the extract are indeed the important optical pigments to the color of the coral in a tank setting. (In all these charts, the vertical scales are unimportant, and data is just scaled for ease of visual comparison. )

The ~451nm peak is the reflected LED light, the 502nm peak is the dominant GFP, and chlorophyll peak is detectable around 675-680nm (chlorophyll shifts a bit when you extract it).

Here's the orange monti cap under normal tank lighting and ~450nm LED photographed with the yellow filter

Note how significant the pigment variation is from one spot to the next on these corals.


and here's the spectral view....

The bright orange pigment has a fluorescent peak around 574nm, and it's likely the chlorophyll fluorescent contributes to the overall red/orange look.

Next best-measured is monti digitata. There's a little bit more going on here. This coral is supposed to have green skin and red polyps. And although my coral is lacking almost all of that color pattern, under the royal blues with the yellow filter, you can see a patch or two where the skin is a bright green compared to the yellower color of the polyps.

Looking at the coral spectrally, the answer is a bit different. The blue data is collected over polyps where there's no green skin. The Green data was collected right over a patch of green skin (but polyp light is present there too.)

What you see from the green and blue spectral data lines is that the polyp and skin have the same GFP with a peak at 520nm - but the polyps have less of it and mixed with a lot of chlorophyll red fluorescence, while the skin would have more GFP and much less chlorophyll red.
The red data showing what's measured in the extract also shows the same GFP (and Chlorophyll.)
Also notice that the data from the extract is getting noisier compared to the first two corals - the detected fluorescence is getting smaller and harder to well-quantify.

The last two corals' GFPs are very noisy and hard to quantify due to low amounts of detected fluorescence.
This is the sinularia I got to replace an extremely bright gorgeous one. This one is has been dull with barely any detected fluorescence. Bottom pic shows the GFP isolated in the royal blue + yellow filter picture.

Here's the coral spectrally. The GFP is so close to the LED, what's shown is the overall measurement (yellow line), the LED by itself (black line), and then the subtraction to mathematically isolate the GFP (blue line).

The subtracted spectra shows that the fluorescent protein in this coral is around 480nm which sometimes gets categorized as a cyan fluorescent protein. The red data from the extract shows that I'm measuring the same fluorescent protein, just with a lot of noise due to a combined difficulty of exciting it and the low amounts of fluorescent protein.

And finally, poccilopora. This is another specimen that shows how strong the variation is in the amounts of fluorescent protein from one part of the coral to another. The GFP is localized not just to the polyps but to the very tip of each polyp tentacle.

And spectrally, you can see that the fluorescent peak around ~496nm is extremely difficult to measure in the extract (red data).

The good news is that if one of the experimental interventions causes this coral to grow a lot of green, then the effect will be very noticeable.

 

[Randy Holmes-Farley]

Nice baseline.

I'm curious about how apparent the most fluorescent signal here (the sarcophyton) is under white lighting. For example, is the polyp color you see under white light in your picture strongly dependent on fluorescence?

I. e., how it looks to you under white light with and without the deep blue end of lighting?

 

[taricha]

Nice baseline.

I'm curious about how apparent the most fluorescent signal here (the sarcophyton) is under white lighting. For example, is the polyp color you see under white light in your picture strongly dependent on fluorescence?

I. e., how it looks to you under white light with and without the deep blue end of lighting?

If you look at the chart for the sarcophyton, you'll notice that the Green emission is actually higher than the reflected blue light of the blue LED that drives it. It's not a measurement artifact, that sort of result has been repeatable in a few different ways of looking at this coral pigment. I had to think about if that was even possible. It is. In this context it means that it captures, converts, and releases more green photons than blue photons that bounce off. It might be converting over 50% of incoming blue photons to green ones, which is pretty insanely efficient (maybe - can't say what fraction is simply absorbed, but it's a pretty pale tissue overall).
So any "white" light will have enough blue in it for the fluorescent pigment to be a big effect on the color.
Here's what it looks like under sunlight +blues (top) and sunlight without blues (bottom)

 

[Randy Holmes-Farley]

Perfect, thanks.

 

[taricha]

Let's talk a little bit about what was done in Phase 1, and then I'll share the Phase 1 results for things that were measured in a subsequent post.

Phase 1: The algal feeding phase, I added enough daily phytofeast to be the recommended amount - it turns the water detectably green. It contains at least one strain - synechococcus - that is actually known to be ingested by hard corals commonly in the wild. The red algae is palmaria palmata and is munched by my foxface and urchins. The red algae has replaced 25% of the food protein input. The idea is that perhaps these algal sources are a good choice for traces as they are guaranteed to provide trace elements in biologically relevant amounts and if algal cells are directly ingested by corals, that might be superior to water trace dosing.

