Microfragment Techniques in Coral Propagation

Introduction

Scleractinian “massive” coral species (‘stony’ corals) are fundamental to coral reef ecosystems, but global restoration efforts have often overlooked them due to slow growth rates. To date, massive coral transplantation is represented by fewer than 5% of studies, as restoration tends to focus on a small group of coral genera chosen for ease of proliferation.
As corals are clonal animals, when microfragmenting corals of the same genotype and arranging them a few centimetres apart, they will fuse when their edges join to create one mass of coral. This technique promotes accelerated growth, a phenomenon that would typically take several years.

We are experimenting with a new coral propagation microfragment technique for the first time at Reefscapers. We use a drill to take core samples from healthy ‘massive’ coral species, and transfer the cores into holes drilled into dead coral skeletons. We hope these cores will then grow over the dead skeleton and form new massive colonies. Usually, microfragmenting methods take coral fragments or cores and grow them out in land-based coral nurseries before being out-planted to the reef. Our method is quicker and cheaper, and we hope to start propagating massive corals in this way into the future.

Our initial trial methodology focuses on the commonly seen lobe coral Porites lobate, which has relatively open corallites and few skeletal elements. The small corallite size also allowed us to experiment with various core diameters based on the availability of drill bits.  To create a seal between the core and the skeleton (for stability, and maximum surface area for encrustation) we are adding a small amount of marine epoxy putty to the edge of the core. Currently, the donor colony is being left with the core holes exposed, but we plan to fill them with epoxy putty to see if this accelerates recovery.

Both the cores/skeleton and the harvested donor colony are being held in our new constant-flow tanks for close observation. We will regularly monitor the health of the cores and donors using the “Coral Watch” health chart, and will also look for early signs of encrusting of the core fragment to the dead skeleton.

January 2020

Started last month, our experimentation with coral cores is progressing well. Most plugs are healthy, with some initial paling likely due to the extraction and transplantation process. There are signs of colonisation of the neighbouring rock, which we initially thought (but no longer think) are ‘Mesenterial filaments’; there is little published research on how massive corals might grow to bridge a small gap.
‘Mesenterial filaments’ are string-like extensions of the internal folds of tissue which create the structure within a coral polyp’s body, typically bright white and full of nematocysts to capture and kill prey.

The parent donor colony now looks less healthy, likely due to an extended period in our storage tanks for (exposing it to sedimentation, fixed water flow and variable light levels). However, the scarring from plug extraction has diminished considerably and tissue recovery on the edges of the holes continues.

We are extending the experiment to an in-situ trial on the reef, with 3 lines of holes/plugs (>5cm apart) made to randomise the positioning (for water flow and light levels). We aim to locate a donor colony and outplant rock within proximity of each other to mimic environmental conditions. We will monitor on a weekly basis, with close attention to predation and to the effect of the epoxy on regrowth.

February 2020

Overall, the coral plugs and the parent colony are showing improved health after we relocated them to increase water flow and reduce direct sunlight. Encrusting has started on the edges of the plugs and the donor holes, and we expect the addition of Epoxy will close the larger gaps and further encourage encrusting.

We have been on several scoping dives to look for new sites. Site#1 consists of 2 large colonies of Porites lobata (2m diameter x 1m high, at depths of 1-5m). They both have areas of healthy living tissue (for donor plugs) interspersed with areas of bare rock (as transplant sites) that will ensure that plugs are subjected to very similar environmental conditions, which should improve the chances of survival.

Upon starting the experiment, we realised the drilling stages were tricky and very time-consuming, so we kept the total number of plugs to a minimum. Further practise and experience with underwater drilling and handling the Epoxy will improve our protocols and decrease the overall time taken.

Drilling and removal of Medium and Large plugs were found to be easier than the Small size, however the Small size will initiate faster encrusting (Medium/Large plugs take 2 months to encrust). Photos will be taken at weekly intervals and added to PhotoQuad software to calculate exact growth rates for each size class at each site. The exposed holes will also be monitored by the photographs for: encrustation, disease, predation, competition, algal growth.

