In the goal to provide ongoing biocontrol options in plant production, several researchers around the world have turned towards the study of lichen extracts as potential biocontrol agents. Lichens present long-living symbiotic systems continuously exposed to pathogens. Indeed, these naturally occurring antagonists play an important role to avoid plant pathogen outbreaks in ecosystems.
According to a research study conducted a few years ago – "Analyzing the antagonistic potential of the lichen microbiome against pathogens by bridging metagenomic with culture studies" – lichens, which are classic examples of self-sustained symbioses, are interesting models for antagonism studies because within these mini-ecosystems the cooperation between microbial partners facilitates stability and longevity under extreme ecological conditions. “Conditioned by the slow growth of many lichens and difficulties in culturing the symbionts, biotechnological exploitation of lichens was lagging behind other natural resources,” noted the researchers in the study. “With the advent of modern technologies, however, the secondary metabolism and antagonistic potentials in lichens receive new impulses, and this will particularly apply to culturable bacterial partners. Although lichens are equipped with various secondary compounds with antagonistic effects, we hypothesize that only a diverse protective microbiome can efficiently maintain stability over longer periods to prevent pathogen attacks.” The objective of this study, undertaken by Tomislav Cernava, Henry Müller, Ines A. Aschenbrenner, Martin Grube and Gabriele Berg with the Institute of Environmental Biotechnology and the Institute of Plant Sciences at Graz University of Technology in Graz, Austria, was to analyze the antagonistic potential of the lichen microbiome against model pathogens by a novel approach bridging metagenomic with culture techniques.
In the study, model pathogens associated with human, lichen and plant diseases were accessed to screen for a broad spectrum of antagonistic activity. The researchers utilized the lung lichen Lobaria pulmonaria, (L.) Hoffm., which is one of the fastest growing leaf-like lichens and used as indicator species of undisturbed forests and air pollution. The researchers also characterized the most active as well as the most abundant lichen-associated antagonists, Stenotrophomonas, which were already identified as versatile antagonists from plant origin. Beneficial Stenotrophomonas strains produced osmoprotectans and spermidine in response to eukaryotic hosts.
“In our study we applied multidisciplinary techniques to link metagenomic data with those obtained from bacterial cultures,” noted the researchers. “Moreover, we could show that lichens are important reservoirs for antagonistic bacteria, which can also be used for biological control approaches to protect plants against biotic and abiotic stress.
To analyze the antagonistic potential in lichens, the researchers studied the bacterial community active against model bacteria and fungi by an integrative approach combining isolate screening, omics techniques and high-resolution mass spectrometry. The highly diverse microbiome of the lung lichen included an abundant antagonistic community dominated by Stenotrophomonas, Pseudomonas and Burkholderia. “While antagonists represent 24.5 percent of the isolates, they were identified with only seven percent in the metagenome, which means that they were overrepresented in the culturable fraction,” stated the researchers. “Isolates of the dominant antagonistic genus Stenotrophomonas produced spermidine as main bioactive component. Moreover, spermidine-related genes, especially for the transport, were identified in the metagenome.” The majority of hits identified belonged to Alphaproteobacteria, while Stenotrophomonas-specific spermidine synthases were not present in the dataset. Evidence for plant growth promoting effects was found for lichen-associated strains of Stenotrophomonas. Linking of metagenomic and culture data was possible but showed partly contradictory results, which required a comparative assessment. “However, we have shown that lichens are important reservoirs for antagonistic bacteria, which open broad possibilities for biotechnological applications,” concluded the researchers. What is lichen? Lichen is a symbiotic organism consisting of a fungus (mycobiont) and a photosynthetic partner (photobiont) which can be either an alga or a cyanobacterium. According to Rajkumar H. Garampalli, a professor of botany at the University of Mysore in India, lichens are inherently resistant to microbial infection due to the production of large numbers of unique secondary metabolites.
“Their flexibility in habitat enables them to synthesize unique, naturally occurring secondary metabolites, which not only are different in their chemical structures but also show differences in their biological activity,” he said. Garampalli and fellow researcher Rashmi Shivanna published a paper in 2014, looking at the efficacy of lichen extracts as biocontrol agents against Fusarium oxysporum F. Sp. Capsici. The aim of the study was to evaluate the antifungal activity of the methanol, ethyl acetate and acetone extract of 10 different lichens against Fusarium wilt, caused by Fusarium oxysporum f. sp. Capsica (FOC), an important plant pathogenic fungus.
