Grassland Restoration: Relative effectiveness of different restoration methods on plant and invertebrate diversity

Semi-natural grasslands in Central Europe can harbor a big variety of plant and invertebrate species. These grasslands have suffered from a strong decline mainly due to agricultural intensification. Agri-environment schemes have been introduced in Europe to promote farmland biodiversity, but they were only little effective, especially so in grasslands. To mitigate this dramatic decline of farmland biodiversity, active grassland restoration is nowadays widely applied and has gained in importance in research. While there is evidence that active grassland restoration is generally effective in promoting plant diversity, little is known about the effectiveness of the factors involved in such restoration operations, such as soil disturbance intensity, seeding methods and seed source. Furthermore, there are no studies showing whether the soil disturbances that are linked to grassland restoration could be harmful to the resident ground-dwelling invertebrates of a grassland that is going to be restored. In this PhD thesis I investigated the effects of different factors involved in active grassland restoration on the restoration outcome. I first conducted a systematic review and meta-analysis to evaluate the relative influence of these factors on restoration efficacy, focusing on plant species richness, and to identify research gaps. In parallel, a field-scale restoration experiment replicated across the Swiss Plateau was launched, with this PhD being part of the restoration experiment. This experiment served to study whether hay transfer can be used to effectively transfer invertebrates from one meadow to another and to which degree soil disturbances linked to grassland restoration are harmful to ground-dwelling invertebrates. Furthermore, I studied the short-term outcome of different restoration methods on the plant community. The restoration methods were: (i) control with no seed addition and no soil disturbance, (ii) hay transfer from a species-rich donor meadow on a harrowed receiver meadow, (iii) hay transfer from a species-rich donor meadow on a ploughed receiver meadow, (iv) sowing of a commercial seed mixture on a ploughed receiver meadow and, (v) sowing of a brush- or vacuum harvested seed mixture on a ploughed receiver meadow. Baseline data of plants and ground-dwelling invertebrates (represented by ground beetles and spiders) was collected in 2018, i.e., one year before restoration took place. In early summer 2019 we restored 48 meadows and additionally sampled invertebrates from the hay transfer treatments. One year after restoration (in 2020) we resampled invertebrates and two years after restoration (in 2021) we reconducted vegetation surveys on the restored and control meadows. The systematic review and meta-analysis revealed the importance of the seed source in grassland restoration, while different soil disturbance intensities or seeding methods (green hay or harvested seeds) do not show differences in the restoration performance on plant species richness. We also identified research gaps, such as little focus on invertebrates in grassland restoration studies and few field scale experiments. With the restoration experiments we could show that invertebrates can be transferred successfully with the hay and that there was no mid-term negative impact on the ground-dwelling invertebrate community due to soil disturbances linked to restoration, no matter of the disturbance intensity. Finally, all four restoration methods that we tested in our experiment significantly increased the plant species richness after two years. All together the present thesis is a contribution to the relatively young research field of restoration ecology with evidence-based recommendations and it comes in a timely moment within the UN decade on restoration

Transfer of invertebrates with hay during restoration operations of extensively managed grasslands in Switzerland

Introduction Hay transfer from a speciose donor meadow to a species-poor receiver grassland is an established method to restore species-rich grassland plant communities. However, it has rarely been investigated to which extent invertebrates can be transferred with hay during such operations, which was the aim of this study. Methods Sampling was conducted in eight sites of the Swiss lowlands with one donor meadow and two receiver sites each. On the receiver sites, three to four white bed sheets of one square meter each were deployed on the ground to receive a standard quantity of fresh hay just transferred from the donor meadow. All living invertebrates were collected from these sheets with an aspirator and subsequently identified to order level. Results On average (± SD), 9.2 ± 11.3 living invertebrates per square meter were transferred with the hay. Beetles were the most abundant species group, representing 46.9% of all transferred invertebrates, followed by true bugs (8.9%) and spiders (7.0%). More individuals were transferred when the donor meadow was mown with a hand motor bar mower than with a rotary disc mower. Similarly, more invertebrates were transferred when the hay was transported loosely with a forage wagon than compacted as bales. Discussion While this study demonstrates that living invertebrates can be transferred with the hay, their subsequent survival and establishment remains to be explored. Implications for insect conservation We recommend using a hand motor bar mower and a forage wagon for increasing the survival probability of invertebrates in hay transfer