March 2020

The coral plugs and parent-colony drill-holes are photographed and monitored weekly, with health assessments using the CoralWatch colour chart. We now have 6 weeks of data, conducted with three different trial plug sizes.

We have observed some predation, probably by parrot fish taking advantage of the increased roughness and accessibility of the coral surfaces.

May 2020

Our in situ coral plug propagation trial has been running for over 3 months now, and by month end, 33% of the original transplanted coral plugs are surviving (8/24 comprising: 4 medium, 3 large, 1 small).

From the current data, we can conclude that Medium and Large fragment sizes offer the greatest chance of success, and these two sizes will be used in future experiments. It is also clear that the chosen site for out-planting plays a vital role in the success of the transplanted fragments.

September 2020

All the parent colonies have displayed zero negative effects from the sampling, which is an encouraging result.
One more coral plug died, giving a 21% success rate with the current methodology. Only 1 plug (R3L1) is encrusting on the surrounding substrate and growing slowly in size; the other surviving plugs are yet to demonstrate outward growth.

In future experiments, we plan to:

• Hold the freshly sampled plugs in a nursery to minimise damage and ensure growth before outplanting.
• To facilitate outward growth, outplant the plugs directly onto the substrate (not in a cavity).
• Outplant plugs from the same parent colony close to one another (shared genetics could lead to better acceptance and larger growth).

Coral Microfragmenting Trial 2021

Studying Surface Area Growth Rates

Introduction: Microfragmenting of coral species as an effective tool of restoration on a Maldivian Reef System in Baa Atoll

Scleractinian “massive” coral species (‘stony’ corals) are some of the most abundant species on coral reefs due to their tolerance of  both disease and environmental disturbance in comparison to non-massive species. However, increasing ocean temperatures and mass coral bleaching events have caused acute thermal stress to all coral species, making them more susceptible to mortality and disease. The loss of massive coral species is of particular concern as they play a vital role in the long-term stability, structural complexity and biodiversity of coral reef ecosystems.

To date, there is no known published research of this technique in the Maldives, however, it is evident (via social media) that various coral restoration programs have adopted this method, but long-term results are unknown. If successful, this technique would be suitable for guest sponsorship, allowing Reefscapers to not only advance our understanding of massive coral growth rates, but allow guests to take part in new cutting-edge research.

We therefore propose to identify the survival rate and success of microfragments on a Maldivian reef system in Baa Atoll. We propose to create a long-term restoration program that incorporates guest education coupled with an exciting new hands-on experience.

Key Research Questions:

1. Is there a difference between surface area growth rates at various sites?
2. Is there a difference between surface area growth rates of various species?
3. Is there a difference between fragment size (‘Small’-‘Medium’) growth rates and survivorship?

Key Literature References:

• Forsman et al. (2015) – Growing Coral Larger and Faster: micro-colony-fusion as a strategy for accelerating coral cover.
  Orbicella faveolata and Pseudodiploria clivosia fused after 139 days when epoxied 1–2cm apart.
• Papke et al. (2021) – Differential Effects of Substrate Type and Genet on Growth of Microfragments of Acropora palmata.
  No difference in growth rates between ceramic vs cement tile (growth rate is predominantly influenced by genetics). Therefore, we will utilise a frame with a cemented centre to enhance microfragment survival and growth.
• Clements & Hay (2019) –  Biodiversity Enhances Coral Growth, Tissue Survivorship and Suppression of Macroalgae.
  Coral growth is enhanced in polycultures vs monocultures.
• Page et al. (2018) – Microfragmenting for the Successful Restoration of Slow Growing Massive Corals.
  Small fragments of massive coral species increased in surface area within the initial growth stages of outplanting, in comparison to large fragments of the Orbicella spp and Montastrea spp. (The higher predation on smaller fragments did not adversely affect survival and growth rates).
• Tortolero-Langarica et al. (2020) – Micro-Fragmentation as a Tool to Restore Remote Reefs.
  Colonies of Pavona clavus directly transplanted to the reef almost doubled in width and length, and live tissue cover increased by 2.5 times over 13 months.
• Broquet (2019) – Applicability of the Microfragmentation Technique to Propagate Corals.
  Fragment volume of Porites lobate increases 23.5%.

