Nitrate Factory

[Subsea]

Overall method to evaluate nutrient processing would be a big help. As we will be using mature substrate from an established healthy tank, I will not try to quantify or qualify the microbial community. It will start out the same in all three tanks.

 

Nutrient input is easily quantified, whether it is ammonia, calcium nitrate, kelp concentrate or fish food. While Tim may not like it, I am considering some mollies. It is boring to feed a tank for two years with no fish. I also am considered the "little people" to maintain cleanliness: Florida Drawf Cerith Snails, micro stars and the pod brothers (amphi & copo). These would be harder to quantify, for that reason, I can be overruled. The macro is easily weighed, either, periodically in the tank or when it is exported. The coral would be quantified in the same way.

 

Have I over simplified the process?

Patrick

[Subsea]

Tibbs,

I will post another thread in biological filtration subforum that is exactly up your alley of expertise. It involves UV sterilizer/clarifier use in my 75G Jaubert Plenum and 135G lagoon tank.

Thanks for tagging along.

Laissez a bonne tems roulee,

Patrick

[tibbsy07]

Overall method to evaluate nutrient processing would be a big help. As we will be using mature substrate from an established healthy tank, I will not try to quantify or qualify the microbial community. It will start out the same in all three tanks.

 

Nutrient input is easily quantified, whether it is ammonia, calcium nitrate, kelp concentrate or fish food. While Tim may not like it, I am considering some mollies. It is boring to feed a tank for two years with no fish. I also am considered the "little people" to maintain cleanliness: Florida Drawf Cerith Snails, micro stars and the pod brothers (amphi & copo). These would be harder to quantify, for that reason, I can be overruled. The macro is easily weighed, either, periodically in the tank or when it is exported. The coral would be quantified in the same way.

 

Have I over simplified the process?

Patrick

I think in terms of nutrient acquisition measure, the general outline you have set up will work. You're going for a proof of principle, qualitative set of data. If you were truly trying to quantify the reactions and products, you'd have to use probes and heavy-duty lab equipment. I don't think it's necessary for this.

 

I also don't think it's necessary to quantify the microbial community. Ensuring that the community is the same would be easy - use the same amount of sand from a single tank to seed the new tanks. Or, do ONE big sand bed, let it age and then spread it out into the three test tanks in identical amounts.

 

That being said, I would not add any other livestock to your tanks. The additional livestock not only makes it more complicated, in my opinion it would call into question the experiment as a whole. Here is why. We want a baseline knowledge of how the substrate and photosynthetic organisms filter the water, right? That means we ONLY want that information. If you add snails, fish, etc. you could completely throw off the balance of the tank. This could shift nitrate use up or down, we don't know.

 

To ensure that your 3 tanks were identical except the test condition (control, algae, coral), you would have to guarantee that the fish/snails/etc. all were genetically identical. They would have to have zero variation. Their gut microbes would have to be identical. They would have to be all the same gender. They would all have to be the same size. The snail shells would have to be the same size/shape. Everything would have had to come from the exact same tank so that literally nothing was different. Any small amount of biological/genetic variation could arguably throw every test you do into the trash. Even if you were able to control for this, you couldn't control whether or not they all eat the same amount, whether they all eat at the same time, how much they release waste, etc. If one died, the entire experiment would be wasted because you couldn't get another identical fish or snail or whatever. As soon as you introduce a new one, you introduce new variables for the tank. New microbes, new pests, etc.

 

Adding anything to the tank other than your water, substrate, and test photosynthetic organism should be a 3rd test, not a 2nd. You need to see what happens without livestock, first. As for the 2 years of boredom, I don't see a reason to go that long. I think that 6 months would actually tell you what you need to know. 2 years would be overkill. I doubt that you'll see a difference in nitrate use at 2 years versus 6 months.

