# Modern research on freshwater biofilters and cycling



## shifty803 (May 17, 2020)

I have been out of the hobby for almost a decade, and due to my background tend to review the scientific literature on topics that interest me. I was surprised to see some very compelling research on the topic of freshwater aquarium biofilters in the last 10 years, and very little discussion about it in aquarium forums I used to be part of. I am not totally sure why that is...

In any case - here are some tentative findings that I have tried to extract from the literature.

1) The long-term primary ammonia-oxidizing component of biofilters is clearly NOT biosomonas and similar bacteria (AOB) as previously thought. Rather, it is ammonia-oxidizing archaea species (AOA). Note that this is not true for marine tanks, only freshwater.

2) Biospira still seems to be one of the dominant forms of nitrite-oxidizing species, even in freshwater tanks.

These two things explain so much about difficulties with cycling and why bottled bacteria never seem to work quite as well as we would hope. Some of the literature on this is pretty dense unfortunately.

If you want to read any paper on this topic, there is a really interesting and much easier to understand thesis by a student at Waterloo:

Szabolcs, Natasha. "Ecology of Ammonia-oxidizing Archaea and Bacteria in Freshwater Biofilters."
https://uwspace.uwaterloo.ca/bitstream/ ... sAllowed=y

She basically shows that the most effective start to cycling a tank is one that includes introduction of AOA populations immediately, which none of the current "bacteria in a bottle" provide. AOB do seem to contribute to the initial process, but her experiments show much faster ammonia reduction with AOA inoculation.

See also (just a couple examples):

Bagchi, S. et. al. "Temporal and Spatial Stability of Ammonia-Oxidizing Archaea and Bacteria in Aquarium Biofilters."
https://dx.plos.org/10.1371/journal.pone.0113515

Sauder, L. et. al. "Aquarium Nitrification Revisited: Thaumarchaeota Are the Dominant Ammonia Oxidizers in Freshwater Aquarium Biofilters."
https://journals.plos.org/plosone/artic ... ne.0023281

My hypothesis is that SafeStart and similar products can "work" for fish-in cycles because nitrite is controlled and ammonia concentrations around, say, 1.0 ppm at normal ph ranges and temperatures are not actually that dangerous to fish over short time periods. The percentage of unionized ammonia (versus total ammonia) is very low until you get into high ph ranges (e.g. 8.0+). Those of us running high pH values therefore can't get away with such methods.


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## DJRansome (Oct 29, 2005)

So fishless cycle with ammonia is still best?


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## shiftyfox (Aug 2, 2019)

If I was to start over I would definitely go down the fishless cycle route with ammonia. 
I went with Fluvals bottled bio and was constantly paranoid that I was risking the fishes health throughout the process.

However throughout the cycling process Ammonia never went over 1ppm and nitrite 0.50ppm 
Would the readings have been much higher without adding the bottled bio at the start - I'm not sure..?


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## DJRansome (Oct 29, 2005)

My fish start gasping when nitrite is 0.5ppm. Still does not sound like it's worth the risk to me. You only have to cycle with ammonia once in your lifetime, better to avoid ALL the toxins.


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## shiftyfox (Aug 2, 2019)

Agree...I guess many of us are guilty of being impatient, you've got this lovely tank set up and your staring at a bunch of rocks for six weeks. 
But yeah do it right and your set for life.


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## willyg1631 (Feb 18, 2020)

shiftyfox --I'm in the middle of a fishless cycle in my new Red Sea Reefer 350 90g. Been keeping a log of my numbers since Day 1 which was May 4. If I were to post it would u be willing to comment on my specific situation. That would be the best help in me understanding what is happening in my water. Need to read the thesis and put together my notes which will take a little time. William
Actually my log is published on another cichlid site but not sure if i can post the location on this site. I;m a little spooked about rules.


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## DJRansome (Oct 29, 2005)

You could send it to shiftyfox via PM.


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## shifty803 (May 17, 2020)

shiftyfox said:


> If I was to start over I would definitely go down the fishless cycle route with ammonia.
> I went with Fluvals bottled bio and was constantly paranoid that I was risking the fishes health throughout the process.
> 
> However throughout the cycling process Ammonia never went over 1ppm and nitrite 0.50ppm
> Would the readings have been much higher without adding the bottled bio at the start - I'm not sure..?


I wanted to add my experience with bottled bacteria this time around, but did not in the original post since it is totally anecdotal and not rigorous by any means.

That being said, I had a similar experience using Tetra Safestart Plus. The nitrite production was totally controlled to the point where I never saw more than trace nitrites during cycling. I dosed ammonia to between 1 and 2 ppm with ammonium chloride and it took 2+ weeks for it to start being used up rapidly.

That time period for ammonia oxidation seems consistent with the findings in these papers. I could speculate that the nitrospira in TSS is the reason I never saw nitrite, which would be fantastic. But honestly, there is no way to actually know if that is a coincidence or not.


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## shifty803 (May 17, 2020)

DJRansome said:


> So fishless cycle with ammonia is still best?


If the tank pH is high enough that unionized ammonia percentage is also high, it seems super risky to do it any other way imho. Nitrite is also potentially a problem that isn't mitigated by low pH.

Nobody in academics has gone out of their way to rigorously study different brands of bacteria in a bottle. Possibly they have some beneficial effect, but it is unclear how and in what circumstances.

