Bacteria in bottle, busting myth, Seneye style.
- Thread starter Dr. Reef
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Becasue there will be no product in the contol is how. To which my concern is addressed and valid. I developed animal vaccines many years ago for Pfizer which included animal testing and imo animal safety should be careful weighed against what is to gain without the sort of justification of "it happens here so it's ok here"
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Dr. Reef
www.drreefsquarantinedfish.com
if its free ammonia NH3 at 0.57 its 3 x the amount of dangerous levels. If its true then clownfish are some strong hardy fish.
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A. ocellaris juveniles (1.20 ± 0.34 g) were exposed to six concentrations of ammonia ranged from 0.23 to 1.63 mg/L NH3-N and eight concentrations of nitrite (26.3–202.2 mg/L NO2−-N). The LC50- 24, LC50-48, LC50-72 and LC50-96 h were estimated to be 1.06, 0.83, 0.75 and 0.75 mg/L for NH3-N and 188.3, 151.01, 124.1 and 108.8 mg/L for NO2−-N. Analysis of gill lesions caused by sublethal concentrations of these nitrogenous compounds showed that both nitrogenous compounds induced tissue lesions such as hyperplasia of epithelium cells, hypertrophy of chloride cells and lamellar lifting to all concentrations tested. However, histopathological alterations were more conspicuous accordingly the increase of ammonia or nitrite in fish exposed to 0.57 mg/L NH3-N or 100 mg/L NO2−-N. Based on our results, we recommend to avoid concentrations higher than 0.57 mg/L of NH3-N and 25 mg/L of NO2-N in water.
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Dr. Reef
www.drreefsquarantinedfish.com
Its best i pull fish out of the control tank before it hits dangerous levels. and continue the rest 4 tanks with fish and bacteria unless they start to show dangerous levels and then fish can be pulled showing product doesnt work.
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Dr. Reef
www.drreefsquarantinedfish.com
tough sons of guns....lol
Seems like a reasonable plan and happy you had the conversation!
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I did address your concern in my first post in response to you. BTW - this is a discussion - And it was addressed with the removal of fish at a level of ammonia < toxic levels....
I was responding to another poster who wanted to abandon fish altogether because it would mess up the study.
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Dr. Reef
www.drreefsquarantinedfish.com
Absolutely, I always post the plans before testing so everyone can weigh in on it and we can find the best route to test. I am glad we can agree and test in safer manner for fish's sake after all the whole idea behind the test is to show if bacteria in bottle can keep ammonia at bay for fish to survive.
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Nitrifying bacteria require ammonia (the type we're talking about) primarily not carbon (they need carbon - but get it from Carbon Dioxide). They also require Phosphate and Mg. My assumption is that those solutions that allow cycling while only adding 'ammonia' have added Phosphate perhaps - or the amount in whatever salt is being used is assumed to have 'some'. In other words 'obligate autotrophs' (nitrifying type) do not require any carbon source only ammonia and a bit of Phosphate and Mg (it is said that Fritz Turbo and Dr. Tims contain this type of bacteria). Heterotrophs can denitrify (reduce ammonia) - but they do it by other pathways using carbon compounds dissolved in the water. They also need Mg and PO4.
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The 'lethal concentration' that killed 50% of the fish in 24 hours was 1.06 ppm. .83ppm killed 50% at 48 hours. etc etc. For this study - I would recommend going up to say .5 in ALL of the tanks - thus the variables analyzed would be : 1. The fall (or rise) in ammonia concentration (the thought being that the ammonia will not rise in the bacteria tanks but it will in the others. 2. The overall survival of the fish (inspire of normal or sublethal ammonia), 3. The length of time fish can remain in the tank before 'lethal' concentrations of ammonia develops.
I think it SHOULD be safe to remove fish from any tank when. the concentration for that tank rises about .5 ppm?
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Dr. Reef
www.drreefsquarantinedfish.com
given that study and numbers quoted being true, 0.5ppm would be a good point to remove fish from the testing and control tanks. Thus preventing harm.
Regarding food, i will put same amount of fish food (weighed on scale) in each tank once a day.
