The All-Pharmaceutical Tank (Satire. Please don't do this.)

[Miami Reef]

Yeah, me too. I was looking forwards to a nice clear result. Even the darned replicates aren’t behaving exactly the same way. Could be tough to unravel this mystery.

If you had the opportunity to add one additional pharmaceutical to the treatment tank, what would you choose?

 

[taricha]

A provisional conclusion to be tested is that the pharmaceutical cocktail has minimal effect on algae growth but stops cyanobacteria. I still need to figure out a way to characterize bacteria growth, possibly by the next report.

I didn't expect that but probably should have. I think this was a good early clue....

The control is running nutrient depleted, no PO4 or any inorganic-N detected 7 days after the weekly media change and daily feeding of fish flakes. The treated aquarium is accumulating tenths of ppm nitrite but no PO4.

The fact that in the first two week check, the tanks were both eating almost all of the available nutrients could have predicted that even with this chemical gauntlet there are plenty of growing photosynthetic organisms that can tolerate the chemicals and grow okay. We can suppress the most sensitive types but there will always be other less sensitive sorts available to fill the niche.

Oh wow! I was under the impression that you only dosed it once in the beginning. I did not expect these results at all!

Dosing once every water change is interesting, Flux Rx is sometimes done every 14 days and Algaefix up to every 3 days. Under those usages, both meds can accumulate in the water, and that is likely helpful in suppressing algae. I think the other meds break down quicker.

 

[Miami Reef]

Under those usages, both meds can accumulate in the water, and that is likely helpful in suppressing algae.

Ah, you think the 100% water changes was not allowing the medications to rise in concentretion?

Perhaps the next step is to only do 50% water changes once a week, and dose the same concentrations as usual. Do you think that would make a difference?

 

[Dan_P]

How would you describe diatom prevalence in the two tanks? I feel like the diatoms and cyano are the two easiest targets to go after with chemical killers.

Diatom population substantially diminished and cyanobacteria absent in microscope slide culture for treatments compared to control. Is this kinda what you expected?

 

[Dan_P]

If you had the opportunity to add one additional pharmaceutical to the treatment tank, what would you choose?

I don’t have a candidate. Besides, 4 pharmaceuticals gives many possible combinations. Add concentration and there are a ton of treatments try. I am not sure that the dosages were determined with much data.

 

[Dan_P]

Ah, you think the 100% water changes was not allowing the medications to rise in concentretion?

Perhaps the next step is to only do 50% water changes once a week, and dose the same concentrations as usual. Do you think that would make a difference?

I think generating a dose-response curve would be very interesting. Maybe a 100% water change every day to ensure a constant concentration and at several levels.

 

[taricha]

Diatom population substantially diminished and cyanobacteria absent in microscope slide culture for treatments compared to control. Is this kinda what you expected?

yes, cyano most sensitive to chemiclean, and diatoms the most sensitive class to algaefix.

Ah, you think the 100% water changes was not allowing the medications to rise in concentretion?

The only one I can point to confidently is that algaefix (etc) used in the hobby rises in concentration in the water. So after a routine of usage, there's maybe 5x doses sloshing around in the water. At least that's what I measured in my display tank, test tanks, and a coral system at my LFS. That is the only one that I think what's happening in Dan's test is different from how it would go with hobby usage. Maybe that's true to a lesser extent with fluconazole? but most people don't do a bunch of repeat dosings of flux rx.

 

[Dan_P]

Week 6 Update

Observing the effects of @taricha “NoUg” super algae eliminator (joke) is starting to feel like watching paint dry. Growth of algae in the control and treatment containers (both replicated) seems to be slowing, possibly indicating that the algae biomass is reaching the carrying capacity of these little ecosystems. Variation within replicates is large, which makes drawing any conclusion about effects tough. Here is this period’s observations.

Visually, the big difference is color. The control aquaria are covered in golden growth. Algae biomass, all the stuff on the sides and on the sand, appears to be somewhat larger than in the control. Microscope slide cultures are frustratingly similar with the exception of no apparent cyanobacteria growth on the treatment slides. Slide cultures all have a greater diversity of microorganisms then before.

