Effect of microplastic pollution on the gut microbiome of anecic and endogeic earthworms.

Microplastic (MP) pollution constitutes an emerging type of pollution threatening both aquatic and terrestrial ecosystems. The impact on aquatic ecosystems has been extensively studied, but the effect on terrestrial ecosystems and their inhabitants is mostly underexplored. In this study, we explored the effect of MP pollution on gut bacterial microbiome of endogeic (Aporrectodea caliginosa) and anecic (Lumbricus terrestris) earthworms. The experiments were performed in sandy soil with 0.2% of low-density polyethylene MPs (LDPE MPs). We observed that the endogeic earthworms had 100% survival, while anecic earthworms survived 25 days in the control (i.e. in absence of MPs) and 21 days in the treatment with LDPE MPs. The main driver of shifts in the diversity and composition of the bacterial communities in the gut of tested earthworms was the lifestyle of the worms, followed by the presence of MPs. The bacterial microbiome diversity was significantly different among the two types of earthworms, and the highest bacterial diversity was found in the gut of the endogeic earthworms. The effect of MPs on gut bacterial microbiome was clearly observed in the changes in the relative abundance of several phyla and families of the bacterial communities in both types of earthworms, although it was most evident in the anecic earthworms. The Actinobacteriota, Proteobacteria, and Firmicutes were the main groups enhanced in the MP treatments, suggesting enrichment of the bacterial communities with potential plastic degraders.

Enhanced surface colonisation and competition during bacterial adaptation to a fungus.

Bacterial-fungal interactions influence microbial community performance of most ecosystems and elicit specific microbial behaviours, including stimulating specialised metabolite production. Here, we use a co-culture experimental evolution approach to investigate bacterial adaptation to the presence of a fungus, using a simple model of bacterial-fungal interactions encompassing the bacterium Bacillus subtilis and the fungus Aspergillus niger. We find in one evolving population that B. subtilis was selected for enhanced production of the lipopeptide surfactin and accelerated surface spreading ability, leading to inhibition of fungal expansion and acidification of the environment. These phenotypes were explained by specific mutations in the DegS-DegU two-component system. In the presence of surfactin, fungal hyphae exhibited bulging cells with delocalised secretory vesicles possibly provoking an RlmA-dependent cell wall stress. Thus, our results indicate that the presence of the fungus selects for increased surfactin production, which inhibits fungal growth and facilitates the competitive success of the bacterium.

Strategies for tailoring functional microbial synthetic communities.

Natural ecosystems harbor a huge reservoir of taxonomically diverse microbes that are important for plant growth and health. The vast diversity of soil microorganisms and their complex interactions make it challenging to pinpoint the main players important for the life support functions microbes can provide to plants, including enhanced tolerance to (a)biotic stress factors. Designing simplified microbial synthetic communities (SynComs) helps reduce this complexity to unravel the molecular and chemical basis and interplay of specific microbiome functions. While SynComs have been successfully employed to dissect microbial interactions or reproduce microbiome-associated phenotypes, the assembly and reconstitution of these communities have often been based on generic abundance patterns or taxonomic identities and co-occurrences but have only rarely been informed by functional traits. Here, we review recent studies on designing functional SynComs to reveal common principles and discuss multidimensional approaches for community design. We propose a strategy for tailoring the design of functional SynComs based on integration of high-throughput experimental assays with microbial strains and computational genomic analyses of their functional capabilities.

Impact of bacterial and fungal inoculants on the resident rhizosphere microbiome and the volatilome of tomato plants under leaf herbivory stress.

