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HOW TO: Clean, easy and highly nutritious greenwater culture for Daphnia and Moina.

louis_last

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These are the best videos I could get that give you a clear sense of scale and the relative sizes of the three species I believe to be Moina macrocopa, the smaller Moina micrura and Ceriodaphnia dubia.
 

Wookii

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So I've changed my lights over to a 32W aquarium LED light (a Fluval Plant 3.0 I had spare in the garage). I can't say whether its increased production any, but the Moina seem to hang higher in the water column. I've also increased the bubble rate from the airpump a little and the additional circulation seems to assist in keeping the yeast mixture suspended.

I think its fair to say the production rate isn't very consistent though. I've not noticed a significant 'crash', in population, but increases in population after removing a batch for feeding seem to be taking longer to bounce back.

I did a large (80% ish) water change at the weekend, and that seemed to create a sudden jump in the population in the couple of days afterwards. I'm hoping that once I start culturing the Chlorella @louis_last from your sample, I might be able to keep the population more stable, as I'm wondering if the build up of detritus from the yeast/dried chlorella overtime is lowering survival rates. Hopefully with the live Chlorella I won't get the same contamination of the water between water changes.

A couple of questions:

1. I'm currently still using RO water (with a little potassium carbonate) for the water changes, as I did when hatching the eggs. Do you use tap water in yours @louis_last? If so, how hard is it? I'd like to try transitioning to tap water, as it would be much more convenient, but my tap water is fairly hard - and I can't find any references to whether Moina will continue to produce in hard water.

2. Similarly with the Chlorella, do you use tap water for that?

3. Also on the Chlorella - I have bought a bottle of the BioBizz fish mix stuff (which stinks incidentally lol) you have linked to in the first post, but have you ever tried fertilising it with inorganic dry salts? Is there a specific reason for going with the BioBizz organic fertilizer?
 

louis_last

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So I've changed my lights over to a 32W aquarium LED light (a Fluval Plant 3.0 I had spare in the garage). I can't say whether its increased production any, but the Moina seem to hang higher in the water column. I've also increased the bubble rate from the airpump a little and the additional circulation seems to assist in keeping the yeast mixture suspended.

I think its fair to say the production rate isn't very consistent though. I've not noticed a significant 'crash', in population, but increases in population after removing a batch for feeding seem to be taking longer to bounce back.

I did a large (80% ish) water change at the weekend, and that seemed to create a sudden jump in the population in the couple of days afterwards. I'm hoping that once I start culturing the Chlorella @louis_last from your sample, I might be able to keep the population more stable, as I'm wondering if the build up of detritus from the yeast/dried chlorella overtime is lowering survival rates. Hopefully with the live Chlorella I won't get the same contamination of the water between water changes.

A couple of questions:

1. I'm currently still using RO water (with a little potassium carbonate) for the water changes, as I did when hatching the eggs. Do you use tap water in yours @louis_last? If so, how hard is it? I'd like to try transitioning to tap water, as it would be much more convenient, but my tap water is fairly hard - and I can't find any references to whether Moina will continue to produce in hard water.
You should see the state of my large master culture, the entire bottom is covered in algae, snail poo, a few leaves and other detritus and I rarely ever siphon much of it off when I perform a water change - occasionally I take a leaf out and let my shrimp clean all the algae off. Once I've drained out about half the water I replace it with old water from my aquarium. My tap water is naturally very soft when it goes into the aquarium but I'm not sure what the water chemistry is like by the time it comes out, there's nothing in there that would make it particularly hard though. When I first set up the culture I added a tiny amount of crushed cuttlebone but mostly for the ramshorns that are in there. It may well be the case that a 'mature' culture with a lot of algae has a sort of buffering capacity not just in terms of harbouring infusoria as a food source but also in processing at least some of the waste produced by the moina and excess food breaking down.
For sustainable production I think the key is letting the population grow enough before you harvest and not being scared to let them reach quite a high population densities. I've also noticed sudden population explosions after water changes though and I think this might be because the water changes stimulate the moina to moult rather than it actually increasing their reproductive rate. The embryos are released from their brood pouch when they moult.
I think you might want to consider adding more snails, the ceriodaphnia dubia that I've sent you kept crashing for me until I added snails to their culture.
2. Similarly with the Chlorella, do you use tap water for that?
I've been using cheap bottled water from coop. Now I'm using 5L bottles. I don't think there would be any problem using tap water but if you use prime or any other dechlorinator I'd also let it stand for a day or two because I think these could potentially interact with the fertilisers in some way.
3. Also on the Chlorella - I have bought a bottle of the BioBizz fish mix stuff (which stinks incidentally lol) you have linked to in the first post, but have you ever tried fertilising it with inorganic dry salts? Is there a specific reason for going with the BioBizz organic fertilizer?
I haven't tried using dry salts although there are some videos on youtube with recipes for exactly that, what you'll find is that some of them also recommend adding molasses which is present in the fishmix. This is because the chlorella can use the sugars present as an energy source as well as light and in theory allows it to actively grow during periods of darkness, I understand this is called a 'mixotrophic' culture. You can also add glucose powder and in theory even grow Chlorella in darkness although it won't be green.
I only recommend the fishmix because it's easy, readily available and cheap relative to any fertiliser I could find marketed specifically for phytoplankton but it may very well not be the best solution and it's well worth experimenting. It also has close to the 6-1 N-P ratio that I've read is best for Chlorella - the fish mix is 5 -1 N- P.
I add half a teaspoon of the fishmix to 2L of water, you'll find that once the bottle has turned green if you add another half teaspoon and wait a little longer it will turn even more green. You could probably just make a more concentrated solution right off the bat but I don't know at what concentration the solution would become too rich for the chlorella. It seems quite obvious when they've used up all the nutrients from the fishmix as there's no trace of the heavy brown colour that it gives the water remaining.
I started my first two bottles with approximately 15ml of very dilute Chlorella culture and I've sent you guys each a pretty concentrated amount from a very mature culture in 330ml bottles so you should have enough to experiment a bit. I'm not remotely emotionally invested in my technique so I would be thrilled if anyone can improve on it. You may get much faster growth with dry salts.

