The Science Of Bottled Bacteria For "cycling" Aquaria


Fish Botherer
Dec 8, 2012
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Many beginners to the world of fishkeeping can be grouped into one of two categories; those who research beforehand and embark on a fishless cycle and those who buy fish first and find themselves deliberately or unwittingly in a fish-in cycle.  Either way, a beginner or experienced aquarist alike needs access to quality information about the fastest way to conduct a cycle.
The majority of online guides to fishless cycling will generally advise the reader to use some combination of household ammonia, fish food, raw shrimp and a generous amount of patience.  For a fish-in cycle the watchword is water changes.  In both cases, bacterial accelerators are nearly universally dismissed as pointless, if they are mentionned at all.
What if there really was a product that can accelerate the cycle time of a new aquarium, protect fish from the toxic build-up of waste chemicals, eliminate new tank syndrome?  Then it should surely deserve a mention in all good advisory guides to cycling.
Statement of Purpose
My goal here is to uncover the truth and expose the validity of the science behind Dr. Tim's One and Only nitrifying bacterial product.  If the product doesn't work or has serious limitations then it would be helpful to firmly quantify them and give it a thorough debunking.  If on the other hand it can be shown to work as advertised then it must be taken seriously as a valuable tool in the aquarist's arsenal.
I would therefore - having gained permission from the moderators - like to ensure that a place remains open where the sensible, scientific analysis of Dr Tim's products can continue for anyone interested.  I would like to remind any participants of the Scientific Section Rules and that personal opinions or unevidenced claims are not relevant in any discussion.  Finally, it is not my intention to open old wounds or stir the proverbial, so I would appeal to any readers to consider this a fresh start, put any past differences aside and advance in the pursuit of better knowledge for all.
My position
To clarify my position, I currently favour the scientific evidence that Dr Tim's One and Only has a sound basis in theory to work as advertised.  I am less convinced by the empirical evidence and my mind is open to be persuaded by well-supported logical arguments from all sides.  I have purchased the product myself and am currently using it, but do not yet know the results so I can offer no personal experience at this time.
I take no credit for the work done in researching this topic.  All this information has been previously researched and discussed by TwoTankAmin and other members of this forum.  All credit for hard work goes to them.  I will merely attempt to summarize the information, evidence and discussion that has been presented so far into a more concise format for ease of reference.
What does One & Only claim to do?
1. Eliminate new tank syndrome (1)
2. Instantly create a bio filter (1)
3. Remove ammonia and nitrite (1)
4. No need to wait (i.e. fish can be added immediately) (1)
5. Ammonia & nitrite should both be dropping to zero within 24 hours after 5-7 days of using the product (2)
(1) Dr Tim's Aquatics One and Only Live Nitrifying Bacteria
(2) Dr Tim's Aquatics Fishless Cycling Guide

Does it contain the right type of bacteria?
The product is based on the results of research conducted by Dr Hovanec and his colleagues while working as employees for Marineland.  Three research papers were published which identified specific species of Nitrosomonas and Nitrospira as the main autotrophic nitrifiying bacteria in aquariums. (3,4,5)
The papers were peer reviewed and have since been verified by independent research. (6,7)  A significant period of time has elapsed since the publishing of all three papers during which none of them have been refuted or supplanted by new research.
As a result of this research Marineland produced Bio-Spira in 2002, a year after the last paper was published. (8)  Marineland was subsequently acquired by Tetra and Dr Hovanec set up his independent company Dr Tim's Aquatics.  BioSpira ceased production and was replaced by Tetra SafeStart and Dr Tim's One & Only respectively.
The evidence seems conclusive that the strains of bacteria in a bottle of One & Only (and Tetra SafeStart) are suitable strains for long-term establishment of filters in home aquaria.  Since Dr Hovanec (and certain of his colleagues) and Tetra hold the patents to these strains of bacteria, they cannot be found in any other commericially available products.
(3) Comparative analysis of nitrifying bacteria associated with freshwater and marine aquaria. (1996) by Hovanec, DeLong
(4) Nitrospira-Like Bacteria Associated with Nitrite Oxidation in Freshwater Aquaria (1998) by Hovanec, Taylor, Blakis, Delong
(5) Identification of Bacteria Responsible for Ammonia Oxidation in Freshwater Aquaria (2001) by Burrell, Phalen, Hovanec