Various studies have addressed the impact of microbial inoculants on the composition of the resident microbiome. How microbial inoculants impact plant metabolism and interact with the resident rhizobiota under herbivory stress remains elusive. Here, we investigated the impact of two bacterial and two fungal inoculants, inoculated as single species and as a synthetic community, on the rhizosphere microbiome and volatilome of tomato plants (Solanum lycopersicum) comparing nonstress conditions to exposed to leaf herbivory by Spodoptera exigua. Based on amplicon sequencing analysis, rhizobacterial community composition was significantly affected by all four inoculants and the magnitude of this effect was dependent on herbivory stress. Fungal community composition was altered by the microbial inoculants but independent of herbivory stress. The rhizosphere volatilome was impacted by the microbial inoculation and differences between treatments were evened under herbivory stress. Each microbial inoculant caused unique changes in the volatilome of stressed plants but also shared similar responses, in particular the enhanced production of dimethyl disulfide and benzothiazole. In conclusion, the introduction of microbial inoculants in the tomato rhizosphere caused unique as well as common changes in the rhizosphere microbiome and volatilome, but these changes were minor compared to the microbiome changes induced by herbivory stress.

Volatile sensation: The chemical ecology of the earthy odorant geosmin.

Geosmin may be the most familiar volatile compound, as it lends the earthy smell to soil. The compound is a member of the largest family of natural products, the terpenoids. The broad distribution of geosmin among bacteria in both terrestrial and aquatic environments suggests that this compound has an important ecological function, for example, as a signal (attractant or repellent) or as a protective specialized metabolite against biotic and abiotic stresses. While geosmin is part of our everyday life, scientists still do not understand the exact biological function of this omnipresent natural product. This minireview summarizes the current general observations regarding geosmin in prokaryotes and introduces new insights into its biosynthesis and regulation, as well as its biological roles in terrestrial and aquatic environments.

Exploring the Interspecific Interactions and the Metabolome of the Soil Isolate Hylemonella gracilis.

Microbial community analysis of aquatic environments showed that an important component of its microbial diversity consists of bacteria with cell sizes of ~0.1 µm. Such small bacteria can show genomic reductions and metabolic dependencies with other bacteria. However, so far, no study has investigated if such bacteria exist in terrestrial environments like soil. Here, we isolated soil bacteria that passed through a 0.1-µm filter. The complete genome of one of the isolates was sequenced and the bacterium was identified as Hylemonella gracilis. A set of coculture assays with phylogenetically distant soil bacteria with different cell and genome sizes was performed. The coculture assays revealed that H. gracilis grows better when interacting with other soil bacteria like Paenibacillus sp. AD87 and Serratia plymuthica. Transcriptomics and metabolomics showed that H. gracilis was able to change gene expression, behavior, and biochemistry of the interacting bacteria without direct cell-cell contact. Our study indicates that in soil there are bacteria that can pass through a 0.1-µm filter. These bacteria may have been overlooked in previous research on soil microbial communities. Such small bacteria, exemplified here by H. gracilis, can induce transcriptional and metabolomic changes in other bacteria upon their interactions in soil. In vitro, the studied interspecific interactions allowed utilization of growth substrates that could not be utilized by monocultures, suggesting that biochemical interactions between substantially different sized soil bacteria may contribute to the symbiosis of soil bacterial communities. IMPORTANCE Analysis of aquatic microbial communities revealed that parts of its diversity consist of bacteria with cell sizes of ~0.1 µm. Such bacteria can show genomic reductions and metabolic dependencies with other bacteria. So far, no study investigated if such bacteria exist in terrestrial environments such as soil. Here, we show that such bacteria also exist in soil. The isolated bacteria were identified as Hylemonella gracilis. Coculture assays with phylogenetically different soil bacteria revealed that H. gracilis grows better when cocultured with other soil bacteria. Transcriptomics and metabolomics showed that H. gracilis was able to change gene expression, behavior, and biochemistry of the interacting bacteria without direct contact. Our study revealed that bacteria are present in soil that can pass through 0.1-µm filters. Such bacteria may have been overlooked in previous research on soil microbial communities and may contribute to the symbiosis of soil bacterial communities.

Antimicrobial volatiles emitted by members of the nest microbiome of social spiders.

