Commercial Production of Zooxanthellae for the Aquarium Industry

REVIVE™: Live Zooxanthellae

​For nearly a decade we have experimented with growing various members of the family Symbiodiniaceae (i.e., zooxanthellae) searching for species suitable for commercial production for the aquarium industry. After much research and trial-and-error experimentation, we finally found a handful of suitable species and worked to produce them in mass culture. In 2017, our selected zooxanthella cultures were ready for commercial distribution, and we released our new product: REVIVE™: Live Zooxanthellae.

REEFreshments®: REVIVE™ (32 oz. Bottle)
​© Aquaholic Aquaculture

Every strain of zooxanthella that we have cultured at Aquaholic Aquaculture® for use in REVIVE™ has been from either the genus Symbiodinium, Breviolum, or Cladocopium (formerly classified as Clade A, B, and C, respectively). These three genera are commonly associated with corals (as well as other common reef inhabitants [e.g., clams, anemones, etc.]) (Muller-Parker et al., 2015) and thus have the best chance of forming a mutualistic relationship with inhabitants of a typical reef aquarium. By including multiple strains from multiple genera in REVIVE™, it increases the likelihood that one (or more) of the strains in our product is a symbiont ‘match’ for the bleached aquarium inhabitant(s). The more unique strains that we add to REVIVE™, the better the product can serve its intended purpose: ‘Reviving’ bleached aquarium inhabitants.

Zooxanthellae in Laboratory Culture

While many members of the family Symbiodiniaceae can live independently of an animal host, it is challenging to find species that can thrive asymbiotically for long-term culture in a laboratory. As phototrophs, laboratory cultures require adequate light and nutrients in order to grow. While zooxanthellae living within an animal host are primarily found in the coccoid (non-motile) stage, in laboratory cultures zooxanthellae tend to alternate between both the coccoid and dinomastigote (motile) stages (Muller-Parker et al., 2015). Like other species of microalgae grown for the aquarium industry, species of Symbiodiniaceae reproduce asexually, increasing their population by creating identical clones of the parent cell. Different genera of Symbiodiniaceae have varied growth rates (Toller et al., 2001). In our experience, cultures of Symbiodinium or Breviolum generally multiply faster than Cladocopium cultures.

Microscopic image of zooxanthellae (Cladocopium sp.)
​© Aquaholic Aquaculture

The Challenges Associated with Bringing New Strains of Zooxanthellae to the Aquarium Industry

​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.

(1) New Starter Cultures
New starter cultures are difficult and often very expensive to obtain. There are very few resources for obtaining symbiont starter cultures, and these resources usually only have a select few strains available at a given time. Moreover, our specific interest in only acquiring symbionts that have the potential to host aquarium inhabitants further amplifies the challenge of finding an appropriate starter culture.

​(2) Commercial-Scale Volumes
Zooxanthellae are notoriously difficult to grow in a laboratory setting separate from their hosts, and it is very challenging to reliably produce these zooxanthellae at commercial-scale volumes. Most obtainable starter cultures are only available in extremely small quantities (e.g., test tube starters), and it is a slow and arduous process growing out these starter cultures to commercial volumes.

Zooxanthellae Starter Cultures
​© Aquaholic Aquaculture

Compared to other microalgae grown for the aquarium industry, zooxanthella cultures are slow-growing, delicate, and fickle, and, therefore, also highly susceptible to contamination. Each zooxanthella strain requires individualized culture conditions, with many species being unreceptive to standard algal growth media and microalgae culturing protocols (Muller-Parker et al., 2015). For every strain that we have cultured, we have had to refine our culture protocols to accommodate the needs of each specific symbiont, adjusting salinity, temperature, light intensity, nutrient densities, etc. so that cultures can thrive. Consistent implementation of these specific culture protocols is crucial, as even a slight deviation from established protocols can quickly result in the loss or contamination of the culture.
 
(3) Testing for Safety and Efficacy
In theory, any new strain that we have selected as a candidate to attempt to culture for REVIVE™ should be safe and effective in a reef aquarium. We have only cultured species that are known coral symbionts, and there is no reason to suspect that they would be anything other than beneficial to a reef aquarium. However, we would not feel comfortable commercially distributing these zooxanthellae without first investigating their safety and usefulness in an aquarium environment. Before any of our strains have been commercially distributed, they have first gone through a ‘Testing Phase’. During the ‘Testing Phase’ we observe the effects that the new symbiont has on bleached aquarium inhabitants (and on the reef aquarium as a whole) to ensure the safety and efficacy of including the strain in REVIVE™.

During the 'Testing Phase' we also assess the zooxanthella strain's capacity for prolonged refrigerated storage. In order to be a viable candidate for REVIVE™, the new strain must be able to achieve a minimum refrigerated shelf life of at least four months.

This 'Testing Phase' is very time-consuming and usually takes several months to over a year.

The Future of REVIVE™: Live Zooxanthellae

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.

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References

​[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).