Ex situ Conservation of Bryophytes

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The project for the ex situ conservation of bryophytes based at the Royal Botanic ... concern was raised about the status of bryophyte conservation in the UK – in.
Ex situ Conservation of Bryophytes M. M. RAMSAY & J. K. ROWNTREE Micropropagation Unit, Royal Botanic Gardens, Kew, Richmond, Surrey, TW9 3AB , UK . [email protected] Introduction Bryophytes are ubiquitous in the terrestrial environment and play important roles in ecosystem function e.g. sequestering nutrients, retaining water, regulating the soil micro-environment and acting as carbon sinks -VITT, 2000). The UK has an exceptional diversity of bryophytes with approximately 600 moss and 300 liverwort species. Of these, 62 are listed for attention under the UK Biodiversity Action Plan -UKBAP), the UK government response to the Rio Convention. The project for the ex situ conservation of bryophytes based at the Royal Botanic Gardens and funded by English Nature, Scottish Natural Heritage and Countryside Council for Wales was established when concern was raised about the status of bryophyte conservation in the UK – in particular that many of the UK’s bryophytes are restricted to a single or handful of sites and are vulnerable to catastrophic events with no strategic centralised ex situ facility to act as an insurance policy. The aim of this unique project has been to deliver the ex situ component of a suite of Species Action Plans within 1 programme through the development and evaluation of experimental protocols for the collection, sterilisation, propagation and basal storage of UK bryophytes and ultimately to develop reintroduction techniques. Collection In accordance with IUCN guidelines, collection protocols have been devised that ensure representative samples are collected from each population, whilst taking into consideration the type of material most suited to initiating into culture. These highlight the requirement for material to be collected from different areas of the population being sampled. The collection protocols ensure: • Collection is undertaken sensitively, ensuring no detrimental effects of collecting on the in situ population • Gaps are not produced in the colony, safeguarding the natural population against invading species. • Genetic variation is represented in the collected sample Collection protocols have been developed for desiccation tolerant mosses, desiccation intolerant mosses and leafy liverworts, and a third for thalloid liverworts and hornworts. Propagation and Storage Sterile -aseptic) culture is essential to prevent cross-contamination of samples if material is being cryopreserved in liquid nitrogen. Novel methods of sterilisation have been developed for bryophytes using NaDCC -Sodium dichloroisocyanurate) and without the addition of detergents. Separate protocols have been developed for:

• sporophytes -1% NaDCC for 3-6 min) • leafy gametophores -0.5% NaDCC for 2-5 min) • gemmae -0.1% or 0.5% NaDCC for 2-8 min) • thallus tissue -0.01% NaDCC for 10-20 min) Gametophytic material -protonemata and gametophores) is grown on one of three standard media:½ strength Murashige & Skoog, ¼ strength Murashige & Skoog or Knops minimal media without sucrose and using Gelrite® as a gelling agent -Burch in press)

Cryopreservation Cryopreservation is the storage of living material at or near the temperature of liquid nitrogen -- 196 °C). At this temperature cellular processes are effectively stopped and the cells or organs are stored in a state of suspended growth, free from pathogens or the risk of genetic drift. It is therefore a valuable tool for plant genetic conservation and allows a tissue bank of vegetative material to be stored over long periods of time. Gametophytic material is cryopreserved as sporophytes are not consistently available for all bryophyte species. A standard protocol has been developed for protonematal material involving: 1. Encapsulation in alginate strips 2. Pre-treatment with ABA and sucrose 3. Dehydration 4. Immersion and storage in liquid nitrogen -BURCH & WILKINSON, 2002) The link between water availability in habitat and desiccation tolerance in bryophytes is well known. Survival after cryopreservation appears to be linked to desiccation tolerance -Burch, 2003) and is currently being assessed for a wide number of species. In addition, cryopreserved samples of Ditrichum cornubicum were shipped to Kings Park and Botanic Garden, Western Australia as part of cryo-exchange project. This offers the potential for duplicate gene-banks The gametophyte samples collected after sterilisation and cryopreservation procedures can be used to investigate any genetic, morphological or cellular changes induced by these procedures. Material will be supplied to Queen Mary University to determine effect of cryopreservation on bryophyte tissue. Reintroduction So far, very few attempts to re-introduce bryophytes have been made. It is therefore critical to develop sound protocols for the choice of candidate species and re-introduction sites, techniques for moving plants from aseptic culture to field conditions and monitoring procedures. The laboratory has already been involved in the successful re-establishment of vascular plants but it is likely that bryophyte physiology, reproductive biology, and life-history strategies may necessitate modification of techniques.

