Clonal forestry in Nordic countries?

Text by: Mari Mette Tollefsrud, Tore Skrøppa, Mikko Tikkinen, Mari Rusanen, Arne Steffenrem. Top photo shows mountainous forest in which spruce forms clusters of clones, photo by Arne Steffenrem, NIBIO/Skogfrøverket.

Commonly used forest reproductive material in the Nordic countries

Across Nordic countries, forest regeneration commonly occurs through various methods. These are: i) Natural regeneration where forests are allowed to regenerate naturally based on the existing tree population. ii) Regeneration based on seed/plants originating from stand seeds. Commonly, stand seed lots are harvested from many trees over a large area. iii) The most prevalent method in the Nordic countries, however, is to use seeds/plants originating from seed orchards. In the seed orchards, seeds are harvested from many clones selected for superiority in one or several traits.

In all the above cases, seeds originate from many open pollinated trees where sexual recombination provides unique combinations of genes and high levels of genetic diversity, demonstrated by differences in growth and adaptability traits.

Production of clonally propagated FMR

Clonal forestry is when the forest reproductive material (FRM) is produced asexually via vegetative propagation from one ancestor stock (ortet/mother plant) of which they are genetically identical to. It is a method that has been used for hundreds of years to propagate requested material for forest plantations. Well known examples are Cryptomeria in Japan and Eucalyptus clones in subtropical areas. Vegetative propagation methods include both “macropropagation” like for instance rooted cuttings and grafting, and “micropropagation” using in vitro tissue culture techniques or somatic embryogenesis.

For production of FRM, rooted cuttings and somatic embryogenesis are the most economically viable methods. For coniferous species however, propagation using cuttings has been problematic due to aging of the ortets. The cuttings develop into branches instead of trees, a problem due to maturation of the starting material. Somatic embryogenesis appears to be the most effective method for vegetative propagation. Through somatic embryogenesis, a single seed gives rise to multiple embryos which are subsequently cultivated into new plants. The technology offers an opportunity to produce FRM both from tested clones as well as mass production of tested families. These two systems are commonly called “clonal forestry” and “family forestry” among professionals.

Before propagation of tested clones, the clones themselves undergo a short-term test cycle in field. The tests might last from a few to maybe 8-10 years. The tests are basis for selection of clones prior to mass production, allowing for the selection of clones with superior performance through the important juvenile phase. The cell lines which are the used for SE, are cryopreserved while waiting for the results from the field tests.

From breeding programs, also full-sib families harboring high genetic gain can be obtained. These families can then be mass produced using somatic embryogenesis. In family forestry, a bulk sample of seeds from the families are mass propagated. It is important to note that in family forestry the parents are tested, not the clones themselves. This means that there can be a greater variation in performance traits (e.g. growth potential or adaptation) compared to tested clones in a clonal forestry.

In family forestry, the number of progenies propagated from each full-sib family, and the relatedness among then, is decisive for the level of genetic diversity maintained in the FRM. For instance, family forestry restricted to clones of 20-40 progenies, would yield the same level of genetic diversity as 20-40 tested clones. If the families being mass-propagated are closely related, genetic diversity may be compromised (e.g. reduced heterozygosity).

In Sweden and Finland, extensive research and development are currently underway to make somatic embryogenesis plants of both spruce and birch more accessible and affordable to the market (Egertsdotter 2018, Tikkinen 2018 and Tikkinen et al. 2019). 

Benefits with clonally propagated FRM

Both clonal forestry and family forestry may bring large benefits when it comes to increased growth, uniformity, and quick transfer of superior material into forest production (e.g. Wu et al. 2019).