Research Materials & Methods

The novel ‘microfragmenting’ technique by scientists at the MOTE Marine Laboratory (Florida, USA) allows a broken piece of massive coral to grow rapidly, for outplanting after just a few months (rather than after several years).

By drawing on our previous research (2019-2020) we hope to create a simple yet effective methodology to microfragment massive corals while reducing time and costs. We also hope to develop a project component that includes guest education and interaction, and possible sponsorship in the future.

Initial ‘scouting’ dives will identify massive coral genera that are prevalent around Landaa Giraavaru. (At some point, we will incorporate ‘Assisted Gene Flow’ and collect corals from neighbouring reefs). Once located, donor colonies will be photographed, health-assessed, and QGIS-located (to monitor recovery).

Typically, a diamond band saw is used to extract samples, but we plan to use a pneumatic drill (as we have already demonstrated success with this technique). Coral cores from each donor colony will be kept separated (using mask boxes). The cores will be cut into smaller sizes (‘Small’ and ‘Medium’) before transferring the microfragment onto a “plug” with marine cement/epoxy.
Coral plugs will be kept in our tanks, for close monitoring before transplantation. From here, guests can choose their own ‘genet’ group (fragments from the same donor coral colony), consisting of 8 – 10 microfragments that they will place onto the centre of the experimental frame. The tag and tribute will then follow the same process currently used in our coral frame sponsorship program.

Whilst in holding tanks, previous studies have added Artemia (weekly) to promote initial growth rates (Forsman et al., 2015) and we plan to trial Rotifera as they are readily available from our Fish Lab.

As cement is readily available and cheap to buy, we plan to create moulded cement plugs that will be placed 1 to 2cm apart. We will note the tile number and individual coral fragment numbers for later growth assessments. Preliminary trials will utilise our flat frames (we have yet to finalise the frame design & size, tile size, and number of fragments per tile).

The completed experimental frames will be placed on areas of rock (for better stability) at several sites around Landaa. The frames will be monitored regularly, including the cleaning, photographing and maintenance of the tile and the microfragments.

Growth Assessment

In order to identify success rates and surface area (SA) growth of microfragments, photographs will be taken of each individual frame. By using a camera with a measuring scale attachment, we will be able to standardise our method. These images will then be analysed for polyp number and surface area growth changes using FIJI by Image J Software. This method will then be replaced with our AI technology, once it has been trained to identify massive coral species and new frame set ups, to improve monitoring efficiency. Four photos of sponsored frames will be taken, following current methods, however, the front facing view of the frame will be replaced by a “top side” view.

Photos (below) showing coral microfragment growth over an 8-month period, from May 2021 to January 2022.

Coral Microfragmenting Trial 2023

Fragment Attachment Success

July 2023

This month, we restarted our coral microfragmentation work with two ‘massive’ coral species, Porites lobia and Favia favus.

A hammer and chisel were used to collect live tissue from parent colonies on the Landaa House Reef, and to remove excess skeleton from the base of the collected fragments. Fragments ranged in size (0.5cm² to 2cm²) and were placed 1cm apart on ceramic tiles. Aquarium glue (“RA Aquatech”) was used to attach the F.favus fragments, and marine epoxy was used to attach P.lobia fragments. Tiles were placed in open-flow water tanks with UV light (6am-6pm daily).

We plan to see how successfully these fragments attach to the tiles and grow, before starting experiments to explore optimal conditions for survival and growth of massive species.

August 2023

Initial observations show that our fragments of P.lobia have paled and appear much less healthy than fragments of F.favus.