Previous experimental work with corals has shown that fusion can reduce size specific mortality among juvenile corals ( Raymundo & Maypa, 2004 ), and controlled conditions can increase survivorship of small colonies ( Raymundo & Maypa, 2004 ; Forsman, Rinkevich & Hunter, 2006 ; Toh et al., 2013 ). Culture of juvenile colonies or small (e.g., ∼1 cm 2 ) fragments combined with fusion of genetically identical colonies (micro-colony fusion) is a potential growth enhancement strategy for coral aquaculture. The ability to promote rapid growth over a pre-determined substrate would be a beneficial tool for a range of applications such as propagation of rare coral species, for the development of standardized growth assays, coral aquaculture, and reef restoration. We examined fusion in Orbicella faveolata and Pseudodiploria clivosa to quantify rates of area increase. Similarly, we conducted an experiment with Porites lobata to characterize tissue spreading and to determine if the rates are influenced by biotic and abiotic factors in two contrasting tank environments. In addition, we compile both qualitative and quantitative examples of isogenic colony fusion across a variety of coral species in both the Atlantic and Pacific Oceans.

A Montipora capitata colony was fragmented and attached to ceramic tiles using the same method as for the Porites lobata fusion experiment. The colonies were photographed after 3 months growth and again after six years of growth ( Figs. S1A–S1C ). Pocillopora meandrina was attached to garden variety plastic mesh fencing material by cutting the mesh, then forcing the fragments between the rigid plastic tabs such that the fragment was secured ( Figs. S1D–S1F ), this method required no adhesive, is very fast, and also worked for Porites compressa ( Figs. S2A and S2B ). For Porites astreoides , 6 fragments were attached to live rock using cyanoacrylate gel and photographed after fragmentation and again after 706 days. Three fragments of Solenastrea bournon i were similarly attached and observed over a period of 511 days ( Table 1 ).

(A) Thirty fragments were epoxied to ceramic tiles on 6/25/2006, yielding 23 cm 2 of area covered by coral tissue; (B) after 38 days of growth, tissue begins to attach and 3 fragments are lost; (C) after 125 days of growth, tissue begins to come in contact with other colonies; (D) after 205 days of growth, most fragments are fused and area covered by tissue is 178 cm 2 ; (E) after 368 days of growth the substrate is completely covered; (F) the resulting colony is approximately a half meter in diameter after one year.

Porites lobata fragments ( n = 240 total fragments from a ca 15 cm portion of a single donor colony) were fragmented to 0.69 ± 0.33 cm 2 (average ± stdev) and epoxied with approximately 2 cm of space between fragments to 30 × 30 cm glossy white ceramic tiles (30 fragments per tile) with marine epoxy (Splash Zone Compound; Woolsey/Z-spar Inc., Rockaway, New Jersey, USA; Fig. 2A ). The tiles were mounted to triangular concrete bases. The eight tiles were divided into two tanks at Kewalo Marine Laboratory that had notable differences in both biotic and abiotic conditions. The ‘cleaned’ tank was exposed to full sun, while the ‘established’ tank was partially shaded. The ‘established’ tank had been continuously running for over 5 years as a mesocosm tank with sand, live rock, and a variety of fish and invertebrates, while the ‘cleaned’ tank was emptied and cleaned prior to the experiment and only a few snails ( Trochus inextus ) and urchins ( Tripneustes gratilla ) were added to control algal growth (these species were also present at similar densities in the ‘established’ tank). Hobo pendant light and temperature loggers (Onset Computer Corporation, Bourne, Massachusetts, USA) recorded data hourly for the duration of the experiment. Digital photographs were taken from a fixed photo frame with a scale, and top down area covered by live coral tissue. Tissue area was measured with ImageJ v 1.0 after fragmentation on 6/24/2006 and after 38 days of growth on 1/15/2007. Fragment tiles were kept in round fiberglass tanks (4 m diameter, 1 m deep) with unfiltered seawater from the same source, each receiving approximately 10 lpm. The fastest growing module (10a) was further monitored and photographed at 125, 205, and 368 days of growth, although after 205 days the area of each individual nubbin was no longer able to be measured since nearly all of the fragments had fused together.

Five ramets of Orbicella faveolata from the same donor colony and of similar size were each fragmented into 0.86 ± 0.22 cm 2 (average ± stdev) pieces and glued to 5 ceramic 20 × 20 cm tiles ( Fig. 1A ). Fragments were attached to tiles using cyanoacrylate gel and were spaced equidistant from one another, separated by approximately 1 cm. The number of fragments per tile ranged from 20 to 23. Similarly 5 separate individual donor colonies of Pseudodiploria clivosa (∼30 cm 2 ) were fragmented into 3.05 ± 1.02 cm 2 (average ± stdev) pieces and glued to 5 separate 20 × 20 cm tiles. Fragments were attached in the same way as above and spaced equidistant from one another, separated by approximately 1.5 cm with 9 fragments mounted to each tile. The tiles were placed in a shallow 340 liter raceway with flowing water drawn from a 24 m deep seawater well at a rate of 2.5 lpm. Temperature was maintained in the raceway between 22 and 26 °C by constant seawater turnover and four air stones (4 cm each) were used for water circulation and aeration. Algal growth was controlled by the shore snail Batillaria minima , daily siphoning of detritus, and manual removal of encroaching algae, particularly in the space between fragments. Additionally, live newly hatched Artemia sp . were broadcast in the raceway on a weekly basis. Top down photographs of each tile were taken at a fixed distance with a 1 cm cube used as a size reference on 9/2/2014, 12/1/2014, and weekly thereafter. One photo of P. clivosa was not taken on 1/19 and therefore was not included in the analysis. Area covered by live coral tissue was measured using these photos with Sigma Scan Pro 5. The tiles were followed for a period of 139 days.