(6) Nitrosomonas Nm143-like ammonia oxidizers and Nitrospira marina-like nitrite oxidizers dominate the nitrifier community in a marine aquaculture biofilm
(7) Identification and Activities In Situ of Nitrosospiraand Nitrospira spp. as Dominant Populations in a Nitrifying Fluidized Bed Reactor
(8) BioSpira Timeline
What about Archaea?
There is some evidence that AOA (ammonia oxidising archaea) may be dominant over AOB (bacteria) in large saltwater settings such as public aquaria and open ocean.  However conditions in small freshwater aquaria are more favourible to AOB. (9,10,11) There are no findings of nitrite oxidising archaea anywhere.
In conclusion, there is currently insufficient evidence that archaea play a significant role in home aquaria.
(9) Nitrosopumilus maritimus genome reveals unique mechanisms for nitrification and autotrophy in globally distributed marine crenarchaea
(10) Differential photoinhibition of bacterial and archaeal ammonia oxidation
(11) Nitrosomonas Nm143-like ammonia oxidizers and Nitrospira marina-like nitrite oxidizers dominate the nitrifier community in a marine aquaculture biofilm

Can the bacteria survive in a bottle?
1. Temperature
The bacteria will be completely killed if the bottle is frozen or if temperatures exceed 95°F (35°C).  The bacteria will remain viable for up to 6 months at room temperature or for 1 year if kept refrigerated. (12)

2. Food
Nitrifying bacteria have mechanisms to deal with periods of starvation including a protective shield called EPS (extracellular polymeric substances).  (12)

3. Oxygen
There is evidence that bacteria will decay more slowly in anoxic conditions although the reasons for this is not understood.  (13, pg 12)

4. Reactivation
Reactivation of bacterial cells that have been starved and preserved above zero temperatures will occur naturally after a short recovery time (13, pg 15)
(12) Dr Tim's Aquatics One and Only Nitrifying Bacteria FAQs
(13) Bacterial Cultivation and collection strategy
Anecdotal Evidence
Bio-Spira was withdrawn from sale as the success rate was not sufficiently high to make the product commercially viable.  This was at least partially due to the exacting demands of the product to be temperature controlled.
Several public aquaria have used large quantities of Dr Tim's One and Only to cycle their tanks including Georgia Aquarium, Arizona SeaLife Aquarium, SeaLife Aquarium at Legoland California and SeaLife London Aquarium. (14)
There are many examples across the internet of people who have both successfully used Dr Tim's One & Only and those who found it to have no effect. 
At least some of the reported malfunctions may be explained by user failure to follow instructions.  There are limited instructions supplied on the bottle and personally I have found the instructions on the website to be unclear.
Susceptibility of the product to temperature extremes during shipping may also account for some malfunctions.
There are also several reported cases that the API ammonia test kit, a commonly used home aquaria testing kit, produces unexpected results when used in combination with Dr Tims ammonium chloride and One & Only, leading some users to overdose their ammonia concentration.
(14) Dr Tim's Aquatics Testimonials
Experimental Evidence
The only experimental evidence that has been presented on this site to date is in the patent application for the strain of AOB developed by Hovanec et al.  The patent was granted in 2005.  It contains details of experiments which show that Hovanec's bacteria are capable of cycling a tank in half the time taken by competing commericial products.  The experiment contains the names of rival products and is not known to have been contested by the manufacturers of those products. (15, Bacterial Additive Test VII, page 26)
(15) Patent Specification for Ammonia Oxiding Bacteria, by Hovanec and Burrell
In closing
I believe that this topic has been well covered on the subject of bacteria species.  Going forward I should like to focus on weaker areas such as methods used to sustain the bacteria in a bottle for up to year and on the new experimental evidence.
I welcome any discussion, additional information, correction of mistakes or challenges of statements made.  I am always keen to learn!  Thanks for reading.

