Culturing Freshwater Pearl Mussel Margaritifera margaritifera in Northern Ireland. A first step toward the reintroduction of a threatened species. Phase 1 - The Development of techniques to mass culture M. margaritifera and the reintroduction of Freshwater Pearl Mussel to a river with historical but no current records of M. margaritifera

April 2002

Executive Summary

1. Introduction
Margaritifera margaritifera L is a large infaunal bivalve which has a complex life cycle. During the reproductive phase, M. margaritifera eggs are retained in the gills of females where they are fertilised and develop into glochidia larvae (Ziuganov et al., 1994). On release from the female, the glochidia must infect an appropriate host, usually a juvenile salmonid, to continue development. Glochidia usually attach to the gills of the host where development continues for between 4 and 12 months at which time they detach from the host and settle into the substratum to complete the life cycle. After a period of four to five years the young M. margaritifera emerge from the substrate (Beasley, 1996).

M. margaritifera was at one time found commonly in many of the river systems in Northern Ireland. However, recent surveys have identified small beds of M. margaritifera in only a few river systems (Beasely et al., 1998). Virtually no mussels below 10 years in age have been found and data would suggest that most individuals are currently in excess of 50 years of age (Beasely et al., 1998). Current research suggests that this species will disappear from Northern Ireland unless adequate protection and management is provided.

Conservation strategies for M. margaritifera vary throughout Europe. However, in river systems where mussel populations are severely depleted stock restoration is the only option for species conservation. Freshwater mussels have been successfully cultivated in Europe and in the U.S.A. In Northern Ireland attempts to conserve M. margaritifera have been centred at the Ballinderry Fish Hatchery which is situated near the Ballinderry river, Co. Tyrone where there is a significant but threatened M. margaritifera population. Here, large numbers of Brown Trout have been successfully infected with glochidial M. margaritifera and these fish have been transplanted into the local river in an attempt to boost the population of mussels.

2. Background to the current project
2.1 Experimental Design
The current project aimed to successfully cultivate and maintain juvenile M. margaritifera under experimental conditions within the Ballinderry Fish Hatchery. All experimental work was carried out at the hatchery. River water was diverted into the hatchery where it was used to maintain a number of experimental tanks. Two main areas were set up. The first area concentrated on the infection of juvenile Brown Trout with glochidial M. margaritifera. The second area was used to set up experimental gravel tanks where glochidial M. margaritifera could excyst from fish gills and burrow into sediment. Two large stainless steel tanks were used for this purpose. The first 'experimental tank' was filled with three different gravel types. These included Ballinderry river gravel, fine gravel (sand) and medium gravel placed randomly in the tank. The second 'river simulation' tank was designed to mimic river conditions by creating variations in water flow through the tank. This tank was filled with Ballinderry river gravel and several artificial meanders were created by varying the depth of gravel along its length. This design allowed water to pass through the tank at a faster rate and prevented the build up of large amounts of sediment. Both tanks were used to hold fish infected with glochidial M .margaritifera. Over the last 2 'experimental' years a total of 3000 fish have been infected with glochidial M. margaritifera and transferred to both tanks.
2.2. Culturing trials 1999 - 2001

2.2.1 Infection of Brown Trout
Successful infection of juvenile Brown Trout with glochidial M. margaritifera was confirmed in both experimental years. Glochidia were seen as small oval bodies scattered along the gills. Most of the fish sacrificed were heavily infected with glochidia.

2.2.2 Glochidial development on artificially infected fish
The number of glochidia found on salmonid hosts was found to significantly decrease over time. Numbers of glochidia peaked in December, 1999 and September, 2000 with an average of approximately 153 glochidia per fish. Numbers of glochidia steadily decreased with an average of only 12 glochidia per fish present in June 2000. A further sample of fish taken in July 2000 revealed that there were no glochidia remaining on the fish gills. This indicated that glochidial excystment was complete.

The process was repeated in the autumn of 2000. Levels of infection were identified as being approximately similar to the previous year. Levels of glochidial infection peaked in September 2000 and were again found to be high in February 2001. It was not possible to obtain further samples due to restrictions in travel imposed because of the outbreak of foot and mouth disease. As a consequence of the limited number of samples taken, numbers of glochida were not found to differ significantly in the second experimental year.

2.2.3 Growth & Development of juvenile M .margaritifera
Samples of gravel were taken to investigate the presence of juvenile M. margaritifera in February and July, 2001. A plastic core measuring 5cm in diameter was used to remove gravel from both tanks. A total of 11 juvenile M. margaritifera were found in the core samples taken. Identifications were verified by Dr Volker Buddensiek, an authority on M. margaritifera in Germany.

The number of M. margaritifera present in the experimental gravel tank has been estimated to be in the region of 7000. However, this estimate should be viewed as an absolute maximum.

2.2.4 Restocking
Restocking rivers with infected fish may represent an effective method of enhancing populations of M. margaritifera in Northern Ireland. Over the course of the current experiment approximately 3000 fish were released into the Ballinderry river where there was suitable salmonid 'nursery' habitat between the beginning of February and the end of March of 2000 and 2001. Using figures generated from both the current and previous investigations, it is possible to estimate the number of M. margaritifera that could have survived in the river subsequent to release. A potential minimum of around 54,000 mussels may have entered the Ballinderry over the past two years of the current experiment. This represents a significant boost to the mussel population on the river since even a survival rate of less than 10% would increase the current population estimates five fold. The current proposal recommends a further survey of the Ballinderry in August 2002 to estimate the success rate of the restocking programme.
3. Outputs

4. Wider Implications

5. Networking and Publicity
Contacts have been made between research groups in Scotland, Norway, Germany, Czechoslovakia and the U.S.A.. In addition, the project has been the subject of local school educational studies on water quality and river ecology. More recently, with the help of the current project, the Millenium Award has funded a project to attempt to re-introduce M. margaritifera on the river Bann, a river with historical, but no recent records of Freshwater Pearl Mussel. This project has attracted interest from bodies in the South of Ireland and was recently the subject of a television production for RTE.
The current project team have liased with various sister projects and formed links with other interested bodies. This has included the presentation of oral papers at both local and international conferences. These have included:

As a result of the meeting in Hof, Dai Roberts coordinated a bid for a European project and has agreed to organise a conference on Pearl mussels in Ireland in 2002
6. Recommendations

Key Words

Freshwater Pearl Mussel, Margaritifera, glochidia, water quality, salmonids, Ireland, distribution, rivers, conservation, culturing, habitat

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