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Did you see my post about endpoints? PS - You could also put in a failsafe using specific criteria for removal - ie even if the ammonia level is .25 - if the fish has (your list of criteria for ammonia toxicity) - they would be removed. Same thing for 'disease' (ie.. if a fish has CI - do you leave them in the study tank or treat. As long as the things are pre-designed it should still be valid (lol unless bottled bacteria contains CI or velvet... etc
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Dr. Reef
www.drreefsquarantinedfish.com
Lol. I got it.
I will order a dozen or more clownfish and place them in QT. Make sure they are healthy and decease free. Then subject them to testing.
Regarding pull out point. 0.5ppm is good.
End point, I guess if bacteria works then as long as ammonia stays 0 after initial cycle for 7 days past, I'll consider it good or else we can pull if ammonia starts to increase.
I will order a dozen or more clownfish and place them in QT. Make sure they are healthy and decease free. Then subject them to testing.
Regarding pull out point. 0.5ppm is good.
End point, I guess if bacteria works then as long as ammonia stays 0 after initial cycle for 7 days past, I'll consider it good or else we can pull if ammonia starts to increase.
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Another point - will you also be testing Nitrite/nitrate.
I dont know if you saw my post about having a tank with just food and NSW - to see how fast ammonia rises with only that. I know thats 6 tanks. but... Its an important control IMO
alex77619
Active Member
Wanted share a translation testing on nitrifying bacteria I found in Taiwan.
Six brand nitrifying bacteria evaluation (finished)
Hello, everyone.
The problem of the efficiency of nitrifying bacteria has always been questioned and concerned about for new salt fish hobbyist . Some laboratory support has done the following tests for your reference.
It’s good to know what you are using.
Please also see the results of the experiment. It is for reference and discussion. It is not intended to sell the brand.
It must be emphasized that the detection method is only the same in the laboratory under the standard conditions, is it the same in the different complex cylinders?
(a) Nitrite test
Test method: Modified according to the nitrite spectrophotometer (NIEA W418.51C) announced by the Environmental Protection Agency .
Test Methods:
1. Take 6 ml of seawater (containing 5 ppm of Nitrite ) and add 0.1 ml (liquid) or 0.1 g (powder) of 6 commercially available products (see Figure 1).
2. Take 1ml of liquid to check if the content of Nitrite is consistent.
3. Set to shake at 27 °C for 20 hours.
4. Take 1 ml of the reacted liquid to detect the content of Nitrite .
Test Results:
1. The CW brand (that is, the C of Figure 1) can almost completely degrade Nitrite .
2. BI brand powder has a partial reaction when it is added, but there is no drop after 20 hours. The reason is not the effect of bacteria.
3. Other brands under this test condition are unable to effectively reduce the Nitrite .
Figure 1. Product
Figure 2. Before the reaction
Figure 3. After the reaction
(2) Total ammonia test
Test method: Modified according to the nitrite spectrophotometer (NIEA W448.51B) announced by the Environmental Protection Agency.
Test Methods:
1. Take 6 ml of seawater (containing 5 ppm of ammonia) and add 0.1 ml or 0.1 g of 6 commercially available products.
2. Take 1ml of liquid to check whether the ammonia content is consistent.
3. Set to shake at 27 °C for 20 hours.
4. Take 1 ml of the liquid after the reaction to detect the ammonia content.
The test results are roughly similar to the Nitrite results .
Fig. 5 before the reaction
Figure 6. After the reaction
(3) Nitrate test
Nitrate : 50ppm
Detection method: using API detection reagent
Test Methods:
1. Take 6 ml of seawater (containing 50 ppm of Nitrate ) and add 0.1 ml or 0.1 g of 6 commercially available products.
2. Take 1ml of liquid to check if the content of Nitrate is consistent.
3. Continuously shake at 44 °C for 44 hours.
4. Take 1 ml of the liquid after the reaction to detect the content of Nitrate .
Test Results:
1. CW brand can effectively degrade Nitrate under this condition, but its decrease is obvious on the second day. The action time is longer than the degradation of Nitrate and total ammonia!
2. The BI brand seems to interfere with the detection of the detection reagent at the beginning, and it is impossible to judge whether it will degrade, similar to the above experimental results.