Excellent start to a scientific discussion! Thank you for your awesomeness. The hounds of hell will not be allowed to descend upon this thread! Let us all so declare in the name of science! (a little melodrama never hurt anyone).
Whenever I discuss this topic I keep a few key things in mind. These are the things that I want to know about a product.
  1. Is it the right kind of bacteria?
  2. How is it kept alive?
  3. How can the aquarist trust the answer given by the retailer to #1 & #2
  4. How long does it live once out of the bottle and in the aquarium?
  5. How well does it reproduce in the new environment into which it has been placed now absent the specially created media that was in the bottle?
  6. Why pay for something nature provides for free?
While #6 isn't a part of the scientific aspect of a product it is one I constantly ask myself so I include it here.
#2 goes toward your closing statement and is what I consider one of the weaker areas of any such product. How is the bacteria fed and how is the waste processed are of paramount importance as is the media in which the bacteria are suspended. Does it degrade over time, if not does this material pose any threat to aquaria due to its longevity?
I see no reason that such a product isn't possible as the theory behind it is sound. So it isn't so much a question of CAN bacteria in a bottle work but rather HOW can it work reliably.
It will take this weekend for me to go over all your links but for now the above frames my interest in this topic and I personally feel it fits into fishkeeping.
It would be great if someone from this forum could just purchase a bottle of this stuff and put it to the test with a fresh tank and fresh filter, ammonia and test kit.
I get paid my bonus at the end of March and have a choice of spare tanks and filters so may put it to the test myself.
That's a good start  for sure.
I feel like for a true experiment we would need to include some longevity study along with it. We would need say 12 bottles from the same lot then use one bottle on a new tank each month to see if the product was as effective at month 12 as it was at month 1 because I think the main issue that is often raised is the shelf-life vs viability.
Livewire - I'm currently using it and my results are being photologged in this thread  - Daize's log
However I would not set too much stock in anecdotal evidence of individual cases, even my own!  My tank was unusually stubborn to start a cycle before I started using the formula, so for all I know I might have inhibitors in my tank that are killing off the bacteria.  There are plenty of other reasons why mileage may vary from person to person, which is why I prefer to analyse on the basis of scientific evidence.  A controlled experiment as tcamos suggests would be interesting, though.
Tcamos - thank you.  Your questions 1 & 3 were the intensive subject of the previous thread and I think these are covered by Hovanec's research, it's peer review and lack of contradictory research, his employment status at the time of the research and the fact that the relevant products were not developed until after all the research had been completed.  Your other questions I would like to know the answer to but I will have a stab at number 6 myself!  For many people, nature can indeed provide the service for free but there are some reasons why others may want to purchase an accelerator.  Reasons I can think of:
1) Difficulties establishing a cycle without mature media (this is my own experience and there are other examples)
2) Desire for expediency of cycle (e.g. during a fish-in situation)
3) If introducing pre-established mature media would risk introducing unwanted diseases (e.g. for a tank intended for wild-caught fish etc).
Now I would like to ask a question of my own regarding the patent experiment in reference 15.  The patent specification presumably must have been written before One & Only existed, so the experiment was comparing Hovanec's cultured bacteria to bottled specimens of rival products.  Could the freshness of Hovanec's own bacteria have contributed to the apparent effectiveness of his bacteria compared to pre-bottled bacteria, thus biasing the experiment?
Your three answers take care of my #6 very well. Thank you for that.
Since your question begins with "Could" we're addressing probability not possibility and so I would answer "Certainly it could". Any variance in the viability of bacteria used in the comparison would color the outcome. For the results to be valid the parameters being compared must be equal otherwise we aren't comparing like:like while the results may still be interesting or even enough to say this unlike thing works better than this unlike thing it isn't necessarily enough to act as a condemnation of the other bottled bacteria.
Wow...that was wordy...
There are a few minor errors in the information presented above.
The product is based on the results of research conducted by Dr Hovanec and his colleagues while working as employees for Marineland.
There are 3 studies with various authors, some of whom are on more than one paper and some who are not. However, not all of the names on each of the studies was employed by Marineland. For example, the Nitrite paper has 4 authors and only Dr H. was affiliated with Marineland and the other three were not. However all four were members of the  Department of Ecology, Evolution and Marine Biology, University of California, Santa Barbara, Santa Barbara, California 93106.
Since Dr Hovanec (and certain of his colleagues) and Tetra hold the patents to these strains of bacteria, they cannot be found in any other commericially available products.
Other companies can use the bacteria as long as they are permitted to by the patent holders. I can not say if any other companies have made arrangements to do so, only that they could if they can reach terms with the patent holders.
BioSpira ceased production and was replaced by Tetra SafeStart and Dr Tim's One & Only respectively.
BioSpira is still being produced and sold by Instant Ocean. It is now only a salt water starter- there is no fresh water BioSpira.
Instant Ocean Bio-Spira Start Up Nitrifying Bacteria Aquarium Additive

