1. Active grassland restoration has gained in importance to mitigate the dramatic decline of farmland biodiversity. While there is evidence that such operations are generally effective in promoting plant diversity, little is known about the effectiveness of the different methods applied. Restoration methods can differ in intensity of seed bed preparation, seed source and technique of seed application.

2. In this systematic literature search and meta-analysis, we screened the literature for studies of the restoration of mesic grasslands in temperate Europe. We focused on active restoration experiments that included a treatment and lasted more than three years. We evaluated the influence of restoration factors on plant species richness relative to non-restored controls.

3. We found 187 articles that investigated the outcome of operations aimed at actively restoring mesic temperate grasslands. Most articles focused on plants, with only 9.6% dealing with other organisms (e.g. beetles, pollinating insects). Many papers had to be excluded due to incomplete data, too short study duration and/or lack of an adequate control. This resulted in 13 articles fulfilling our criteria for inclusion, yielding a total of 56 data points for the meta-analysis.

4. Restoration actions increased plant species richness by, on average, 17.4%, compared to controls. The seed source explained a significant amount of variation in plant species richness: seeds originating from a speciose donor grassland had a positive effect. This effect was even enhanced when combined with a commercial seed mix, whereas commercial seed mixes alone had no significant effect. We did not observe any effect of other factors, such as the type of seed bed preparation or the seed application method.

5. A seed-source obtained from species-rich grasslands seems to be key to efficient grassland restoration in mesic grasslands of temperate Europe. Even though seeds from a speciose donor grassland should be preferred over commercial seeds, associating natural and commercial seed mixes increases plant species richness. This systematic literature search further revealed two major research gaps in grassland restoration ecology: a deficit in long-term investigations as well as a deficit in studies focusing on non-plant organisms.

We followed the guidelines of the Collaboration for Environmental Evidence (Pullin & Stewart, 2006) and the ROSES standard (see ROSES form in Table S1) to conduct our systematic literature search. By doing so, we ensure repeatability of our search and screening process (Romanelli, Meli, Naves, Alves, & Rodrigues, 2021; Romanelli, Silva, et al., 2021). We prepared a protocol that was peer-reviewed and published (Slodowicz, Humbert, & Arlettaz, 2019). As some points of the original protocol had to be amended due to some unexpected issues, the following section contains the updated protocol.

Systematic literature search

We formulated our research question according to the PICO-structure (population, intervention, control, outcome): Do different seed addition methods for the restoration or re-creation of species-rich grasslands differ in their effectiveness to enhance the diversity of plants? (See Table S2 for details on the question components). Based on the question components, we developed an initial search string that went through a scoping process. This initial string was used in a search in the Web of Science database. We compared the search result with the reference lists of two reviews on the same topic (Hedberg & Kotowski, 2010; Kiehl, Kirmer, Donath, Rasran, & Hölzel, 2010). To achieve adequate sensitivity, we adapted the search string, until no references of the reviews were missed. We used the final search string as a template for our database searches and adapted it accordingly to the requirements of the respective databases: (grassland* OR meadow* OR pasture*) AND (restor* OR seed addition OR seed transfer OR hay transfer OR sow* OR strew*) AND (*diversity OR enhance* OR success OR richness OR establish*). We conducted the database searches between 26 November 2019 and 16 March 2020 in Web of Science Core Collection, Scopus, Directory of open access journals (DOAJ) and eThOs. In addition, we used the ‘Publish or perish’ software (Harzing, 2007) to search articles in Google Scholar and retained the first 1000 hits. A detailed overview of the search string development and database searches can be found in the supplementary material (Table S3, Appendix S1).

To complement the database search, we looked for other publications and grey literature in Google, organisational websites and through direct requests to authors. The searches and requests were done in English, French, German and Polish. We removed duplicates automatically using the JabRef Reference Manager (JabRef Development Team, 2021).