The growth on the aquarium walls was examined this period. The biofilm on the control wall was thick and resilient like skin whereas the treatment biofilm was thin and fragile that easily broke into pieces. Both showed a complex ecosystem but only the control biofilms contained cyanobacteria. The control biofilms could be described as cyanobacteria biofilms with other organisms living in or on it. The cyanobacteria were the likely reason for the strength and golden color of the biofilm.

The biofilms samples shown below were thick and prevented focus on all organisms at once. The control biofilm on the left is a mat of fine filament cyanobacteria studded with dinoflagellates and interwoven algae filaments. The treatment biofilm on the right consisted of algae fibers ( ) and a matrix of single cell organisms and probably extracellular polymer. Dinoflagellates studded this biofilm but cyanobacteria were not observed.

A provisional conclusion to be tested is that the pharmaceutical cocktail has minimal effect on algae growth but stops cyanobacteria. I still need to figure out a way to characterize bacteria growth, possibly by the next report.

Week 8 Update

The @taricha “NoUg“ continues to disappoint. Take a look at the control and treatment aquaria. I am returning the product for a full refund

At a very high level, both the controls and treatments are equivalent messes with the exception that the controls appear to be more yellow. This is likely from the fine filament cyanobacteria that is growing on the filamentous algae attached to the sand. The color of the walls are undoubtedly from cyanobacteria growth. The white tufts (I refer to them as clear algae) growing on the sand in the treatments appear to be the same species as growing in the controls. The slight coloration of the tufted growth on the right treatment is from what looks to be an infestation of encapsulated dinoflagellates. I find this difference in the treatment replicates interesting and frustrating.

The collection of photos below are from microscope slides placed in each aquaria on day 40 for two weeks. Notice the lack of similarity between the replicates. Clearly, one has to be careful about concluding cause-and-effect relationships from observations of one aquarium.

Overall, cyanobacteria seem to be missing on all slides, while diatoms and dinoflagellates are plentiful. Filamentous algae and single cell algae also populate the microscope slide cultures.

The dosing is scheduled to last two more weeks and I will try to draw some sort of conclusion from the visual data at week 10

 

[Randy Holmes-Farley Verified Community Expert]

Notice the lack of similarity between the replicates. Clearly, one has to be careful about concluding cause-and-effect relationships from observations of one aquarium.

Perhaps that difference is akin to releasing a ball on the very top of a hill. Which way it goes depends strongly on the local starting conditions.

 

[Miami Reef]

Week 8 Update

The @taricha “NoUg“ continues to disappoint. Take a look at the control and treatment aquaria. I am returning the product for a full refund

At a very high level, both the controls and treatments are equivalent messes with the exception that the controls appear to be more yellow. This is likely from the fine filament cyanobacteria that is growing on the filamentous algae attached to the sand. The color of the walls are undoubtedly from cyanobacteria growth. The white tufts (I refer to them as clear algae) growing on the sand in the treatments appear to be the same species as growing in the controls. The slight coloration of the tufted growth on the right treatment is from what looks to be an infestation of encapsulated dinoflagellates. I find this difference in the treatment replicates interesting and frustrating.

The collection of photos below are from microscope slides placed in each aquaria on day 40 for two weeks. Notice the lack of similarity between the replicates. Clearly, one has to be careful about concluding cause-and-effect relationships from observations of one aquarium.

Overall, cyanobacteria seem to be missing on all slides, while diatoms and dinoflagellates are plentiful. Filamentous algae and single cell algae also populate the microscope slide cultures.

The dosing is scheduled to last two more weeks and I will try to draw some sort of conclusion from the visual data at week 10

Are you eventually going to try raising the dose to see what concentrations are needed to eliminate the algae?

 

[Dan_P]

Perhaps that difference is akin to releasing a ball on the very top of a hill. Which way it goes depends strongly on the local starting conditions.

That is the way it seems. Local conditions plus dozens or hundreds of biotic and abiotic interactions. Quite chaotic.

I have observed at least a dozen replicate algae ecosystems go in different directions each time. There is a general “direction” in which they head, but at the species level, there seems to be no pattern beyond a “it’s sorta the same if you squint at the data”.