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Makes marine aquariums fish-safe instantly
  • Prevents new tank syndrome
  • Live nitrifying bacteria
  • Contains patented nitrifying bacteria proven to reduce fish loss due to ammonia and nitrite toxicity
The bacteria will be completely killed if the bottle is frozen or if temperatures exceed 95°F (35°C).
Actually this is not the proper quote from Dr. Hovanec's site. The above quote should be:
If the liquid in the bottle freezes, the nitrifying bacteria are killed. It doesn’t matter the brand – freezing kills the nitrifying bacteria. High temperatures also can kill or damage nitrifying bacteria. If the bottle is exposed to 110°F for a day or so, the bacteria can be killed. Prolonged exposure to temperatures over 95°F drastically reduces the shelf life of nitrifying bacteria.
Finally, on the topic of the Archaea there is a great deal more research now available. What I have found is that the assumptions made in many studies about the potential role of AOA in nitrification has been based on gene counts, especially for amo. It is turning  out that this is likely not a reliable method for determining what is actually doing the oxidizing. If you really want to delve into this topic do a Google Scholar search using the term "AOA vs AOB." Then sort the results by date to see the most recent research first. I am also happy to PM my links on the subject to anyone who wants them.
The only experimental evidence that has been presented on this site to date is in the patent application for the strain of AOB developed by Hovanec et al.
I have posted links to a couple of other studies/products, here is one:
I know there is one more I posted which compared several commercial starters, I can not find my link and can't find the post where I also had it. It was actually a part of another study but not the primary focus. Also, the nitrite paper by Hovanec et. al. used Hagen's Cycle in some tanks. The conclusion was it had the wrong bacteria- i.e. Nitrobacter. Basically in the end, the Nitrobacter was not found in the tanks but Notrospira was.
Look at the list of questions posed by tcamos. I have posted the answers to every one of those questions and included links to many studies which answer all of those questions. Yet here I am seeing the same questions being asked yet again. So let me just put up all the links and let those who want to read do so:
Growth at Low Ammonium Concentrations and Starvation Response as Potential Factors Involved in Niche Differentiation among Ammonia-Oxidizing Bacteria
Cell density-regulated recovery of starved biofilm populations of ammonia-oxidizing bacteria.
Strategies of aerobic ammonia-oxidizing bacteria for coping with nutrient and oxygen fluctuations
Strategies of aerobic ammonia-oxidizing bacteria for coping with nutrient and oxygen fuctuations
Influence of Starvation on Potential Ammonia-Oxidizing Activity and amoA mRNA Levels of Nitrosospira briensis
FOne of the more interesting reads deals with soil bacteria, some of which is still viable after being stored for over 90 years:
In a preliminary study, to establish if dried soils can provide a historical record of bacterial communities, samples from the Broadbalk soil archive dating back to 1868 were investigated and plots treated with either farmyard manure (FYM) or inorganic fertilizer (NPK) were compared. As anticipated, the processes of air-drying and milling
greatly reduced bacterial viability whilst DNA yields declined less and may be preserved by desiccation. A higher proportion of culturable bacteria survived the archiving process in the FYM soil, possibly protected by the increased soil organic matter. The majority of surviving bacteria were firmicutes, whether collected in 2003 or in 1914, but a wide range of genera was detected in DNA extracted from the samples using PCR and DGGE of 16S rRNA genes.