[Subsea]

Tibbs,

Thank you for the affirmation of methodology and completeness of details for Phase 2. Testing and documentation for 6 months work for me. It suites me to leave the tanks set up for an extended time frame of 2 year or longer to advertise my company with a live webcam feed. After the nutrient testing documentation is completed, I will focus on aesthetics and bring the "little people" in to clean up the tanks.

Laissez la bonne temps roulee,

Patrick

[Subsea]

Due to inferior computer savvy, I am having difficulty posting Ward Lab analysis.

 

So I will discuss some concerns and facilitate the lab analysis later.

 

At present, I am drawing ground water from Middle Trinity Aquifer at 900' depth. Iron bacteria are present in low quantities in aquifer and in my macro grow-out systems. Bio available iron in water is negligible at .01ppm in both aquifer and in growout system. As I have been dosing with kelp concentrate, which includes iron at .08% and nitrogen at 16%, I know that iron was added to grow out system, yet analysis shows less than .01ppm. At that concentration, it is on the lowest end of the lab sensitivity at $5 per test. One other reason that I know iron is in my growout system, even when lab analysis shows negligible, I saw macro respond to it. Yes, I saw it and I will supply pictures when I get a better photographer here. Both Randy Holmes Farley and Russ Kronewetter of live-plants described iron uptake as a storage mechanism of macro. I saw the storage mechanism develop on Gracilaria Hayi which I am cultivating in a tumble culture.

 

As I am not interested in growing iron bacteria at the expense of depriving macro algae of a necessary nutrient, I am a little concerned with this knowledge. One other aspect of iron bacteria in my ground water is the unplesant smell of sulphur/h2s? In addition to the smell, any place that sees water sprinklers is stained brownish red. I am implementing a chlorine shock of my well at the pump suction up through the production casing. For many aquifer users, twice a year is sufficient. However, this will not deal with the iron bacteria that went from a fresh water aquifer to a macro growout system at a specific gravity of 1.026.

 

Any thoughts?

[tibbsy07]

Here are the test results. Sorry if the quality isn't great - I had to do some tinkering to block out information, etc.

 

 

Overall, you have pretty low bacterial problems. It's hard to know exactly what you define and what they define as "iron bacteria" because virtually all bacteria require some level of iron - it just depends on the species and the amount required. At 500 CFU/ml, that's not much at all. Hardly what I would consider a detriment to your macroalgae and trying to reduce them or combat them will likely be more trouble than it's worth. They found the exact same amount of iron bacteria in your aquifer and your tank, which could be because of an artifact of their testing process (entirely likely). Taking all this into account, I don't think you have anything to worry about. Your macro will grow just fine.

 

The sulfur/H₂S could be a problem. The brownish red could be iron or it could be biofilms of microbes. Hard to know without pictures or testing.

[Subsea]

Thanks Tibbs.

 

I think that some test are on the extreme low end of there sensitivity. Considering both nitrate and phosphate at .01ppm, I see macro as consuming nutrients as fast as I add them in this 1000G growout system.

 

As I have both iron and phosphate concentrate, I will load my system up with both of those nutrients later today. We shall see what grows.

Laissez la bonne temps roulee,

Patrick

[Subsea]

Tibbs,

Getting back to nitrate study, what level of ammonia in tank would be considered harmful to the development of de-nitrifying bacteria. I have several tanks between 6-8ppm. However, the live rock has processed ammonia down to .5ppm with all tanks receiving the same volume of ammonia. Initially, I planned to control ammonia addition using nitrate as a control parameter between 50-100ppm nitrate. However, I do not want to damage the development of the de-nitrifying bacteria.

Patrick

[tibbsy07]

I'm not sure, to be honest. It would depend on the microbe, amount of each type of bacteria, biofilm formation, etc. If the bacteria can use it as a nutrient source, you're going to have to go WAY high to kill them. In theory, because they use it for a food source, you may just be giving them unlimited nutrients by continuing to add ammonia in high amounts. The only way to truly know would be to do a MIC test, but that's not easy to do at home, and it's not easy to do in my lab as we don't have stuff to do that for marine microbes.

