Impact of mixed microalgal and bacterial species on organic micropollutants removal in photobioreactors under natural light.

Single microalgae species are effective at the removal of various organic micropollutants (OMPs), however increased species diversity might enhance this removal. Sixteen OMPs were added to 2 continuous photobioreactors, one inoculated with Chlorella sorokiniana and the other with a microalgal-bacterial community, for 112 d under natural light. Three media were sequentially used in 3 Periods: I) synthetic sewage (d 0-28), II) 10x diluted anaerobically digested black water (AnBW) (d 28-94) and III) 5x diluted AnBW (d 94-112). Twelve OMPs were removed > 30 %, while 4 were < 10 % removed. Removal efficiencies were similar for 9 OMPs, yet the mixed community showed a 2-3 times higher removal capacity (µg OMP/g dry weight) than C. sorokiniana during Period II pseudo steady state. The removal decreased drastically in Period III due to overgrowth of filamentous green algae. This study shows for the first time how microbial community composition and abundance are key for OMPs removal.

Impact of wastewater characteristics on the removal of organic micropollutants by Chlorella sorokiniana.

Microalgae-based technologies can be used for the removal of organic micropollutants (OMPs) from different types of wastewater. However, the effect of wastewater characteristics on the removal is still poorly understood. In this study, the removal of sixteen OMPs by Chlorella sorokiniana, cultivated in three types of wastewater (anaerobically digested black water (AnBW), municipal wastewater (MW), and secondary clarified effluent (SCE)), were assessed. During batch operational mode, eleven OMPs were removed from AnBW and MW. When switching from batch to continuous mode (0.8 d HRT), the removal of most OMPs from AnBW and MW decreased, suggesting that a longer retention time enhances the removal of some OMPs. Most OMPs were not removed from SCE since poor nutrient availability limited C. sorokiniana growth. Further correlation analyses between wastewater characteristics, biomass and OMPs removal indicated that the wastewater soluble COD and biomass concentration predominantly affected the removal of OMPs. Lastly, carbon uptake rate had a higher effect on the removal of OMPs than nitrogen and phosphate uptake rate. These data will give an insight on the implementation of microalgae-based technologies for the removal of OMPs in wastewater with varying strengths and nutrient availability.

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.

Removal processes of individual and a mixture of organic micropollutants in the presence of Scenedesmus obliquus.

Organic micropollutants (OMPs) need to be removed from wastewater as they can negatively affect aquatic organisms. It has been demonstrated that microalgae-based technologies are efficient in removing OMPs from wastewater. In this study, the removal processes and kinetics of six persistent OMPs (diclofenac, clarithromycin, benzotriazole, metoprolol, carbamazepine and mecoprop) were studied during cultivation of Scenedesmus obliquus in batch mode. These OMPs were added as individual compounds and in a mixture. Short experiments (8 days) were performed to avoid masking of OMP removal processes by light and nutrient limitation. The results show that diclofenac, clarithromycin, and benzotriazole were mainly removed by photodegradation (diclofenac), biodegradation (benzotriazole), or a combination of these two processes (clarithromycin). Peroxidase was involved in intracellular and extracellular biodegradation when benzotriazole was present as individual compound. Carbamazepine, metoprolol and mecoprop showed no biodegradation or photodegradation, and neglectable removal (<5%) by bioadsorption and bioaccumulation. Using an OMP mixture had an adverse effect on the photodegradation of clarithromycin and diclofenac, with reduced first-order kinetic constants compared to the individual compounds. Benzotriazole biodegradation was inhibited by the presence of the OMP mixture. This indicates that the presence of OMPs inhibits the photodegradation and biodegradation of some individual OMPs. These results will improve our understanding of removal processes of individual and mixtures of OMPs by microalgae-based technologies for wastewater treatment.

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.

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.

Effect of the amount of organic trigger compounds, nitrogen and soil microbial biomass on the magnitude of priming of soil organic matter.

Priming effects (PEs) are defined as short-term changes in the turnover of soil organic matter (SOM) caused by the addition of easily degradable organic compounds to the soil. PEs are ubiquitous but the direction (acceleration or retardation of SOM decomposition) and magnitude are not easy to predict. It has been suggested that the ratio between the amount of added PE-triggering substrate to the size of initial soil microbial biomass is an important factor influencing PEs. However, this is mainly based on comparison of different studies and not on direct experimentation. The aim of the current study is to examine the impact of glucose-to-microbial biomass ratios on PEs for three different ecosystems. We did this by adding three different amounts of 13C-glucose with or without addition of mineral N (NH4NO3) to soils collected from arable lands, grasslands and forests. The addition of 13C-glucose was equivalent to 15%, 50% and 200% of microbial biomass C. After one month of incubation, glucose had induced positive PEs for almost all the treatments, with differences in magnitude related to the soil origin and the amount of glucose added. For arable and forest soils, the primed C increased with increasing amount of glucose added, whereas for grassland soils this relationship was negative. We found positive correlations between glucose-derived C and primed C and the strength of these correlations was different among the three ecosystems considered. Generally, additions of mineral N next to glucose (C:N = 15:1) had little effect on the flux of substrate-derived C and primed C. Overall, our study does not support the hypothesis that the trigger-substrate to microbial biomass ratio can be an important predictor of PEs. Rather our results indicate that the amount of energy obtained from decomposing trigger substrates is an important factor for the magnitude of PEs.