As to question 6: Why pay for something nature provides for free? I have answered that in posts on this site. I used my own personal experiences to illustrate. It involves the acquisition of recently caught and imported P Altum angels. Because these fish live in acid waters with very little hostile bacteria, their immune systems are somewhat repressed and they virtually have no resistance to the bacteria in most established tanks. Putting Altums into a well established tank is basically a death sentence. So how can one get a tank cycled without all the potentially dangerous bacteria? Well you start with a bottled bacteria (one that actually works) and cycle with that. Coupled with ro water and UV sterilization (turned off while the bacteria is being added and for a time thereafter), the bottled bacteria lets you cycle without "contaminating" the water. This applies to any of the acid water fish that folks may keep and which is freshly imported.
I should have started my post above by saying. Nice work, daize.
I am a nudge about making sure statements presented as fact are accurate. I really hate when I do misstate something and I try to point it out when others do it. So my gut instinct was to be the editor instead of somebody happy to see a decent organization of the information, even though it was not 100% perfect in my eyes. Considering I hold a black belt in the ancient art of Tai Po, I am not one to talk about perfection.
Again, sorry daize.
I haven't had time to read all the links yet, would you mind a quick summary of how the inaccurate information might affect the science? Also, for clarity of presentation would you mind listing my questions with the answer under it? I think that would really help organize the data since there is so much of it so that new comers to the topic and have it in a simple Q&A format.
I can see us getting far afield if we don't funnel each new post in some way.
Perhaps a good place to start is to define all the relevant questions. Are there any we need to answer besides the one's I asked?
A question that I would like answered is what are the differences in bacterial storage methods between Dr Tim's One and Only, Tetra's SafeStart and Instant Ocean's BioSpira (allowing for differences due to saltwater & freshwater).  Assuming that they all contain the same strains of bacteria then can we treat them all as one and the same?  Otherwise any differences in the bottling process might indicate a difference in performance and viability.
I haven't yet checked to see if any of these companies publish their methods - I assume that Dr Hovanec does somewhere but I've not seen it yet.  I will have a look later unless anyone has a link immediately to hand? ;)
TTA thanks for your information and links.  I can try to read through them all and provide a short summary later, they all sound like exactly the kind of information I was hoping for.  It must be frustrating to keep posting the same things but I'm hoping if we can analyse all the information in one place you may not need to again!  No need to apologise, it's good to have the right information and as I said before I welcome being corrected!
  1. Is it the right kind of bacteria?
  2. How is it kept alive?
  3. How can the aquarist trust the answer given by the retailer to #1 & #2
  4. How long does it live once out of the bottle and in the aquarium?
  5. How well does it reproduce in the new environment into which it has been placed now absent the specially created media that was in the bottle?
  6. What are the differences in bacterial storage methods between various products?
    Dr Tim's One and Only
  7. Tetra's SafeStart
  8. Instant Ocean's BioSpira

