Biocontrol Potential of Beneficial Fungus Aureobasidium pullulans Against Botrytis cinerea and Colletotrichum acutatum.

Biological control is a promising approach to reduce plant diseases caused by fungal pathogens and ensure high productivity in horticultural production. In the present study, we evaluated the biocontrol potential and underlying mechanisms of the beneficial fungus Aureobasidium pullulans against Botrytis cinerea and Colletotrichum acutatum, casual agents of gray mold and anthracnose diseases in strawberry. Notably, this is the first time that A. pullulans has been tested against C. acutatum in strawberry. A. pullulans strains (AP-30044, AP-30273, AP-53383, and AP-SLU6) showed significant variation in terms of growth and conidia production. An inverse relationship was found between the growth and conidiation rate, suggesting a trade-off between resource allocation for growth and conidial production. Dual plate co-culturing assays showed that mycelial growth of B. cinerea and C. acutatum was reduced by up to 35 and 18%, respectively, when challenged with A. pullulans compared with control treatments. Likewise, culture filtrates of A. pullulans showed varying levels of antifungal activity against B. cinerea and C. acutatum, reducing the mycelial biomass by up to 90 and 72%, respectively. Furthermore, milk powder plate assays showed that A. pullulans produced substantial amounts of extracellular proteases, which are known to degrade fungal cuticle. Ultra-high performance liquid chromatography-mass spectrometry (UHPLC-MS) analyses revealed that A. pullulans produced exophilins, liamocins, and free fatty acids known to have antifungal properties. A. pullulans shows high potential for successful biological control of strawberry diseases and discuss opportunities for further optimization of this beneficial fungus.

Bee-Vectored Aureobasidium pullulans for Biological Control of Gray Mold in Strawberry.

Gray mold caused by Botrytis cinerea is a common postharvest disease in strawberries, reducing shelf life considerably. We investigated the potential of the yeast-like biocontrol fungus Aureobasidium pullulans (AP-SLU6) vectored by bumblebees (Bombus terrestris) in the Flying Doctors® system to inhibit the pathogen and increase the shelf life of harvested strawberries (cultivar Sonata). Using bumblebees as vectors of various biocontrol agents is becoming increasingly popular, but any potentially negative effects on bee performance have been understudied. Our results show that, over the 4-week period of the trial, the performance and activity of the bees were not negatively affected by A. pullulans. The bees successfully picked up the powder formulation; then, they carried and deposited it on the flowers. The vectoring of the biocontrol agent significantly reduced gray mold development on the harvested fruits by 45% and increased shelf life by 100% in comparison with control treatments. This suggests that the biocontrol fungus applied during flowering successfully reduced Botrytis infection and thus, effectively protected the fruits from gray mold. In addition, the bee-vectored application of the biocontrol agent was found to be significantly more effective than spray application because the latter may temporarily increase humidity around the flower, thereby creating a suitable environment for the pathogen to thrive. In summary, our study demonstrates that A. pullulans vectored by bumblebees can decrease gray mold infection and improve the shelf life of strawberries without adversely affecting the bees, thus providing a basis for the sustainable and efficient control of gray mold on strawberry.[Formula: see text] Copyright © 2022 The Author(s). This is an open access article distributed under the CC BY 4.0 International license.

How far away is the next basket of eggs? Spatial memory and perceived cues shape aggregation patterns in a leaf beetle.

Gregarious organisms need to handle the trade-off between increasing food competition and the positive effects of group living, and this is particularly important for ovipositing females. We hypothesized that insect females consider how many conspecifics previously visited a host plant. In a no-choice assay, we show that the gregarious blue willow leaf beetle (Phratora vulgatissima) laid the most eggs and the largest clutches on plants where a sequence of few individual females was released, compared to plants where one or many different females were repeatedly released. Therefore, this species is more sensitive to the indirectly perceived number of conspecifics than the directly perceived number of eggs on a plant. We further hypothesized that females adjust their own intra-plant egg clutch distribution to that of conspecifics and discovered a new behavioral component, i.e., the modulation of distances between clutches. Females adjusted these distances in ways indicating the use of spatial memory, because the largest distance increases were observed on plants with their own clutches, compared to plants with clutches from conspecifics. However, adjustment of aggregation level and distance between clutches occurred only on a suitable, and not on an unsuitable, Salix genotype. We conclude that both behaviors should reduce competition between sibling and non-sibling larvae.

