Help with experiment parameters discussion on DOC saltwater demand and microbial evaluation.

[Dan_P]

I totally see your points and I really appreciate you taking the time.

Unfortunately the experiment is not going the way most folks would think it would.

Taking that I’ve started this system around two weeks ago it is already showing signs of stability.

On the 10 of the October my parameters were as follow.

No3 5 mg/l
Po4 0.4 mg/l
Kh 8
Ph 8
O2 10 mg/

At that date my daily dose was

0.002 grams DOC
4 ml of nitrogen solution
0 ml of phosphates

Since then, dosage has been consistent.

Today the on the 16 of October

No3 5 mg/l
Po4 0.4 mg/l
Kh 9.5
Ph 8.2
O2 10 mg/l

At first observation, phosphate has been stable at 0.4 mg/l without any additions since the 10 of October. (Barely any phosphate has been used in the last 6 days) there was a exception on week one most likely due to colonisation of the surface area.

The daily consumption of nitrogen/nitrogen has been around 0.8 mg/l

There has not been any impact on oxygen the colour has darkened today so probably above 10 mg/l

Under the microscope, on the protein skimmer content, I can see that there is many different types of bacteria and Protozoa flourishing in the tank that leaves space for discussion if diversity is really better, so far I can’t understand the ph rise in 0.2 and the Kh rise in 1.5 as I don’t have any photosynthetic organisms in the system (no water changes have been performed). In addition the DOC used has had no impact on phosphate since the system has been stable or 6 days.

Hard to say what’s going on in such a short time and with little experimental detail. It could be something as simple as nothing is happening and there was a mistake made with the nitrogen addition. Here are additional ideas.

If the analytical measurements are not being made with a photometer, the results can have a large variation possibly making the comparison of results meaningless.

Since the measured concentration of the added reagents i mediately after addition is not reported, at least once, there is no way to confirm what is being added.

The hourly consumption rate is not reported. That could be useful information about the functioning of the system over time. Infrequent measurements mean we cannot tell the difference between a system consuming nutrients one hour v 24 hours. That tells us something about whether the system is growing or contracting.

Why was the phosphate dose stopped?

What reagents are being added for N, P, and C additions? What is the reagent concentration? How often are reagents prepared?

 

[sixty_reefer]

Hard to say what’s going on in such a short time and with little experimental detail. It could be something as simple as nothing is happening and there was a mistake made with the nitrogen addition. Here are additional ideas.

If the analytical measurements are not being made with a photometer, the results can have a large variation possibly making the comparison of results meaningless.

Since the measured concentration of the added reagents i mediately after addition is not reported, at least once, there is no way to confirm what is being added.

The hourly consumption rate is not reported. That could be useful information about the functioning of the system over time. Infrequent measurements mean we cannot tell the difference between a system consuming nutrients one hour v 24 hours. That tells us something about whether the system is growing or contracting.

Why was the phosphate dose stopped?

What reagents are being added for N, P, and C additions? What is the reagent concentration? How often are reagents prepared?

A lab environment, equipment, lots of free time and $$$$ would certainly help and explain some of what has happened much better.
The phosphate was stopped as this type of DOC doesn’t affect phosphate maybe It’s not available to all bacteria.

 

[Dan_P]

A lab environment, equipment, lots of free time and $$$$ would certainly help and explain some of what has happened much better.

What about my questions?

The phosphate was stopped as this type of DOC doesn’t affect phosphate maybe It’s not available to all bacteria.

Got it.

 

[sixty_reefer]

What about my questions?

I’ve missed that part, are you asking the concentration of the daily dose or the solution concentration?

At the beginning I was testing daily to adjust the balance, the last 6 days have been just every two days. The nitrogen dose is spread 12 hours apart and it seems that is all being converted by nitrifying bacteria to nitrate.
I should be able to do some hourly testing over the weekend if you believe it could be beneficial to the results.

This is a 4x view of the current Protozoa being harvested, taking that it’s still a young project.

 

[Dan_P]

I’ve missed that part, are you asking the concentration of the daily dose or the solution concentration?

What is the nitrogen compound being dosed?
What is the measured concentration in the aquarium immediately after dosing?
Howare you measuring the nitrogen compound in the aquarium?

Same three questions for phosphate and carbohydrate.

At the beginning I was testing daily to adjust the balance, the last 6 days have been just every two days. The nitrogen dose is spread 12 hours apart and it seems that is all being converted by nitrifying bacteria to nitrate.
I should be able to do some hourly testing over the weekend if you believe it could be beneficial to the results.

OK thanks..

This is a 4x view of the current Protozoa being harvested, taking that it’s still a young project.

This doesn’t look like protozoa. Did you send the correct link.

 

[sixty_reefer]

What is the nitrogen compound being dosed?
What is the measured concentration in the aquarium immediately after dosing?
Howare you measuring the nitrogen compound in the aquarium?