[*]Do the above products contain the same kind of bacteria as each other?
Note: I left out my original #6 because I realized it's more of a philosophical question rather than a scientific one so doesn't belong in this discussion. I also edited the thread title because the topic shouldn't be just one brand name but rather a comparison of various products to be fair the old title skewed the thread.
1.Is it the right kind of bacteria?
& 7.Do the above products contain the same kind of bacteria as each other?
The answer to this can be found in references 4 & 5 in my original post.  The answer is Yes :)
I think we can assume that those three products do contain the same strains of bacteria as they were all derived from the same research.

2.How is it kept alive?
From his product FAQ (ref 12), Dr Hovanec states that the AOB and NOB are grown together as a microbial community of bacteria on a microstructure, not as “free” cells in water, which helps the bacteria stay active for a longer time period.  I believe that he holds a patented method of bacteria storage, which I would like to see more information about - I've been unable to track it down (still hoping TTA might have a link for this).
3.How can the aquarist trust the answer given by the retailer to #1 & #2
Peer review of work, independent verification and lack of contradictory research.  Personally I'm satisfied that all these criteria have been met.

4.How long does it live once out of the bottle and in the aquarium?
I think the question needs to be not how long it lives but whether it can resume nitrification activity after a period of prolonged dormancy and starvation.  The answer to this is in the links TTA provided, which I will try to summarize very briefly.
Growth at Low Ammonium Concentrations and Starvation Response as Potential Factors Involved in Niche Differentiation among Ammonia-Oxidizing Bacteria
This study involved comparing two strains of bacteria for growth at low levels of ammonia and their relative response after starvation for periods of up to 10 weeks.  The conclusion is that the strain of bacteria which was out-competed for ammonia (Nitrosomonas europa) was specially adapted to survive periods of ammonia deprivation and was therefore more adept at reactivating when a fresh source of ammonia became available.  This confirms that bacteria can reactivate after periods of starvation and that reactivation time is dependent upon species.
Cell density-regulated recovery of starved biofilm populations of ammonia-oxidizing bacteria
This study concludes that there is no delay in recovery of starved nitrifiers, as long as they exist in a biofilm rather than free cell suspension.  This might support Dr Hovanec's method of cultivating microbial communities.
Strategies of aerobic ammonia-oxidizing bacteria for coping with nutrient and oxygen fluctuations
More of the same and also suggests that AOB can survive periods of oxygen deprivation by switching from nitrifying to denitrifying activitity, although it seems this still is not clearly understood.
Influence of Starvation on Potential Ammonia-Oxidizing Activity and amoA mRNA Levels of Nitrosospira briensis
Another study demonstrating the diverse lag times of different strains of AOB when recovering from varying periods of starvation.
(I'm sure that TTA will correct me if I have misunderstood any of these articles or missed anything pertinent!)
5.How well does it reproduce in the new environment into which it has been placed now absent the specially created media that was in the bottle?
Without knowing exactly what that media is and how it is affected upon introduction to the aquarium, I think that this question might perhaps be best answered by experimental evidence, i.e. the presence of nitrifying activity after introduction of the contents into an aquarium.  Again I will try to summarize the information from TTA's link:
Elevated salinity selects for a less diverse ammonia-oxidizing population in aquarium biofilters
ABIL was used to seed biological filters with bacteria during an experiment on saltwater AOB.  The ammonia levels were shown to drop immediately, suggesting that the ABIL product contained live ammonia oxidising bacteria which immediately began nitrification upon introduction to the test environment.  Thus dispelling the general myth that bacteria-in-a-bottle products never work.
I can't find much information about ABIL but presumably it has no connection with Dr Hovanec's research so I find it interesting that there is more than one strain of AOB that can claim an immediate reduction of ammonia levels.  I thought perhaps it might be one of those species adapted for much higher levels of ammonia but the ammonia levels in this experiment were around the same as you'd find in a normal fishless cycle (5 mg/L NH4-N).  So I wonder, what would happen in a contest of survival between ABIL and One & Only?  If you put both into a home aquarium environment then presumably only one strain of bacteria would eventually become dominant, yet both are independantly able to cycle a tank.  This suggests to me that the contents of a bottle o' bacteria may not be the stuff that eventually colonises the biomedia in a mature aquarium, even if it is the stuff that initially cycles your tank and makes it safe.  But now I am hypothesizing and digressing rapidly, so I will stop :)
daize- I am going to point you in a tangential but very relevant direction. In fact, you touched on it earlier. And that is the biofilm in which the bacteria live. You noted it by another term for it, EPS.
Extracellular polymeric substances or EPS are biosynthetic polymers from prokaryotic (bacteria, archaea) and eukaryotic (algae, fungi) microorganisms, which either form (loose or tight) slimes around the microbial cells or excreted as discrete gels to the surrounding environment. Typically, EPS are heterogeneous mixtures of polysaccharides, proteins, nucleic acids, lipids and other polymeric compounds. The highly diverse chemical composition of EPS is a result of the different processes related to their production and their immediate environment: active microbial secretion, shedding of cell surface materials, cell lysis and adsorption from the environment (Wingender et al., 1999).

