All-in-One Wonderbacteria: PNS Probio

AfishionadoBy Afishionado 3 weeks agoNo Comments

I’m going to make a bold statement. In my personal opinion, bacteria are one of the most important things in our aquariums.

Bacteria??

Ok, hear me out.

For starters, in order to have fish in our own personal slice of ocean, we need strains of bacteria and for them to thrive in our closed system in order to properly jump start the nitrifying cycle so that the fish won’t suffer and perish from the ammonia burns on their gills. This is saltwater aquarium basics 101 right? This is a huge deal and something that everyone has to go through in our wonderful and diverse hobby.

Are you with me so far?                                                              (Picture Credit : pinkyfilters.com)

 

But once that initial cycle is established, and the fish are doing well and everything seems to be getting settled, thoughts of bacteria or the importance of bacteria often takes a back seat in our mind and it seldomly ever gets talked about. And most of us are ok with that.

Is this it though? Not really.

How many times have you heard that vast biodiversity is so important in our hobby? If you have heard that, understand and agree with it, and know that bacteria are in the same category. Having all sorts of different beneficial microbes in our aquarium is something that we should strive to do. We are trying to mimic nature, after all.

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Going back a bit, does completion of the nitrogen cycle mean that the significance and the importance of the bacteria has diminished? Not in the slightest bit. With all the new animals that are constantly being introduced to their forever home in your aquarium, the need for good bacteria is ever increasing and it’s something that you should definitely look into. What is going in our tanks on the microscopic level? Fascinated yet?

There are multiple companies out there with several different strains of bacteria from reputable companies such as Fritz Aquatics, Dr Tim’s Aquatics, Two Little Fishies, and Brightwell Aquatics, to name a few. Today, I want to talk about something that’s completely different- a new strain that I came across in a product that was recommended to me. PNS PROBIO from Hydrospace LLC.

What’s in it? 

The strain of bacteria in this product is R. palustris, which is one of the oldest strain of Purple Nonsulfur Bacteria (PNB).

PNB was referenced in Julian Sprung and Charles Delbeek’s book : The Reef Aquarium, Vol. 3 (Page 585 for you guys who have the book). To be referenced in a book that is considered to be one of the reef bibles of our time has certainly piqued my interest in this product and I wanted to find out the legitimacy of the product.

From the book, The Reef Aquarium, Vol. 3: Science, Art, and Technology:

 

Bacteria and photosynthetic bacteria.

Ocean researchers have only recently recognized that bacterioplankton make up a significant amount of the carbon fixation in the sea, since they make up about 11% or more of the microbes found near the surface in seawater (Kolber et al., 2003). This fact suggests that their importance as a food source for coral reefs has probably also been underestimated, though early and recent work on the subject does recognize bacteria as a food source (Sorokin, 1973a and b, Bak et al., 1998).

Purple non-sulfur photosynthetic bacteria (PSB) may be a useful food for such notoriously difficult creatures as Dendronephthya and other filter feeders that specialize on eating microscopic plankton. Kolber et al. (2003) found that PSB occur from the surface down to approximately 150 meters (420ft.), with a maximum concentration at approximately 35 meters (98ft.) PSB are a non-taxonomic group of several different organisms that have a versatile lifestyle, being able to grow as photoheterotrophs, photoautotrophs, or chemotrophs, depending on the existing environmental conditions. PSB can switch from a metabolism based on the consumption of organic carbon to one based on its production by photosynthesis, depending on the concentration of organic matter in ocean water. In environments that are rich in dissolved organic matter, PSB stop making photosynthetic pigment and simply consume organic matter.

In the aquaculture industry, PSB are raised to help condition water and boost the health of shrimp and other aquaculture products. They can be cultured and maintained for extended periods in closed jars, so the use of them as a feed for marine invertebrates is feasible.

 

Fascinating stuff right? What does it do EXACTLY for our aquariums? I had so many questions and to learn more about this, I reached out to Kenneth Wingerter who cultures this bacteria for our aquarium hobby.