Microbes produce and respond to a range of structurally and functionally diverse volatiles. Many microbial volatiles have antimicrobial properties. Since volatiles can diffuse through complex 3D systems like spider nests, they are promising pathogen protection for social arthropods. Here, we analyzed the volatilomes of five nest microbiome members of the Namibian, social spider Stegodyphus dumicola, namely the bacteria Massilia sp. IC2-278, Massilia sp. IC2-477, Sphingomonas sp. IC-11, Streptomyces sp. IC-207, and the fungus Aureobasidium sp. CE_32, and tested their antimicrobial activity against two putative spider pathogens, namely Bacillus thuringiensis and Purpureocillium lilacinum. Most nest microbiome members released volatilomes with antibacterial and/or antifungal activities under in vitro conditions. The analysis of their volatilomes using GC/Q-TOF revealed that they include numerous antimicrobial volatiles. We tested the antimicrobial activity of five pure volatile compounds found in the volatilomes and revealed that all of them were antibacterial and/or antifungal. We could not identify the same antimicrobial volatiles as in a previous in situ study, but our results indicate that social spider-associated microorganisms as a source of antimicrobial volatiles are important for pathogen inhibition. Additionally, we showed the influence of the volatilomes on the antibiotic sensitivity of B. thuringiensis offering novel approaches to counter antibiotic resistance.

Exploring the Volatiles Released from Roots of Wild and Domesticated Tomato Plants under Insect Attack.

Plants produce volatile organic compounds that are important in communication and defense. While studies have largely focused on volatiles emitted from aboveground plant parts upon exposure to biotic or abiotic stresses, volatile emissions from roots upon aboveground stress are less studied. Here, we investigated if tomato plants under insect herbivore attack exhibited a different root volatilome than non-stressed plants, and whether this was influenced by the plant's genetic background. To this end, we analyzed one domesticated and one wild tomato species, i.e., Solanum lycopersicum cv Moneymaker and Solanum pimpinellifolium, respectively, exposed to leaf herbivory by the insect Spodoptera exigua. Root volatiles were trapped with two sorbent materials, HiSorb and PDMS, at 24 h after exposure to insect stress. Our results revealed that differences in root volatilome were species-, stress-, and material-dependent. Upon leaf herbivory, the domesticated and wild tomato species showed different root volatile profiles. The wild species presented the largest change in root volatile compounds with an overall reduction in monoterpene emission under stress. Similarly, the domesticated species presented a slight reduction in monoterpene emission and an increased production of fatty-acid-derived volatiles under stress. Volatile profiles differed between the two sorbent materials, and both were required to obtain a more comprehensive characterization of the root volatilome. Collectively, these results provide a strong basis to further unravel the impact of herbivory stress on systemic volatile emissions.

Plastic mulch film residues in agriculture: impact on soil suppressiveness, plant growth, and microbial communities.

Plastic mulch film residues have been accumulating in agricultural soils for decades, but so far, little is known about its consequences on soil microbial communities and functions. Here, we tested the effects of plastic residues of low-density polyethylene and biodegradable mulch films on soil suppressiveness and microbial community composition. We investigated how plastic residues in a Fusarium culmorum suppressive soil affect the level of disease suppressiveness, plant biomass, nutrient status, and microbial communities in rhizosphere using a controlled pot experiment. The addition of 1% plastic residues to the suppressive soil did not affect the level of suppression and the disease symptoms index. However, we did find that plant biomasses decreased, and that plant nutrient status changed in the presence of plastic residues. No significant changes in bacterial and fungal rhizosphere communities were observed. Nonetheless, bacterial and fungal communities closely attached to the plastisphere were very different from the rhizosphere communities with overrepresentation of potential plant pathogens. The plastisphere revealed a high abundance of specific bacterial phyla (Actinobacteria, Bacteroidetes, and Proteobacteria) and fungal genera (Rhizoctonia and Arthrobotrys). Our work revealed new insights and raises emerging questions for further studies on the impact of microplastics on the agroecosystems.

Air Ambulance: Antimicrobial Power of Bacterial Volatiles.

