Mycoviruses are viruses that infect fungi. In recent years, an increasing number of mycoviruses have been reported in a wide array of fungi. With the growing interest of scientists and society in ...reducing the use of agrochemicals, the debate about mycoviruses as an effective next-generation biocontrol has regained momentum. Mycoviruses can have profound effects on the host phenotype, although most viruses have neutral or no effect. We speculate that understanding multiple transmission modes of mycoviruses is central to unraveling the viral ecology and their function in regulating fungal populations. Unlike plant virus transmission via vegetative plant parts, seeds, pollen, or vectors, a widely held view is that mycoviruses are transmitted via vertical routes and only under special circumstances horizontally via hyphal contact depending on the vegetative compatibility groups (i.e., the ability of different fungal strains to undergo hyphal fusion). However, this view has been challenged over the past decades, as new possible transmission routes of mycoviruses are beginning to unravel. In this perspective, we discuss emerging studies with evidence suggesting that such novel routes of mycovirus transmission exist and are pertinent to understanding the full picture of mycovirus ecology and evolution.
•We studied phyllosphere application of glyphosate on barley rhizosphere.•Application of glyphosate caused an increased turn-over of the root system.•Application of glyphosate stimulated bacteria and ...protists abundance.•Application of glyphosate modified bacterial community profiles.•Glyphosate on sensitive plants increased culturability of bacteria and protists.
Glyphosate is extensively used for weed control and to ripen crops. Despite a number of studies on the direct effect of glyphosate on plants and soil organisms, only little is known about indirect effect of glyphosate on rhizosphere microbial communities, following the accelerated turnover of the fast-dying root biomass. In microcosms we studied the indirect effect of glyphosate on the microbial community in the rhizosphere of barley with phyllosphere application of glyphosate in comparison to leaving the plant intact or cutting off the shoot. Attempting to link the response of bacterial and protist communities to foliar application of glyphosate, we measured bacterial and protist abundance, diversity and physiological status, as well as soil organic carbon. Foliar application of glyphosate doubled bacterial abundance of the culturable fraction present in the rhizosphere compared to the other treatments with no effect on total abundance. Also the abundance of culturable protists increased as an effect of glyphosate and the bacterial genetic diversity as revealed by 16S rDNA DGGE analysis was affected. Overall, the results indicate that when barley leaves are treated with glyphosate, the availability of organic carbon in the rhizosphere of the dying roots is altered, which in turn may alter the bacterial and protist communities and their interactions. This can have implications for general soil carbon turnover processes and CO2 release in arable systems.
Anaerobic
fermentation is widely used to simulate rumen kinetics and study the microbiome and metabolite profiling in a controlled lab environment. However, a better understanding of the interplay ...between the temporal dynamics of fermentation kinetics, metabolic profiles, and microbial composition in
rumen fermentation batch systems is required. To fill that knowledge gap, we conducted three
rumen fermentations with maize silage as the substrate, monitoring total gas production (TGP), dry matter degradability (dDM), and methane (CH
) concentration at 6, 12, 24, 36, and 48 h in each fermentation. At each time point, we collected rumen fluid samples for microbiome analysis and volatile fatty acid (VFA) analysis. Amplicon sequencing of 16S rRNA genes (V4 region) was used to profile the prokaryotic community structure in the rumen during the fermentation process. As the fermentation time increased, dDM, TGP, VFA concentrations, CH
concentration, and yield (mL CH
per g DM at standard temperature and pressure (STP)) significantly increased. For the dependent variables, CH
concentration and yield, as well as the independent variables TGP and dDM, polynomial equations were fitted. These equations explained over 85% of the data variability (
> 0.85) and suggest that TGP and dDM can be used as predictors to estimate CH
production in rumen fermentation systems. Microbiome analysis revealed a dominance of Bacteroidota, Cyanobacteria, Desulfobacterota, Euryarchaeota, Fibrobacterota, Firmicutes, Patescibacteria, Proteobacteria, Spirochaetota, and Verrucomicrobiota. Significant temporal variations in Bacteroidota, Campylobacterota, Firmicutes, Proteobacteria, and Spirochaetota were detected. Estimates of alpha diversity based on species richness and the Shannon index showed no variation between fermentation time points. This study demonstrated that the
fermentation characteristics of a given feed type (e.g., maize silage) can be predicted from a few parameters (CH
concentration and yield, tVFA, acetic acid, and propionic acid) without running the actual
trial if the rumen fluid is collected from similar donor cows. Although the dynamics of the rumen prokaryotes changed remarkably over time and in accordance with the fermentation kinetics, more time points between 0 and 24 h are required to provide more details about the microbial temporal dynamics at the onset of the fermentation.
