In the wild when a coral bleaches (i.e., expels its zooxanthellae) it has the opportunity to uptake new zooxanthellae from the water column (via indirect transmission) and heal. In contrast, when corals bleach in aquariums the expelled zooxanthellae are effectively removed via modern mechanical filtration, inhibiting corals from reuptaking the zooxanthellae. The main goal of REVIVE™ is to add live zooxanthellae back into the aquarium, providing corals with the opportunity to uptake the newly introduced zooxanthellae and recover (assuming that the stressor that caused the bleaching event has since been remedied).
 
REVIVE™ currently contains symbiont species from the genera Symbiodinium, Breviolum, and Cladocopium (formerly classified as Clade A, B, and C, respectively). While its current formulation has been undoubtedly successful in ‘reviving’ bleached aquarium inhabitants (see our Testimonials page), at Aquaholic Aquaculture® we are always trying to improve our products, ensuring that we continue to provide aquarists with superior live feeds. The more unique strains from various genera that REVIVE™ contains, the likelier that one (or more) of the strains is a symbiont ‘match’ for the bleached aquarium inhabitant(s). As such, we have spent the last few years searching for new unique strains of zooxanthellae that would be appropriate for us to mass culture and add to our current REVIVE™ blend in order to enhance its overall efficacy.
 
New zooxanthella starter cultures are difficult to acquire, and zooxanthellae are notoriously challenging to grow in a laboratory setting separate from their hosts, making them exceptionally challenging to reliably produce at commercial-scale volumes. Since the release of our product REVIVE™: Live Zooxanthellae in 2017, we have worked to acquire and commercially produce new strains of zooxanthellae, but, despite numerous culture attempts with various species of symbionts, we have had no tangible outcomes until…

After years of trial-and-error experimentation, we are excited to announce that we finally have four new promising zooxanthella strains for REVIVE™!
 
All four of the new strains of zooxanthellae are theoretically great candidates for aquarium-purposes, as all four of them are known coral symbionts. To date, all four new strains have been responding well to our attempts to gradually scale them up for mass culture. While we are still perfecting our culture protocols for each of them, based on their recorded growth under our current culture methods over the course of the last several months, we feel confident that with patience and further refinement of our protocols that all four of these strains have the capacity for commercial production.

As we continue to ramp up production of these new zooxanthella strains and approach the ‘Testing Phase’, we will be on the lookout for aquarists who would like to be among the first to test these four new strains of zooxanthellae described above in our newly formulated blend of REVIVE™. Interested in being one of the first to try the new and improved REVIVE™? Reach out to us at [email protected] to join our beta-tester waitlist.

[1] Baker, A. C. (1999). Symbiosis ecology of reef-building corals. Ph.D. dissertation. University of Miami.
 
[2] Baker, A. C., Starger, C. J., McClanahan, T. R., & Glynn, P. W. (2004). Corals' adaptive response to climate change. Nature.
 
[3] LaJeunesse, T. C. (2002). Diversity and community structure of symbiotic dinoflagellates from Caribbean coral reefs. Marine Biology.
 
[4] LaJeunesse, T. C., Smith, R. T., Finney, J., & Oxenford, H. (2009). Outbreak and persistence of opportunistic symbiotic dinoflagellates during the 2005 Caribbean mass coral 'bleaching' event. Proceedings of The Royal Society. 276(1676).
 
[5] LaJeunesse, T. C., Parkinson, J. E., Gabrielson, P. W., Jeong, H. J., Reimer, J. D., Voolstra, C. R., & Santos, S. R. (2018). Systematic revision of Symbiodiniaceae highlights the antiquity and diversity of coral endosymbionts. Current Biology, 28(16).
 
[6] Manzello, D. P., Matz, M. V., Enochs, I. C., Valentino, L., Carlton, R. D., Kolodziej, G., Serrano, X., Towle, E. K., & Jankulak, M. (2019). Role of host genetics and heat-tolerant algal symbionts in sustaining populations of the endangered coral Orbicella faveolata in the Florida Keys with ocean warming. Global Change Biology. 25(3).
 
[7] Mashini, A. G., Parsa, S., & Mostafavi, P. G. (2015). Comparison of Symbiodinium populations in corals from subtidal region and tidal pools of northern coasts of Hengam Island, Iran. Journal of Experimental Marine Biology and Ecology, 473.
 