We are currently facing an antimicrobial resistance crisis, which means that a lot of bacterial pathogens have developed resistance to common antibiotics. Hence, novel and innovative solutions are urgently needed to combat resistant human pathogens. A new source of antimicrobial compounds could be bacterial volatiles. Volatiles are ubiquitous produced, chemically divers and playing essential roles in intra- and interspecies interactions like communication and antimicrobial defense. In the last years, an increasing number of studies showed bioactivities of bacterial volatiles, including antibacterial, antifungal and anti-oomycete activities, indicating bacterial volatiles as an exciting source for novel antimicrobial compounds. In this review we introduce the chemical diversity of bacterial volatiles, their antimicrobial activities and methods for testing this activity. Concluding, we discuss the possibility of using antimicrobial volatiles to antagonize the antimicrobial resistance crisis.

Biosynthesis, evolution and ecology of microbial terpenoids.

Covering: through June 2021Terpenoids are the largest class of natural products recognised to date. While mostly known to humans as bioactive plant metabolites and part of essential oils, structurally diverse terpenoids are increasingly reported to be produced by microorganisms. For many of the compounds biological functions are yet unknown, but during the past years significant insights have been obtained for the role of terpenoids in microbial chemical ecology. Their functions include stress alleviation, maintenance of cell membrane integrity, photoprotection, attraction or repulsion of organisms, host growth promotion and defense. In this review we discuss the current knowledge of the biosynthesis and evolution of microbial terpenoids, and their ecological and biological roles in aquatic and terrestrial environments. Perspectives on their biotechnological applications, knowledge gaps and questions for future studies are discussed.

Antimicrobial Compounds in the Volatilome of Social Spider Communities.

Social arthropods such as termites, ants, and bees are among others the most successful animal groups on earth. However, social arthropods face an elevated risk of infections due to the dense colony structure, which facilitates pathogen transmission. An interesting hypothesis is that social arthropods are protected by chemical compounds produced by the arthropods themselves, microbial symbionts, or plants they associate with. Stegodyphus dumicola is an African social spider species, inhabiting communal silk nests. Because of the complex three-dimensional structure of the spider nest antimicrobial volatile organic compounds (VOCs) are a promising protection against pathogens, because of their ability to diffuse through air-filled pores. We analyzed the volatilomes of S. dumicola, their nests, and capture webs in three locations in Namibia and assessed their antimicrobial potential. Volatilomes were collected using polydimethylsiloxane (PDMS) tubes and analyzed using GC/Q-TOF. We showed the presence of 199 VOCs and tentatively identified 53 VOCs. More than 40% of the tentatively identified VOCs are known for their antimicrobial activity. Here, six VOCs were confirmed by analyzing pure compounds namely acetophenone, 1,3-benzothiazole, 1-decanal, 2-decanone, 1-tetradecene, and docosane and for five of these compounds the antimicrobial activity were proven. The nest and web volatilomes had many VOCs in common, whereas the spider volatilomes were more differentiated. Clear differences were identified between the volatilomes from the different sampling sites which is likely justified by differences in the microbiomes of the spiders and nests, the plants, and the different climatic conditions. The results indicate the potential relevance of the volatilomes for the ecological success of S. dumicola.

The Chemistry of Stress: Understanding the 'Cry for Help' of Plant Roots.

Plants are faced with various biotic and abiotic stresses during their life cycle. To withstand these stresses, plants have evolved adaptive strategies including the production of a wide array of primary and secondary metabolites. Some of these metabolites can have direct defensive effects, while others act as chemical cues attracting beneficial (micro)organisms for protection. Similar to aboveground plant tissues, plant roots also appear to have evolved "a cry for help" response upon exposure to stress, leading to the recruitment of beneficial microorganisms to help minimize the damage caused by the stress. Furthermore, emerging evidence indicates that microbial recruitment to the plant roots is, at least in part, mediated by quantitative and/or qualitative changes in root exudate composition. Both volatile and water-soluble compounds have been implicated as important signals for the recruitment and activation of beneficial root-associated microbes. Here we provide an overview of our current understanding of belowground chemical communication, particularly how stressed plants shape its protective root microbiome.