The nitrogen is a off the shelf product, it’s a mix of nitrogen ingredients that 10ml is equivalent to 1 ppm nitrate in 24 hours in a 100 litres. (Currently adding 4ml day in 50 litres)
The only way I’ve tested this was with a No2 test kit that showed 0.1 ppm after a couple hours from dosing 2ml it’s all transformer by morning before second dose.

The phosphate solution is a of the shelf product also, 1ml is equivalent to 0.01 ppm in 100 litres a few minutes after dosing. I have tested this before and is accurate. (Currently not adding phosphate as demand is minimal)

The Carbon is a carbohydrate extracted from algae I believe it’s a semi labile dissolved organic carbon, I can only have a educated guess of the strength of it although it stays from a few days to weeks in the water column. ( currently adding 0.002 grams day) In the video that I’ve shared, shows the carbon as the large still matter at 4x and 10x magnifications (new video)

Same three questions for phosphate and carbohydrate.



OK thanks..


This doesn’t look like protozoa. Did you send the correct link.

Yes, this is the correct video at 4 magnifications. The still matter is the carbon used in this test and I assume the Protozoa are the organisms moving faster in the video, there is also millions of smaller bacteria although they only start being visible from 10 magnifications ( I have a video of that if you like to have a look) my 40x is not working properly need to order a new one.
In addition everything on this videos it’s mainly in the water column not from the tank surfaces.

Edit:

10 magnification video


And 4 magnification

 

[sixty_reefer]

There would be a option here to send a sample to DNA testing and identify the particular strain of bacteria in the medium that is breaking down the carbon, but I’ve spent more than I wanted with this and identifying a certain species of bacteria wouldn’t really make a big difference imo.

 

[Dan_P]

The nitrogen is a off the shelf product, it’s a mix of nitrogen ingredients that 10ml is equivalent to 1 ppm nitrate in 24 hours in a 100 litres. (Currently adding 4ml day in 50 litres)
The only way I’ve tested this was with a No2 test kit that showed 0.1 ppm after a couple hours from dosing 2ml it’s all transformer by morning before second dose.

The phosphate solution is a of the shelf product also, 1ml is equivalent to 0.01 ppm in 100 litres a few minutes after dosing. I have tested this before and is accurate. (Currently not adding phosphate as demand is minimal)

The Carbon is a carbohydrate extracted from algae I believe it’s a semi labile dissolved organic carbon, I can only have a educated guess of the strength of it although it stays from a few days to weeks in the water column. ( currently adding 0.002 grams day) In the video that I’ve shared, shows the carbon as the large still matter at 4x and 10x magnifications (new video)




Yes, this is the correct video at 4 magnifications. The still matter is the carbon used in this test and I assume the Protozoa are the organisms moving faster in the video, there is also millions of smaller bacteria although they only start being visible from 10 magnifications ( I have a video of that if you like to have a look) my 40x is not working properly need to order a new one.
In addition everything on this videos it’s mainly in the water column not from the tank surfaces.

Edit:

10 magnification video


And 4 magnification

Thanks, this helps me understand the nature if this experiment.

 

[sixty_reefer]

Thanks, this helps me understand the nature if this experiment.

No worries, and the video? Should I be looking at something other than Protozoa?

 

[Dan_P]

No worries, and the video? Should I be looking at something other than Protozoa?

Tough question to answer. You might need to kill, fix and count motile species. Here are some parameters to consider when deciding.

Where you sample in the aquarium may have a huge impact on which species are observed. This means many samples could be required for every time point to understand if the community structures of the system are changing because of the experimental treatment and whether the change is location dependent.

Every square millimeter will likely contain a different bacteria community structure. Not sure how to deal with this.

Any microorganism community structure might not be stable over time. If a species your monitoring drops in population size, it might not be observable and you lose your link to what is happening in the experiment. Many species might need to be observed at first until you become familiar with the system and learn which species are useful to follow.

 

[sixty_reefer]

Tough question to answer. You might need to kill, fix and count motile species. Here are some parameters to consider when deciding.

Where you sample in the aquarium may have a huge impact on which species are observed. This means many samples could be required for every time point to understand if the community structures of the system are changing because of the experimental treatment and whether the change is location dependent.

Every square millimeter will likely contain a different bacteria community structure. Not sure how to deal with this.

Any microorganism community structure might not be stable over time. If a species your monitoring drops in population size, it might not be observable and you lose your link to what is happening in the experiment. Many species might need to be observed at first until you become familiar with the system and learn which species are useful to follow.

For the time being I’ve just been taking samples from the protein skimmer, for my personal goals with this experiment, I need to keep a focus on the pelagic organisms rather to the films developing on the tank surfaces as they are key elements.
Although it started as a vast research I believe I stumbled during this test on what I have been looking for a while that could be more beneficial that a tank evaluation and testing.