Exploring bacterial interspecific interactions for discovery of novel antimicrobial compounds.

Recent studies indicated that the production of secondary metabolites by soil bacteria can be triggered by interspecific interactions. However, little is known to date about interspecific interactions between Gram-positive and Gram-negative bacteria. In this study, we aimed to understand how the interspecific interaction between the Gram-positive Paenibacillus sp. AD87 and the Gram-negative Burkholderia sp. AD24 affects the fitness, gene expression and the production of soluble and volatile secondary metabolites of both bacteria. To obtain better insight into this interaction, transcriptome and metabolome analyses were performed. Our results revealed that the interaction between the two bacteria affected their fitness, gene expression and the production of secondary metabolites. During interaction, the growth of Paenibacillus was not affected, whereas the growth of Burkholderia was inhibited at 48 and 72 h. Transcriptome analysis revealed that the interaction between Burkholderia and Paenibacillus caused significant transcriptional changes in both bacteria as compared to the monocultures. The metabolomic analysis revealed that the interaction increased the production of specific volatile and soluble antimicrobial compounds such as 2,5-bis(1-methylethyl)-pyrazine and an unknown Pederin-like compound. The pyrazine volatile compound produced by Paenibacillus was subjected to bioassays and showed strong inhibitory activity against Burkholderia and a range of plant and human pathogens. Moreover, strong additive antimicrobial effects were observed when soluble extracts from the interacting bacteria were combined with the pure 2,5-bis(1-methylethyl)-pyrazine. The results obtained in this study highlight the importance to explore bacterial interspecific interactions to discover novel secondary metabolites and to perform simultaneously metabolomics of both, soluble and volatile compounds.

Soil networks become more connected and take up more carbon as nature restoration progresses.

Soil organisms have an important role in aboveground community dynamics and ecosystem functioning in terrestrial ecosystems. However, most studies have considered soil biota as a black box or focussed on specific groups, whereas little is known about entire soil networks. Here we show that during the course of nature restoration on abandoned arable land a compositional shift in soil biota, preceded by tightening of the belowground networks, corresponds with enhanced efficiency of carbon uptake. In mid- and long-term abandoned field soil, carbon uptake by fungi increases without an increase in fungal biomass or shift in bacterial-to-fungal ratio. The implication of our findings is that during nature restoration the efficiency of nutrient cycling and carbon uptake can increase by a shift in fungal composition and/or fungal activity. Therefore, we propose that relationships between soil food web structure and carbon cycling in soils need to be reconsidered.

A fragrant neighborhood: volatile mediated bacterial interactions in soil.

There is increasing evidence that volatile organic compounds (VOCs) play essential roles in communication and competition between soil microorganisms. Here we assessed volatile-mediated interactions of a synthetic microbial community in a model system that mimics the natural conditions in the heterogeneous soil environment along the rhizosphere. Phylogenetic different soil bacterial isolates (Burkholderia sp., Dyella sp., Janthinobacterium sp., Pseudomonas sp., and Paenibacillus sp.) were inoculated as mixtures or monoculture in organic-poor, sandy soil containing artificial root exudates (ARE) and the volatile profile and growth were analyzed. Additionally, a two-compartment system was used to test if volatiles produced by inter-specific interactions in the rhizosphere can stimulate the activity of starving bacteria in the surrounding, nutrient-depleted soil. The obtained results revealed that both microbial interactions and shifts in microbial community composition had a strong effect on the volatile emission. Interestingly, the presence of a slow-growing, low abundant Paenibacillus strain significantly affected the volatile production by the other abundant members of the bacterial community as well as the growth of the interacting strains. Furthermore, volatiles released by mixtures of root-exudates consuming bacteria stimulated the activity and growth of starved bacteria. Besides growth stimulation, also an inhibition in growth was observed for starving bacteria exposed to microbial volatiles. The current work suggests that volatiles produced during microbial interactions in the rhizosphere have a significant long distance effect on microorganisms in the surrounding, nutrient-depleted soil.

Microbial Small Talk: Volatiles in Fungal-Bacterial Interactions.