Bayesian species delimitation reveals generalist and specialist parasitic wasps on Galerucella beetles (Chrysomelidae): sorting by herbivore or plant host.

BACKGROUND: To understand the ecological and evolutionary consequences of species interactions in food webs necessitates that interactions are properly identified. Genetic analyses suggest that many supposedly generalist parasitoid species should rather be defined as multiple species with a more narrow diet, reducing the probability that such species may mediate indirect interactions such as apparent competition among hosts. Recent studies showed that the parasitoid Asecodes lucens mediate apparent competition between two hosts, Galerucella tenella and G. calmariensis, affecting both interaction strengths and evolutionary feedbacks. The same parasitoid was also recorded from other species in the genus Galerucella, suggesting that similar indirect effects may also occur for other species pairs. METHODS: To explore the possibility of such interactions, we sequenced mitochondrial and nuclear genetic markers to resolve the phylogeny of both host and parasitoid and to test the number of parasitoid species involved. We thus collected 139 Galerucella larvae from 8 host plant species and sequenced 31 adult beetle and 108 parasitoid individuals. RESULTS: The analysis of the Galerucella data, that also included sequences from previous studies, verified the five species previously documented as reciprocally monophyletic, but the Bayesian species delimitation for A. lucens suggested 3-4 cryptic taxa with a more specialised host use than previously suggested. The gene data analyzed under the multispecies coalescent model allowed us to reconstruct the species tree phylogeny for both host and parasitoid and we found a fully congruent coevolutionary pattern suggesting that parasitoid speciation followed upon host speciation. CONCLUSION: Using multilocus sequence data in a Bayesian species delimitation analysis we propose that hymenopteran parasitoids of the genus Asecodes that infest Galerucella larvae constitute at least three species with narrow diet breath. The evolution of parasitoid Asecodes and host Galerucella show a fully congruent coevolutionary pattern. This finding strengthens the hypothesis that the parasitoid in host search uses cues of the host rather than more general cues of both host and plant.

Quantitative proteomics reveals regulatory differences in the chondrocyte secretome from human medial and lateral femoral condyles in osteoarthritic patients.

BACKGROUND: Osteoarthritis (OA) is a destructive joint disease and there are no known biomarkers available for an early diagnosis. To identify potential disease biomarkers and gain further insight into the disease mechanisms of OA we applied quantitative proteomics with SILAC technology on the secretomes from chondrocytes of OA knees, designated as high Mankin (HM) scored secretome. A quantitative comparison was made between the secretomes of the medial and lateral femur condyle chondrocytes in the same knee since the medial femur condyle is usually more affected in OA than the lateral condyle, which was confirmed by Mankin scoring. The medial/lateral comparison was also made on the secretomes from chondrocytes taken from one individual with no clinically apparent joint-disease, designated as low Mankin (LM) scored secretome. RESULTS: We identified 825 proteins in the HM secretome and 69 of these showed differential expression when comparing the medial and lateral femoral compartment. The LM scored femoral condyle showed early signs of OA in the medial compartment as assessed by Mankin score. We here report the identification and relative quantification of several proteins of interest for the OA disease mechanism e.g. CYTL1, DMD and STAB1 together with putative early disease markers e.g. TIMP1, PPP2CA and B2M. CONCLUSIONS: The present study reveals differences in protein abundance between medial/lateral femur condyles in OA patients. These regulatory differences expand the knowledge regarding OA disease markers and mechanisms.

Performance of an herbivorous leaf beetle (Phratora vulgatissima) on Salix F2 hybrids: the importance of phenolics.