In the study, a total of 10 lichens were screened for in vitro antifungal activity against Fusarium oxysporum. The total yield of the extract varied from 22-1,330 mg/ml (Figure 1). Highest yield was obtained from Flavoparmelia caperata with 1,330 mg/ml of methanol, Roccella montagnei yielded 880 mg/ml of ethyl acetate extract, while the lowest yield was from Usnea Sp. with only 22 mg/ml. “The results showed that the tested lichen extracts proved to be a significant biocontrol agent with the tested fungi,” noted the researchers. “But further investigations on the antimicrobial activity as well as the economical and fast isolation of the metabolite from the lichens are needed.” A year later, Garampalli and Shivanna conducted a similar study, to evaluate the fungitoxic effect of lichens in the management of rhizome rot of ginger caused by Fusarium solani (Mart.) Sacc. Ginger (Zingiber officinale Rosc.) is an important commercial crop cultivated throughout India for its rhizome as a spice. India is the largest producer of ginger, accounting for about one-third of total world output. But ginger is affected by several fungal pathogens among which rhizome rot caused by Fusarium solani is most common. The researchers stated a total of eight lichens were identified in which six lichens were of foliose growth form and two lichens were of fruticose growth form. “The results obtained in the study showed lichens have strong antifungal activity,” said the researchers. “Evaluation of eight lichen extracts showed to be effective in inhibiting the growth of the fungus.”
Amongst these, the most promising were Parmotrema tinctorum and Flavoparmelia caperata, which showed highest zone of inhibition in diffusion assay, least MIC value and good inhibition zones against the growth of Fusarium solani due to the fungicidal principle of metabolites present in the lichens. “The present work concentrated on finding out the effect of natural products, which are eco-friendly and less harmful than commercial synthetic compounds like fungicides,” stated the researchers. “Consequently, the investigated lichens could be used as a natural fungicide in the management of the diseases caused by plant pathogens. Further studies on the fractionation of solvent extracts and characterization may reveal the compounds responsible for the antifungal potentials.” ●
Individual lichens can have up to three fungi, study shows Individual lichens may contain up to three different fungi, according to new research from an international team of researchers. This evidence provides new insight into another recent discovery that showed lichen are made up of more than a single fungus and alga, overturning the prevailing theory of more than 150 years. The new study was a collaboration between the University of Alberta in Canada and Uppsala University in Sweden, and was led by Veera Tuovinen, a postdoctoral fellow under the supervision of Toby Spribille, assistant professor in University of Alberta's department of biological sciences.A classic example of symbiosis, lichens have long been known to be the result of a mutually beneficial relationship between fungi and algae. "With the microscopy, we could visualize the mosaic of different organisms within the lichen," said Tuovinen, who completed her PhD at Uppsala University. “We're realizing that interactions are much more complex than previously thought." The research team used advanced DNA sequencing to examine the genomes of the wolf lichen, a brilliant chartreuse-yellow lichen that grows on trees across Western Canada, the United States and Europe. While the species is well-studied, the researchers found that, almost regardless of where they were sampled, the wolf lichens contained not one or two fungi, but three." Our findings from two years ago challenged the long-held view that lichens were made up of a single fungus and alga," explained Spribille. "This new research complicates the nature of these relationships even further. For one thing, it means that no two lichens necessarily have the same medley of partners.""What this means in concrete terms to the overall symbiosis is the big question," added Hanna Johannesson, associate professor at Uppsala University and joint supervisor of the research. "What we are finding now is basically what researchers since the 1800s would have liked to know – who are the core players, what function do they perform, all the cards on the table."With the roster of players present in wolf lichens becoming clear, Johannesson and Spribille want to figure out how each member benefits in the give-and-take world of symbiosis. The scientists are particularly interested in the ability of fungi and algae to construct architectural structures from microscopic building blocks."The fungi and algae that make lichens are doing very interesting things in a confined space," says Spribille. "Knowing that there might not be any one way to pigeonhole the relationship is very helpful moving forward."The research was conducted with collaborators from Uppsala University and the Swedish University of Agricultural Sciences, as well as Indiana University in the United States. The paper, "Two basidiomycete fungi in the cortex of wolf lichens," was published in Current Biology (doi: 10.1016/j.cub.2018.12.022).