The possible implication of this notion is that when faced with fixing a nuisance organism situation, the solution might not work fir your aquarium and there is no way to know. I think there are also implications for adding ”beneficial” bacteria to an aquarium. Maybe it’s a crap shoot for whether they grow or not, whether they have an effect ir not.

 

[taricha]

I am returning the product for a full refund

I am certain this was user error. Just read the forums for all of the happy customers who used these products to rid themselves of unsightly algae... for a little while.... until it came back.

A notable thing in this pic to me is the color/texture difference between the controls and treatments.
A yellow-er slimy-er more sheet-like growth pattern dominates the surface of the controls, while the treatments are paler/fluffier.
The slimy sheets I'd label as cyano if I just have to eyeball it.

This could be taken as a suggestion that this kind of approach rather than preventing algae, merely shifts the population so that you grow an approximately equivalent mess of a slightly different community.

I wonder at this late stage - if you applied the cocktail to one of the controls, would it find a bunch of susceptible organisms and thus make everything look "better" temporarily - in a way that it certainly doesn't help with the treatments.

 

[Dan_P]

I am certain this was user error. Just read the forums for all of the happy customers who used these products to rid themselves of unsightly algae... for a little while.... until it came back.

OK, I will order another case

A notable thing in this pic to me is the color/texture difference between the controls and treatments.
A yellow-er slimy-er more sheet-like growth pattern dominates the surface of the controls, while the treatments are paler/fluffier.
The slimy sheets I'd label as cyano if I just have to eyeball it.

You are correct about the slimy appearance of the controls being caused by cyanobacteria. The stuff forms an amazingly tough fiber network.

This could be taken as a suggestion that this kind of approach rather than preventing algae, merely shifts the population so that you grow an approximately equivalent mess of a slightly different community.

Since I am very interested in what nudges ecosystem populations, I might be adding these and other pharmaceuticals to my study.

I wonder at this late stage - if you applied the cocktail to one of the controls, would it find a bunch of susceptible organisms and thus make everything look "better" temporarily - in a way that it certainly doesn't help with the treatments.

I am indeed looking for some ideas on what to do after dosing stops. I could switch control and treatment. We could see how the treatment ecology recovers and how the control improves. I was also thinking of heavy feeding to see what would grow again in the control and treatment aquaria after living under nutrient depletion.

 

[Dan_P]

Week 8 Update

The @taricha “NoUg“ continues to disappoint. Take a look at the control and treatment aquaria. I am returning the product for a full refund

At a very high level, both the controls and treatments are equivalent messes with the exception that the controls appear to be more yellow. This is likely from the fine filament cyanobacteria that is growing on the filamentous algae attached to the sand. The color of the walls are undoubtedly from cyanobacteria growth. The white tufts (I refer to them as clear algae) growing on the sand in the treatments appear to be the same species as growing in the controls. The slight coloration of the tufted growth on the right treatment is from what looks to be an infestation of encapsulated dinoflagellates. I find this difference in the treatment replicates interesting and frustrating.

The collection of photos below are from microscope slides placed in each aquaria on day 40 for two weeks. Notice the lack of similarity between the replicates. Clearly, one has to be careful about concluding cause-and-effect relationships from observations of one aquarium.

Overall, cyanobacteria seem to be missing on all slides, while diatoms and dinoflagellates are plentiful. Filamentous algae and single cell algae also populate the microscope slide cultures.

The dosing is scheduled to last two more weeks and I will try to draw some sort of conclusion from the visual data at week 10

Week 10 Update. Experiment Conclusion

Week ten marks the conclusion of this experiment in which I observed how a cocktail of algicides affect the growth of algae. @taricha proposed a satirical approach to starting an aquarium that treated nuisance alga as it arose with one of several algicides. My version of this idea was to combine all the chemicals and use them continuously for ten weeks in a simple model aquarium.