The interview is below and my questions are in bold:

Hello Kenneth, how are you? This is some fascinating stuff that you have going on here. From what I have been told from a few respected members of our industry, this bacteria is sort of all-in-one bacteria that not only helps with nitrification process, but also helps to get rid of unwanted sludges in our aquarium and also acts as an extremely nutritious food and probiotic bacteria for the animals in our tank. Can you give me some background information on yourself, what PNSB and what R. palustris is, and how you got into all this?

Hi Richard. 

I’ve been an aquarium hobbyist, first and foremost, since the mid-1980s. Saltwater aquarium husbandry instruction and saltwater livestock itself was pretty hard to come by back then. My first reef tanks were what we would now call FOWLR tanks; I remember being particularly proud of a few Aiptasia anemones I’d gotten on my first live rock. Ha!

I stress that I’m primarily a hobbyist because few of the many jobs I’ve had in this field have ever felt like “work.” This has always been a bit of an obsession for me; I’m totally sure that you and many of our fellow hobbyists can identify with that.  🙂  The longest I’ve ever gone without an aquarium since I was a kid was the couple years I lived on a fishing boat haha

Most of my early work in the industry was in LFSs, first in North Dakota, then in the PNW while studying at the University of Oregon. A pivotal point was obtaining a gig as a humble lab tech at the enormous UofO Zebrafish Facility (2002-2007). The bulk of my work there was raising live foods. To be honest, I thought it would be boring, and was just happy to be breeding fish. To my surprise, I found the special challenges of live aquaculture food culture to be extremely rewarding. Fun even, maybe? 

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After obtaining my degree in Biology, I moved out to the coast to attend the Aquarium Science Program (2010-2011 cohort). While there, I focused much of my study on live food culture (including practicums culturing phyto for an oyster farm and rotifers for a fish hatchery). After completing the program, I continued to work in the industry in Tennessee, Connecticut and Colorado in hatcheries as well as in retail and wholesale operations. 

Another pivotal moment came when I took a job as a production biologist at a small (at least then!) company called AlgaeBarn. At that time, they only offered pods and phyto. I was initially recruited to develop various expansions, including their fish, invert and macroalgae departments. Along with all that I created most of the content for their blog. In researching/writing about phyto and planktonic coral foods, I stumbled upon some information about purple non-sulfur bacteria (PNSB). The apparent importance of these organisms in marine ecosystems and their potential for home aquaria intrigued me. I did make a couple of proposals to culture PNSB there, but that project just kept getting pushed back as other new projects and expansions fell upon us. 

When AlgaeBarn made a major move to the Denver area a few years ago, I opted to stay behind and build a business of my own. The early “version” of Hydrospace LLC included a broader list of planktonic aquarium food items, including the purple non-sulfur bacterium Rhodopseudomonas palustris. That product, PNS ProBio, gradually gained attention in the hobby; the other products, not so much haha. So ProBio kind of became the flagship product. Eventually we developed a companion product, PNS YelloSno, which is a simulated marine snow that is photofermented with the bacteria in ProBio. YelloSno is for suspension-feeding inverts that utilize larger particle sizes, but there’s an another feature of this product that we’ll touch upon in a bit.

https://www.youtube.com/watch?v=6D_8AX8v7Vw

Brewing of YelloSno.

To return to your opening comments, yes, these are indeed “all-in-one” bacteria. They are extremely unusual in that they possess the enzymes required to perform all nitrogen transformations, including (1) assimilation of both organic and inorganic nitrogen, (2) nitrification, (3) denitrification and (4) nitrogen fixation. While not all aquarium hobbyists are familiar with that last function, nitrogen fixation, it’s probably the most important thing these guys do.

Basically, nitrogen-fixing bacteria (often referred to as diazotrophs) take nitrogen gas from the environment and convert it into ammonia, which they then use to create new biomass (i.e. grow). Nitrogen fixation is essentially the opposite of denitrification; it is how biologically available nitrogen is reintroduced into all ecosystems. Without nitrogen-fixers, the Earth would become increasingly infertile until all life would cease. 