Disentangling soil microbiome functions by perturbation.

Soil biota contribute to diverse soil ecosystem services such as greenhouse gas mitigation, carbon sequestration, pollutant degradation, plant disease suppression and nutrient acquisition for plant growth. Here, we provide detailed insight into different perturbation approaches to disentangle soil microbiome functions and to reveal the underlying mechanisms. By applying perturbation, one can generate compositional and functional shifts of complex microbial communities in a controlled way. Perturbations can reduce microbial diversity, diminish the abundance of specific microbial taxa and thereby disturb the interactions within the microbial consortia and with their eukaryotic hosts. Four different microbiome perturbation approaches, namely selective heat, specific biocides, dilution-to-extinction and genome editing are the focus of this mini-review. We also discuss the potential of perturbation approaches to reveal the tipping point at which specific soil functions are lost and to link this change to key microbial taxa involved in specific microbiome-associated phenotypes.

Dissecting Disease-Suppressive Rhizosphere Microbiomes by Functional Amplicon Sequencing and 10× Metagenomics.

Disease-suppressive soils protect plants against soilborne fungal pathogens that would otherwise cause root infections. Soil suppressiveness is, in most cases, mediated by the antagonistic activity of the microbial community associated with the plant roots. Considering the enormous taxonomic and functional diversity of the root-associated microbiome, identification of the microbial genera and mechanisms underlying this phenotype is challenging. One approach to unravel the underlying mechanisms is to identify metabolic pathways enriched in the disease-suppressive microbial community, in particular, pathways that harbor natural products with antifungal properties. An important class of these natural products includes peptides produced by nonribosomal peptide synthetases (NRPSs). Here, we applied functional amplicon sequencing of NRPS-associated adenylation domains (A domains) to a collection of eight soils that are suppressive or nonsuppressive (i.e., conducive) to Fusarium culmorum, a fungal root pathogen of wheat. To identify functional elements in the root-associated bacterial community, we developed an open-source pipeline, referred to as dom2BGC, for amplicon annotation and putative gene cluster reconstruction through analyzing A domain co-occurrence across samples. We applied this pipeline to rhizosphere communities from four disease-suppressive and four conducive soils and found significant similarities in NRPS repertoires between suppressive soils. Specifically, several siderophore biosynthetic gene clusters were consistently associated with suppressive soils, hinting at competition for iron as a potential mechanism of suppression. Finally, to validate dom2BGC and to allow more unbiased functional metagenomics, we performed 10× metagenomic sequencing of one suppressive soil, leading to the identification of multiple gene clusters potentially associated with the disease-suppressive phenotype. IMPORTANCE Soil-borne plant-pathogenic fungi continue to be a major threat to agriculture and horticulture. The genus Fusarium in particular is one of the most devastating groups of soilborne fungal pathogens for a wide range of crops. Our approach to develop novel sustainable strategies to control this fungal root pathogen is to explore and exploit an effective, yet poorly understood naturally occurring protection, i.e., disease-suppressive soils. After screening 28 agricultural soils, we recently identified four soils that were suppressive to root disease of wheat caused by Fusarium culmorum. We also confirmed, via sterilization and transplantation, that the microbiomes of these soils play a significant role in the suppressive phenotype. By adopting nonribosomal peptide synthetase (NRPS) functional amplicon screening of suppressive and conducive soils, we here show how computationally driven comparative analysis of combined functional amplicon and metagenomic data can unravel putative mechanisms underlying microbiome-associated plant phenotypes.

Intraspecific variation in multiple trait responses of Alexandrium ostenfeldii towards elevated pCO2.