Article screening

Screening was done on title, abstract and full-text level by two reviewers (DS, AD). A third reviewer (JYH, co-author of this paper) checked for inclusion consistency using Cohen’s Kappa (Pullin & Stewart, 2006) on a subsample of 500 articles from each reviewer, respectively. A Kappa score of > 0.6 indicated high consistency between reviewers. At title and abstract level, we included all restoration studies that were conducted within temperate Europe. At full-text level, the articles had to fulfill our eligibility criteria for inclusion (Table 1). We distinguished three grassland habitat types: dry, wet and mesic. We considered grassland habitats to be “dry” if the substrate was coarse or sandy with low water retention capacity and low amount of nutrients in the soil (e.g., Wolff et al., 2017). We considered grasslands to be “wet” if they were peat or fen meadows (e.g., Klimkowska et al., 2010), or alluvial meadows (e.g., Schmiede et al., 2009). All other grasslands were considered mesic and therefore eligible populations. In our meta-analysis, we have intentionally excluded short-term studies of less than three years duration to reduce confounding factors. In effect, the first two years after restoration are typically characterized by a rise in species richness (Albert et al., 2019; Baasch, Engst, Schmiede, May, & Tischew, 2016; Freitag et al., 2021). This is often due to the presence of ruderal species, which have become dominant in the seed bank after perturbation (Valkó, Rádai, & Deák, 2022). Once the grassland species become more dominant, the number of ruderal species diminishes (Albert et al., 2019). This is reflected in a slight decline in species richness after the second year of restoration (Freitag et al., 2021). For this reason, we focused on the mid-term, thus ensuring that the plant community had become more stable. Yet, to identify research gaps in a later phase, we compiled a separate list of all excluded European studies at full-text screening. All articles on European grassland restoration excluded during the full-text screening step are provided in Table S4.

Data extraction and moderators

The geographic location of each restoration site was recorded and, if necessary, changed into decimal degrees. If the site coordinates were not provided, we looked for a locality (such as a city, village, or a protected area) in the site description of the respective article and determined the coordinates from Google Maps. As potential moderator variables (effect modifiers) we included the control type, type of study design, seed source, seed material, seed bed preparation (seed bed preparation is requisite for efficient seed addition), former land-use, restoration duration, number of experimental replicates, as well as vegetation survey plot size (see Table 2 for a detailed definition of each moderator). These moderator variables are either linked to applied aspects of grassland restoration (e.g., seed source), which are relevant for practitioners, or to experimental aspects (e.g., control type). As response variables, we extracted the mean plant species richness and a measure of variance (which was converted to standard deviation if necessary) from the restored and control plots. We extracted these from summary tables, calculated it from raw data or extracted it from the figures using the WebPlotDigitizer (Rohatgi, 2021). We contacted the authors by e-mail to request missing data if relevant data was missing or not extractable from the study. A list of data sources used in the study is provided in the Data sources section.