I don't know why we seem to be having different experiences when it comes to over/under feeding the cultures but so far my experience has been that the ceriodaphnia dubia I've sent you are even easier to overfeed than the Moina, even feeding with chlorella. I actually found a paper backing this up too where they found a negative correlation between their reproductive rate and the concentration of food in the water but they didn't offer any explanation why as it was beyond the scope of their research. Snails and daily very light feeding seems to be the key for these guys. They're so small that I've still got them going in several 32oz plastic cups and I feed them appoximately 6ml of chlorella every other day which barely clouds the water. I'm still getting to grips with the best protocol for C. dubia as they behave somewhat unpredictably - it's easy to crash the cultures but on the other hand I originally isolated them from a moina culture that crashed, all the moina had died but I noticed significant numbers of the even smaller C. dubia still apparently thriving.
 

dw1305

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Wookii

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Thanks Louis, thats all good info. I only asked on the dry salts, as I have pretty much a lifetime supply in the garage bought for the main tank EI plant fert dosing. I didn't realise that the BioBizz contained molasses, though that does explain the dark brown colouration. Once I have the Chlorella culture up and running from the sample you're sending me, I'll try to see if I can make a suitable fert from dry salts.

In terms of snails in the tank, I do have about 4 x decent sized Ramshorns, and 6-7 Bladder snails for the 20 litres. I've recently seen a load of new babies - as is often the case with snails - so I'll have many more shortly. Does that sound sufficient? To be fair the snails appear to generate more waste than they actually clean up. Would I be better leaving all this in the base of the tank then, rather than hoovering it up at the water change?

I agree on the culture population, I'm probably harvesting too frequently. That paper I linked to previously suggests:

Moina cultures, however, routinely reach densities of 19,000 individuals per gallon (5,000/L) and are, therefore, better adapted for intensive culture.

So the maximum would be 100,000 individuals in my 20 litre tank :oops: . . .whilst I don't think I'll ever see that many, I'd wager I'm probably at less than 2,000 - its impossible to count of course, so it could be even lower. So I should perhaps see how high I can get the population before regularly harvesting. I couldn't imagine what, say, 50,000 individuals would look like - it'd be solid with them.

Its difficult to say on the feeding. I'm not sure if I'm feeding too much, or if its having a negative effect such as a reduction in lifespan of the older Moina - I may well be as I always seem to have hundreds more babies than I do adults. Hopefully the Chlorella will help with this, as presumably you can't overfeed the Chlorella - being live it wil presumably continue to grow in the Moina culture? Do you ever dose the BioBizz directly into the Moina culture?
 

Wookii

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Hi all,

It is usual to have snails in the culture <"to act as tank janitors">.

Hard is fine.

cheers Darrel

Thanks Darrel. I do have some snails, but they make more mess than they clear up.

Is RO/rainwater just required for the egg hatching then? I did wonder if the adult Moina might need minerals from the water column for development/moulting, that the RO water might not be providing - is that likely?

If I transition the culture from RO to tap water a litre every few days, I assume that will be fine?
 

Wookii

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You should see the state of my large master culture, the entire bottom is covered in algae, snail poo, a few leaves and other detritus and I rarely ever siphon much of it off when I perform a water change - occasionally I take a leaf out and let my shrimp clean all the algae off.