The genotype of the plant determines, through the expression of the phenotype, how well it is suited as food for herbivores. Since hybridization often results in profound genomic alterations with subsequent changes in phenotypic traits, it has the potential to significantly affect plant-herbivore interactions. In this study, we used a population of F2 hybrids that originated from a cross between a Salix viminalis and a Salix dasyclados genotype, which differed in both phenolic content and resistance to the herbivorous leaf beetle Phratora vulgatissima. We screened for plants that showed a great variability in leaf beetle performance (i.e., oviposition and survival). By correlating leaf phenolics to the response of the herbivores, we evaluated the importance of different phenolic compounds for Salix resistance to the targeted insect species. The performance of P. vulgatissima varied among the F2 hybrids, and two patterns of resistance emerged: leaf beetle oviposition was intermediate on the F2 hybrids compared to the parental genotypes, whereas leaf beetle survival demonstrated similarities to one of the parents. The findings indicate that these life history traits are controlled by different resistance mechanisms that are inherited differently in the hybrids. Salicylates and a methylated luteolin derivative seem to play major roles in hybrid resistance to Phratora vulgatissima. Synergistic effects of these compounds, as well as potential threshold concentrations, are plausible. In addition, we found considerable variation in both distributions and concentrations of different phenolics in the F2 hybrids. The phenolic profiles of parental genotypes and F2 hybrids differed significantly (e.g., novel compounds appeared in the hybrids) suggesting genomic alterations with subsequent changes in biosynthetic pathways in the hybrids.

Political deafness may impede transition to biological control.

Biological control has developed into a realistic alternative to replace chemical pesticides. A long-awaited paradigm shift is now adopted by the European Commission through a proposed new Regulation on sustainable use of plant protection products. Unfortunately, the scientific framework underpinning biocontrol is seriously neglected, impeding transition to sustainable plant production.

Evolutionary Ecology of Plant-Arthropod Interactions in Light of the "Omics" Sciences: A Broad Guide.

Aboveground plant-arthropod interactions are typically complex, involving herbivores, predators, pollinators, and various other guilds that can strongly affect plant fitness, directly or indirectly, and individually, synergistically, or antagonistically. However, little is known about how ongoing natural selection by these interacting guilds shapes the evolution of plants, i.e., how they affect the differential survival and reproduction of genotypes due to differences in phenotypes in an environment. Recent technological advances, including next-generation sequencing, metabolomics, and gene-editing technologies along with traditional experimental approaches (e.g., quantitative genetics experiments), have enabled far more comprehensive exploration of the genes and traits involved in complex ecological interactions. Connecting different levels of biological organization (genes to communities) will enhance the understanding of evolutionary interactions in complex communities, but this requires a multidisciplinary approach. Here, we review traditional and modern methods and concepts, then highlight future avenues for studying the evolution of plant-arthropod interactions (e.g., plant-herbivore-pollinator interactions). Besides promoting a fundamental understanding of plant-associated arthropod communities' genetic background and evolution, such knowledge can also help address many current global environmental challenges.

Focused Identification of Germplasm Strategy (FIGS): polishing a rough diamond.

Focused Identification of Germplasm Strategy (FIGS) has been advocated as an efficient approach to predict and harness variation in adaptive traits in genebanks or wild populations of plants. However, a weakness of the current FIGS approach is that it only utilizes a priori knowledge of one evolutionary factor: natural selection. Further optimization is needed to capture elusive traits, and this review shows that nonadaptive evolutionary processes (gene flow and genetic drift) should be incorporated to increase precision. Focusing on plant resistance to insect herbivores, we also note that historic selection pressures can be difficult to disentangle, and provide suggestions for successful mining based on eco-evolutionary theory. We conclude that with such refinement FIGS has high potential for enhancing breeding efforts and hence sustainable plant production.

Pollinators and herbivores interactively shape selection on strawberry defence and attraction.