Granulobasidium vellereum (Ellis &Cragin) Jülich, a promising biological control agent

Studies involving biological control of wood decay fungi and pests in forest areas are scarce. The basidiomycete Granulobasidium vellereum (Ellis & Cragin) Jülich has been isolated as wood endophyte from London Plane trees (Platanus acerifolia (Ait.) Willd). This basidiomycete produces a variety of sesquiterpenes with antifungal activity. In this study we evaluated the potential activity of G. vellereum against wood decay fungi and fungi associated with the ambrosia beetle Megaplatypus mutatus Chapuis, an important forest pest in Populus sp. A combination of in vitro assays was made, in cultures and on wood blocks. Granulobasidium vellereum’s Volatile Organic Compounds (FVOCs) were characterized and their potential role in biocontrol was assessed. Granulobasidium vellereum did not cause a significant loss of weight on P. acerifolia and Populus wood and inhibited the growth of the target fungi, mainly when inoculated first. Up to nineteen volatile compounds were determined in G. vellereum strains. The growth of all target fungi was inhibited by FVOCs and in some cases the morphology of the fungi was altered. These results indicate that G. vellereum can be used as a Biological Control Agent (BCA) of xylophagous fungi and fungi related to forest pests. Further investigations should focus on developing application strategies for M. mutatus management.Fil: Robles, Carolina Analía. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Micología y Botánica. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Micología y Botánica; ArgentinaFil: Ceriani Nakamurakare, Esteban Daniel. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Biodiversidad y Biología Experimental. Laboratorio de Micología Experimental; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Slodowicz, Mariel Pamela. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones Científicas y Técnicas para la Defensa. Centro de Investigación de Plagas e Insecticidas; ArgentinaFil: Gonzalez Audino, Paola Andrea. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones Científicas y Técnicas para la Defensa. Centro de Investigación de Plagas e Insecticidas; ArgentinaFil: Carmaran, Cecilia Cristina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Micología y Botánica. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Micología y Botánica; Argentin

The relative effectiveness of seed addition methods for restoring or re‑creating species rich grasslands: a systematic review protocol

Background: Extensively managed grasslands in temperate biomes are capable of harboring a big variety of plant and invertebrate species. Yet, they have suffered from a strong decline in the past decades mainly due to agricultural intensification. Agri-environment schemes have been introduced in Europe in order to promote farmland biodiversity, but they were only little effective, especially so in grasslands. Not surprisingly, grassland restoration and recreation through active seed addition has thus gained in importance in the recent years. The most common methods used rely either on the addition of commercial seed mixes, on the addition of seeds collected from a speciose donor grassland or on transferring hay from a speciose donor grassland after the soil of the receiver site has been prepared (ploughing, harrowing or topsoil removal). While there is evidence that these restoration methods may contribute to improve the biodiversity of grasslands, especially plant diversity, their relative effectiveness remains poorly known. The aim of this systematic review is to scrutinize the peer-reviewed literature for scientific evidence about comparative effectiveness. Methods: We will search for peer-reviewed journal articles in bibliographic databases and grey literature in search engines and organizational websites dealing with at least one of the above mentioned seed addition methods. We will only include studies which were carried out in temperate Europe. Through a scoping exercise a search string was developed which was based on a previously prepared test-list. The search string was then tested for validity with two independent reference lists. Screening will be done on the title, abstract and full-text level and consistency checking will be done on a random subsample by a second reviewer. After critically appraising internal validity of the retained studies, data on the responses of plants and invertebrates as well as all relevant meta-data will be extracted and coded. A meta-analysis will be conducted on studies with high internal validity whereas a narrative analysis will be done with descriptive statistics on studies with medium internal validity. Potential effect modifiers like study duration, former land use or local climate will be included in the analysis as moderators

The relative effectiveness of different grassland restoration methods: A systematic literature search and meta-analysis

1. Active grassland restoration has gained importance in mitigating the dramatic decline of farmnland 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 method 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 for more than 3 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

L'efficacité relative des différentes méthodes de restauration des prairies : une recherche systématique de la littérature et une méta‐analyse