Ultimately, seeding the tank with anything from an established tank appears to be the best way, but we already knew that! The thesis also speculates that her home tanks behaved differently than the lab tanks, because she wasn't rigorous about cleanliness and cross-contamination. For example, it is possible AOA can get into a tank from our skin. Or maybe if you have well water instead of municipal water you are better off, etcetera. There are just so many factors.


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## willyg1631 (Feb 18, 2020)

forgot about that. So not allowed right.


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## Fogelhund (Dec 3, 2002)

I think it should be obvious, that the best cycling method, is not to cycle at all, with a new tank. Simply put the filters for the new tank, on an existing tank... easy enough for those of us with Multiple tank syndrome. I do wonder, if people would pay a premium, to purchase a "cycled" filter for a new tank, from a vendor?


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## shiftyfox (Aug 2, 2019)

I'm surprised they don't..I asked a few vendors at the time for some mature media and no one was willing to part with any.

Willy my first tank was cycled with adding fish straight away along with bottled bacteria added over 3 days. Your definitely doing it the correct way. Have you decided on what fish your adding once cycled..?

I remember it seemed to take ages to cycle and could never seem to get the ammonia test tube from a light green to yellow 
At the time I didn't realise that the API master kit tested for both NH3 and NH4 so I'm certain it was showing ammonium


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## Bd79 (Apr 11, 2011)

Thanks for posting this. You make an excellent point. Unfortunately, most people still refer to the "beneficial bacteria" in the filter. In fact, about a year ago I went to a local fish club meeting where the speaker was a salesperson from an aquarium products company, which shall remain nameless. He spent a lot of time on the topic of beneficial bacteria, aquarium cycling, and bottled bacteria, supposedly basing his presentation on the advice from the scientists at his company. He had a lot of very complicated dos and don'ts in terms of water changes and the like, due to the needs of filter bacteria.

Near the end of the presentation, he started taking questions, so I asked him if the fact that archaea were the predominant ammonia oxidizers in aquarium filters changed his advice at all. He asked me where I got that idea from and I told him it came from scientific literature. He asked me to send it to him and promised he would get back to me. I got his email address and sent one of the articles to him, but he never replied, perhaps not surprisingly.


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## DJRansome (Oct 29, 2005)

So we have to say beneficial archaea now? Maybe beneficial organisms would work better?


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## Cosi (Apr 1, 2020)

Interesting. I personally would like to see more applied research on denitrifying bacteria in freshwater aquariums. Not as a replacement for water changes but as a supplement to improve water quality. Its possible that there isnt much interest due to the ease freshwater water changes or the complexity of maintaining denitrifying colonies. Searching the forums I find wildly varying and largely unsupported opinions ranging from XXX media eliminates nitrates to denitrificaiton is a myth. Specific research of interest to me includes:

Can media in a typical canister filter support denitrifying bateria colonies?
Denitrification using only naturally occuring organic matter as proton donors (metabolic products) vs dosing proton donor.
What specific conditions (eg low nitrates, zero oxygen) promote bacterial reduction of sulfates to sulfides?
Bacterial reduction of phosphates (vs nitrates) to phosphorous.

There was an interesting International Space Station experiment with freshwater fish that used a simultaneuos nitrification and denitrification biological filter. Ethanol was used as proton doner.
https://www.sciencedirect.com/science/a ... 241400008X


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## Cosi (Apr 1, 2020)

DJRansome said:


> So we have to say beneficial archaea now? Maybe beneficial organisms would work better?


DJ the correct technical term is apparently prokaryotes which includes both the Bacteria and Archaea domains.


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## DJRansome (Oct 29, 2005)

I can see us eventually getting people to say organisms instead of bacteria, but prokaryotes may be a stretch.


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## Cosi (Apr 1, 2020)

DJRansome said:


> I can see us eventually getting people to say organisms instead of bacteria, but prokaryotes may be a stretch.


Ha! I think prokaryotes rolls off the tongue nicely.


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## Bd79 (Apr 11, 2011)

DJRansome said:


> So we have to say beneficial archaea now? Maybe beneficial organisms would work better?


I prefer beneficial microbes, just to be clear that we are talking about tiny critters, technically speaking.


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## BC in SK (Aug 11, 2012)

DJRansome said:


> Maybe beneficial organisms would work better?


That is way too vague and general. It's just going to confuse. What are you referring to, plants? Algae? Fungi? Nematodes? Arachnids? Shrimp? Fish?


Bd79 said:


> DJRansome said:
> 
> 
> > I prefer beneficial microbes, just to be clear that we are talking about tiny critters, technically speaking.


I agree. 
We are talking about something that is so small we can not see it with our own eyes. At least that part is accurate. 
Though specifically which ones we do not necessarily know. May very well be completely different species, in different tanks.


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## nowheretroupe (9 mo ago)

I have an aquarium too. I'm happy to see that the aquarium owners are responsible for the fish's health and control level of Ph, temperature, ammonia, and nitrites. But, unfortunately, it doesn't apply to our natural water sources. We all must cooperate on this question, isn't it? There are many ways in making the water clearest. I wrote a paper about this, so you can find a lot of useful info here. I think all of us want to drink clear water. Let's do it together!


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