Coral biologists and oceanographers have for long been perplexed by the high productivity of tropical coral reefs, which characteristically are found in extremely nutrient-poor environments (a dilemma they call the “Darwin’s Paradox”). It was only in the 1970s that we discovered how the rapid growth of reef-building corals owes largely to the activity of diazotrophic bacteria. Specifically, the zooxanthellae fix carbon (i.e. make organic substances out of CO2) that the coral can utilize, whilst the diazotrophs provide a fixed source of nitrogen for the zooxanthellae. Corals particularly favor rhizobial diazotrophs such as Rhodopseudomonas. Coral biologists now often refer to the bacterium-dinoflagellate-coral superorganism as the “coral holobiont.” 

Saw couple of things that grabbed my attention.


1.) Nutrition for corals?

2.) Provides Probiotics for the animals in our aquarium? 

Just as they eat excess zooxanthellae, corals eat excess bacterial symbionts. And yes, these bacteria are quite nutritious. One study found that they are over 70% protein, contain several essential fatty acids and are highly digestible. Moreover, they are rich in healthful carotenoid pigments. Even more, they’re shown to possess numerous probiotic properties (that’s a deep, deep rabbit hole worthy of its own discussion). In short, as part of an animal’s gut flora, they increase feed conversion and inhibit pathogenic microbes such as Vibrio.

Are there any cons? Possible overdose which can lead to catastrophic failures such as bacterial blooms or white slimes that are associated with bacteria overdoses?  From my understanding, R. palustris also can create ammonia from nitrogen gases. Isn’t this usually the stuff that we run away from?

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As with any natural filtration method, the only con to using these microbes is the time and effort required to promote the establishment of impactful population sizes. To start, they undergo a lag phase (essentially an acclimation period) of around a week and a half before starting exponential growth phase. To nudge them along, we suggest cutting protein skimmers, UV sterilizers and ozonizers for 12-24 hours after each application (at least the first application). Because these bacteria prefer illuminated anaerobic microhabitats, they readily colonize substrates such as sand beds and anaerobic biofilter media. One may facilitate colonization by soaking the first dose into a common filter sponge, ceramic biofilter block, etc. and then dropping it into the sump in an area of low flow (preferably illuminated). 

To say the least, they cannot easily be overdosed. We’ve actually tried using obscene amounts of PNS ProBio on all major types of coral, and failed to cause any apparent harm. To this day, I honestly don’t know how much it would take, as we gave up at 40x overdoses in the last trial. And that was a system without a skimmer!

Because these guys are primarily anaerobic, you won’t encounter the hazy blooms or gross slimes associated with aerobic heterotrophs. Nor do they pose any significant risk of oxygen depletion. 

With respect to nitrogen fixation, there is no risk of them dangerously elevating your ammonia levels. The reason for this is simple: Diazotrophy is energetically expensive. They only produce ammonia when NH4/NO2/NO3 is critically depleted in their environment; and even then, they don’t generate much more than they require for their own growth. In fact, the very presence of ammonia in their environment automatically shuts down the expression of genes responsible for running their diazotrophic machinery. Think of it this way: The coral (i.e. its zooxanthellae) will never run out of biologically available nitrogen internally, even while the water column is sufficiently nitrogen depleted to limit growth of competing turf and film algae. Just as in nature.

I saw this in Reef2Reef while I was doing some light reading on your product with R. palustris. I was fascinated with the bacteria’s flexibility with interacting with and adapting to carbon dosed surroundings that many of us aquarists tend to use in order to control the nutrients in our glass-enclosed reef systems. Here is a post from the thread that really intrigued me:

“R. palustris is definitely responsive to carbon dosing. It can utilize ethanol, but prefers acetate. In the absence of a suitable organic carbon, it additionally can switch to an autotrophic mode and utilize CO2. It’s pretty adaptable in that respect. It even possesses enzymes necessary to degrade cellulose (unlike most organisms), and so can even consume the poorly digestible detritus that accumulates from dead algae.

It prefers, but doesn’t require, anaerobic conditions. It’s known as one of the most oxygen-tolerant purple non-sulfur bacteria. In fact, it can live quite happily as an aerobic heterotroph. It’s flagellated and highly motile, and swims quite effectively in the water column. It’s even chemotactic toward sources of organic carbon, and swims right after them. So again, it certainly could be used for carbon dosing in the traditional sense. However, depending upon the unique microbiological community in a given aquarium system, it may or may not be able to compete with other, obligately aerobic, bacteria in the plankton.

