Can the size of your nitrifying bacteria colony be measured?

[brandon429]

Here are applications for Dom's question related to common posts:

-during fallow if you dont have fish present, and you dont add food during the wait, can you add your fish back all at once and still pass all ammonia safety?

-I have ten tangs among rocks and sand for 5 years. if i remove all sand instantly, does the live rock need time to take on more bacteria, is removal in sections of sand required to allow new growth time?

-when I buy live rocks at the pet store that have living coralline, some fanworms, a little algae attached, do I still add bottle bac and add ammonia to 2 ppm? Why do 100% of all cycle materials in books or online not differentiate live rock transfer vs new rock cycling?

100% of all training materials says add shrimp, dose to 2 ppm ammonia, add bottle bac, wait till api shows zero ammonia. dated cycling info doesnt differentiate, it has you burn your animals paid extra for. if we add true live rock with worms and pods and attached corals, why in the world would we burn them with dosed ammonia, we know filter bacteria are there by association to living creatures

cycling rules for the hobby actually dont make sense when work thread patterns are analyzed.

 

[Dom]

and yet their reefs are doing fine without this useless info.

This info isn't useless, it is about expanding your understanding of how your tank works.

If it is so useless, why participate in the thread?

 

[brandon429]

I struggle to find another subject as applicable to our tanks where no forums across the internet have a suitable umpire. amazing.
we cant get peer reviewed material for aquarium science, dangit. we, as peers currently provide the best reviews I guess.

google scholar has oceanic studies, which aren't aquarium studies.

fish disease prevention has suitable referees

chemistry has a hundred qualified reef refs

and cycling has people with sales at stake writing the rulebooks.

 

[HB AL]

I haven't had to cycle a new tank in decades, whether downsizing or upsizing using the rock and some sand of previous tank I haven't had to cycle. Now I dont go over stocking on fish and corals on day 1. I think the only way to measure it is by going with how much bioload its processing successfully either by testing or observation of corals health.

 

[Aqua Man]

2 tanks 10 g

Tank 1 - 10 lbs live rock from a seasoned reefer. 1 cup live sand

Tank 2 - 10 lbs DRY rock, Dry sand
T2 used bottle bac

Both tanks have same amount of sand.

Started on the same day with the same fish/bio load.

Are the the bacteria colonies going to be the same size? I want to say no.

 

[brandon429]

I would agree to that. My caveat prior was post cycle.


at the end of the cycle, both systems can endure a full water change, redose ammonia, and it will be oxidized next day


they perform equal in ways you can measure, post cycle. The fact some surfaces had a ramp up time from total dry is accepted.

my main detail to watch out for is us thinking wet surfaces given time to meet a cycle, let’s say the universal 30 days on all cycling graphs printed and online, do not have total coverage relative to surface area.


there are no sterile unused zones post cycle, all vital space has its biofilm layering and adding any more decreases presentation to wastewater vs increasing it. That specific detail of already cycled surfaces reserving room to take on more bac, if more fish are added, is where the public totally diverges from what we can reliably demo in work threads.


two Tanks

one given maximum half a ppm ammonia during 30 day cycle once or twice.


one given 2 ppm several times. Much more feed input.

at the end of cycle, both get a full water change and no form of testing can distinguish filter efficiency differences among the two if the surface area is the exact same between both tanks. They keep the same starting bioload and function in the thousandths ppm on seneye is the bet, weaker cycled one converts the same it won’t run in hundredths ppm, slightly slower.


once cycled, surface area is the modulator not bacteria

 

[Aqua Man]

What about bacteria being over burdened?

Could slightly slower allow enough free ammonia to become lethal?

 

[brandon429]

I truly don’t know but the best way to branch out past api measures / stuck cycles is by watching what seneye cycles say and by watching the sheer volume of fish-in cycle posts in the new tank forum

rtr probably has two hundred fish-in cycles and I can’t remember one that died due to ammonia toxicity which has classic signs in fish and in the system. when a couple clowns added wait two weeks to die on separate days it’s not ammonia that did it, ammonia toxicity is a fast killer just like loss of kidney or nephridial function in higher order animals is quickly lethal. Hours

im sure there is a degree our bac can’t keep up with but lethality from lack of bac or surface area in displays is so rare I can’t think of a single work example that fits the bill

 

[DrTim]

I am not exactly sure what you are asking when you say 'size' so will present a few different thoughts.

If you mean the numbers of ammonia-oxidizing bacteria (AOB) versus the number of nitrite-oxidizing bacteria (NOB versus the number of non-nitrifying bacteria? That technique is easy. What is hard is to know how representative your sample is compared to the rest of the tank but let's put that aside for now.