[8] Muller-Parker, G., D’Elia, C.F., & Cook, C.B. (2015). Interactions between corals and their symbiotic algae. In: Birkeland, C. (eds) Coral Reefs in the Anthropocene.
 
[9] Riddle, D. (2016). An update on Symbiodinium species and their hosts. Advanced Aquarist.
 
[10] Toller, W. W., Rowan, R., & Knowlton, N. (2001a). Zooxanthellae of the Montastraea annularis species complex: patterns of distribution of four taxa of Symbiodinium on different reefs and across depths. Biological Bulletin, 201(3).
 
[11] Toller, W. W., Rowan, R., & Knowlton, N. (2001b). Repopulation of zooxanthellae in the Caribbean corals Montastraea annularis and M. faveolata following experimental and disease-associated bleaching. Biological Bulletin. 201(3).
 
[12] Wang, C., Zheng, X., Li, Y., Sun, D., Huang, W., & Shi, T. (2022). Symbiont shuffling dynamics associated with photodamage during temperature stress in coral symbiosis. Ecological Indicators, 145.

​One of the more interesting microalgae that we grow at Aquaholic Aquaculture is Phaeodactylum tricornutum. P. tricornutum is a pennate diatom that has a unique pleomorphic nature. Unlike most diatoms, P. tricornutum is weakly silicified, and, because of this, its cell wall has exceptional plasticity, enabling the diatom to change its shape (Tesson et al., 2009). P. tricornutum has four distinct morphotypes: (1) Ovoid (also referred to as Oval or Round), (2) Fusiform, (3) Triradiate, and (4) Cruciform (He et al., 2014; Lewin et al., 1958).

​The ovoid morphotype is either oval or round in appearance and possesses a raphe that enables motility (Tesson et al., 2009). This shape is preferentially benthic, with ovoid cells tending to clump together and sink to the bottom (Lewin et al., 1958). In laboratory cultures the ovoid morph usually predominates cultures grown on solid media (e.g., agar plates) or is found adhering to the walls of its culture vessel (Barker, 1935; Lewin et al., 1958; Tesson et al., 2009). Research has shown that the elongated versus rounded shape of the ovoid morphotype is dependent upon culture conditions, with a tendency for ovoid cells to take on a more circular shape when stressed (e.g., nutrient depleted) and an elongated shape when healthy and thriving (Tesson et al., 2009).

​The cruciform morphotype is an irregular four-armed cell in the shape of a perfect cross (Wilson, 1946). This morphotype is extremely rare (He et al., 2014; Wilson, 1946). The cruciform morphotype was first described in depth by Douglas Wilson in 1946 (under the name of Nitzschia closterium f. minutissima). Since Wilson's publication in 1946, this morphotype has rarely been reported being observed in laboratory cultures (He et al., 2014; Wilson, 1946). Wilson noted that while this morphotype is extremely rare, it is relatively stable and reproduces true to type (e.g., asexual clonal division) (Wilson, 1946).

​While it was previously thought that P. tricornutum morphology may be due to passive cellular mutation, recent research has demonstrated that morphological shifts are an active response to external factors that activate P. tricornutum’s morphogenetic mechanisms (Tesson et al., 2009). Current research now suggests that P. tricornutum’s pleomorphism is an adaptation that evolved in response to the ever-changing environmental conditions of the coastal waters where P. tricornutum is predominately found, with each of the four morphotypes representing distinct ecophenotypes adapted for specific environmental conditions (He et al., 2014; Tesson et al., 2009). In support of this theory, research by He et al. (2014) demonstrated that morphological shifts in P. tricornutum could be triggered in the laboratory by manipulating culture conditions such as temperature, salinity, light, and culture media.

​The morphology of P. tricornutum cells impacts their biomacromolecule contents, with nutritional composition (e.g., lipid, protein dry weight, and carbohydrate content) varying depending on which of the four morphotypes the cell exhibits (He et al., 2014; Lewin et al., 1958). Of the four P. tricornutum morphotypes, He et al. (2014) demonstrated the cruciform morphotype to be the most nutritious, with an exceptional fatty acid profile. In their research, He et al., (2014) found that an abundance of cruciform morphotypes correlated with a significant increase in lipid content, with their predominately cruciform culture demonstrating both the maximum content of neutral lipid in a single cell and total yield.