All that being said, R. palustris is most competitive as an anaerobic photoheterotroph. It thrives in anoxic areas of the aquascape that receive some light (deep pits in live rock, just beneath the upper layer of the sand bed, within pockets of detritus, etc.). This means that it consumes organics (whether dosed or existing as wastes) along with ammonia/nitrite/nitrate and phosphate WITHOUT consuming oxygen. In other words, the presence of this bacterium not only enhances carbon dosing, but also reduces the risk of oxygen depletion.

This bacterium is extremely effective at sequestering nitrogen and phosphorus. Actually, it’s used in wastewater treatment plants to remove phosphate at crazy high levels we would never see in an aquarium. Cool thing about nitrogen use… In nutrient-poor environments, it can actually fix nitrogen as you indicated. This species has indeed been found living in association with reef-building corals. The corals actively take up these bacteria, and in some regions have a higher preference for them seasonally–where ammonium is less available to their zooxanthellae in certain months than in others. They may grow in both the coral gut and mucus, feeding on organic exudates from the coral as well as the zooxanthellae. It has well-known anti-vibriotic properties and thus almost certainly reduces the incidence of diseases such as RTN.

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Despite its overall versatility, this bacterium does have an Achilles heel. It cannot synthesize its own B vitamins, but requires them for growth, and therefore must obtain them from its environment. Depending upon what you feed, what you supplement, what kind of chem filtration you use, etc., your aquarium may or may not have abundant reserves of free vitamin B floating around. This is why the companion product PNS YelloSno is useful. In addition to serving as a coral food, it’s rich in B vitamins, and so promotes growth of PNS ProBio. (AlgaeBarn just made the combo available in a reduced-price package.)

On a final note… You mentioned that you wish to target phosphate. One advantage of using this particular microbe for that purpose is its ability to fix nitrogen. Specifically, you don’t need to worry about Redfield ratios; if nitrogen becomes the limiting nutrient, R. palustris will simply fix its own and continue to aggressively sequester phosphorus without interruption.”

Something that I want to find out later from people that are far more knowledgeable than myself in chemistry:

Acetate uses oxygen (very small increments) to turn into salt form with calcium and alkalinity but because that the bacteria utilizes CO2, does that mean it will oxygenate the water, negating the negative side effects of acetate? Furthermore, even IF people didn’t use acetate, but if they were to have a high CO2 environment battling low pH, can they use this potential rise of pH in their aquariums due to the bacteria using the CO2? Higher pH = faster coral growth. Mind boggling possibilities. I know.

Going back to the interview. Can this bacteria be put on a doser for us lazy reefers?  What is the shelf life of bottled bacteria and once exposed to oxygen, how long does it last before going bad?

We’ve never tested ProBio in dosers, and don’t really believe dosers are necessary or even of much benefit for using the product. The main reason is that these bacteria simply are so hard to overdose. But yeah, for the “lazy” reefer, it might be possible. For that, we’d just suggest dispensing it in a manner that would maintain anoxic storage conditions and minimize contamination by other microbes.

The shelf life of these bacteria is pretty impressive. We provide an expiration date on each bottle that is six months out from the bottling date (most bottles hit the market immediately after bottling). They store at room temp, but can briefly tolerate temperatures ranging from just above freezing to at least 120 degrees F. They easily tolerate exposure to oxygen, though oxygen does degrade their carotenoids (they cannot perform photosynthesis in the presence of oxygen). Actually, the worst part of exposing them to air is the potential for introducing contaminant organisms–so keep those caps clean and those bottles closed as much as possible! On a side note, PNS YelloSno is not a live product (it’s pasteurized midway through the fermentation process) and does require refrigeration, particularly after opening!

Give us some recommendations. How can we start this in our aquariums? Is there a different way to introduce this to a new tank and existing, mature tanks or it does the age really matter? What is the most effective way to do this?