References

• Albert, Á. J., Mudrák, O., Jongepierová, I., Fajmon, K., Frei, I., Ševčíková, M., … Doležal, J. (2019). Grassland restoration on ex-arable land by transfer of brush-harvested propagules and green hay. Agriculture, Ecosystems and Environment, 272(November 2018), 74–82. doi: 10.1016/j.agee.2018.11.008
• Baasch, A., Engst, K., Schmiede, R., May, K., & Tischew, S. (2016). Enhancing success in grassland restoration by adding regionally propagated target species. Ecological Engineering, 94(2016), 583–591. doi: 10.1016/j.ecoleng.2016.06.062
• Freitag, M., Klaus, V. H., Bolliger, R., Hamer, U., Kleinebecker, T., Prati, D., … Hölzel, N. (2021). Restoration of plant diversity in permanent grassland by seeding: Assessing the limiting factors along land-use gradients. Journal of Applied Ecology, 58(8), 1681–1692. doi: 10.1111/1365-2664.13883
• Harzing, A. W. (2007). Publish or Perish. Retrieved from https://harzing.com/resources/publish-or-perish/
• Hedberg, P., & Kotowski, W. (2010). New nature by sowing? The current state of species introduction in grassland restoration, and the road ahead. Journal for Nature Conservation, 18(4), 304–308. doi: 10.1016/j.jnc.2010.01.003
• JabRef Development Team. (2021). JabRef --- An open-source, cross-platform citation and reference management software. Retrieved from https://www.jabref.org/
• Kiehl, K., Kirmer, A., Donath, T. W., Rasran, L., & Hölzel, N. (2010). Species introduction in restoration projects - Evaluation of different techniques for the establishment of semi-natural grasslands in Central and Northwestern Europe. Basic and Applied Ecology, 11(4), 285–299. doi: 10.1016/j.baae.2009.12.004
• Klimkowska, A., Kotowski, W., Van Diggelen, R., Grootjans, A. P., Dzierz̈a, P., & Brzezińska, K. (2010). Vegetation Re-development After Fen Meadow Restoration by Topsoil Removal and Hay Transfer. Restoration Ecology, 18(6), 924–933. doi: 10.1111/j.1526-100X.2009.00554.x
• Pullin, A. S., & Stewart, G. B. (2006). Guidelines for systematic review in conservation and environmental management. Conservation Biology, 20(6), 1647–1656. doi: 10.1111/j.1523-1739.2006.00485.x
• Rohatgi, A. (2021). Webplotdigitizer: Version 4.5. Retrieved from https://automeris.io/WebPlotDigitizer
• Romanelli, J. P., Meli, P., Naves, R. P., Alves, M. C., & Rodrigues, R. R. (2021). Reliability of evidence-review methods in restoration ecology. Conservation Biology, 35(1), 142–154. doi: 10.1111/cobi.13661
• Romanelli, J. P., Silva, L. G. M., Gonçalves, M. C. P., Naves, R. P., de Almeida, D. R. A., de Resende, A. F., & Rodrigues, R. R. (2021). Repeatability of the searching process in reviews of restoration outcomes. Restoration Ecology, 29(8), 1–10. doi: 10.1111/rec.13496
• Schmiede, R., Donath, T. W., & Otte, A. (2009). Seed bank development after the restoration of alluvial grassland via transfer of seed-containing plant material. Biological Conservation, 142(2), 404–413. doi: 10.1016/j.biocon.2008.11.001
• Slodowicz, D., Humbert, J. Y., & Arlettaz, R. (2019). The relative effectiveness of seed addition methods for restoring or re-creating species rich grasslands: A systematic review protocol. Environmental Evidence, 8(1), 1–7. doi: 10.1186/s13750-019-0174-2
• Valkó, O., Rádai, Z., & Deák, B. (2022). Hay transfer is a nature-based and sustainable solution for restoring grassland biodiversity. Journal of Environmental Management, 311(October 2021), 114816. doi: 10.1016/j.jenvman.2022.114816
• Wolff, C., Gilhaus, K., Hölzel, N., & Schneider, S. (2017). Status and restoration potential of heathlands and sand grasslands in the southwest of Luxembourg. Tuexenia, 37(1), 179–200. doi: 10.14471/2017.37.007