[Subsea]

We shalll limit ammonia to what is easy to measure with API test kits: 0.5 - 8.0 ppm. In the case of tank with live rock, I have increased dosage rate to twice what is going into the other tanks.

 

We shall feed the bugs. Microbial overlords. I like the term.

Bonzai,

Patrick

[Subsea]

Just a quick note.

As I measured ammonia concentrations this morning, I see an early trend with three media; coarse diver collected substrate, dry oolite sand and live rock.

The dry oolite sand is processing ammonia and is tied in second place with corse live sand. At this time, live rock is processing 8ml of ammonia per day with next two at less than 4ml per day.

[Subsea]

Musings from the porch: about substrate.

 

I continue to see a widening gap between live rock, at quadrupled the amount of ammonia processed to the next two closest, coarse live sand and dry oolite sand. Why is the sand not processing more ammonia? Even coarse substrate, 2mm-5mm, is several hundred times more surface area than most live rock. Oolite sand would be several thousand times the surface area of most live rock.. Why are the superior surface areas favoring bacteria colonization not processing ammonia? It is the same processes that happen in our enclosed eco systems. The pore spaces are not being penetrated by advection current. These spaces are becoming stagnant with bio films further preventing the nutrient rich water from reaching the bacteria. In looking at the slight laminar flow from air bubblers I see this as a temporary study in biofiltration. If this was a personnel system to be maintained with good husbandry, I would bring in the janitors: reproducing snails, worms, pods, micro stars with some mollies to clean up the algae.

 

At the same time that I am monitoring eight 10G study tanks, I cycled four 40G tanks in 10 days. The fish were added last. Each day each tank was feed 4ml of clear ammonia. With bacteria and pods introduced with live rock, lights were brought in with differrent macro algae. Iron & iodine were added individually on a 2 times a week cycle. Ammonia concentrate at 4ml is added each day with no measured ammonia 24 hrs later. This tanks will be an experiment in red macro algae.

Bonzai,

Patrick

[tibbsy07]

I was thinking about this the other day, and I think our hypothesis about the sand may be incorrect. The sand, especially deep, can be anaerobic. There may be more surface area overall, but it only matters if the surface area is equal in its value to the organisms processing the ammonia. Nitrosomonas spp. (ammonia to nitrite) can undergo annamox (anaerobic ammonia oxidation to nitrite) but Nitrobacter spp.(nitrite to nitrate) are aerobic. Have you looked at nitrITE levels in the tanks at all? You might find that your sand tanks have a build up of nitrite and are limited by the inability to oxidize that to nitrate because of a lack of aerobic surface area.

[Subsea]

I was thinking about this the other day, and I think our hypothesis about the sand may be incorrect. The sand, especially deep, can be anaerobic. There may be more surface area overall, but it only matters if the surface area is equal in its value to the organisms processing the ammonia. Nitrosomonas spp. (ammonia to nitrite) can undergo annamox (anaerobic ammonia oxidation to nitrite) but Nitrobacter spp.(nitrite to nitrate) are aerobic. Have you looked at nitrITE levels in the tanks at all? You might find that your sand tanks have a build up of nitrite and are limited by the inability to oxidize that to nitrate because of a lack of aerobic surface area.

 

I agree that even with more surface area per unit weight, it does not guarantee that nutrient rich water will feed the bacteria. Due to lack of advective flow, lower sand bed particles are not receiving nutrient rich water. Considering that all motion in tank is laminar with one air pump, the flow is insufficient to drive water thru pore spaces between sand grains. I can only postulate that pore spaces in live rock is sufficiently large enough to allow thus passive circulation to feed sufficient bacteria.