A study by Liziniewicz et al. (2018), based on 3–4-year-old nursery plants, showed that selected clones produced almost 30% more compared to non-bred plant material. In another study, Liziniewicz and Berlin (2019) evaluated growth and development of different genetically improved material 13-16 years after plating. They showed that the genetically improved groups all produced higher volumes than the unimproved local provenances. Comparing i) single clones, ii) clonal mixtures and iii) a mixture of bulk-formed families, the increases were 12, 16 and 21 % in height; 10, 17 and 21% for diameter and 23, 44 and 56% for volume production, respectively. Notably, in this study clonal mixtures show higher genetic gain over single clones which varied substantially in performance, suggesting that clonal mixes rather than one or a few clones should be deployed in clonal forestry. Improvements in genetic gain have been documented in other Swedish experiments and through simulations using various models and genetic parameters from trials (Rosvall 2019). In Wu (2019), genetic gain from clonal forestry relative to family forestry is reviewed showing that an extra genetic gain (5-25%) is possible from clone testing and deployment relative to deployment of family forestry.

Vegetative propagation by somatic embryogenesis offers the opportunity for a quick transfer of consistent breeding gain into the forest. For instance, FRM with superior growth, adaptation to specific climatic conditions, or resistance to specific pathogens can be more quickly obtained through somatic embryogenesis than normal seed production in seed orchards. Utilization of gene-based knowledge, through marker-based selection in combination with somatic embryogenesis is a method that may speed up the transfer of genetic gain even more (Edesi et al. 2021). Field tests, however, are important to check for maladaptation and negative traits. How long the field test should last is a trade-off between risk of maladaptation at later stage versus production delay of the FRM, and costs related to cryopreservation of the candidate clones under testing. Integrating genomic selection with somatic embryogenesis is suggested to significantly speed up the advancement of clonal forestry by reducing or even eliminating the need for prolonged clonal testing (Wu 2019). 

Risks with clonally propagated FRM

Genetic diversity is crucial for future adaptation because it provides the raw material for populations to respond to environmental changes, such as climate change, disease outbreaks, and habitat alterations. The highest risk with clonal forestry is the reduction of genetic diversity.

Population fitness has been shown to be positively correlated related to both population size, heterozygosity and quantitative genetic variation (Reed and Frankham 2003). A forest with few genotypes/clones will have a much higher vulnerability to unforeseen abiotic events, as well as biotic factors like new pathogen attacks. If only a few clones are planted, and the stands are left to regenerate naturally for the next rotations with little gene flow from the outside, a reduction of effective populations size will also increase the probability for accumulation of deleterious alleles through genetic drift in the following generations (Kliman et al. 2008; Kimura and Ohta 1969).

Forest trees often carry a significant genetic load, meaning that they have many harmful recessive alleles within their populations. If the clones are highly related, and left to cross with each other, such alleles are more likely to become exposed leading to inbreeding depression negatively affecting the fitness of the population (explained in Ingvarsson and Dahlberg 2018). However, forest trees grow over large geographical areas and have large population sizes. Especially for species such as spruce, pine and birch which are wind pollinated and wind dispersed, gene flow from the surroundings will contribute to increase the effective population size in the next generation.

Clonal forestry may also experience negative effects of the genotype – environment interaction. Even if the clonal material is tested, it is impossible to test for all types of environments. In the Nordic region, climatic gradients are evident both longitudinally, latitudinally and altitudinally, and we know that the microclimate varies largely over short distances, so here this genotype-environment interaction that may be particularly relevant. Risk of plantation failure may therefore be high in a Nordic heterogeneous environment.

Moreover, we lack knowledge related to clonal forestry and its effects on biodiversity, ecosystem services and ecosystem interactions. If forest trees, as key species of the ecosystems, lack genetic diversity and struggle to adapt, it can disrupt the balance of the ecosystem, affecting other species and ecological processes. Basically, we have very little knowledge on what is “sufficient” amount of genetic diversity to secure a resilient future forest (but see e.g. Wu 2019 that has reviewed theoretical models on ”safe” number of clones). 

Regulation of vegetatively propagated material across Nordic countries

European Union Directive 1999/105 emphasizes the importance of clonal mixtures containing an adequate level of genetic diversity. Similarly, the OECD forest seed and plant scheme specifies that clonal mixtures must exhibit sufficient genetic variation. When marketing clonally propagated material, it is imperative to specify the number of clones employed in the mixture along with their proportions on the certificate. It rests within individual countries to regulate factors such as number of clones and/or their shares, duration of clonal use in the production, or over how large areas clonally propagated material can be deployed.