Initial laboratory based trials have commenced investigating the gradual acclimatisation of plants to lower humidity, increased airflow, variable temperatures and lower nutrient availability. Four species have weaned onto natural substrates using these weaning techniques. Wakehurst Place, Kew’s garden in West Sussex has a Site of Special Scientific Interest of UK and European significance for its bryophyte assemblages on sandstone outcrops. It contains the Francis Rose Reserve for Cryptogamic Plants, one of the first of its kind in Europe with extensive interpretation of lower plants and their significance. These in situ facilities will be used as a testing ground for reestablishment of laboratory grown material of Orthodontium gracile -enabled in part by the Friends of Kew Threatened Plant Appeal). Material of O. gracile is being bulked up for genetic analysis , to determine the genetic variability of the species from 16 sites around the UK The re-introduction guidelines produced will be promoted through both the IUCN SSC Reintroduction Specialist and Bryophyte Specialist Groups Global Stategy for Plant Conservation Target Delivery The Plant Diversity Challenge report -CHEFFINGS et al, 2004) is the UK’s response to the Global Strategy for Plant Conservation -GSPC). The Strategy includes 16 outcome-oriented targets to be met by 2010. The report was prepared following an extensive two-stage consultation process and compiled on behalf of the Government by a partnership between the Joint Nature Conservation Committee, Plantlife International and the Royal Botanic Gardens, Kew. Relevant targets include: Target 3. Development of models with protocols for plant conservation and sustainable use, based on research and practical experience; Target 8. 60 per cent of threatened plant species in accessible ex situ collections, preferably in the country of origin, and 10 per cent of them included in recovery and restoration programmes Within the UK, there has been considerable progress towards Target 8 particularly for vascular plants. The Project is making a major contribution to ongoing actions which include developing methodologies for the ex situ conservation and re-introduction of bryophytes. European Habitats and Species Directive and Bern Convention taxa will also be considered for inclusion in the Project. Of the 26 bryophytes on the Bern Convention, four are found in Britain: Marsupella profunda, Petalophyllum ralfsii, Hamatocaulis vernicosus and Buxbaumia viridis. The project is working with the European Committee for Conservation of Bryophytes to prioritise the remaining 22 H&S Directive species, threatened throughout mainland Europe, in order to deliver target 2.6 of the European Plant Conservation Strategy : “At least 12 priority species of bryophytes brought into ex situ conservation and methodology promoted internationally”.

Species Currently in Ex Situ Collection Species

Status

Aplodon wormsjoldii Bartramia stricta Cyclodictyon laetevirons Ditrichum cornubicum Ditrichum plumbicola Leptodontium gemmescens Orthodontium gracile Orthotricum obtusifolium Seligera carnicolica Weissia multicapsularis Weissia rostellata Zygodon forsteri Zygodon Gracilis

Sterile

Critically Endangered

Sterile Cryo Sterile

Critically Endangered

Sterile Cryo Sterile Sterile Cryo Sterile Cryo Non-sterile

World Red List

Critically Endangered

European Red List ECCB -1995)

British Red List Church et al -2001)

Rare

Endangered

Endangered

Endangered

Vulnerable

Near threatened

Rare

Vulnerable

Endangered

Vulnerable Endangered

Sterile

Critically Endangered

Sterile

Endangered

Endangered

Sterile

Rare

Near Threatened

Sterile

Vulnerable

Endangered

Sterile

Vulnerable

Endangered

References BURCH, J. -in press- How gelling agent and sucrose affect the growth of bryophytes. The Botanic Gardens Micropropagation News. BURCH, J. -2003- Some mosses survive cryopreservation without prior pre-treatment. The Bryologist 106-2): 270-277. BURCH, J. & T. WILKINSON -2002- Cryopreservation of protonemata of Ditrichum cornubicum -Paton) comparing the effectiveness of four cryoprotectant pretreatments. Cryo-Letters 23: 197-208. CHEFFINGS, C.; M. HARPER & A. JACKSON -(Eds.) -2004- Plant Diversity Challenge - The UK's response to the Global Strategy for Plant Conservation. JNCC , Peterborough Church JM, Hodgetts NG, Preston CD, Stewart NF -2001- British red data books – mosses and liverworts. JNCC, Peterborough. EUROPEAN COMMITTEE FOR CONSERVATION OF BRYOPHYTES -1995- Red data book of European bryophytes. ECCB. Trondheim. VITT, D.H. -2000- Peatlands: ecosystems dominated by bryophytes. In Bryophyte Biology, 312-343. Eds. AJ Shaw and B Goffinet. Cambridge University Press, Cambridge.