As every tank is different, the ideal dosage depends upon a lot of things–amount of available nutrients/organics/vitamins/trace elements, amount of habitable living space, intensity of competition from other microbes, degree of grazing by corals and other filter-feeders, loss to skimming/sterilization, availability of light, etc. We recommend 1.25 ml/gallon/day to start. This might seem excessive to some, but many of our users consider any excess to be a source of nutritious live coral food.  🙂  For sensitive systems, disrupted systems, etc. we may recommend a half dose every other day just to be on the safe side (despite the fact that these guys are pretty safe as it is). For those using YelloSno, we suggest switching between the two on a daily basis (one product per day on an alternating basis). 

While the main benefits of using these bacteria are a bit different between new and old tanks (e.g. nitrogen cycling for new tanks, phosphorus cycling for old tanks), usage is the same.

In the R2R response above, you said that it needs vitamin B to grow; does our aquarium for the most part (if kept maintained with regular water changes, etc.) lack vitamin B and has to be supplemented or can the bacteria grow on its own with photosynthesis once introduced in the system if the tank is kept up with regular maintenance?

Actually, they’re capable of synthesizing most if not all of the B vitamin complex, at least under certain conditions. Based on lab studies, their B vitamin requirements definitely appear to vary between strains or environments. What does seem pretty clear is that synthesizing B vitamins is energetically costly for them and that they benefit greatly from having them provided. That’s one big benefit of using YelloSno with this product, as YelloSno is quite rich in B vitamins.

Is there any research done by others or by you on how effective this bacteria is on nitrates and phosphates?

Yes, lots. It’s been used and tested extensively in aquaculture and in wastewater treatment for decades. There’s abundant peer reviewed literature on this subject, so I’d encourage the curious reader to explore specific topics on Google (for example, “Rhodopseudomonas+aquaculture+phosphate”).

How much does this cost and where can you find it?

ProBio and YelloSno (16 oz.) are $28 and $34 respectively. They’re available online through AlgaeBarn . They’re increasingly showing up in LFSs across the U.S. (if your favorite stores don’t have them yet, please encourage them to contact Aquarium Supply Distribution www.aquariumsupplydistribution.com). Additionally, we’re presently exploring options for distributing ProBio internationally.

I know that every tank is different but what would be an realistic expectation for the average user, and how much does something like this cost when you break it down? 

For someone who wants to use this to export high concentrations of NO3/PO4 but their system just cannot support big populations, or for someone who simply wants to use it to feed a very densely stocked collection of corals, clams, etc., the maximum recommended daily dosage for a 100-gallon tank costs approximately seven bucks per day. That’s a maximum–not too bad when you consider that it’s a nitrate remover, phosphate remover, organics remover, live food, probiotic and carotenoid supplement all rolled up in one. 

Some other aquarists might successfully establish a large population with just a few doses–again, it depends upon a lot of factors. We don’t peddle this as a miracle elixir; it cannot dramatically fix a diseased or poorly maintained system overnight. Like most natural approaches to reefkeeping, using these and other beneficial microbes yields relatively slow and modest results. Like one great reefkeeper once said, nothing good in reefkeeping happens fast! When considering cost, as well as building expectations, it’s best to look at the long-term, holistic benefits in adding these bacteria.

There you go guys, I hope you guys learned something new and fascinated by this product as I certainly did. Being exposed in the hobby, I tend to come across plethora of products and to say that this is possibly one of the most fascinating product that I have came across in 2021, it’s quiet an accomplishment. I will be trying this product on a doser and test my theory out while seeking out the truth from amazing Dr. Craig Bingman regards to actuality of my thought process in reef chemistry. I am very grateful to Kenneth for taking his time out of his busy schedule to answer all of these questions for me and for you guys who are reading this to the end. I know it’s been a while but I promise. I will do this more often.

Happy reefing guys!

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Afishionado : a person who likes, knows about, and appreciates a usually fervently pursued interest or activity. (In this case, fish, corals and reef keeping). We are group of passionate hobbyists who are all about educating, creating innovative contents and providing eye popping visuals for hobbyists of all levels.

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