3. Other brands under the test conditions, can not degrade Nitrate (there is almost no comparison with the control group, the same is true with spectrophotometer.)
Figure 7. Before the reaction (from left to right AP SE ST BI 50 CW control group)
Figure 8. After the reaction (1st day)
Figure IX. After the reaction (2nd day)
Again, these results are of course limited, and their reference value is judged by the viewer.
(4) Growth of bacteria
This part is for simple presentation to everyone.
I use the simplest picture to discriminate without using the usual petri dish or the way of counting the nine squares.
This stage of the experiment used two repetitions
test method
1. Take 6 ml of sterilized seawater (to avoid interference from bacteria) and add 0.1 ml or 0.1 g of 6 commercially available products.
2. Place at 27 °C for 48 hours. Photo record~
Test Results
1. Before the reaction, the SE and BI groups showed white turbidity and yellow turbidity when they entered the water.
The CW group has a little turbidity, and the rest are clear.
2. After the reaction, the CW group became obviously cloudy and cloudy .
It shows that the fungi may grow in a large amount, and the number of bacteria is confirmed by a microscope.
No other significant changes in the remaining groups
Figure 10. Before the reaction (from left to right AP / SE / SI / BI / 50 / CW)
Figure 11. After the reaction (from left to right AP / SE /SI /50 /BI /CW Sorry, BI and 50 are reversed at this time)
Six brand nitrifying bacteria evaluation (finished)
Hello, everyone.
The problem of the efficiency of nitrifying bacteria has always been questioned and concerned about for new salt fish hobbyist . Some laboratory support has done the following tests for your reference.
It’s good to know what you are using.
Please also see the results of the experiment. It is for reference and discussion. It is not intended to sell the brand.
It must be emphasized that the detection method is only the same in the laboratory under the standard conditions, is it the same in the different complex cylinders?
(a) Nitrite test
Test method: Modified according to the nitrite spectrophotometer (NIEA W418.51C) announced by the Environmental Protection Agency .
Test Methods:
1. Take 6 ml of seawater (containing 5 ppm of Nitrite ) and add 0.1 ml (liquid) or 0.1 g (powder) of 6 commercially available products (see Figure 1).
2. Take 1ml of liquid to check if the content of Nitrite is consistent.
3. Set to shake at 27 °C for 20 hours.
4. Take 1 ml of the reacted liquid to detect the content of Nitrite .
Test Results:
1. The CW brand (that is, the C of Figure 1) can almost completely degrade Nitrite .
2. BI brand powder has a partial reaction when it is added, but there is no drop after 20 hours. The reason is not the effect of bacteria.
3. Other brands under this test condition are unable to effectively reduce the Nitrite .
Figure 1. Product
Figure 2. Before the reaction
Figure 3. After the reaction
(2) Total ammonia test
Test method: Modified according to the nitrite spectrophotometer (NIEA W448.51B) announced by the Environmental Protection Agency.
Test Methods:
1. Take 6 ml of seawater (containing 5 ppm of ammonia) and add 0.1 ml or 0.1 g of 6 commercially available products.
2. Take 1ml of liquid to check whether the ammonia content is consistent.
3. Set to shake at 27 °C for 20 hours.
4. Take 1 ml of the liquid after the reaction to detect the ammonia content.
The test results are roughly similar to the Nitrite results .
Fig. 5 before the reaction
Figure 6. After the reaction
(3) Nitrate test
Nitrate : 50ppm
Detection method: using API detection reagent
Test Methods:
1. Take 6 ml of seawater (containing 50 ppm of Nitrate ) and add 0.1 ml or 0.1 g of 6 commercially available products.
2. Take 1ml of liquid to check if the content of Nitrate is consistent.
3. Continuously shake at 44 °C for 44 hours.
4. Take 1 ml of the liquid after the reaction to detect the content of Nitrate .
Test Results:
1. CW brand can effectively degrade Nitrate under this condition, but its decrease is obvious on the second day. The action time is longer than the degradation of Nitrate and total ammonia!