So the technique is called FISH (Fluorescence In-SItu Hydridization) combined with MARS (Microautoradiography) or you can do FISH combined with a Laser Scanning Confocal Microscope. FISH using molecular probes that are specific for the species, genus, group of bacteria of interest. You use different colors to distinguish different species. Below is a FISH picture I took of a nitrifying biofilm. The AOB are green the NOB are red. The individual cells are colored so with some software and right scope you can count the individual cells. As I mentioned you can target species so you could count different species of AOB or AOB versus AOA etc.

For more info about this see Ecophysiological Interaction between Nitrifying Bacteria and Heterotrophic Bacteria in Autotrophic Nitrifying Biofilms as Determined by Microautoradiography-Fluorescence In Situ Hybridization Applied and Environmental Microbiology (AEM ) 2004: Vol 70.

To see it applied to aquariums you can download my peer-reviewed paper - Identification of Bacteria Responsible for Ammonia Oxidation in Freshwater Aquaria AEM 2001: Vol 67. Yes Freshwater but the techniques are the same and a google scholar search will yield a lot of papers.

So yes, you can count the cells. But things are not that simple even in an aquarium.

Under perfect conditions the conversion of ammonia or nitrite by AOB and NOB, respectively, is considered a zero order reaction. Meaning, with some limits, the amount of ammonia or nitrite does not matter as the cell can only convert so much so fast. More ammonia does not produce more nitrite from the individual cell. You need more cells all working at the same rate which is why ammonia (or nitrite) builds-up until more cells are produced.

Aquariums are not perfect conditions - not only does the amount of ammonia and nitrite vary but so does the pH along with the amount of micro-nutrients like phosphate, copper and iron. What this means is that the individual cell may not be working at maximum efficiency (its not) and may not divide at a fix rate of time (say every 20 hours). Since the cellular conversion rate now depends on the levels of the variety of reactants this is now called a first order reaction.

How is this important to aquariums? Because it means that the increased rate of conversion of ammonia to nitrite or nitrite to nitrate does not necessarily mean an increased number of cells only that the existing cells are doing more work.

So you could measure a doubling the ammonia conversion but not count twice as many AOB cells.

That's why biology is so interesting - the answers are not easy!


Cheers

 

[Dom]

I am not exactly sure what you are asking when you say 'size' so will present a few different thoughts.

If you mean the numbers of ammonia-oxidizing bacteria (AOB) versus the number of nitrite-oxidizing bacteria (NOB versus the number of non-nitrifying bacteria? That technique is easy. What is hard is to know how representative your sample is compared to the rest of the tank but let's put that aside for now.

So the technique is called FISH (Fluorescence In-SItu Hydridization) combined with MARS (Microautoradiography) or you can do FISH combined with a Laser Scanning Confocal Microscope. FISH using molecular probes that are specific for the species, genus, group of bacteria of interest. You use different colors to distinguish different species. Below is a FISH picture I took of a nitrifying biofilm. The AOB are green the NOB are red. The individual cells are colored so with some software and right scope you can count the individual cells. As I mentioned you can target species so you could count different species of AOB or AOB versus AOA etc.

For more info about this see Ecophysiological Interaction between Nitrifying Bacteria and Heterotrophic Bacteria in Autotrophic Nitrifying Biofilms as Determined by Microautoradiography-Fluorescence In Situ Hybridization Applied and Environmental Microbiology (AEM ) 2004: Vol 70.

To see it applied to aquariums you can download my peer-reviewed paper - Identification of Bacteria Responsible for Ammonia Oxidation in Freshwater Aquaria AEM 2001: Vol 67. Yes Freshwater but the techniques are the same and a google scholar search will yield a lot of papers.

So yes, you can count the cells. But things are not that simple even in an aquarium.

Under perfect conditions the conversion of ammonia or nitrite by AOB and NOB, respectively, is considered a zero order reaction. Meaning, with some limits, the amount of ammonia or nitrite does not matter as the cell can only convert so much so fast. More ammonia does not produce more nitrite from the individual cell. You need more cells all working at the same rate which is why ammonia (or nitrite) builds-up until more cells are produced.

Aquariums are not perfect conditions - not only does the amount of ammonia and nitrite vary but so does the pH along with the amount of micro-nutrients like phosphate, copper and iron. What this means is that the individual cell may not be working at maximum efficiency (its not) and may not divide at a fix rate of time (say every 20 hours). Since the cellular conversion rate now depends on the levels of the variety of reactants this is now called a first order reaction.

How is this important to aquariums? Because it means that the increased rate of conversion of ammonia to nitrite or nitrite to nitrate does not necessarily mean an increased number of cells only that the existing cells are doing more work.