For months, we had no success eliciting the formation of cruciform cells; our cultures continued to consist solely of fusiform and triradiate cells. However, we did start to notice that as we continued to decrease the temperature of the cultures that there was an increase in the proportion of triradiate cells to fusiform cells.

Finally! It happened! In February 2025, we pulled a sample from our experimental cultures to examine under the microscope, and we found our first cruciform cells!

​[1] Barker, H. A. (1935). Photosynthesis in diatoms. Arch. Mikrobiol. 6. 141.
 
[2] DeMartino, A., Meichenin, A., Shi, J., Pan, K. H., & Bowler, C. (2007). Genetic and phenotypic characterization of Phaeodactylum tricornutum (Bacillariophyceae) accession. J Phycol., 43. 992-1009.
 
[3] He, L., Han, X., & Yu, Z. (2014). A rare Phaeodactylum tricornutum cruciform morphotype: culture conditions, transformation and unique fatty acid characteristics. PLoS One, 9(4).
 
[4] Lewin, J. C., Lewin, R. A., & Philpott, D. E. (1958). Observations on Phaeodactylum tricornutum. J. Gen Microbiol., 18(2).
 
[5] Tesson, B., Gaillard, C., & Martin-Jezequel, V. (2009). Insights into the polymorphism of the diatom Phaeodactylum tricornutum Bohlin. Botanica Marina, 52. 104-116.
 
[6] Wilson, D. P. (1946). The triradiate and other forms of Nitzschia. Journal of the Marine Biological Association of the United Kingdom, 26(3), 235-270.

​Over the last decade, we have cultured numerous strains of zooxanthellae in the hopes of finding as many species as possible to add to REVIVE™, but very few have proven suitable for commercial production. Our pursuit to bring zooxanthellae to the aquarium industry has been wrought with numerous hurdles, with the main obstacles being: (1) Procuring new reef-aquarium-appropriate symbiont starter cultures, (2) Achieving reliable commercial-scale volumes, and (3) Testing for safety and efficacy.

While our current strains of zooxanthellae for REVIVE™ have proven themselves to undoubtedly be valuable for helping bleached aquarium inhabitants (see our Testimonials), we continue to look for new symbionts that we can attempt to mass culture and bring to the aquarium industry. Over time, we plan to add as many unique strains of zooxanthellae to REVIVE™ as possible to increase its efficacy and to continue to provide aquarists with the best tool for ‘reviving’ their bleached aquarium inhabitants.

​[1] Muller-Parker, G., D’Elia, C.F., & Cook, C.B. (2015). Interactions between corals and their symbiotic algae. In: Birkeland, C. (eds) Coral Reefs in the Anthropocene.
 
[2] Toller, W. W., Rowan, R., & Knowlton, N. (2001). Repopulation of zooxanthellae in the Caribbean corals Montastraea annularis and M. faveolata following experimental and disease-associated bleaching. Biological Bulletin. 201(3).

REVIVE™ can be broadcast-fed or target-fed depending on the extent of the bleaching event. The necessary quantity and frequency of dosage is dependent on the severity and extent of the bleaching event as well as the water volume of the aquarium housing the affected corals.
 
When using REVIVE™ as a broadcast-feed, we recommend a starting dosage of ¼ oz per 10 gallons added every day to the aquarium in a high flow area. Mechanical filtration should be disabled for approximately 2 hours after dosing REVIVE™ to allow the corals time to uptake the microalgae.

In addition to zooxanthellae, REVIVE™ also contains the microalgae Rhodomonas sp. and Phaeodactylum tricornutum. Rhodomonas and Phaeodactylum are added to REVIVE™ as a supplemental live coral feed intended to aid with the recovery of bleached corals through providing high quality nutrition.
 
Aquarists often overlook the important role that live feeds play in the growth, coloration, and overall health of corals. While the zooxanthellae that reside within the tissues of the coral provide the coral with food, these zooxanthellae are only one component of nourishment for the coral. In order to sustain vibrancy and optimal health, corals require regular planktonic feedings in addition to the food created by the zooxanthellae.
 