Toby Spribille examines lichen growing on a tree. New findings by scientists show that individual lichens can have up to three fungi.
Photo: John Ulan
What is a lichen? A lichen is not a single organism; it is a stable symbiotic association between a fungus and algae and/or cyanobacteria. Like all fungi, lichen fungi require carbon as a food source; this is provided by their symbiotic algae and/or cyanobacteria, that are photosynthetic. The lichen symbiosis is thought to be a mutualism, since both the fungi and the photosynthetic partners, called photobionts, benefit. Which fungi form lichens? Many unrelated and very different fungi form lichens, including mushroom-forming fungi, and especially cup-fungi. Ninety-eight percent of lichen fungi are cup-fungi, or ascomycetes. Fully half of all ascomycetes and one in five of all known fungi form lichens. Lichenization is an ecological strategy, or a common nutritional mode among unrelated fungi. What are lichen photobionts? Lichen photobionts are the green algae or cyanobacteria that provide the simple sugars to their fungal partners. Ninety percent of all lichens associate with a green-algal photobiont. About 100 species of photobionts are known, and the commonest ones are from four main groups. Lichen fungi specialize on particular photobionts. Typically, they only associate with a small group of related species, though they may associate flexibly with different photobionts according to their environmental situation. Source: The British Lichen Society
Fig 1: Total yield of the extracts obtained by different solvents in mg/ml. Source: Rashmi Shivanna and Rajkumar H. Garampalli
A recent study finds that the invasive spotted wing drosophila (Drosophila suzukii) prefers to lay its eggs in places that no other spotted wing flies have visited. The finding raises questions about how the flies can tell whether a piece of fruit is virgin territory – and what that might mean for pest control. D. suzukii is a fruit fly that is native to east Asia, but has spread rapidly across North America, South America, Africa and Europe over the past 10-15 years. The pest species prefers to lay its eggs in ripe fruit, which poses problems for fruit growers, since consumers don’t want to buy infested fruit. To avoid consumer rejection, there are extensive measures in place to avoid infestation, and to prevent infested fruit from reaching the marketplace. “Ultimately, we’re talking about hundreds of millions of dollars in potential crop losses and increases in pest-management costs each year in the United States,” says Burrack.
Burrack, co-author of a paper on the study and a professor of entomology at North Carolina State University (U.S.). “These costs have driven some small growers out of business. “The first step toward addressing an invasive pest species is understanding it. And two fundamental questions that we had are: Which plants will this species attack? And why does it pick those plants?” One of the things that researchers noticed when observing infestations on farms was that the species’ egg-laying behavior was different, depending on the size of the infestation. When D. suzukii populations were small, there would only be a few eggs laid in each piece of fruit, and they would only be in ripe fruit. If there were more D. suzukii present, more eggs would be laid in each piece of fruit. The researchers had also noticed that large populations of D. suzukii were also more likely to lay eggs in fruit that wasn’t ripe. To better understand the egg-laying behavior of D. suzukii, the researchers conducted a series of experiments. And the results surprised them. Specifically, the researchers found that, given a choice, female D. suzukii preferred to lay their eggs in fruit that other flies had never visited. “It doesn’t matter if the other flies lay eggs,” Burrack says. “It doesn’t even matter if the other flies are male or female. It only matters if other flies have touched a piece of fruit. If untouched fruit is available, D. suzukii will reject fruit that other flies have visited. “We’re not sure if the flies leave behind a chemical or bacterial marker, or something else entirely – but the flies can tell where other flies have been.” The researchers say the next step is to determine what, exactly, the D. suzukii are detecting. “If we can get a better understanding of what drives the behaviour of this species, that could inform the development of new pest-control techniques,” Burrack says. “We’re not making any promises, but this is a significant crop pest – and the more we know, the better.”
The paper, “Social signals mediate oviposition site selection in Drosophila suzukii,” is published in the journal Scientific Reports. Corresponding author of the paper is Johanna Elsensohn, who did the work while completing her Ph.D. at NC State. The paper was co-authored by Marwa F. K. Aly, a faculty member of Minia University, and Coby Schal, the Blanton J. Whitmire Distinguished Professor of Entomology at NC State. ●
Invasive spotted wing drosophila (Drosophila suzukii) on a raspberry.