 

I have not been measuring nitrite. This is a good time to send eight samples to Ward Lab for ammonia, nitrite and nitrate. As it is Friday, I will ship samples on Monday. As I have been feeding both phosphate and iron along with ammonia to my grow out tanks, I will consolidate sample shipment cost and detail test for phosphate, iron and total nitrogen in grow out tanks.

Patrick

[tibbsy07]

I agree that even with more surface area per unit weight, it does not guarantee that nutrient rich water will feed the bacteria. Due to lack of advective flow, lower sand bed particles are not receiving nutrient rich water. Considering that all motion in tank is laminar with one air pump, the flow is insufficient to drive water thru pore spaces between sand grains. I can only postulate that pore spaces in live rock is sufficiently large enough to allow thus passive circulation to feed sufficient bacteria. I have not been measuring nitrite. This is a good time to send eight samples to Ward Lab for ammonia, nitrite and nitrate. As it is Friday, I will ship samples on Monday. As I have been feeding both phosphate and iron along with ammonia to my grow out tanks, I will consolidate sample shipment cost and detail test for phosphate, iron and total nitrogen in grow out tanks. Patrick

I don't think it's access to nutrients, necessarily. What I mean is that if your sand bed is 50% aerobic, meaning it has oxygen, and 50% anaerobic, no oxygen - then Nitrosomonas spp. will be fine because they can undergo anaerobic and aerobic ammonia oxidation into nitrite. Nitrobacter spp. on the other hand require oxygen. They will die without it, so they can only be in the 50% with oxygen. If 100 parts of ammonia are being broken down into 100 parts of nitrite, only 50 units of nitrate can come out of it because there is only 50% of the space that Nitrobacter spp. can grow. They are the rate limiting step. In the live rock, the live rock exists as the sole source of oxidation of ammonia to nitrate, and both bacterial organisms can grow, so in theory 100 ammonia can be 100 nitrite and then 100 nitrate. These are all made up numbers and not accurate in terms of actual chemical equations, etc., but in general I think this might be what's happening.

 

Nitrobacter might be able to undergo annamox reactions, but I don't think so.

[Subsea]

As a means to increase circulation in eight study tanks, I have increased air pump volume from two 1/8" lines to ten 1/8" air supply lines. A dramatically increase in turbulent flow conditions was seen equally in all eight tanks. I expect a dramatic increase in ammonia processing to be ramped up in all tanks. We shall evaluate ammonia processing with increased circulation rates. This will generate more evaporation. With increased ammonia additions, I expect more make up water usage with influx of nutrients. This is my method of using high TDS in Aquifier as good nutrient recycling and zero discharge operation as a sustainable mariculture facility. I consider environmental stewardship as a first tier "core value" in the operation of Aquaculture Ranch. Discussions during these post provide much food for thought in operation of 10K gallon mariculture facility during the worst drought in 200 years in Texas Hill Country.

[Subsea]

I don't think it's access to nutrients, necessarily. What I mean is that if your sand bed is 50% aerobic, meaning it has oxygen, and 50% anaerobic, no oxygen - then Nitrosomonas spp. will be fine because they can undergo anaerobic and aerobic ammonia oxidation into nitrite. Nitrobacter spp. on the other hand require oxygen. They will die without it, so they can only be in the 50% with oxygen. If 100 parts of ammonia are being broken down into 100 parts of nitrite, only 50 units of nitrate can come out of it because there is only 50% of the space that Nitrobacter spp. can grow. They are the rate limiting step. In the live rock, the live rock exists as the sole source of oxidation of ammonia to nitrate, and both bacterial organisms can grow, so in theory 100 ammonia can be 100 nitrite and then 100 nitrate. These are all made up numbers and not accurate in terms of actual chemical equations, etc., but in general I think this might be what's happening.

 

Nitrobacter might be able to undergo annamox reactions, but I don't think so.