Tripartite interactions between plants, herbivores, and pollinators hold fitness consequences for most angiosperms. However, little is known on how plants evolve in response-and in particular what the net selective outcomes are for traits of shared relevance to pollinators and herbivores. In this study, we manipulated herbivory ("presence" and "absence" treatments) and pollination ("open" and "hand pollination" treatments) in a full factorial common-garden experiment with woodland strawberry (Fragaria vesca L.). This design allowed us to quantify the relative importance and interactive effects of herbivore- and pollinator-mediated selection on nine traits related to plant defence and attraction. Our results showed that pollinators imposed stronger selection than herbivores on traits related to both direct and indirect (i.e., tritrophic) defence. However, conflicting selection was imposed on inflorescence density: a trait that appears to be shared by herbivores and pollinators as a host plant signal. However, in all cases, selection imposed by one agent depended largely on the presence or ecological effect of the other, suggesting that dynamic patterns of selection could be a common outcome of these interactions in natural populations. As a whole, our findings highlight the significance of plant-herbivore-pollinator interactions as potential drivers of evolutionary change, and reveal that pollinators likely play an underappreciated role as selective agents on direct and in direct plant defence.

'Resistance Mixtures' Reduce Insect Herbivory in Strawberry (Fragaria vesca) Plantations.

The transition toward more sustainable plant protection with reduced pesticide use is difficult, because there is no "silver bullet" available among nonchemical tools. Integrating several plant protection approaches may thus be needed for efficient pest management. Recently, increasing the genetic diversity of plantations via cultivar mixing has been proposed as a possible method to reduce pest damage. However, previous studies have not addressed either the relative efficiency of exploiting cultivar mixing and intrinsic plant herbivore resistance or the potential utility of combining these approaches to increase cropping security. Here, using a full factorial experiment with 60 woodland strawberry plots, we tested for the relative and combined effect of cultivar mixing and intrinsic plant resistance on herbivore damage and yield. The experiment comprised two levels of diversity ("high" with 10 varieties and "low" with two varieties) and three levels of resistance ("resistant" comprising only varieties intrinsically resistant against strawberry leaf beetle Galerucella tenella; "susceptible" with susceptible varieties only; and "resistance mixtures" with 50:50 mixtures of resistant and susceptible varieties). The experiment was carried out over two growing seasons. Use of resistant varieties either alone or intermixed with susceptible varieties in "resistance mixtures" reduced insect herbivory. Interestingly, resistant varieties not only reduced the mean damage in "resistance mixtures" by themselves being less damaged, but also protected intermixed susceptible varieties via associational resistance. The effect of higher genetic diversity was less evident, reducing herbivory only at the highest level of herbivore damage. In general, herbivory was lowest in plots with high diversity that included at least some resistant varieties and highest in low diversity plots consisting only of susceptible varieties. Despite this, no significant difference in yield (fruit biomass) was found, indicating that strawberry may be relatively tolerant. Our results demonstrate that combined use of high genetic diversity and resistant varieties can help reduce pest damage and provide a useful tool for sustainable food production. "Resistance mixtures" may be particularly useful for sensitive food crops where susceptible varieties are high yielding that could not be completely replaced by resistant ones.

A dual-tag microarray platform for high-performance nucleic acid and protein analyses.

DNA microarrays serve to monitor a wide range of molecular events, but emerging applications like measurements of weakly expressed genes or of proteins and their interaction patterns will require enhanced performance to improve specificity of detection and dynamic range. To further extend the utility of DNA microarray-based approaches we present a high-performance tag microarray procedure that enables probe-based analysis of as little as 100 target cDNA molecules, and with a linear dynamic range close to 10(5). Furthermore, the protocol radically decreases the risk of cross-hybridization on microarrays compared to current approaches, and it also allows for quantification by single-molecule analysis and real-time on-chip monitoring of rolling-circle amplification. We provide proof of concept for microarray-based measurement of both mRNA molecules and of proteins, converted to tag DNA sequences by padlock and proximity probe ligation, respectively.

Making sense of Integrated Pest Management (IPM) in the light of evolution.