Photo: Hannah Burrack
Researchers have developed a new technique to protect rice seeds against fungal infections that can ruin up to half of all rice crops in the world. The biocontrol method, which involves inoculation of flowers with a different fungus that doesn't cause disease and using seeds harvested from the flower to grow crops, is even better at protecting rice plants from diseases than existing fungicide approaches, and could also be used against similar pathogens that affect other staple crops. The destructive seedborne bakanae disease, which affects rice plants, is currently typically combatted with either chemical fungicides or by hot-water treatment of seeds, all of which face growing challenges to their effectiveness. However, researchers have developed a new anti-bakanae technique that actively encourages the spread of a different, non-pathogenic variety of fungus which has been shown to outcompete the disease-causing fungus on rice seeds. This biocontrol method not only delivers protection against bakanae disease as effectively as traditional methods but can also prevent bakanae disease from affecting the seeds, which current techniques cannot. The researcher's findings are reported in the journal Applied and Environmental Microbiology. The pathogenic fungus Fusarium fujikuroi produces gibberellic acid, a plant growth hormone, on rice plants and drives abnormal elongation and etiolation. The affected plants appear pale yellow or white, produce no edible grains, and suffer from weak stems that topple over, hence the name "bakanae," Japanese for "foolish seedling." Losses in the field are substantial wherever the disease emerges, but particularly severe in Asian countries, where the disease can hit 20 to 50 percent of crops.
Throughout agriculture, efforts to reduce conventional pesticide use are widespread in order to limit negative impacts on other organisms, but the additional problems that conventional methods of tackling bakanae disease face only add to the need to come up with an alternative. None of these techniques have been very stable, and thus lead to disease outbreaks. They are also not very efficient at combating deeply infected seed stocks. On top of this, existing chemical fungicides also increasingly face challenges from fungicide-resistant strains of the fungus. The new biocontrol technique, developed by plant pathologists at Tokyo University of Agriculture and Technology, involves spraying rice flowers with a non-pathogenic strain of the Fusaria fungus and produces rice seeds carrying the non-pathogenic Fusaria. Testing against conventional techniques showed roughly the same level of effectiveness, both against transmission of the disease to seeds, but also transmission among offsprings. "Investigation under the microscope suggests that the non-pathogenic strain out-competes its cousin, preventing the pathogenic fungi from colonizing the seed, while the growth of the 'good' fungus causes no harm," said Tsutomu Arie, professor, and Hiroki Saito, graduate student at the laboratory of plant pathology, Graduate School of Agriculture, Tokyo University of Agriculture and Technology. As the spread of the good fungus appears to completely replace the bad fungus, the technique should also work on heavily affected seed stocks.
Because rice seeds are usually stored for about six months over the winter before sowing in Japan, the non-pathogenic Fusaria in seeds needed to survive at least this amount of time. So, to track how long the protection lasted, the researchers genetically tweaked the fungus to make them fluorescent. Six months later, microscopic investigations found that fungal mycelia were still fluorescent, demonstrating they were still there and outcompeting their "bad" fungus cousins. The reproductive mechanics of other staple crops in the Poaceae family of grasses the rice plant belongs to, such as wheat, barley and corn, are similar enough that the technique could work on fungal infestations that affect these plants as well. The researchers now aim to test their new method on these crops, as well as on tomatoes, spinach, lettuce and carrots.●
Koppert Biological Systems acquired Geocom, headquartered in Lençóis Paulista, São Paulo, Brazil. Operating in the field of agricultural technology, Geocom is a pioneer in image geoprocessing and the first to develop an exclusive technology for the application of biological agents via drones in Brazil. “Our goal is to accelerate precision agri-farming which is crucial for bringing biologicals to the fields,” says Martin Koppert, agri division director with Koppert Biological Systems. “The Geocom brand is synonymous with high-tech developments. The acquisition in Brazil is a strategic decision which can favour the whole corporation by exchanging experiences and technologies with other subsidiaries around the world.” The founders of Geocom, Antonio Donizeti de Oliveira and Gláucio Carrit Antiga, will remain involved as consultants. “We see this acquisition not only as a milestone in the company's history, but also a source of great pride. Throughout its history, Geocom has established itself in the market as a leader in its segment within digital agriculture, and now it can maintain that level led by Koppert.” ●
The European Crop Protection Association has changed its name to CropLife Europe, and is expanding its mandate to include digital and precision farming, plant biotech innovation and biopesticides alongside conventional pesticides. The launch of CropLife Europe comes as EU policymakers and governments are increasingly calling on the agri-food system to transition to a more sustainable model. “We believe this is best achieved through a holistic approach, so an agile association representing a host of technologies under one roof will be better equipped to represent the integrated solutions needed to deliver sustainable agriculture and respond to the rapidly changing demands from society and evolving policy frameworks,” said Géraldine Kutas, CropLife Europe director-general. “We believe that producing enough food sustainably cannot be achieved by simply reducing the availability of solutions to farmers. Instead we need to accelerate the development of new, better solutions and enhanced farming practices that utilize more technology to produce our food while using less resource.”