As above, I used phytofeast and for the Red algae I ended up using a mix of Two Little Fishies "red Sea veggies" (palmaria palmata) and omega one "Red Nori" (no species info).
The red algae is eaten by a fish, urchins, and some hermits - so remineralization and release of traces will depend to some extent on what is released (or not fully digested) by the animals. The phyto is a bit more interesting because it could be eaten directly by corals, and thus provide nutrients/traces directly to the target organism and bypass the water. The phytofeast also contains a little alginate as a thickener and 3 different organic acids as preservatives, so it has some carbon dose potential for growing bacteria that some corals may eat, or maybe not.
To me personally, this has a chance to be the most humorous result - if the phyto meets the coral nutritional/trace needs and increases coral color while leaving the trace elements in the water still undetectable/"depleted" on ICP-OES/MS. From a tank management perspective - it would be the opposite of the highly involved testing and dosing of a large number of elements - instead of testable water chemistry ... "just add some phyto".

Here's a very (very) rough analysis of what some element additions might look like from these Algal additions.
The last two columns are how much ppb per week is being added from the algae, and how much ppb was measured in my aquarium water (ICP-MS).
This is from a paper published on palmaria palmata element profile, and values are assumed to be inaccurate - they are merely here for a sense of the scale of these inputs.

The color coding in the last column reflects recommended targets for the elements. Red = too much in the water, Blue = not enough, Green = fine, yellow = recommendations from different sources disagree.

And here's the same table done for phytofeast, using published element profile of nannochloropsis - one of the ingredients as a representative for the rest of the microalgae in the product.

The amounts don't look very impressive, if one is trying to meet trace deficiencies in these elements. It's not going to meet the needs for Iodine, or Mn - the two elements in this list that are actually agreed as too low in my water.
They might meet needs for Fe and Zn, but probably not much else. (Cu too, but mine's already high)
The more interesting angle is that these values are divided by tank water volume, but if phyto is ingested by corals directly rather than uptake from the water, then it might be delivering these elements in much more beneficial concentrations to the corals.

So that's the logic behind phase 1. But of course it may not work at all . Results in a subsequent post.

 

[Dan_P]

Let's talk a little bit about what was done in Phase 1, and then I'll share the Phase 1 results for things that were measured in a subsequent post.

As above, I used phytofeast and for the Red algae I ended up using a mix of Two Little Fishies "red Sea veggies" (palmaria palmata) and omega one "Red Nori" (no species info).
The red algae is eaten by a fish, urchins, and some hermits - so remineralization and release of traces will depend to some extent on what is released (or not fully digested) by the animals. The phyto is a bit more interesting because it could be eaten directly by corals, and thus provide nutrients/traces directly to the target organism and bypass the water. The phytofeast also contains a little alginate as a thickener and 3 different organic acids as preservatives, so it has some carbon dose potential for growing bacteria that some corals may eat, or maybe not.
To me personally, this has a chance to be the most humorous result - if the phyto meets the coral nutritional/trace needs and increases coral color while leaving the trace elements in the water still undetectable/"depleted" on ICP-OES/MS. From a tank management perspective - it would be the opposite of the highly involved testing and dosing of a large number of elements - instead of testable water chemistry ... "just add some phyto".

Here's a very (very) rough analysis of what some element additions might look like from these Algal additions.
The last two columns are how much ppb per week is being added from the algae, and how much ppb was measured in my aquarium water (ICP-MS).
This is from a paper published on palmaria palmata element profile, and values are assumed to be inaccurate - they are merely here for a sense of the scale of these inputs.

The color coding in the last column reflects recommended targets for the elements. Red = too much in the water, Blue = not enough, Green = fine, yellow = recommendations from different sources disagree.

And here's the same table done for phytofeast, using published element profile of nannochloropsis - one of the ingredients as a representative for the rest of the microalgae in the product.

The amounts don't look very impressive, if one is trying to meet trace deficiencies in these elements. It's not going to meet the needs for Iodine, or Mn - the two elements in this list that are actually agreed as too low in my water.
They might meet needs for Fe and Zn, but probably not much else. (Cu too, but mine's already high)
The more interesting angle is that these values are divided by tank water volume, but if phyto is ingested by corals directly rather than uptake from the water, then it might be delivering these elements in much more beneficial concentrations to the corals.

So that's the logic behind phase 1. But of course it may not work at all . Results in a subsequent post.

Am I right in assuming that we should add the two columns of weekly added trace elements and compare to the pre-phase one ICP?

Will there be an ICP-MS of the water after weeks of dosing algae?

 

[taricha]

Am I right in assuming that we should add the two columns of weekly added trace elements and compare to the pre-phase one ICP?

Sure. That would represent a plausible total amount of traces added per week on a ug/L basis. How much of that is available for any particular organism to use is anyone's guess.

Will there be an ICP-MS of the water after weeks of dosing algae?

You betcha.