EPS are often associated with the formation of biofilms and microbial aggregates. In biofilm systems, they are mainly responsible for binding cells and other particulate  materials together and to the solid-liquid interface (Characklis and Wilderer, 1989).
For a true understanding of how and why the bacteria can survive starvation and drying, why Dr. H. cultures and packs them as he does, why they can resists the effects of chlorine, chloramine and antibiotics, one needs to look at biofilms. The answers are found there. There is no method of storage other than how the bacteria are set up before they go into the bottle. There is nothing one can add that will extend the lifespan etc. Remember, the bacteria need ammonia/nitrite, oxygen and carbon to survive. Even if one could add in ammonia and nitrite and a carbonate, how much oxygen could be contained? Also, since we know both high levels of free ammonia or nitrite (nitrous acid is the actual culprit) can inhibit or even kill the bacteria, how much of either could be added when its packaged?
If the bacteria are ensconced in a biofilm on a solid surface and in good shape, the removal of food or oxygen is detected and they respond by going "dormant." I would be surprised if there is anything in the bottle besides the bacteria and the medium it is on and some ro water. The label does say it is 100% natural.
And there is more to the survival issue in terms of bacteria in a bottle. if one reads Dr. Hovanec's site carefully you discover a few more interesting statements. I was a bit confused by what he was saying about bacteria in general vs those in his nitrifier product. So I wrote and asked him. The upshot is there is a difference between the product not being of great help to the user vs its being dead and totally useless in 6 months to a year.
Also, in your articles on fishless cycling and bacteria living in bottle you state:
"The only way a bacterial cell truly ‘dies’ is if the cell wall breaks apart (called cell lyses) and the cell contents spill out or if the cell in poisoned by some
substance that penetrates the cell wall and kills the cell."  
However, you also indicate there is a maximum useful time that the bacteria can survivie in a bottle:
"They can live in a bottle but under optimal conditions the time period is about one year. The nitrifying bacteria don’t die in the bottle, their activity level
drops and eventually it becomes so low that there is little measurable positive effect when they are poured into the aquarium water. "
I am again confused. Especially in light of your statement which says:
"Nitrifiers belong to a very old line of bacteria (millions of years) and they have developed ways to cope with very long periods of “drought."
This would seem to indicate that they can "shut down" for extended periods and come back to "life" once better conditions are present. So why wouldn't the bacteria in the bottle also be able to "revive" after many years as well?
Again this is semantics. The cell may be alive but it is not useful to use in the sense that it cannot rapidly (in hours/days) convert ammonia to nitrite.
Bacteria in a bottle can be revived and even an old bottle of bacteria is better than no bacteria (as long as they were the right bacteria in the first place). But people expect things in different time periods.

The reason for the "old wives tale" that bacteria in a bottle don't work is due more to the fact that the bacteria in the bottle were the WRONG bacteria
in the first place so they didn't even work before placed in the bottle.

But for the cells to stay active they need to be fed and this cannot happen in a bottle.  So once placed in the bottle the bacteria activity slowly fades -
they are not dead but they are not real active