My aquarium was an acrylic box with a working volume of 0.2 L containing 20 g of clean sand with an established population of nitrifying bacteria (BioSpira). Each of four aquaria was inoculated with 0.2 L aquarium water for 24 hours. Then the aquarium water in two aquaria was replaced with Instant Ocean saltwater (1.026 spg) and served as replicate controls. In the other two aquaria Instant Ocean spiked with Algaefix, Chemiclean, Metroplex and Reef Flux at the recommended dose was added. A weekly 100% water change was performed for ten weeks, the control receiving fresh Instant Ocean and the treatments fresh Instant Ocean spiked with the algicide cocktail. The model aquaria were aerated with humidified air to minimize evaporation and the aquaria were gently swirled on an orbital mixer. 300 PAR light was supplied by a 6300K LED grow light on a 12/12 schedule. A clean microscope slide was placed in each aquarium every two weeks to observe the algae species that were actively settling and growing. Daily feeding consisted of marine fish flakes at a nitrogen input equivalent to 1 ppm nitrate and P input of 0.06 ppm phosphate.

Results

The aquaria were photographed through the front wall about every two weeks and the compiled photographs from day 29 to 68 are presented below. While the appearance of the replicates differ, it is probably safe to say that the overall appearance of the treated aquaria differs from the control primarily in color and to a lesser extent in the amount of visible algae. Growth, i.e., the increase in the visible amount of algae, slows and probably ceases around 54 days in all aquaria, a consequence of constant nutrient addition. Development of the yellow coloration occurs between days 29 and 44 when the demand for nitrogen likely exceeds the input.

Based on a microscopic examination of samples of the visible algae and biofilm on the aquarium walls, the treatment with algicides prevented cyanobacteria growth that otherwise grew well in the control aquaria. This likely accounts for the lighter yellow coloration in the treatment aquaria. Coloration that is observed in the treatment aquaria seems to be from dinoflagellate growth. The near white appearance of treatment 1 coincides with the lack of micro algae growth on the tuft filaments. A slightly colored tuft of algae in treatment 1 was infested with dinoflagellates.

Observation of microscope slide cultures consisted of recording five evenly space photomicrographs in the middle of the slide along its length. Diatoms (Nitzschia) were counted in each photomicrograph and summed for the slide. Thin filament cyanobacteria population for each photomicrograph was assigned a number between 0 (none observed) and 3 (heavy growth) and the estimates averaged for each slide. The trends are shown in the charts below. The trend for diatoms in the control replicates peaked around day 25 and then declined. Treatment replicates trends differed from the control and each other, but still showed a peak followed by a decline in population. The pharmaceutical cocktail seems to have delayed the population peak in treatment 2 and delayed and diminished the peak population in treatment 1. Diatom cells in the treatment aquaria were curved and twisted rather than straight as the cells were in the controls. The late population peak in the treatment aquaria seems to coincide with the increase in visible algae growth. Cyanobacteria also reached a peak at the same time as the diatoms in the control replicates before declining. Cyanobacteria growth was suppressed in the treatments.

Heterotrophic bacteria activity in the water of each aquarium at the end of the experiment was assessed by inoculating an Instant Ocean based medium containing methylene blue with water samples taken just before the weekly water change. The blue color of the medium fades when bacteria consume the oxygen in the media. Larger initial amounts of bacteria result in faster oxygen consumption and quicker color fading (Fading Onset Time). The results of triplicate measurements are presented below and show that the amount of bacteria in the treatment water is lower (longer Fading Onset Time) than in the control. It is not clear whether the difference reflects lower algae growth and less algae exudates, or an inhibition of bacteria growth by the algicides.

Conclusion. The satirical solution to nuisance algae growth as applied in this experiment may not be of any use to an aquarist but the cocktail or one of its components could be useful for studying algae growth in the absence of cyanobacteria. Application of higher doses might have interesting effect on algae and might suppress diatom growth. Testing each component separately at different doses is on the agenda.

 

[taricha]

One of the interesting ideas here @Dan_P is the question of how it is that the treatment tanks overcame the various chemicals to "ugly up" to near the same level as the control after a slow start.
Two ways come to mind: one is that the treatments simply cultured up less susceptible uglies in place of the more common initial uglies.

The second way is that maybe the doses of chemicals got diluted across all the increasing amount of growth and the susceptible common types eventually could grow.