Data sources

• Albert, Á.-J., Mudrák, O., Jongepierová, I., Fajmon, K., Frei, I., Ševcíková, M., … Dolezal, J. (2019). Grassland restoration on ex-arable land by transfer of brush-harvested propagules and green hay. Agriculture, Ecosystems and Environment, 272(November 2018), 74–82. doi: 10.1016/j.agee.2018.11.008
• Auestad, I., Austad, I., & Rydgren, K. (2015). Nature will have its way: Local vegetation trumps restoration treatments in semi-natural grassland. Applied Vegetation Science, 18(2), 190–196. doi: 10.1111/avsc.12138
• Baasch, A., Engst, K., Schmiede, R., May, K., & Tischew, S. (2016). Enhancing success in grassland restoration by adding regionally propagated target species. Ecological Engineering, 94(2016), 583–591. doi: 10.1016/j.ecoleng.2016.06.062
• Chevalier, R., Le Bris, C., Swiderski, C., Baron, S., Isselin-Nondedeu, F., Lesage, C., & Michau, F. (2018). Restauration de la biodiversité floristique des bordures de champs par semis de fleurs sauvages : premiers résultats obtenus en Beauce. Sciences Eaux & Territoires, Numéro 25(1), 52. doi: 10.3917/set.025.0052
• Dolnik, C., Jansen, D., & Rickert, B.-H. (2020). Praxisleitfaden BlütenMeer 2020. Molfsee. Retrieved from https://www.stiftungsland.de/fileadmin/pdf/Bluetenmeer2020/20-2841_Praxisleitfaden_Naturschutz_Internet.pdf
• Freitag, M., Klaus, V. H., Bolliger, R., Prati, D., Schäfer, D., Hinderling, J., … Hölzel, N. (2021). Data and Code for the manuscript Freitag et al. “Restoration of plant diversity in permanent grassland by seeding: assessing the limiting factors along land-use gradients.” doi: https://doi.org/10.25829/bexis.30915-10
• Fritch, R. A., Sheridan, H., Finn, J. A., Kirwan, L., & hUallacháin, D. Ó. (2011). Methods of enhancing botanical diversity within field margins of intensively managed grassland: A 7-year field experiment. Journal of Applied Ecology, 48(3), 551–560. doi: 10.1111/j.1365-2664.2010.01951.x
• Gentili, R., Montagnani, C., Gilardelli, F., Guarino, M. F., & Citterio, S. (2017). Let native species take their course: Ambrosia artemisiifolia replacement during natural or “artificial” succession. Acta Oecologica, 82, 32–40. doi: 10.1016/j.actao.2017.05.007
• Hovd, H. (2008). Occurrence of meadow herbs in sown and unsown ploughed strips in cultivated grassland. Acta Agriculturae Scandinavica Section B: Soil and Plant Science, 58(3), 208–215. doi: 10.1080/09064710701593046
• Prach, K., Jongepierová, I., Řehounková, K., & Fajmon, K. (2014). Restoration of grasslands on ex-arable land using regional and commercial seed mixtures and spontaneous succession: Successional trajectories and changes in species richness. Agriculture, Ecosystems and Environment, 182, 131–136. doi: 10.1016/j.agee.2013.06.003
• Smith, B. M., Diaz, A., & Winder, L. (2017). Grassland habitat restoration: Lessons learnt from long term monitoring of Swanworth Quarry, UK, 1997-2014. PeerJ, 2017(11), 1997–2014. doi: 10.7717/peerj.3942
• Veen, G. F., van der Putten, W. H., & Bezemer, T. M. (2018). Biodiversity-ecosystem functioning relationships in a long-term non-weeded field experiment. Ecology, 99(8), 1836–1846. doi: 10.1002/ecy.2400
• Woodcock, B. A., Vogiatzakis, I. N., Westbury, D. B., Lawson, C. S., Edwards, A. R., Brook, A. J., … Mortimer, S. R. (2010). The role of management and landscape context in the restoration of grassland phytophagous beetles. Journal of Applied Ecology, 47(2), 366–376. doi: 10.1111/j.1365-2664.2010.01776.x
• Woodcock, B. A., Edwards, A. R., Lawson, C. S., Westbury, D. B., Brook, A. J., Harris, S. J., … Mortimer, S. R. (2010). The Restoration of Phytophagous Beetles in Species-Rich Chalk Grasslands. Restoration Ecology, 18(5), 638–644. doi: 10.1111/j.1526-100X.2008.00472.x
• Woodcock, B. A., Westbury, D. B., Brook, A. J., Lawson, C. S., Edwards, A. R., Harris, S. J., … Mortimer, S. R. (2012). Effects of seed addition on beetle assemblages during the re-creation of species-rich lowland hay meadows. Insect Conservation and Diversity, 5(1), 19–26. doi: 10.1111/j.1752-4598.2011.00132.x

Note that the three Articles by Woodcock, Vogiatzakis, et al., (2010), Woodcock, Edwards, et al., (2010) and Woodcock et al., (2012) shared several study sites, which we included in our meta-analysis. Their effect sizes are summarized in Fig. 3 as “Woodcock et al., 2010”.

All analyses were performed with R version 4.1.1 (R Core Team, 2021) using the metafor (Viechtbauer, 2010) and puinform packages (van Aert, 2018).