There is increasing evidence that volatile organic compounds (VOCs) play an important role in the interactions between fungi and bacteria, two major groups of soil inhabiting microorganisms. Yet, most of the research has been focused on effects of bacterial volatiles on suppression of plant pathogenic fungi whereas little is known about the responses of bacteria to fungal volatiles. In the current study we performed a metabolomics analysis of volatiles emitted by several fungal and oomycetal soil strains under different nutrient conditions and growth stages. The metabolomics analysis of the tested fungal and oomycetal strains revealed different volatile profiles dependent on the age of the strains and nutrient conditions. Furthermore, we screened the phenotypic responses of soil bacterial strains to volatiles emitted by fungi. Two bacteria, Collimonas pratensis Ter291 and Serratia plymuthica PRI-2C, showed significant changes in their motility, in particular to volatiles emitted by Fusarium culmorum. This fungus produced a unique volatile blend, including several terpenes. Four of these terpenes were selected for further tests to investigate if they influence bacterial motility. Indeed, these terpenes induced or reduced swimming and swarming motility of S. plymuthica PRI-2C and swarming motility of C. pratensis Ter291, partly in a concentration-dependent manner. Overall the results of this work revealed that bacteria are able to sense and respond to fungal volatiles giving further evidence to the suggested importance of volatiles as signaling molecules in fungal-bacterial interactions.

Volatiles in Inter-Specific Bacterial Interactions.

The importance of volatile organic compounds for functioning of microbes is receiving increased research attention. However, to date very little is known on how inter-specific bacterial interactions effect volatiles production as most studies have been focused on volatiles produced by monocultures of well-described bacterial genera. In this study we aimed to understand how inter-specific bacterial interactions affect the composition, production and activity of volatiles. Four phylogenetically different bacterial species namely: Chryseobacterium, Dyella, Janthinobacterium, and Tsukamurella were selected. Earlier results had shown that pairwise combinations of these bacteria induced antimicrobial activity in agar media whereas this was not the case for monocultures. In the current study, we examined if these observations were also reflected by the production of antimicrobial volatiles. Thus, the identity and antimicrobial activity of volatiles produced by the bacteria were determined in monoculture as well in pairwise combinations. Antimicrobial activity of the volatiles was assessed against fungal, oomycetal, and bacterial model organisms. Our results revealed that inter-specific bacterial interactions affected volatiles blend composition. Fungi and oomycetes showed high sensitivity to bacterial volatiles whereas the effect of volatiles on bacteria varied between no effects, growth inhibition to growth promotion depending on the volatile blend composition. In total 35 volatile compounds were detected most of which were sulfur-containing compounds. Two commonly produced sulfur-containing volatile compounds (dimethyl disulfide and dimethyl trisulfide) were tested for their effect on three target bacteria. Here, we display the importance of inter-specific interactions on bacterial volatiles production and their antimicrobial activities.

The neonicotinoid imidacloprid shows high chronic toxicity to mayfly nymphs.

The present study evaluated the acute and chronic toxicity of imidacloprid to a range of freshwater arthropods. Mayfly and caddisfly species were most sensitive to short-term imidacloprid exposures (10 tests), whereas the mayflies showed by far the most sensitive response to long-term exposure of all seven arthropod species tested (28-d EC10 values of approximately 0.03 µg/L). The results indicated a high aquatic risk of chronic exposure of imidacloprid to mayflies.

Determination of the neurotoxins BMAA (beta-N-methylamino-L-alanine) and DAB (alpha-,gamma-diaminobutyric acid) by LC-MSMS in Dutch urban waters with cyanobacterial blooms.

We aimed to determine concentrations of the neurotoxic amino acids beta-N-methylamino-L-alanine (BMAA) and alpha-,gamma-diaminobutyric acid (DAB) in mixed species scum material from Dutch urban waters that suffer from cyanobacterial blooms. BMAA and DAB were analysed in scum material without derivatization by LC-MSMS (liquid chromatography coupled to tandem mass spectrometry) using hydrophilic interaction chromatography (HILIC). Our method showed high selectivity, good recovery of added compounds after sample extraction (86% for BMAA and 85% for DAB), acceptable recovery after sample hydrolysation (70% for BMAA and 56% for DAB) and acceptable precision. BMAA and DAB could be detected at an injected amount of 0.34 pmol. Free BMAA was detected in nine of the 21 sampled locations with a maximum concentration of 42 microg/g DW. Free DAB was detected in two locations with a maximum concentration of 4 microg/g DW. No protein-associated forms were detected. This study is the first to detect underivatized BMAA in cyanobacterial scum material using LC-MSMS. Ubiquity of BMAA in cyanobacteria scums of Dutch urban waters could not be confirmed, where BMAA and DAB concentrations were relatively low; however, co-occurrence with other cyanobacterial neurotoxins might pose a serious health risk including chronic effects from low-level doses.