•A standardized protocol yields reproducible bulkDNA metabarcoding results.•Highly abundant and large species are shared most often between laboratories.•Need for standardization when comparing alpha ...diversity between different studies.•Betadiversity patterns are robust to changes in the metabarcoding wetlab protocol.
DNA metabarcoding can be used in marine environmental monitoring if results are reproducible between labs and robust against modifications to the lab protocol. In this interlaboratory study, we conducted a ring test where subsamples of blended macrobenthos samples were distributed to four laboratories located in Belgium, the Netherlands, Germany and Denmark. Samples were processed by a standardized lab protocol and by an adapted protocol, and the resulting datasets were analyzed with the same bioinformatics pipeline. Different biodiversity indicators were calculated. Our results show that bulkDNA metabarcoding of marine macrobenthos offers a highly reproducible assessment of alpha diversity patterns when using a standardized protocol, since comparable species numbers, Shannon indices and Inverse Simpson indices were found between laboratories. Especially high abundant species and species with large body sizes were shared between the laboratories. The need for using a standardized protocol to enhance comparability in alpha diversity between different studies was shown. Beta diversity patterns are less subjected to changes in the metabarcoding protocol and were almost identical between different laboratories, as the main clustering was always based on the macrobenthic community, independent of the used protocol or the laboratory that conducted the work. We conclude that DNA metabarcoding for marine environmental monitoring is an appropriate method when the aim is to study changes in community patterns and advocate its implementation in routine monitoring programs of national and European authorities, providing that a standardized protocol is implemented and/or a detailed description of the protocol is available.
The use of molecular methods to investigate protist communities in soil is in rapid development this decade. Molecular analysis of soil protist communities is usually dependant on direct genomic DNA ...extraction from soil and inefficient or differential DNA extraction of protist DNA can lead to bias in downstream community analysis. Three commonly used soil DNA extraction methods have been tested on soil samples from three European Long-Term Observatories (LTOs) with different land-use and three protist cultures belonging to different phylogenetic groups in different growth stages. The methods tested were: ISOm-11063 (a version of the ISO-11063 method modified to include a FastPrep ®-24 mechanical lysis step), GnS-GII (developed by the GenoSol platform to extract soil DNA in large-scale soil surveys) and a commercial DNA extraction kit — Power Lyzer™ PowerSoil® DNA Isolation Kit (MoBio). DNA yield and quality were evaluated along with DNA suitability for amplification of 18S rDNA fragments by PCR. On soil samples, ISOm-11063 yields significantly higher DNA for two of the three soil samples, however, MoBio extraction favors DNA quality. This method was also more effective to recover copies of 18S rDNA numbers from all soil types. In addition and despite the lower yields, higher DNA quality was observed with DNA extracted from protist cultures with the MoBio method. Likewise, a bead-beating step shows to be a good solution for DNA extraction of soil protists, since the recovery of DNA from protist cultures and from the different soil samples with the ISOm method proved to be efficient in recovering PCR-amplifiable DNA. This study showed that soil DNA extraction methods provide biased results towards the cyst stages of protist organism.
•Commercial kit favors DNA quality at the cost of lower DNA yields.•Commercial kit recovered higher quantities of eukaryotic 18S rDNA from soil.•Non-commercial methods recovered higher quantities of DNA from in vitro protist.•Soil DNA isolation biased towards trophozoites of protists at the expense of cysts.
Advances in sequencing technologies allow deeper studies of the soil protist diversity and function. However, little attention has been given to the impact of the chosen soil DNA extraction procedure ...to the overall results. We examined the effect of three acknowledged DNA recovery methods, two manual methods (ISOm-11063, GnS-GII) and one commercial kit (MoBio), on soil protist community structures obtained from different sites with different land uses. Results from 18S rRNA gene amplicon sequencing suggest that DNA extraction method significantly affect the replicate homogeneity, the total number of operational taxonomic units (OTUs) recovered and the overall taxonomic structure and diversity of soil protist communities. However, DNA extraction effects did not overwhelm the natural variation among samples, as the community data still strongly grouped by geographical location. The commercial DNA extraction kit was associated with the highest diversity estimates and with a corresponding higher retrieval of Excavata, Cercozoa and Amoebozoa-related taxa. Overall, our findings indicate that this extraction offers a compromise between rare and dominant taxa representation, while providing high replication reproducibility. A comprehensive understanding of the DNA extraction techniques impact on soil protist diversity can enable more accurate diversity assays.