hope that makes sense
This is part of the reason behind the 6 months or 12 if refrigerated statement regarding the bacteria once in the bottle. If you look into the links above for how the bacteria react to being starved and how fast they revive, it shows why the age of the bottle will determine, to some extent, how rapidly its contents work. But I do not think we are talking a material differences relative to a normal fishless cycle. Fresh bacteria put into a good environment can mean one might cycle in 5 days. Bacteria in the bottle for lets say 3 months and/or a tank not quite as perfect, and it might be 7 days. While for older bottles and/or even less ideal tank conditions and it may run over to 9 days. This all assumes that bacteria was not frozen nor exposed to too much heat along the way.
Under all of this science is still the need for the information to have practical application for the average fish keeper. But for one to be able to write a simple, easy to understand guide to cycling, the author really needs to understand the science in order to provide the proper information. If you don't know why, when or how much ammonia can be a bad thing, how can you tell a newbie the best way to be cycling a tank or when to do a water change during a fish in cycle?
Or what about when somebody makes the oft seen post that goes something like this: "All (or almost all) the good bacteria are in your filter. The amount in the gravel or on the decor (plants/rocks/wood) is minimal." Is this an accurate statement and why or why not? When you pour a bacterial additive into your tank, where does it end up? And wouldn't you know it, photosensitivity comes into play- oops more science. Aren't virtually all of the filters you have seen made with a non-transparent (i.e. not clear) material?
It takes a while to discover the gestalt of aquarium nitrifying bacteria.
I see.  So there are only two main criteria that determine whether a bottled bacteria product will work.
(1) The bottle have to be suitable for the conditions in a home aquarium (I know there are exceptions such as extremely low pH tanks, but my point is mainly about the products that contain strains of bacteria that prefer high-ammonia conditions such as wastewater treatment)
(2) The bacteria have to be cultivated into a strong, healthy biofilm before bottling so that they can enter a dormant state during starvation.
So we know there are only a few products that can be shown to have the 'right' bacteria.  Dr Tim's, Tetra and Instant Ocean were all developed from Dr Hovanec's experiments which are still the most up-to-date accepted research.  But there may also be others, such as ABIL which has been shown to have viable AOB but we don't know what strain they are.
Then the success of the product is very much dependent on the cultivation method of the bacteria before they are bottled, as far as I'm aware only Dr Tim provides much information about this.
We know that bacteria can survive without food in a bottle and we know they can reactivate when given a new food source.  I'm less clear on how they survive without oxygen for so long.  Do they simply stop metabolising during dormancy, or do they switch to a denitrifying metabolism? 
We also know that different strains of bacteria can survive bottled for different periods.  The information I've seen (e.g ref 13) suggests that most bacteria can survive for only a few weeks or months.  However Dr Tim's One and Only has a shelf life of 6-12 months, that's a very long time for live bacteria.  Is it just happy coincidence that the bacteria which Dr Hovanec identified as being the dominant strain in freshwater aquaria, also happen to be extremely good at surviving long periods of dormancy?
tcamos are there any questions which you feel are outstanding from your list?
daize they all survive for more than a few weeks. The issues are more one of how many survive and then how long it takes to recover any ability to oxidize and then to recover to the same level as before they went dormant.
It is not only being well established in a biofilm that is important, it is also important that they be well fed and healthy when ammonia and/or oxygen are greatly reduced. When they sense the lack of food or O, they go dormant and wait for better times to "wake up."
There is some indication that the AOB may be able to change what they do in order to survive- its in one of the studies I posted.
It has been reported that nitrifiers are able to survive under anaerobic conditions, for example in fish-pond sediments (Diab et al., 1992) and in the anaerobic hypolimnion of wastewater reservoirs (Abeliovich, 1987). Diab et al. (1992) suggested that nitrifying bacteria survive anaerobic conditions either by switching their metabolism to a very low rate resulting in a state of resting cells or by switching from a nitrifying to a denitrifying activity.

In other studies, Nitrosomonas europaea was found to be capable of nitrite denitrification with molecular hydrogen, hydroxylamine or organic matter (pyruvate, formate) as
electron donors, resulting in the production of N2O and N2 (Ritchie & Nicholas, 1972; Abeliovich & Vonshak, 1992; Stüven et al., 1992; Bock et al., 1995). Although the denitrification genes nirK and norB, encoding for the nitrite reductase (Nir) and nitric oxide reductase (Nor) enzyme respectively, have been identified in the genome of Nitrosomonas europaea (Chain et al., 2003), the mechanism of denitrification by AOB has yet to be unravelled and its role is still a point of discussion. It has been suggested that this AOB denitrification activity is a protection mechanism against the negative effects of high nitrite concentrations (Poth & Focht, 1985; Stein & Arp, 1998b). Alternatively, it has been recognized as a process of high importance for anaerobic growth (Poth & Focht, 1985; Bock et al., 1995; Schmidt et al., 2001) as well as for the supply of NO necessary for ammonia oxidation (Schmidt et al., 2004a, c). There are no studies that support the hypothesis that nitrifier dentrification is a strategy to withstand an anaerobic environment.

Alternatively, under oxygen-limited or anoxic conditions, ammonium could act as an electron donor that is oxidized with nitrite instead of oxygen as electron acceptor (Bock et al., 1995; Philips et al., 2002). The first evidence for anaerobic ammonia oxidation coupled to cell growth by Nitrosomonas eutropha was published by Schmidt & Bock (1997). In this reaction, molecular oxygen is replaced by nitrogen dioxide or nitrogen tetroxide (Schmidt & Bock, 1998). This anoxic metabolism by ammonia oxidizers has recently been reviewed in detail and will not be described here (Schmidt et al., 2002). Besides its role in the coexistence/competition between ammonia oxidizers and anaerobic ammonia-oxidizing Planctomycetes (anammox bacteria) in oxygen-limited environments (Schmidt et al., 2002), this metabolic feature might be pivotal for the survival and maintenance of AOB during oxygen-limited or anoxic periods.
From Influence of Starvation on Potential Ammonia-Oxidizing Activity and amoA mRNA Levels of Nitrosospira briensis

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