Dissolved oceanic CO2 concentrations are rising as result of increasing atmospheric partial pressure of CO2 (pCO2), which has large consequences for phytoplankton. To test how higher CO2 availability affects different traits of the toxic dinoflagellate Alexandrium ostenfeldii, we exposed three strains of the same population to 400 and 1,000 µatm CO2, and measured traits including growth rate, cell volume, elemental composition, 13C fractionation, toxin content, and volatile organic compounds (VOCs). Strains largely increased their growth rates and particulate organic carbon and nitrogen production with higher pCO2 and showed significant changes in their VOC profile. One strain showed a significant decrease in both PSP and cyclic imine content and thereby in cellular toxicity. Fractionation against 13C increased in response to elevated pCO2, which may point towards enhanced CO2 acquisition and/or a downscaling of the carbon concentrating mechanisms. Besides consistent responses in some traits, other traits showed large variation in both direction and strength of responses towards elevated pCO2. The observed intraspecific variation in phenotypic plasticity of important functional traits within the same population may help A. ostenfeldii to negate the effects of immediate environmental fluctuations and allow populations to adapt more quickly to changing environments.

Microbial volatile organic compounds in intra-kingdom and inter-kingdom interactions.

Microorganisms produce and excrete a versatile array of metabolites with different physico-chemical properties and biological activities. However, the ability of microorganisms to release volatile compounds has only attracted research attention in the past decade. Recent research has revealed that microbial volatiles are chemically very diverse and have important roles in distant interactions and communication. Microbial volatiles can diffuse fast in both gas and water phases, and thus can mediate swift chemical interactions. As well as constitutively emitted volatiles, microorganisms can emit induced volatiles that are triggered by biological interactions or environmental cues. In this Review, we highlight recent discoveries concerning microbial volatile compounds and their roles in intra-kingdom microbial interactions and inter-kingdom interactions with plants and insects. Furthermore, we indicate the potential biotechnological applications of microbial volatiles and discuss challenges and perspectives in this emerging research field.

A non-invasive soil-based setup to study tomato root volatiles released by healthy and infected roots.

The role of root exudates in mediating plant-microbe interactions has been well documented. However, the function of volatile organic compounds (VOCs) emitted by plant roots has only recently begun to attract attention. This newly recognized relevance of belowground VOCs has so far mostly been tested using systems limited to a two-compartment Petri-dish design. Furthermore, many of the plant-microbe interaction studies have only investigated the effects of microbial VOCs on plant growth. Here, we go two steps further. First we investigated the volatile profile of healthy and pathogen (Fusarium oxysporum) infected tomato roots grown in soil. We then used a unique soil-based olfactometer-choice assay to compare the migration pattern of four beneficial bacteria (Bacillus spp.) towards the roots of the tomato plants. We demonstrate that the blend of root-emitted VOCs differs between healthy and diseased plants. Our results show that VOCs are involved in attracting bacteria to plant roots.

Microbial and volatile profiling of soils suppressive to Fusarium culmorum of wheat.

In disease-suppressive soils, microbiota protect plants from root infections. Bacterial members of this microbiota have been shown to produce specific molecules that mediate this phenotype. To date, however, studies have focused on individual suppressive soils and the degree of natural variability of soil suppressiveness remains unclear. Here, we screened a large collection of field soils for suppressiveness to Fusarium culmorum using wheat (Triticum aestivum) as a model host plant. A high variation of disease suppressiveness was observed, with 14% showing a clear suppressive phenotype. The microbiological basis of suppressiveness to F. culmorum was confirmed by gamma sterilization and soil transplantation. Amplicon sequencing revealed diverse bacterial taxonomic compositions and no specific taxa were found exclusively enriched in all suppressive soils. Nonetheless, co-occurrence network analysis revealed that two suppressive soils shared an overrepresented bacterial guild dominated by various Acidobacteria. In addition, our study revealed that volatile emission may contribute to suppression, but not for all suppressive soils. Our study raises new questions regarding the possible mechanistic variability of disease-suppressive phenotypes across physico-chemically different soils. Accordingly, we anticipate that larger-scale soil profiling, along with functional studies, will enable a deeper understanding of disease-suppressive microbiomes.