We investigated the use of eDNA metabarcoding for supplementing traditional diver-based monitoring of biodiversity of marine boulder reefs within the photic zone. The applied sampling design made it ...possible to evaluate the usefulness of eDNA monitoring as a supplement for traditional monitoring. Specifically, this study aimed to (1) assess the local influence of boulder reefs on biodiversity across the North Sea to Baltic Sea transition zone and (2) investigate the importance of environmental gradients for patterns in community structure. On samples collected during August 2020, we compared the composition and abundance of species associated with nine reefs, representing an environmental gradient of salinity (16–33 psu), water temperature (16–21°C) and water depth (6–29 m). At each reef site, water was sampled near the bottom just above the reef and on average 2.6 km upstream and downstream (location) and sequenced with metabarcoding using COI, 18S and 12S rDNA primers. eDNA identified 400 species, diver-based observations identified 184 with an overlap of 70 species (12%) and 81 genera (18%). While eDNA identified many infaunal species, it did not detect several macroalgal species which dominated in the diver-based observations. Multivariate analysis of eDNA and diver-based community structure both distinguished between reef communities, with a significant match between patterns observed by the two methods (
r
= 0.37,
p
= 0.02). Furthermore, the eDNA approach made it possible to identify significant differences in species composition between upstream, above-reef and downstream locations, suggesting that eDNA leaves a local footprint in benthic habitats. Patterns in both eDNA and diver-based species composition and richness were significantly related to geographical distance, salinity, water temperature and water depth. Despite of low detection of macroalgae, the eDNA sampling provided a substantial supplement to traditional diver-based monitoring of biodiversity around benthic hotspots in the Danish marine waters and therefore we recommend to add eDNA methods to conventional monitoring programs in the future.
Microorganisms interact with plant roots through colonization of the root surface, i.e., the rhizoplane or the surrounding soil, i.e., the rhizosphere. Beneficial rhizosphere bacteria such as ...Pseudomonas spp. can promote plant growth and protect against pathogens by producing a range of bioactive compounds, including specialized metabolites like cyclic lipopeptides (CLPs) known for their biosurfactant and antimicrobial activities. However, the role of CLPs in natural soil systems during bacteria-plant interactions is underexplored. Here, Pseudomonas fluorescens SBW25, producing the CLP viscosin, was used to study the impact of viscosin on bacterial root colonization and microbiome assembly in two cultivars of winter wheat (Heerup and Sheriff). We inoculated germinated wheat seeds with SBW25 wild type or a viscosin-deficient mutant and grew the plants in agricultural soil. After 2 weeks, enhanced root colonization of SBW25 wild type compared to the viscosin-deficient mutant was observed, while no differences were observed between wheat cultivars. In contrast, the impact on root-associated microbial community structure was plant-genotype-specific, and SBW25 wild type specifically reduced the relative abundance of an unclassified oomycete and Phytophthora in Sheriff and Heerup, respectively. This study provides new insights into the natural role of viscosin and specifically highlights the importance of viscosin in wheat root colonization under natural soil conditions and in shaping the root microbial communities associated with different wheat cultivars. Furthermore, it pinpoints the significance of microbial microdiversity, plant genotype, and microbe-microbe interactions when studying colonization of plant roots.IMPORTANCEUnderstanding parameters governing microbiome assembly on plant roots is critical for successfully exploiting beneficial plant-microbe interactions for improved plant growth under low-input conditions. While it is well-known from in vitro studies that specialized metabolites are important for plant-microbe interactions, e.g., root colonization, studies on the ecological role under natural soil conditions are limited. This might explain the often-low translational power from laboratory testing to field performance of microbial inoculants. Here, we showed that viscosin synthesis potential results in a differential impact on the microbiome assembly dependent on wheat cultivar, unlinked to colonization potential. Overall, our study provides novel insights into factors governing microbial assembly on plant roots, and how this has a derived but differential effect on the bacterial and protist communities.
Here we show that a commercial blocking reagent (G2) based on modified eukaryotic DNA significantly improved DNA extraction efficiency. We subjected G2 to an inter-laboratory testing, where DNA was ...extracted from the same clay subsoil using the same batch of kits. The inter-laboratory extraction campaign revealed large variation among the participating laboratories, but the reagent increased the number of PCR-amplified16S rRNA genes recovered from biomass naturally present in the soils by one log unit. An extensive sequencing approach demonstrated that the blocking reagent was free of contaminating DNA, and may therefore also be used in metagenomics studies that require direct sequencing.
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