CropLife Europe’s members will continue to invest in research and development, expanding its focus on innovation to support the production of food for all in a fair and sustainable way. “The European Commission’s Green Deal is a real game changer,” said Livio Tedeschi (BASF), CropLife Europe chair. “Together with the EU Farm to Fork Strategy and the EU Biodiversity Strategy for 2030, it provides us all with a great opportunity to deliver more sustainable agriculture; to ensure that the EU supports the delivery of the UN Sustainable Development Goals; and to further strengthen food security not only in Europe, but globally. We want our industry to be part of the solution that will help deliver the EU Green Deal. We will play our part and hope that other stakeholders will join us in driving these crucial topics forward.” The expanded CropLife Europe organization will encompass a wider range of topics including pesticides and biopesticides used in organic, conservation (low till, no till), agroforestry and conventional agriculture models; precision and digital applications which enable delivery of the minimum amount of product, at the right place, at the right time; and plant biotech traits that will enable crops to thrive in difficult conditions using less resources or provide greater benefit in people’s diets. ●
By Marianne Loison
IBMA France has reaffirmed the biocontrol target of 30 percent of the PPP market in France by 2030, at the IBMA France web symposium held 26 January 2021. Biocontrol products represented 11 percent of plant protection sales in France in 2019. The approval period by ANSES (Agence nationale de sécurité sanitaire de l'alimentation, de l'environnement et du travail) tends to be 15-16 months rather than the prescribed six months. Speaking at the symposium, Julien Denormandie, minister from the Ministry of Agriculture and Food, said the French government would offer some financial support for the national strategy, as already announced on 10 November 2020. “Research will also be supported through the France-relaunch plan, which totals 15 million euros in grants for 2021 and 2022,” added Denormandie. Denormandie said he hoped the definition of biocontrol products could be fully shared by all counties of the EU. The same objective is already supported by France and by the European Union through the guidelines of the Green Deal and the Farm to Fork strategy. The French presidency of the EU in 2022 is expected to support this goal. In France, biocontrol has avoided the legislation that impacted the rest of the PPP market, namely the “separation of advice and selling roles for PPPs,” effective since January 2021 and from the EGalim law adopted by the National Assembly in October 2018. French Cooperatives and retailers had to choose between sales (95 percent made this choice) and advice for conventional PPPs, but this does not apply to biocontrol products. The situation might have a positive impact on biocontrol deployment. In France, a quarter of CEPP files (savings certificates for plant protection products) are already related to biocontrol. According to the French Ecophyto II plan, distributors must reach 20 percent of their sales with CEPP from 2016 to 2021 in order not to pay penalties.
The fruit and vegetables sector has switched much more quickly towards biocontrol than the arable crops sector. “We have reached 50 percent of biocontrol for the protection of specialized crops,” said Claude Bizieux, supply director at vegetable cooperative CAMN, Nantes. “Twenty percent of the areas we harvest in the north of France use biocontrol,” estimates Daphné Souliez, project manager at Bonduelle, a main player in the vegetable industry. The rebalancing is expected before 2025. “Within two years we will have more products for arable crops, but the deployment remains longer,” said Céline Barthet, president of IBMA France. “IBMA France's ambition is to bring to the market two biocontrol solutions with complementary modes of action for the majority of uses by 2030.” ●