Rhodomonas and Phaeodactylum have some of the highest microalgal polyunsaturated fatty acid concentrations making them a perfect addition to REVIVE™ for the purpose of nourishing your livestock back to health. Through dosing REVIVE™, bleached corals have the opportunity to uptake newly introduced zooxanthellae, and then these corals can feed on Rhodomonas and Phaeodactylum receiving the nourishment that they need to thrive.

[1] LaJeunesse, T. C., Parkinson, J. E., Gabrielson, P. W., Jeong, H. J., Reimer, J. D., Voolsrra, C. R., & Santos, S. R. (2018). Systematic revision of Symbiodiniaceae highlights the antiquity and diversity of coral endosymbionts. Current Biology, 28(16).

[2] Robinson, J. D., & Warner, M. E. (2006).  Differential impacts of photoacclimation and thermal stress on the photobiology of four phylotypes of symbiodinium (pyrrhophyta). Journal of Phycology, 42​(3), 568-579.

​Technically the term “zooxanthellae” has no taxonomic meaning, but it is used colloquially to describe the symbiotic golden-colored dinoflagellates that reside within animals (such as corals, anemones, clams, etc.). Zooxanthellae’s small size (the majority are <11 µm in diameter) and similar morphology have made them difficult to study and catalogue, as it is inordinately challenging to distinguish between different species of zooxanthellae without relying on genetic testing (LaJeunesse et al., 2018; Muller-Parker et al., 2015; Toller et al., 2001a).
 
Up until 2018, zooxanthellae were all classified as members of the family Symbiodinium. Members of this family were sorted into “clades” to help differentiate and organize the various zooxanthellae. Seven main clades (i.e., clades A-G) were established, and zooxanthellae were catalogued first by clade and then by “sub-clade” (i.e. “type” or “strain”) using alpha-numeric designations. For example, the designation “A1” indicated that the zooxanthella belonged to clade "A" and sub-clade "1" (LaJeunesse et al., 2018).
 
In 2018, LaJeunesse et al. released a paper redefining these loose clades into seven official genera under the newly established family name: Symbiodiniaceae. In recent years, refinement of DNA analyses revealed that the clades of zooxanthellae were more genetically diverse than previously recognized and that these clades warranted being reclassified as unique new genera in order to help better describe and organize members of the family. The original family name of “Symbiodinium” was repurposed for labeling the new genus that was designated to encompass members specifically of the former clade A. The genera representing the former clades B-G were designated as follows: Breviolum, Cladocopium, Durusdinium, Effrenium, Fugacium, and Gerakladium. Below we describe some of the main characteristics of each of these genera.

[1] Baker, A. C. (1999). Symbiosis ecology of reef-building corals. Ph.D. dissertation. University of Miami.
 
[2] Baker, A. C., Starger, C. J., McClanahan, T. R., & Glynn, P. W. (2004). Corals' adaptive response to climate change. Nature.
 
[3] LaJeunesse, T. C. (2002). Diversity and community structure of symbiotic dinoflagellates from Caribbean coral reefs. Marine Biology.
 
[4] LaJeunesse, T. C., Smith, R. T., Finney, J., & Oxenford, H. (2009). Outbreak and persistence of opportunistic symbiotic dinoflagellates during the 2005 Caribbean mass coral 'bleaching' event. Proceedings of The Royal Society. 276(1676).
 
[5] LaJeunesse, T. C., Parkinson, J. E., Gabrielson, P. W., Jeong, H. J., Reimer, J. D., Voolstra, C. R., & Santos, S. R. (2018). Systematic revision of Symbiodiniaceae highlights the antiquity and diversity of coral endosymbionts. Current Biology, 28(16).
 
[6] Manzello, D. P., Matz, M. V., Enochs, I. C., Valentino, L., Carlton, R. D., Kolodziej, G., Serrano, X., Towle, E. K., & Jankulak, M. (2019). Role of host genetics and heat-tolerant algal symbionts in sustaining populations of the endangered coral Orbicella faveolata in the Florida Keys with ocean warming. Global Change Biology. 25(3).
 
[7] Mashini, A. G., Parsa, S., & Mostafavi, P. G. (2015). Comparison of Symbiodinium populations in corals from subtidal region and tidal pools of northern coasts of Hengam Island, Iran. Journal of Experimental Marine Biology and Ecology, 473.
 