Post a picture of it up, it would be a useful reference. What leaves do you add?
 

dw1305

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Hi all,
Is RO/rainwater just required for the egg hatching then? I did wonder if the adult Moina might need minerals from the water column for development/moulting, that the RO water might not be providing - is that likely?
That would be my guess. I used to use a mix of tap water and rain water for mine.
To be fair the snails appear to generate more waste than they actually clean up. Would I be better leaving all this in the base of the tank then, rather than hoovering it up at the water change?
Yes, you can just leave it. It is all fairly inert after it has been through the snails. When you start a new culture I'd actually transfer a bit of mulm to the new vessel.

cheers Darrel
 

Wookii

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I found a paper here on culturing Chlorella using inorganic ferts:


The composition of inorganic fertilizers 3:3:1 produces the highest population density of 2,348 × 104 cells·mL-1 at day of 9.7. The composition of inorganic fertilizers 3:3:1 generates the highest growth rate of 25.9 %/day.

So they found a ratio of 3:3:1 (N : P : K presumably) produced the best results. They used urea, but I assume I could substitute with KNO3.

One other interesting point was that they quote the ideal temperature range for Chlorella growth as 26-31 degrees C. I assume you don't heat your culture bottle @louis_last?
 
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louis_last

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I found a paper here on culturing Chlorella using inorganic ferts:




So they found a ratio of 3:3:1 (N:p:K presumably) produced the best results. They used urea, but I assume I could substitute with KNO3.

One other interesting point was that they quote the ideal temperature range for Chlorella growth as 26-31 degrees C. I assume you don't heat your culture bottle @louis_last?
I don't heat mine at all, they stay at a pretty steady 22c. It would be really interesting to see a side by side comparison of two bottles with the fish mix and inorganic salts. I have some garden fert pellets that I think are about that ratio, I'm going to go dig them out and set up a side by side in two 5L bottles under LEDs right now. I got the 6-1 N-P ratio I mentioned from the parosphromenus project.
 

louis_last

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Post a picture of it up, it would be a useful reference. What leaves do you add?
20210325_181851.jpg

20210325_181906.jpg

A great source of algae covered leaves for shrimp or otocinclus. The clean ones in the first picture have recently been scoured by the shrimp and put back in the moina culture to ripen again.
I count 17 large mature ramshorns in here by the way so about 2 per litre as this is probably slightly under 10L.
 

dw1305

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Hi all,
So they found a ratio of 3:3:1 (N:p:K presumably) produced the best results.
I've had some <"emoticon smiley issues as well">. "3" : N will be nitrogen, the "3" : P maybe phosphorus (P) <"but it may be P2O5">, and the "1" might be K but it might be K2O.
They used urea, but I assume I could substitute with KNO3.
You would need another nitrogen source to get the 3:3:1 ratio, so either ammonium nitrate (NH4NO3) or urea (CO(NH₂)₂.). Personally I'd just go down the <"Miracle Gro"> route.

cheers Darrel
 

Wookii

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And a video to try and give you some sense of the population density it runs at. I've been harvesting a lot recently because I'm also feeding newt larvae as well as my fish.

This is mine - it’s really hard to film them on the phone, the detail isn’t good enough to pick up all the tiny babies in amongst all the dots that are actually visible, and the camera struggles to focus:

 
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louis_last

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This is mine - it’s really hard to film them on the phone, the detail isn’t good enough to pick up all the tiny babies in amongst all the dots that are actually visible, and the camera struggles to focus:

The bottom of the tank is getting there now! I'm really pretty sure you have a different species that is slightly smaller than what is in the master culture I've shown you here. I think the intermediate sized ones, between my M. macrocopa and the C. dubia, that I recently aquired and believe to be M. micrura, are what you've got here and what I sent to Miranda B with her Chlorella starter. I sent you the C. dubia but if at some point you want some of the larger species to mess around with I'll fire some at you too.
 

Wookii

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The bottom of the tank is getting there now! I'm really pretty sure you have a different species that is slightly smaller than what is in the master culture I've shown you here. I think the intermediate sized ones, between my M. macrocopa and the C. dubia, that I recently aquired and believe to be M. micrura, are what you've got here and what I sent to Miranda B with her Chlorella starter. I sent you the C. dubia but if at some point you want some of the larger species to mess around with I'll fire some at you too.

To be honest, the stuff on the bottom of the tank has been generated since Sunday! The Ramshorns turd like it’s going out of fashion! Lol

I think the species I have are large enough - if anything the adults are a touch too large for the nano fish I keep. I’ll send you some eggs of the ones I have, and you can hatch them and compare directly - I have loads of eggs left over.
 
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