The observation about diatoms being the same common type (nitzschia) in the control and treatment, but with some cell damage/irregularities in the treatment suggests the latter. It seems like the diatoms were initially prevented, but later the growth of other things probably diluted the surface-active algaecide enough that the same diatoms could grow - albeit misshapen. In practice, the concentrations used by dosing every 3 days according to bottle label would be more effective against the diatoms longer term, probably.

The cyano growth might point the other way - it seems like the treatment for cyano (chemi-clean) seems to have essentially excluded filamentous cyano from the community for almost the entirety of the experiment. Instead, the treatmet simply grew other less susceptible things in its place. Both the treatment and control seemed to consume whatever nutrients were available, so plenty of things grew in each to deplete all available resources.

 

[Dan_P]

One of the interesting ideas here @Dan_P is the question of how it is that the treatment tanks overcame the various chemicals to "ugly up" to near the same level as the control after a slow start.
Two ways come to mind: one is that the treatments simply cultured up less susceptible uglies in place of the more common initial uglies.

The second way is that maybe the doses of chemicals got diluted across all the increasing amount of growth and the susceptible common types eventually could grow.

The observation about diatoms being the same common type (nitzschia) in the control and treatment, but with some cell damage/irregularities in the treatment suggests the latter. It seems like the diatoms were initially prevented, but later the growth of other things probably diluted the surface-active algaecide enough that the same diatoms could grow - albeit misshapen. In practice, the concentrations used by dosing every 3 days according to bottle label would be more effective against the diatoms longer term, probably.

The cyano growth might point the other way - it seems like the treatment for cyano (chemi-clean) seems to have essentially excluded filamentous cyano from the community for almost the entirety of the experiment. Instead, the treatmet simply grew other less susceptible things in its place. Both the treatment and control seemed to consume whatever nutrients were available, so plenty of things grew in each to deplete all available resources.

We are arm in arm on this ^^^

I am curious about the cyanobacteria. Is it wiped out or just hunkering down as a cyst or single cells. In part II of this experiment (yawn), all aquaria will get weekly 100% water changes with Instant Ocean and fed as usual.

 

[taricha]

I am curious about the cyanobacteria. Is it wiped out or just hunkering down as a cyst or single cells.

At the risk of extrapolating too far, when I looked at algaefix, dinox and similar, I found that the concentrations or potency of the algaecide needed to stop algae growth were much easier to achieve than those needed to actually kill the algae. That is algaestatic concentrations/potency much lower than algaecidal ones. Fluconazole might be the same way, both of those seem to the chemicals that stick around for some time.
The macrolide antibiotics used against cyano, I think are much shorter lived in the system, so they might actually achieve cell death instead of just halting growth while their concentration is high.

 

[Dan_P]

At the risk of extrapolating too far, when I looked at algaefix, dinox and similar, I found that the concentrations or potency of the algaecide needed to stop algae growth were much easier to achieve than those needed to actually kill the algae. That is algaestatic concentrations/potency much lower than algaecidal ones. Fluconazole might be the same way, both of those seem to the chemicals that stick around for some time.
The macrolide antibiotics used against cyano, I think are much shorter lived in the system, so they might actually achieve cell death instead of just halting growth while their concentration is high.

Thanks for clarifying. I had not gotten this far in thinking about these chemicals in terms of -staitic v -cide. Should save me some work

 

[Randy Holmes-Farley Verified Community Expert]

When I was working on a team developing antimicrobial polymers, we would sometimes test against a panel of dozens of microbes. Some were strains of the same species, and between strains there could still be substantial differences in susceptibility.

Thus, seeing some particular organism survive does not necessarily mean others of even that same species necessarily survived.

Thus, surviving diatom species may be a subset of those that were originally there, rather than a whole species that are less susceptible.

 

[Dan_P]

When I was working on a team developing antimicrobial polymers, we would sometimes test against a panel of dozens of microbes. Some were strains of the same species, and between strains there could still be substantial differences in susceptibility.

Thus, seeing some particular organism survive does not necessarily mean others of even that same species necessarily survived.

Thus, surviving diatom species may be a subset of those that were originally there, rather than a whole species that are less susceptible.

This could make the “number needed to treat” in the hobby rather large for algicides. The strains of nuisance algae being passed around the reefing community might have been already narrowed down to the very resistant strains.