2. The BI brand seems to interfere with the detection of the detection reagent at the beginning, and it is impossible to judge whether it will degrade, similar to the above experimental results.
3. Other brands under the test conditions, can not degrade Nitrate (there is almost no comparison with the control group, the same is true with spectrophotometer.)
Figure 7. Before the reaction (from left to right AP SE ST BI 50 CW control group)
Figure 8. After the reaction (1st day)
Figure IX. After the reaction (2nd day)
Again, these results are of course limited, and their reference value is judged by the viewer.
(4) Growth of bacteria
This part is for simple presentation to everyone.
I use the simplest picture to discriminate without using the usual petri dish or the way of counting the nine squares.
This stage of the experiment used two repetitions
test method
1. Take 6 ml of sterilized seawater (to avoid interference from bacteria) and add 0.1 ml or 0.1 g of 6 commercially available products.
2. Place at 27 °C for 48 hours. Photo record~
Test Results
1. Before the reaction, the SE and BI groups showed white turbidity and yellow turbidity when they entered the water.
The CW group has a little turbidity, and the rest are clear.
2. After the reaction, the CW group became obviously cloudy and cloudy .
It shows that the fungi may grow in a large amount, and the number of bacteria is confirmed by a microscope.
No other significant changes in the remaining groups
Figure 10. Before the reaction (from left to right AP / SE / SI / BI / 50 / CW)
Figure 11. After the reaction (from left to right AP / SE /SI /50 /BI /CW Sorry, BI and 50 are reversed at this time)
Last edited:
I saw this thread today and I can be to late with this information. I have not read the whole thread so excuse me if I´m repeating anything.
Fish do not excrete NH3/NH4 in an even fashion through 24 hours. 80% of the excretion occurs within 2 hours after eating. It means that you can use batch of NH3/NH4 a day to mimic real life.
Further on - If you give a fish 1 g of dry feed (dry feed with protein concentration of 45 %) - you will feed the fish with around 0.072 g N (72 mg N) 1 X 0.45 (protein content) x 0.16 (average N concentration in protein). Of this the fish will take around 20 % to new biomass and 80 % will go as waste. In this case with dry food it means that around 58 mg will enter the water as NH3/NH4 - N. If you measure in NH4 - it will be 74 mg NH4 -or 70 mg NH3 If you use 100 liter tanks it correspond to around 0.74 ppm NH4. If you use frozen food - the protein content is around 12 %. It means that 1 gr frozen food will give you around 15 mg NH3/NH4 - N -> around 20 mg NH4 -> 100 liter water -> 0,2 ppm NH4. In my case with using fish and nitrification bacteria I use around 10 mg frozen artemia every third day the first week. It means that a 100 l tank will have around 0.002 ppm NH3/NH4 every third day (see). If you should mimic a fish cycling method - you should add between 0,2 mg/100 l to 20 mg NH3/NH4 /100l
Sincerely Lasse
Fish do not excrete NH3/NH4 in an even fashion through 24 hours. 80% of the excretion occurs within 2 hours after eating. It means that you can use batch of NH3/NH4 a day to mimic real life.
Further on - If you give a fish 1 g of dry feed (dry feed with protein concentration of 45 %) - you will feed the fish with around 0.072 g N (72 mg N) 1 X 0.45 (protein content) x 0.16 (average N concentration in protein). Of this the fish will take around 20 % to new biomass and 80 % will go as waste. In this case with dry food it means that around 58 mg will enter the water as NH3/NH4 - N. If you measure in NH4 - it will be 74 mg NH4 -or 70 mg NH3 If you use 100 liter tanks it correspond to around 0.74 ppm NH4. If you use frozen food - the protein content is around 12 %. It means that 1 gr frozen food will give you around 15 mg NH3/NH4 - N -> around 20 mg NH4 -> 100 liter water -> 0,2 ppm NH4. In my case with using fish and nitrification bacteria I use around 10 mg frozen artemia every third day the first week. It means that a 100 l tank will have around 0.002 ppm NH3/NH4 every third day (see). If you should mimic a fish cycling method - you should add between 0,2 mg/100 l to 20 mg NH3/NH4 /100l
Sincerely Lasse
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