So you could measure a doubling the ammonia conversion but not count twice as many AOB cells.

That's why biology is so interesting - the answers are not easy!


Cheers

As I mentioned earlier, I understood that the Nitrifying Bacteria Colony in a tank is proportional to the bio load.

More bio load, more bacteria required to process the load. Less load, less bacteria required to process.

I wanted to know if there was a way to test for nitrifying bacteria so that I could compare the results to the bio load in the tank.

 

[DrTim]

As I mentioned earlier, I understood that the Nitrifying Bacteria Colony in a tank is proportional to the bio load.

More bio load, more bacteria required to process the load. Less load, less bacteria required to process.

I wanted to know if there was a way to test for nitrifying bacteria so that I could compare the results to the bio load in the tank.

I was trying to explain that "More bioload = more bacteria" is not necessarily true. It is not axiomatic that more ammonia or nitrite means more bacteria.

Yes there are ways to measure the number of nitrifiers but you need specialized equipment.

Regards

 

[blasterman]

I see what the

ramp up time doesn't exist, looking for a ref that says it does/excellent way to vet claims.

Once Nitrobacter have established and the nitrite back cycle gets worked through the growth of the bacteria bed is exponential. So yeah, you can ramp up a bio load and the colonies can increase quite rapidly so theres no issue. After a period of time a healthy ecosystem will establish lots of low activity bacteria as opposed to specific regions of high activity because the later system likely has flow problems, lots of stagnant areas, etc. The sewage plant guy knows all about this.

However, it is not the same event going from a zero bioload environment to a low or moderate one. I can run zero nutrient SPS tanks, or low nutrient SPS tanks, but going from low to none, waiting too long and going back to low can cause issues because the bacteria are no longer present. This why I harp on reef stores to keep fish in their live rock rubble tanks. Without at least a small source of ammonia the rock becomes inert. The exact time frame this occurs is what is debatable.

 

[brandon429]

Disagreed first paragraph Blaster
Cannot go to inert state, we have multi-year fallow testing on file from DJ City. he kept live rocks wet for 36 mos with no feed

on year three we did ammonia testing and they passed just like rocks kept with fish the whole time. They handled instantly more bioload than they were adapted or ramped up to


Daniel our mod measured ammonia on a mindstream device during mature reef tank instant sand removal and the conversion rate with fish didn’t change from ppb pre sandbed removal or after. Rocks had zero ramp up time to carry more bioloading and the ammonia conversion rate never changed, directly opposite of the claim that ramp up time is required or even occurs in post-cycle work testing. Keeping the conversion rate in ppb was the amazing ability of live rock tank to tank.

I do agree that bac colonies are active and dynamic and make cell count changes based on boons and deficits of feed in an aquarium but they dont retro scale to zero bacteria due to natural feed mechanisms, bac get fed even if we don’t provide directly for them. DJ’s fallow nitrifiers were still getting feed, they weren’t truly starved since they stayed in water.

 

[Darsh]

I don't see how it matters what size/quantity there are? If you need more, put more surface area in the tank. Am I missing something?

As usual. Yes haha

 

[Plecosam]

This info isn't useless, it is about expanding your understanding of how your tank works.

If it is so useless, why participate in the thread?

Because there is always at least one, and people complain about Facebook, secondly, some people just want to parrot the "hear say" and not actually think for themselves.

My 2c what about tumbling media like K1 where the old bacteria gets dislodged?

 

[Plecosam]

Ppl

As usual. Yes haha

Personally I think surface area is only a small piece of the puzzle but is used as a big marketing tool to sell expensive medi.

 

[pdxmonkeyboy]

Aquabiomics can tell you the diversity and relative abundance of each species of bacteria in the tank.

As stated above "total number" is not actually a useful measurement as each species does things at different rates and different effeciencies.

 

[pdxmonkeyboy]

Personally I think surface area is only a small piece of the puzzle but is used as a big marketing tool to sell expensive medi.

You are free to think that but surface area is needed for biofilms to propagate. And since when is media expensive anyways?

 

[Darsh]

Ppl

Personally I think surface area is only a small piece of the puzzle but is used as a big marketing tool to sell expensive medi.

I was referring to the "missing something" part.

 

[Plecosam]

Yes surface area is needed, but in my opinion we are trained to think expensive media is needed by marketing departments. The exception is where volume is limited such as in a canister filter, but in a sump many things have and can be substituted for purchased media.

I have run substituted media in heavily stocked tanks and nothing bad happened, in my 200 gallon aquarium I do not have any purchased media except for some very cheap imitation tumbling k1 and different porosity foam.