EUFORGENs recommendation number 32 encourages clonal mixtures and controls on clone status in clonal stands. Generally, clones of the ‘Tested’ category should be preferred, although to minimize ecological risks and increase landscape diversity, mosaics of a variety of different monoclonal stands are recommended.

Clonally propagated reproductive material has so far had limited use within the Nordic countries, resulting in a corresponding scarcity of regulations governing its utilization. In Finland, the first somatic embryogenesis based basic material was registered in 2017 and since then, somatic embryogenesis germinant have been marketed to forest tree nurseries in commercial pilot projects (Tikkinen et al. 2019). Neither Sweden nor Norway have approved any somatic embryogenesis based basic material for forestry use.

In Finland the possibility to use individual clones or accepted combinations of several clones depends on the species and the category of the basic material. There are specifications on the number of vegetatively produced plants certified under the category “qualified”, but no restriction on the numbers if they are certified under the category “tested”. E.g., for basic material from “parents of family” certified under qualified, a maximum number of 4 million plants from each family can be produced. For clones certified under qualified, a maximum of 1 million plants can be produced from a single ortet, i.e., 1 million copies of a clone allowed. There are also specifications on how much the percentage of single clones in a clonal mixture can vary. The rules and regulations are found in the Decree on Trade in Forest Reproductive Material as referred to below. 

Sweden does not have specific rules on a minimum number of clones in a clonal mixture, or maximum number of copies used per clone. Sweden has on the other hand set a limit that states a maximum of five percent of the area of productive forest land within a utilization unit can be regenerated with vegetatively propagated material, up to a limit of 20 hectares (up to 400 ha) within a utilization unit. A utilization unit encompasses the productive forests, belonging to the same owner, within one or more municipalities. The use of vegetative material over areas larger than half a hectare must be pre-notified to the Swedish Forest Agency at least six weeks before planting.

Norway’s current regulation emphasizes the use of clonal mixtures, preferably in combination with seed plants. It also regulates a minimum number of clones to be used regenerating a forest site. Clonal mixtures should include a minimum of 30 tested clones, and the plants should originate from at least 10 unrelated parent couples. Furthermore, there is a restriction allowing a maximum of 50 plants from each clone to be deployed in a single location. If the clonal mixtures are combined with seed plants, the number of plants from each clone can be increased to 100. From bulk-formed families, there are also specific requirements in place. It is worth nothing that Norway’s regulations are currently undergoing a review process and are expected to be updated soon. 

In Denmark, there are currently no specific regulations governing the minimum number of clones. Instead, the primary focus revolves around quality criteria for cuttings of Populus spp. As for now, there is only one approved clone of Populus. The decision to rely on a single clone is driven by the need to strike a balance between genetic variability and the assurance of short-term performance.  

Basically, regulations related to the use of FRM and clones are old and not adjusted to extensive use of somatic embryogenesis plants. Nordic countries should thus review and if needed update their regulations in line with the development of new technologies and new types of FRM. In evaluation of regulations, special focus should be given to the traceability of clonally propagated materials to be able to take actions if needed. Moreover, an increase in use of clonal or family forestry should release a corresponding increase in the conservation of genetic resources. 