Active grassland restoration has gained importance in mitigating the dramatic decline of farmnland 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 method of seed application. 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 for more than 3 years. We evaluated the influence of restoration factors on plant species richness relative to non‐restored controls. 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. 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. 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.La restauration active des prairies a gagné en importance pour atténuer le déclin dramatique de la biodiversité des terres agricoles. Bien qu'il soit prouvé que de telles opérations sont généralement efficaces pour promouvoir la diversité végétale, on sait peu de choses sur l'efficacité des différentes méthodes appliquées. Les méthodes de restauration peuvent différer selon l’intensité de la préparation du lit de semence, la source des graines et la méthode d’application des semis. Dans cette recherche systématique de la littérature et cette méta‐analyse, nous avons examiné la littérature pour trouver des études sur la restauration des prairies mésiques en Europe tempérée. Nous nous sommes concentrés sur des expériences de restauration active incluant un témoin et un suivi d'au moins 3 ans. Nous avons évalué l'influence des facteurs de restauration sur la richesse en espèces végétales par rapport aux témoins non restaurés. Nous avons trouvé 187 articles examinant les résultats des opérations visant à restaurer activement les prairies mésiques tempérées. La plupart des articles se concentraient sur les plantes, avec seulement 9,6 % traitant d'autres organismes (par exemple les coléoptères, les insectes pollinisateurs). De nombreux articles ont dû être exclus en raison de données incomplètes, d'une durée d'étude trop courte et/ou d'un manque de témoin adéquat. Cela a abouti à 13 articles répondant à nos critères d'inclusion, produisant un total de 56 points de données pour la méta‐analyse. Les actions de restauration ont augmenté la richesse en espèces végétales de 17,4 % en moyenne par rapport aux témoins. La source des graines explique une variation significative de la richesse en espèces végétales : les graines provenant d’une prairie donneuse d’espèces ont eu un effet positif. Cet effet était même renforcé lorsqu’il était combiné avec un mélange de graines commerciales, alors que les mélanges de graines commerciales seuls n’avaient aucun effet significatif. Nous n’avons observé aucun effet d’autres facteurs, tels que le type de préparation du lit de semence ou la méthode d’application des semis. Une source de graines obtenue à partir de prairies riches en espèces semble être la clé d'une restauration efficace des prairies mésiques d'Europe tempérée. Même si les graines provenant d’une prairie donneuse d’espèces doivent être préférées aux graines commerciales, associer des mélanges de graines naturelles et commerciales augmente la richesse en espèces végétales. Cette recherche systématique de la littérature a en outre révélé deux lacunes majeures dans la recherche sur l'écologie de la restauration des prairies : un déficit dans les enquêtes à long terme ainsi qu'un déficit dans les études axées sur les organismes non végétaux

Sex pheromone component produced by microbial associates of the forest pest Megaplatypus mutatus

Megaplatypus mutatus (Chapuis) (Coleoptera: Platypodidae) is an ambrosia beetle native to South America that causes economic loss and was recently introduced to Italy, where it attacks and damages live poplar trees. Sulcatol and sulcatone are male-produced pheromone components involved in the mating process of M. mutatus. Their relative proportions are highly variable among insects, although the temporal pattern shows that initially only sulcatol is present, and sulcatone increases with time, until they are finally both depleted. Sulcatol and sulcatone may be produced de novo by the beetles, they may be produced by fungi, or both pathways may contribute to their production. Sulcatol is stored in the males’ hindgut but sulcatone is only present in emissions, so there is an oxidation process to transform the alcohol to the ketone before or during pheromone release. It is our hypothesis that fungi associated with M. mutatus are responsible for this process. In this work, we studied a possible contribution of associated microorganisms in the conversion of sulcatol into sulcatone and its consequent role in the temporal release pattern of these sex pheromone components observed in male insects. Moreover, we inhibited the postulated enzymes involved in this pheromone conversion process – 3-hydroxy-3-methyl-glutatyl-CoA reductase (HMGR) and P450 enzymes of a fungal strain – and added an antibiotic and a fungicide to the homogenate during sulcatol-sulcatone conversion. Among the fungal species, particular interest was given to Graphium basitruncatum (Matsush.) Seifert & Okada (Microascales), as it is present in male but not in female exoskeletons and in insect gallery samples, suggesting a possible different role in pherome production, as the male is the pheromone-producing sex. Several isolated strains were able to convert sulcatol to sulcatone, whereas the fungus G. basitruncatum showed the highest production of this ketone. Additionally, inhibition of P450 enzymes and HMGR from G. basitruncatum on this alcohol-ketone conversion demonstrated that HMGR is involved in sulcatone generation using sulcatol as precursor, and that P450 enzymes are not. Finally, sulcatone production diminished significantly in homogenized tissues of male and female M. mutatus following addition of an antibiotic and a fungicide. The results suggest that fungi associated with M. mutatus are involved in pheromone production.Fil: Slodowicz, Mariel Pamela. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones Científicas y Técnicas para la Defensa. Centro de Investigación de Plagas e Insecticidas; ArgentinaFil: Ceriani Nakamurakare, Esteban Daniel. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Biodiversidad y Biología Experimental; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria; ArgentinaFil: Carmaran, Cecilia Cristina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Biodiversidad y Biología Experimental; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria; ArgentinaFil: Gonzalez Audino, Paola Andrea. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones Científicas y Técnicas para la Defensa. Centro de Investigación de Plagas e Insecticidas; Argentin