 

I see the zone that de-nitrification occurs in to be somewhat broader and grouped under faculative bacteria. Bob Goemns in his books and articles on live sand secrets separates the anaerobic bacteria from the faculative in efficiency as faculative bacteria chemistry is 100 times more efficient at nitrate processing. I did not understand the chemistry equation that he used in his book. In NNR techniques, Bob Goemns relied exclusively on bacteria while Ron Schimck and Rob Toonen relied on micro fauna and fana in sand beds.

[Subsea]

With increased circulation, the difference between ammonia processing of the other substrates are approaching the live rock. One other impediment to ammonia processing by substrate is nuisance microalgae covering some of the sand bed. This same micro algae is growing in all tanks due to filtered sunlight inside porch. Samples sent to measure ammonia, nitrite and nitrate.

 

I fell that insufficient circulation was the big problem. However, nuisance micro algae can skew ammonia processing by bacteria in two ways. It can inhibit nutrient rich water from reaching substrate surfaces and it will remove ammonia from bulk water column. As long as the algae film is consistent in all study tanks, we will press on.

Patrick

[Subsea]

Some specific updates/ my measurements

 

8. Control tank is maxed out with 8ppm NH4 and 150ppm nitrate after adding 34ml of NH4

7. Rock rubble is maxed out on nitrate at 100ppm. NH4 is 0ppm after adding 56ml of NH4

6. Live rock has processed 94 ml of NH4. NH4 at <.5ppm and nitrate is at 15ppm

5. Ceramic rock has processed 28ml of NH4. NH4 at 4ppm and nitrate at 20ppm

4. Sponge Bob has processed 72ml of NH4. NH4 at 0ppm and nitrate at 20ppm

3. Bioballs have processed 42ml of NH4. NH4 is at .5ppm and nitrate is at 20 ppm

2. Oolite sand has processed 54ml of NH4. NH4 is at 4ppm and nitrate is at 15ppm.

1. Coarse sand has processed 56ml of NH4. NH4 at 1ppm and nitrate at 20ppm.

[tibbsy07]

Great, thanks for the update!

[Subsea]

I expect Ward Lab results back today: ammonia, nitrite, nitrate.

 

General comments:

 

Bioballs and rock rubble both started out slow. In fact each contributed ammonia into the water. The bioballs were used and not thoroughly cleaned. The rock rubble was purchased from BRS and was a 10lb package with an assortment of all the dry rock they sell. At present it is processing 20ml of ammonia a day.

 

Tied for second place is bioballs and Sponge Bob, each at 12 ml/day.

 

Live rock fluctuates between 10-12 ml/day.

 

Coarse diver collected substrate is processing 8 ml/day. Dry oolite sand is processing 4-6 ml/day. I contribute this poor showing to nuisance micro algae which is clogging up the substrate. Early in the study, I decided I would not remove this complication. In a healthy aquarium the janitors would do this.

 

The four study tanks with the least amount of micro algae are rock rubble, live rock, bio balls and Sponge Bob, in that order. Sponge Bob has moved ahead of live rock in processing ammonia per day at 12ml/day compared to 10 ml/day and I expect it to exceed live rock in total ammonia processed later this week.

 

If the trend continues, I expect bioballs to exceed live rock in ammonia processing and perhaps nitrate processing. I find this very surprising with 1 lb of bioballs to 10 lbs of live rock.

 

Untill later,

Patrick

[tibbsy07]

What exactly is sponge bob, again?

[Subsea]

Sponge Bob is represented by two sponge filters. One was the sponge removed from AC50 HOB and one that you would see in a 10G fresh water tank with an air uplift tube.

 

Without a doubt, nitrate is being processed in this study tank.

[tibbsy07]

Sponge Bob is represented by two sponge filters. One was the sponge removed from AC50 HOB and one that you would see in a 10G fresh water tank with an air uplift tube.

 

Without a doubt, nitrate is being processed in this study tank.

Are they just in the tank or are they in an HOB?

[Subsea]

In the tank. The sponge with the air uplift tube has water passing thru it. The other sponge has passive circulation which is the same as the other media.