[8] Muller-Parker, G., D’Elia, C.F., & Cook, C.B. (2015). Interactions between corals and their symbiotic algae. In: Birkeland, C. (eds) Coral Reefs in the Anthropocene.
 
[9] Riddle, D. (2016). An update on Symbiodinium species and their hosts. Advanced Aquarist.
 
[10] Toller, W. W., Rowan, R., & Knowlton, N. (2001a). Zooxanthellae of the Montastraea annularis species complex: patterns of distribution of four taxa of Symbiodinium on different reefs and across depths. Biological Bulletin, 201(3).
 
[11] Toller, W. W., Rowan, R., & Knowlton, N. (2001b). Repopulation of zooxanthellae in the Caribbean corals Montastraea annularis and M. faveolata following experimental and disease-associated bleaching. Biological Bulletin. 201(3).
 
[12] Wang, C., Zheng, X., Li, Y., Sun, D., Huang, W., & Shi, T. (2022). Symbiont shuffling dynamics associated with photodamage during temperature stress in coral symbiosis. Ecological Indicators, 145.

​Zooxanthellae form a symbiotic relationship with corals in a partnership known as mutualism, in which the phototrophic zooxanthellae partner with the heterotrophic host coral. Through photosynthesis the zooxanthellae convert sunlight, inorganic nutrients (derived from either the coral’s metabolic waste products or through uptake of dissolved inorganic compounds from the water column), and carbon dioxide into carbon and energy sources for the coral, and, in return, the coral provides a safe habitat for the zooxanthellae. The zooxanthellae consume some of their photosynthetically fixed carbon for their own respiratory and growth requirements, but the rest of the carbon is made available for utilization by the host coral. While corals are able to acquire some nutrition through holozoic feeding (e.g., capturing and consuming zooplankton), the rest of the coral’s nutrition is dependent upon the photosynthetic products derived from its zooxanthellae (Muller-Parker et al., 2015; Toller et al., 2001). Through photosynthesis, zooxanthellae provide up to 90% of a coral’s energy demand, playing a vital role in coral nutrition and health (Mashini et al., 2015).
 
Recent research by LaJeunesse et al. (2018) estimates that this mutualistic relationship between corals and their microalgal symbionts has existed for over 140 million years and has survived multiple extinction events, with rDNA sequencing tracing ancestors of modern-day corals and their symbionts back to the middle to late Jurassic period of the Mesozoic Era. It is theorized that this algal-animal relationship originally evolved in response to corals being unable to attain sufficient nutrition solely from holozoic feeding, especially in harsh environments (Muller-Parker et al., 2015).

​Coral bleaching is a phenomenon in which a host dissociates from its symbionts when stressed. When corals bleach, they lose the zooxanthellae that give them their color, and they turn white. Coral bleaching is an indicator that the coral is under significant stress, and it is correlated with coral death (LaJeunesse et al., 2018; Muller-Parker et al., 2015).
 
In order for corals and zooxanthellae to maintain their symbioses, the benefits from the zooxanthellae’s photosynthetic production must outweigh the metabolic cost of maintaining the zooxanthellae. When the cost of sustaining the zooxanthellae becomes too great and the mutualistic relationship is no longer advantageous to both parties, the mutualism is disrupted, and the coral expels its zooxanthellae (LaJeunesse et al., 2018; Muller-Parker et al., 2015). There are many factors that can influence the balance of this symbiotic relationship and thus affect coral bleaching, including but not limited to: Fluctuations in temperature, salinity, nutrients, sediments, turbidity, air exposure, rainfall, and light intensity, as well as respective growth rates of the host coral and its symbionts. Additionally, both the severity of the stressor as well as the duration of the stressor affect whether or not a coral succumbs to bleaching (LaJeunesse et al., 2018; Muller-Parker et al., 2015; Toller et al., 2001).

​Yes, corals can recover from bleaching. Bleaching alone is not a death sentence. When the stressor that caused the bleaching event is removed and favorable conditions return, corals can reestablish symbioses with zooxanthellae and get their colors back (i.e., re-brown). Corals can reestablish symbioses with zooxanthellae in one of two ways: (1) Residual background zooxanthellae populations that survived through the bleaching event can multiply and/or (2) The coral can establish new symbiotic relationships with free-living zooxanthellae from the water column (i.e., indirect transmission) (Muller-Parker et al., 2015). Once symbioses are reestablished, it can take some time for the coral to re-brown and appear healthy again; augmentation of zooxanthella populations to standard healthy coral densities usually takes 1.5 – 3 months (LaJeunesse et al., 2009).
 