Concluding remarks

Links to the regulations on forest reproductive material in the different countries

Denmark: https://www.retsinformation.dk/eli/lta/2023/1418

Sweden: The Swedish Forest Agency's regulations on production for marketing, marketing, and importation for marketing of forest reproductive material' SKSFS 2002:2, from 2002, which follows the EU directive. https://www.skogsstyrelsen.se/globalassets/lag-och-tillsyn/foreskrifter-efter-amne/skogsvard/sksfs-2002-2-skogsstyrelsens-foreskrifter-om-produktion-for-saluforing-saluforing-samt-inforsel-for-saluforing-av-skogsodlingsmaterial.pdf

In addition, Sweden regulates the use of vegetatively propagated material in the Swedish Forest Agency's regulations (SKSFS 2011:7) https://www.skogsstyrelsen.se/globalassets/lag-och-tillsyn/grundforeskrifter-samt-andringar/sksfs-2011-7/sksfs-2014-7-skogsstyrelsens-foreskrifter-och-allmanna-rad-till-skogsvardslagen.pdf

Finland: Food and safety institution, compilation of the relevant regulations https://www.ruokavirasto.fi/en/plants/forest-tree-seed-and-seedling-production

Decree on Trade in Forest Reproductive Material / Maa- ja metsätalousministeriön asetus metsänviljelyaineiston kaupasta 1055/2002 (2002) http://www.finlex.fi/fi/laki/alkup/2002/20021055]

Norway: Regulation on forest seeds and forest plants https://lovdata.no/dokument/SF/forskrift/1996-03-01-291

References

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Egertsdotter U. 2018. Plant physiological and genetical aspects of the somatic embryogenesis process in conifers. Scandinavian Journal of Forest Research 34: 360-369. https://doi.org/10.1080/02827581.2018.1441433

Ingvarsson PK, Dahlberg H. (2018) The effects of clonal forestry on genetic diversity in wild and domesticated stands of forest trees. Scandinavian Journal of Forest Research 5: 370-379. https://doi.org/10.1080/02827581.2018.1469665

Kimura M, Ohta T. (1969) The average number of generations until fixation of a mutant gene in a population. Genetics 61:763-771 https://doi.org/10.1093/genetics/61.3.763

Kliman R, Sheehy B, Schultz J (2008) Genetic Drift and Effective Population Size. https://www.nature.com/scitable/topicpage/genetic-drift-and-effective-population-size-772523/

Lelu-Walter, M-A, et al. 2013. Somatic embryogenesis in forestry with a focus on Europe: state of-the-art, benefits, challenges and future direction. Tree Genetics & Genomes 9: 883-899. https://doi.org/10.1007/s11295-013-0620-1

Liziniewicz M & Berlin M. 2019. Differences in growth and areal production between Norway spruce (Picea abies L. Karst) regeneration material representing different levels of genetic improvement. Forest Ecology and Management 435: 158-169. https://doi.org/10.1016/j.foreco.2018.12.044 

Liziniewicz M, Berlin M & Karlsson B. 2018. Early assessments are reliable indicators for future volume production in Norway spruce (Picea abies L. Karst) genetic field trials. Forest Ecology and Management 411:75-81. https://doi.org/10.1016/j.foreco.2018.01.015

Reed DH, Frankham R. 2003. Correlation between fitness and genetic diversity. Conservation Biology 17:230-237. https://www.jstor.org/stable/3095289

Rosvall O. 2019. Using Norway spruce clones in Swedish forestry: Swedish forest conditions, tree breeding program and experiences with clones in field trials. Scandinavian Journal of Forest Research 34: 324-351. https://doi.org/10.1080/02827581.2018.1562566

Tikkinen M. 2018. Improved propagation efficiency in a laboratory–nursery interface for somatic embryogenesis in Norway spruce. Dissertationes Forestales 265. 35 p. https://doi.org/10.14214/df.265

Tikkinen M, Varis S, Välimäki S, Nikkanen T & Aronen T. 2019. Somatic embryogenesis of Norway spruce in Finland – seven years from start to first commercial pilots. The Fifth International Conference of the International Union of Forest Research Organizations Unit 2.09.02: Somatic Embryogenesis and Other Vegetative Propagation Technologies, September 10-15, 2018, Coimbra, Portugal. https://www.iufro.org/fileadmin/material/publications/proceedings-archive/20902-coimbra18.pdf

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Wu HK. 2019. Benefits and risks of using clones in forestry – a review. Scandinavian Journal of Forest Research 34: 352-359. https://doi.org/10.1080/02827581.2018.1487579