Integrated Pest Management (IPM) is a holistic approach to combat pests (including herbivores, pathogens, and weeds) using a combination of preventive and curative actions, and only applying synthetic pesticides when there is an urgent need. Just as the recent recognition that an evolutionary perspective is useful in medicine to understand and predict interactions between hosts, diseases, and medical treatments, we argue that it is crucial to integrate an evolutionary framework in IPM to develop efficient and reliable crop protection strategies that do not lead to resistance development in herbivores, pathogens, and weeds. Such a framework would not only delay resistance evolution in pests, but also optimize each element of the management and increase the synergies between them. Here, we outline key areas within IPM that would especially benefit from a thorough evolutionary understanding. In addition, we discuss the difficulties and advantages of enhancing communication among research communities rooted in different biological disciplines and between researchers and society. Furthermore, we present suggestions that could advance implementation of evolutionary principles in IPM and thus contribute to the development of sustainable agriculture that is resilient to current and emerging pests.

Wild strawberry shows genetic variation in tolerance but not resistance to a generalist herbivore.

Plants' defenses against herbivores usually include both resistance and tolerance mechanisms. Their deployment has predominantly been studied in either single-plant genotypes or multiple genotypes exposed to single herbivores. In natural situations, however, most plants are attacked by multiple herbivores. Therefore, aims of this study were to assess and compare the effects of single and multiple herbivores on plant resistance and tolerance traits, and the consequences for overall plant performance. For this, we exposed multiple genotypes of wild woodland strawberry (Fragaria vesca) to jasmonic acid (JA), to mimic chewing herbivory and induce the plants' defense responses, and then introduced the generalist herbivore Spodoptera littoralis to feed on them. We found that woodland strawberry consistently showed resistance to S. littoralis herbivory, with no significant genetic variation between the genotypes. By contrast, the studied genotypes showed high variation in tolerance, suggesting evolutionary potential in this trait. Prior JA application did not alter these patterns, although it induced an even higher level of resistance in all tested genotypes. The study provides novel information that may be useful for breeders seeking to exploit tolerance and resistance mechanisms to improve strawberry crops' viability and yields, particularly when multiple herbivores pose significant threats.

Plant resistance does not compromise parasitoid-based biocontrol of a strawberry pest.

Plant nutritional  quality can influence interactions between herbivores and their parasitoids. While most previous work has focused on a limited set of secondary plant metabolites, the tri-trophic effects of overall phenotypic resistance have been understudied. Furthermore, the joint effects of secondary and primary metabolites on parasitoids are almost unexplored. In this study, we compared the performance and survival of the parasitoid species Asecodes parviclava Thompson on wild woodland strawberry (Fragaria vesca L.) genotypes showing variation in resistance against the parasitoid's host, the strawberry leaf beetle (Galerucella tenella L.). Additionally, we related the metabolic profiles of these plant genotypes to the tritrophic outcomes in order to identify primary and secondary metabolites involved in regulating plant potential to facilitate parasitism. We found that parasitoid performance was strongly affected by plant genotype, but those differences in plant resistance to the herbivore were not reflected in parasitoid survival. These findings could be explained in particular by a significant link between parasitoid survival and foliar carbohydrate levels, which appeared to be the most important compounds for parasitism success. The fact that plant quality strongly affects parasitism should be further explored and utilized in plant breeding programs for a synergistic application in sustainable pest management.

Delivering Integrated Pest and Pollinator Management (IPPM).

The need to reduce pollinator exposure to harmful pesticides has led to calls to expedite the adoption of integrated pest management (IPM). We make the case that IPM is not explicitly 'pollinator friendly', but rather must be adapted to reduce impacts on pollinators and to facilitate synergies between crop pollination and pest control practices and ecosystem services. To reconcile these diverse needs, we introduce a systematic framework for 'integrated pest and pollinator management' (IPPM). We also highlight novel tools to unify monitoring and economic decision-making processes for IPPM and outline key policy actions and knowledge gaps. We propose that IPPM is needed to promote more coordinated, ecosystem-based strategies for sustainable food production, against the backdrop of increasing pesticide regulation and pollinator dependency in agriculture. VIDEO ABSTRACT.

MLGA--a rapid and cost-efficient assay for gene copy-number analysis.

Structural variation is an important cause of genetic variation. Whole genome analysis techniques can efficiently identify copy-number variable regions but there is a need for targeted methods, to verify and accurately size variable regions, and to diagnose large sample cohorts. We have developed a technique based on multiplex amplification of size-coded selectively circularized genomic fragments, which is robust, cheaper and more rapid than current multiplex targeted copy-number assays.