Recent research has demonstrated that the unique adaptations of different species of Symbiodiniaceae affect a coral host’s ability to persevere through and/or recover from bleaching events, with specific species of Symbiodiniaceae being better suited to survive and thrive in particular situations. As such, a coral may “shuffle” or “switch” symbionts to meet its needs within its specific environment (LaJeunesse et al., 2009; Muller-Parker et al., 2015).

[1] Baker, A. C., Starger, C. J., McClanahan, T. R., & Glynn, P. W. (2004). Corals' adaptive response to climate change. Nature.
 
[2] LaJeunesse, T. C., Parkinson, J. E., Gabrielson, P. W., Jeong, H. J., Reimer, J. D., Voolstra, C. R., & Santos, S. R. (2018). Systematic revision of Symbiodiniaceae highlights the antiquity and diversity of coral endosymbionts. Current Biology, 28(16).
 
[3] LaJeunesse, T. C., Smith, R. T., Finney, J., & Oxenford, H. (2009). Outbreak and persistence of opportunistic symbiotic dinoflagellates during the 2005 Caribbean mass coral 'bleaching' event. Proceedings of The Royal Society. 276(1676).
 
[4] Mashini, A. G., Parsa, S., & Mostafavi, P. G. (2015). Comparison of Symbiodinium populations in corals from subtidal region and tidal pools of northern coasts of Hengam Island, Iran. Journal of Experimental Marine Biology and Ecology, 473.
 
[5] Muller-Parker, G., D’Elia, C.F., & Cook, C.B. (2015). Interactions between corals and their symbiotic algae. In: Birkeland, C. (eds) Coral Reefs in the Anthropocene.
 
[6] Toller, W. W., Rowan, R., & Knowlton, N. (2001). Zooxanthellae of the Montastraea annularis species complex: patterns of distribution of four taxa of Symbiodinium on different reefs and across depths. Biological Bulletin, 201(3).

When we released REEF RECIPE™ (our premium dry coral food) in December of 2021, we made the claim that when REEF RECIPE is fed according to our instructions (see the REEF RECIPE label on the jar or the REEF RECIPE website page), no negative phosphate (PO4) affects should be observed. This claim that we made at REEF RECIPE's product release was based off of data collected from feedings here at our aquaculture facility coupled with data collected from beta-tester reef aquariums. Phosphate parameters were measured with a combination of both Hanna and Salifert test kits. Data collected both on- and off-site confirmed that when dosed according to our instructions, REEF RECIPE had no significant effect on phosphate levels in aquaria.

In recent months, we have taken our REEF RECIPE phosphate experiment a step further. While the data previously collected via Hanna and Salifert test kits is undeniably valuable, we wanted to run a new feeding trial here out our aquaculture facility in which we dosed REEF RECIPE and sent off weekly water samples for Inductively Coupled Plasma - Optical Emission Spectrometry (ICP-OES) analysis to get the most accurate phosphate readings possible. Our methods and results are outlined below.

The Experimental System

Our experimental trial began on 01/04/2022 and was conducted through 02/08/2022. The system selected for the experiment was one of our coral systems here at our aquaculture facility. This particular coral system is approximately 850 gallons, and it houses a variety of corals (including soft corals, LPS, and SPS corals).

Feeding Protocol

The experimental coral system was fed once daily at a consistent dosage of 8.5 teaspoons, strictly following the instructions listed on the REEF RECIPE jar and the REEF RECIPE website page. For our experiment, we chose to broadcast feed (rather than target feed) the REEF RECIPE. Once the REEF RECIPE was hydrated in water from our coral system, we dumped the solution into a high-flow area of the coral system (i.e., in front of an Ecotech Vortech powerhead) to be dispersed to the corals.

Data Collection

Two water samples were collected from the experimental coral system between the hours of 10:00am EST and 3:00pm EST every Tuesday during the experimental trial. These water samples were mailed off same-day for ICP-OES analysis by Reef*Labs in Bradenton, FL. Specifically, we were interested in the ICP-OES readings for phosphate (PO4) and phosphorus.