Physicochemical Properties and Field Evaluation of Monolithic Wax Formulations for the Controlled Release of a Forest Pest Pheromone

This work shows the way to develop a monolithic device for the release of an active agent. Different waxes to prepare monolithic dispensers were used. Studies to characterize the release rate of the pheromone dispensers, and to determine many physicochemical properties of the active components and waxes utilized as matrices, were conducted. The advantages of using monolithic devices as an insect pheromone dispenser in order to control an insect that attacks commercial poplar plantations were demonstrated. Also, the practical application employing monolithic dispensers manufactured in this paper with the sex pheromone of the ambrosia beetle Megaplatypus mutatus (Coleoptera: Platypodidae) was verified. This forest pest is known to attack tree plantations in South America. The release rates were mainly dependent of matrix hardness and viscosity, as well as the particle size of fillers. A good performance of the formulated monolithic dispensers was observed during the field trials, with a reduction of damage levels of 40%.Fil: Slodowicz, Mariel Pamela. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones Científicas y Técnicas para la Defensa. Centro de Investigación de Plagas e Insecticidas; ArgentinaFil: Ceriani Nakamurakare, Esteban Daniel. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Biodiversidad y Biología Experimental; ArgentinaFil: Carmaran, Cecilia Cristina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Biodiversidad y Biología Experimental; ArgentinaFil: Gonzalez Audino, Paola Andrea. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones Científicas y Técnicas para la Defensa. Centro de Investigación de Plagas e Insecticidas; Argentina. Universidad Nacional de San Martín; Argentin

Mycobiota associated with the ambrosia beetle Megaplatypus mutatus: Threat to poplar plantations

Megaplatypus mutatus (syn. Platypus sulcatus) is an ambrosia beetle native to South America, which represents the main forest pest in Argentina of Poplar plantations and is also an emerging pest in Europe, representing a potential risk to forest and fruit plantations globally due to its low specificity. Knowledge of the interactions this insect has with microbes will be important in understanding its impacts and management. In this study, we characterized the fungal diversity associated with M. mutatus in Populus trees in Argentina. The fungal community of 28 attacked trees was studied by evaluating 1104 gallery fragments and 110 fragments of insects. Fungal isolates were identified using morphology and phylogenetic analysis of the internal transcribed spacer region of nDNA. Nineteen taxa were identified, the most relevant Fusarium solani species complex, three species of Raffaelea and Graphium basitruncatum. Despite the lower frequency occurred by Raffaelea spp. and G. basitruncatum, we detected a specificity between male/female and location in the gallery. Additionally, the topology of the galleries is described based on analyses with computed tomography and nuclear magnetic resonance. Enhancing these techniques, an image combining both data was produced, suggesting that the water circulation across outer sections might be involved in the modulation of the gallery topology.Fil: Ceriani Nakamurakare, Esteban Daniel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Biodiversidad y Biología Experimental; ArgentinaFil: Slodowicz, Mariel Pamela. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones Científicas y Técnicas para la Defensa. Centro de Investigación de Plagas e Insecticidas; ArgentinaFil: Gonzalez Audino, Paola Andrea. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones Científicas y Técnicas para la Defensa. Centro de Investigación de Plagas e Insecticidas; ArgentinaFil: Dolinko, Andrés Ezequiel. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Biodiversidad y Biología Experimental; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria; ArgentinaFil: Carmaran, Cecilia Cristina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Biodiversidad y Biología Experimental; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria; Argentin