The ‘cleaned’ and ‘established’ tanks had significant differences in irradiance and temperature ( Table 2 and Fig. 5 ). The irradiance values for the ‘cleaned’ tank were more than twice as high as the ‘established’ tank, while temperature values were significantly different, they only differed by a few tenths of a degree Celsius ( Table 2 and Fig. 5 ). Corals in the ‘cleaned’ tank increased in average area covered by coral tissue by 122%, while the ‘established’ tank increased by 217% over 38 days. The ‘established’ tank had higher attachment failure (10.2% of the fragments were missing, compared to only 1 out of 109 fragments missing for the ‘cleaned’ tank), the missing fragments were most likely caused by sea urchin grazing, since grazing marks were observed near the colonies in the ‘established’ tank, and no grazing marks were observed in the ‘cleaned’ tank. The ‘established’ tank had red coralline algae covering a large proportion of the tiles after 38 days ( Figs. 2B – 2D ), while the ‘cleaned’ tank had almost no coralline algae. The initial fragment size was related to the rate of growth, with a linear equation explaining nearly 26% of the variability, with 1 cm 2 fragments nearly doubling in area over 38 days ( Fig. 4 ).

For P. lobata , the overall rate of increase was 5.6 cm 2 per month; however the rates of tissue spreading differed according to tank, which contrast in both biotic and abiotic conditions. Module 10A had the highest rates of growth and was followed for additional time intervals after the tank comparison experiment. After 205 days, the rate of tissue increase was 22.7 cm 2 per month, a 357% increase in area covered by tissue ( Fig. 2 ). The majority of coral colonies were fused and it was no longer possible to measure the growth of individual colonies since the entire 0.6 m module became a single fused colony ( Fig. 2F ).

After 139 days O. faveolata fragments increased in size by 329% with 13.5% of the fragmented colonies fusing together, while P. clivosa fragmented colonies increased in area by 154% with 31.1% of colonies fusing ( Table 1 and Fig. 1 ). No fragments of either species detached or perished during the experiment. The growth rates of both species appeared to be approximately linear, explaining 86% of the variation for P. clivosa , and 88% for O. faveolata ( Fig. 3 ). A second order polynomial regression explained 94% of the variance for P. clivosa and 97% of the variance for O. faveolata ( Fig. 3 ). A linear regression between initial fragment size and final fragment size further indicated that growth rates are related to colony size, with larger fragments growing at a faster rate, explaining 56% of the variability in O. faveolata and 79% in P. clivosa ( Fig. 4 ).

The overall rate of growth for common Atlantic and Pacific corals across all observations was ∼20 cm 2 /month ± 25 cm 2 /month (average ± standard deviation), with a minimum of 0.2 cm 2 /month for Solenastrea bournoni and a maximum of 63.2 cm 2 /month for Orbicella faveolata ( Table 1 ). These observations occurred over variable sampling periods and under various sampling conditions and some of these factors were examined in more detailed experiments for Orbicella faveolata, Pseudodiploria clivosa, and Porites lobata .

Discussion

Orbicella faveolata and Pseudodiploria clivosa fusion experiments In less than 4 months micro-fragments of O. faveolata increased a total of 293 cm2 while P. clivosa fragments grew 222 cm2. This corresponds to ∼11 cm and ∼9 cm of increased colony diameter respectively assuming circular colonies; however the present study measures changes in area covered by thin sheets of encrusted tissue, therefore these rates are not directly comparable to most field studies which measure change in maximum diameter or linear extension, for example, Caribbean corals grow at a rate of 0.5–1 cm per year (Hubbard & Scaturo, 1985; Logan, Yang & Tomascik, 1994; Cruz-Piñón, Carricart-Ganivet & Espinoza-Avalos, 2003). Nevertheless, starting with only 89 cm2 of O. faveolata tissue and 136 cm2 of P. clivosa tissue, four months of growth yielded a 329 and 154% increase in area respectively. Growth rates for both species fit the expectations of linear rates of growth, explaining between 86 and 88% of the variance; however, a second order polynomial curve explained between 94 and 97% of the variation indicating that growth rates likely accelerated towards the end of the experiment (Fig. 3). The differences in growth rates through time could be due to a variety of factors; however the initial size of fragment is clearly important, with smaller sized fragments growing at a slower rate than larger fragments (Fig. 4). It was beyond the scope of this experiment to determine if rates of tissue spreading correspond to seasonality, temperature, colony age, or other biotic or abiotic factors; however, previous work has also found clear relationships between size and growth rates, with larger fragments growing at a faster rate (Edwards & Clark, 1999; Lirman et al., 2010; Lirman et al., 2014).