 

[BeanAnimal]

Thoughts:

-Biofilm accounts for 90-95% of the biomass (excluding fish and coral if they are present).
-No quantification of C
-You can't measure DOC
-makeup of biofilm and water column is significantly different.
-The skimmer samples are not representative of the water column. They have a different concentration and likely show different bioactivity.

If ~5% of the biomass is in the water column and it greatly differs from the bulk of the biomass on surfaces, I am not sure what is being quantified to what conclusion.

Is there a hypothesis?
What are the controls?
Can you explain how the results intend to be interpreted?

 

[sixty_reefer]

Can you explain how the results intend to be interpreted?

There isn’t any results posted…

 

[BeanAnimal]

There isn’t any results posted…

That is a rather odd response. Given the specific thoughts and questions presented.

 

[Dan_P]

There isn’t any results posted…

I think you could discuss, at least theoretically, how you would use the protozoa videos to determine whether a change in nitrogen source had an effect on the aquarium after equilibrium was reached. For example were you expecting the number of protozoa to decline or the species to change? When I looked at mixed micro algae cultures, I had a difficult time seeing a difference in algae communities using nitrate v ammonia.

 

[Dan_P]

Thoughts:

-Biofilm accounts for 90-95% of the biomass (excluding fish and coral if they are present).
-No quantification of C
-You can't measure DOC
-makeup of biofilm and water column is significantly different.
-The skimmer samples are not representative of the water column. They have a different concentration and likely show different bioactivity.

If ~5% of the biomass is in the water column and it greatly differs from the bulk of the biomass on surfaces, I am not sure what is being quantified to what conclusion.

Is there a hypothesis?
What are the controls?
Can you explain how the results intend to be interpreted?

Results from single aquarium experiments, like the ones BRS performs, are difficult to interpret because so many factors are uncontrolled. Unless the experimental treatment has a very large effect, the effort is likely to be a strike out. This type of experimentation is analogous to swinging for the fences in baseball instead of just getting on base.

 

[sixty_reefer]

I think you could discuss, at least theoretically, how you would use the protozoa videos to determine whether a change in nitrogen source had an effect on the aquarium after equilibrium was reached. For example were you expecting the number of protozoa to decline or the species to change? When I looked at mixed micro algae cultures, I had a difficult time seeing a difference in algae communities using nitrate v ammonia.

The discussion already happened, I believe it got closed down
But folks could explain why carbon dosing is not affecting phosphate. I’ve just retested tonight and no changes since the 10th of this month.

No3. 5 mg/l
Po4. 0.4 mg/l
Ph. 8.2
Kh. 9

Regarding current conditions I hope nothing changes and that just stays like that infinitely wile the tank is running.

 

[MnFish1]

Imho the problem is too many variables and no control. IMHO you can’t do this the way you’re trying to do it. It might help if you started by verifying the method
Using a normal carbohydrate. I think you have a specific idea which you are trying to prove and you’re designing an experiment that is more likely to prove what you want it to as compared to prove or disprove your theory

 

[BeanAnimal]

The discussion already happened, I believe it got closed down
But folks could explain why carbon dosing is not affecting phosphate. I’ve just retested tonight and no changes since the 10th of this month.

No3. 5 mg/l
Po4. 0.4 mg/l
Ph. 8.2
Kh. 9

Regarding current conditions I hope nothing changes and that just stays like that infinitely wile the tank is running.

Hi - you have still not answered my questions or Dan's. In any case, You appear to be counting Protozoa and hope that parameters stay the same - Forgive, me but what is the correlation to explaining how carbon dosing affects phosphate levels?

Repeating prior thoughts and questions in case you missed
Thoughts:

-Biofilm accounts for 90-95% of the biomass (excluding fish and coral if they are present).
-No quantification of C
-You can't measure DOC
-makeup of biofilm and water column is significantly different.
-The skimmer samples are not representative of the water column. They have a different concentration and likely show different bioactivity.

If ~5% of the biomass is in the water column and it greatly differs from the bulk of the biomass on surfaces, I am not sure what is being quantified to what conclusion.

Is there a hypothesis?
What are the controls?
Can you explain how the results intend to be interpreted?

Dan_P (among other things asked):
For example were you expecting the number of protozoa to decline or the species to change?

 

[MnFish1]

The discussion already happened, I believe it got closed down
But folks could explain why carbon dosing is not affecting phosphate. I’ve just retested tonight and no changes since the 10th of this month.

No3. 5 mg/l
Po4. 0.4 mg/l
Ph. 8.2
Kh. 9

Regarding current conditions I hope nothing changes and that just stays like that infinitely wile the tank is running.

There is simply no way to make conclusions on this post except 10 hypotheticals which will not help imho. I applaud your method. And though I don’t necessarily think Dan p s experiments can adequately explain in a full tank with fish etc. I would consider going along his route with smaller volumes and more experimental tanks. Don’t take this as a criticism. It’s not meant that way. But I think it’s the only way to do what you’re trying to do. Note this is also not a criticism against Dan p. I just wonder how well these micro aquaria translate to mature 500 gallon tanks