Shared signals -'alarm calls' from plants increase apparency to herbivores and their enemies in nature.

The attraction of natural enemies of herbivores by volatile organic compounds as an induced indirect defence has been studied in several plant systems. The evidence for their defensive function originates mainly from laboratory studies with trained parasitoids and predators; the defensive function of these emissions for plants in natural settings has been rarely demonstrated. In native populations and laboratory Y-tube choice experiments with transgenic Nicotiana attenuata plants unable to release particular volatiles, we demonstrate that predatory bugs use terpenoids and green leaf volatiles (GLVs) to locate their prey on herbivore-attacked plants. By attracting predators with volatile signals, this native plant reduces its herbivore load - demonstrating the defensive function of herbivore-induced volatile emissions. However, plants producing GLVs are also damaged more by flea beetles. The implications of these conflicting ecological effects for the evolution of induced volatile emissions and for the development of sustainable agricultural practices are discussed.

Herbivore-induced "rent rise" in the host plant may drive a diet breadth enlargement in the tenant.

Inter- and intraspecies variations in host plant traits are presumably involved in many host shifts by insect herbivores, and elucidating the mechanisms involved in such shifts has been a crucial goal in insect-plant research for several decades. Here we propose that herbivore-induced evolutionary increases in host plant resistance may cause oligophagous insect herbivores to shift to other sympatric plants as currently preferred host plants become increasingly unpalatable. We tested this hypothesis in a system based on the perennial herb Filipendula ulmaria (Rosaceae), whose herbivory defense has become gradually stronger due to prolonged selection by Galerucella tenella (Coleoptera: Chrysomelidae) herbivory in a boreal archipelago. We show that Galerucella gradually increases its use of the alternative host plant Rubus arcticus (Rosaceae) in parallel to gradually increased resistance in Filipendula. Our results imply that, by driving the evolutionary increase in Filipendula resistance, Galerucella is also gradually making the original host species more unpalatable and thereby driving its own host-breadth enlargement. We argue that such self-inflicted "rent rises" may be an important mechanism behind host plant shifts, which in turn are believed to have preceded the speciation of many phytophagous insects.

Folivory has long-term effects on sexual but not on asexual reproduction in woodland strawberry.

Plant fitness is often a result of both sexual and asexual reproductive success and, in perennial plants, over several years. Folivory can affect both modes of reproduction. However, little is known about the effects of folivory on resource allocation to the two modes of reproduction simultaneously and across years. In a 2-year common garden experiment, we examined the effects of different levels of folivory by the strawberry leaf beetle, Galerucella tenella, on current growth, as well as current and future sexual and asexual reproduction (runners) of perennial woodland strawberry, Fragaria vesca. In addition, we measured the chlorophyll content in leaves in the year of experimental damage to determine whether there was increased photosynthetic activity, and, thus, a compensatory response to herbivory. Finally, we tested whether the previous year's folivory, as a result of its effect on plant fitness, affected the level of natural herbivory the plant experienced during the subsequent year. In the year of experimental damage, plants that were exposed to moderate and high levels of folivory (25% and 50% leaf area consumed, respectively) increased their photosynthetic activity compared to control plants. However, only plants exposed to high folivory exhibited negative effects, with a lower probability of flowering compared to control plants, indicating that plants exposed to low or moderate folivory were able to compensate for the damage. Negative effects of folivory were carried over to the subsequent year. Plants that were exposed to moderate folivory (25% leaf area consumed) during first year produced fewer flowers and fruits in the subsequent year. Runner production was consistently unaffected by folivory. The effects of experimental folivory on the level of natural herbivory were mediated via its effects on plant fitness. Our results show that the negative effects of folivory only influence sexual reproduction in woodland strawberry. Furthermore, even though woodland strawberry can tolerate moderate amounts of folivory in the short term, the negative effects on fitness appear later; this highlights the importance of studying the effects of herbivory over consecutive years in perennial plants.