Comparative fungal community analyses using metatranscriptomics and ITS-amplicon sequencing from Norway spruce

2021 ◽  
Author(s):  
Andreas Schneider ◽  
John Sundh ◽  
Görel Sundström ◽  
Kerstin Richau ◽  
Nicolas Delhomme ◽  
...  
Keyword(s):  
Community Structure ◽  
Microbial Communities ◽  
Norway Spruce ◽  
Environmental Samples ◽  
Marker Gene ◽  
Amplicon Sequencing ◽  
Fungal Communities ◽  
Individual Species ◽  
Rna Seq ◽  
Sequencing Data

<p>Microbial communities are major players in carbon and nitrogen cycling globally and are of particular importance for plant communities in the nutrient poor soils of boreal forests. Especially relevant are the fungal communities in the soil that interact with the plants in multiple ways, indirectly through their pivotal role in the breakdown of organic matter and, more directly, through mycorrhizal symbiosis with plant roots. Large-scale disturbances of these complex microbial communities can lead to shifts in soil carbon storage with unknown and global-scale long-term consequences. To understand the dynamics of these communities and their relationship to associated plants in response to climate change and anthropogenic influence, we need a better understanding of how modern “omics” methods can help us to understand compositional and functional shifts of these microbiomes. Microbial gene expression and functional activity can be assayed with RNA sequencing (RNA-Seq) data from environmental samples. In contrast, currently phylogenetic marker gene amplicon sequencing data is generally used to assess taxonomic composition and community structure of the microbiome. Few studies have considered how much of this structural and taxonomic information is included in RNA-Seq transcriptomic data from matched samples. Here we describe fungal communities using both RNA-Seq and fungal ITS1 DNA amplicon sequencing to compare the outcomes between the methods. We used a panel of root and needle samples from mature stands of the coniferous tree species Picea abies (Norway spruce) growing in untreated (nutrient deficient) and nutrient enriched plots at the Flakaliden forest research site in boreal northern Sweden. We created an assembly-based, reproducible and hardware agnostic workflow to taxonomically and functionally annotate fungal RNA-Seq data obtained from Norway spruce roots, which we compared to matching ITS amplicon sequencing data.<strong> </strong>We show that the community structure indicated by the fungal transcriptome is in agreement with that generated by the ITS data, while also identifying limitations imposed by current database coverage. Furthermore, we show examples to demonstrate how metatranscriptomics data additionally provides biologically informative functional insight at the community and individual species level. These findings highlight the potential of metatranscriptomics to advance our understanding of interaction, response and effect both between host plants and their associated microbial communities, and among the members of microbial communities in environmental samples in general.</p>

scholarly journals Comparative Fungal Community Analyses Using Metatranscriptomics and Internal Transcribed Spacer Amplicon Sequencing from Norway Spruce

mSystems ◽  
2021 ◽  
Vol 6 (1) ◽  
Author(s):  
Andreas N. Schneider ◽  
John Sundh ◽  
Görel Sundström ◽  
Kerstin Richau ◽  
Nicolas Delhomme ◽  
...  
Keyword(s):  
Gene Expression ◽  
Microbial Communities ◽  
Norway Spruce ◽  
Marker Gene ◽  
Amplicon Sequencing ◽  
Rna Seq ◽  
Sequencing Data ◽  
Plant Component ◽  
Alpha And Beta Diversity ◽  
Field Samples

ABSTRACT The health, growth, and fitness of boreal forest trees are impacted and improved by their associated microbiomes. Microbial gene expression and functional activity can be assayed with RNA sequencing (RNA-Seq) data from host samples. In contrast, phylogenetic marker gene amplicon sequencing data are used to assess taxonomic composition and community structure of the microbiome. Few studies have considered how much of this structural and taxonomic information is included in transcriptomic data from matched samples. Here, we described fungal communities using both host-derived RNA-Seq and fungal ITS1 DNA amplicon sequencing to compare the outcomes between the methods. We used a panel of root and needle samples from the coniferous tree species Picea abies (Norway spruce) growing in untreated (nutrient-deficient) and nutrient-enriched plots at the Flakaliden forest research site in boreal northern Sweden. We show that the relationship between samples and alpha and beta diversity indicated by the fungal transcriptome is in agreement with that generated by the ITS data, while also identifying a lack of taxonomic overlap due to limitations imposed by current database coverage. Furthermore, we demonstrate how metatranscriptomics data additionally provide biologically informative functional insights. At the community level, there were changes in starch and sucrose metabolism, biosynthesis of amino acids, and pentose and glucuronate interconversions, while processing of organic macromolecules, including aromatic and heterocyclic compounds, was enriched in transcripts assigned to the genus Cortinarius. IMPORTANCE A deeper understanding of microbial communities associated with plants is revealing their importance for plant health and productivity. RNA extracted from plant field samples represents the host and other organisms present. Typically, gene expression studies focus on the plant component or, in a limited number of studies, expression in one or more associated organisms. However, metatranscriptomic data are rarely used for taxonomic profiling, which is currently performed using amplicon approaches. We created an assembly-based, reproducible, and hardware-agnostic workflow to taxonomically and functionally annotate fungal RNA-Seq data obtained from Norway spruce roots, which we compared to matching ITS amplicon sequencing data. While we identified some limitations and caveats, we show that functional, taxonomic, and compositional insights can all be obtained from RNA-Seq data. These findings highlight the potential of metatranscriptomics to advance our understanding of interaction, response, and effect between host plants and their associated microbial communities.

scholarly journals Rbec: a tool for analysis of amplicon sequencing data from synthetic microbial communities

2021 ◽  
Vol 1 (1) ◽  
Author(s):  
Pengfan Zhang ◽  
Stjin Spaepen ◽  
Yang Bai ◽  
Stephane Hacquard ◽  
Ruben Garrido-Oter
Keyword(s):  
Microbial Communities ◽  
Amplicon Sequencing ◽  
Fungal Communities ◽  
Polymorphic Variation ◽  
Sequencing Data ◽  
Sequencing Errors ◽  
Extensive Evaluation ◽  
Culture Independent ◽  
Reference Sequences ◽  
Synthetic Microbial Communities

AbstractSynthetic microbial communities (SynComs) constitute an emerging and powerful tool in biological, biomedical, and biotechnological research. Despite recent advances in algorithms for the analysis of culture-independent amplicon sequencing data from microbial communities, there is a lack of tools specifically designed for analyzing SynCom data, where reference sequences for each strain are available. Here we present Rbec, a tool designed for the analysis of SynCom data that accurately corrects PCR and sequencing errors in amplicon sequences and identifies intra-strain polymorphic variation. Extensive evaluation using mock bacterial and fungal communities show that our tool outperforms current methods for samples of varying complexity, diversity, and sequencing depth. Furthermore, Rbec also allows accurate detection of contaminants in SynCom experiments.

scholarly journals Reference-based error correction of amplicon sequencing data from synthetic communities

2021 ◽  
Author(s):  
Pengfan Zhang ◽  
Stjin Spaepen ◽  
Yang Bai ◽  
Stephane Hacquard ◽  
Ruben Garrido-Oter
Keyword(s):  
Microbial Communities ◽  
Amplicon Sequencing ◽  
R Package ◽  
Fungal Communities ◽  
Polymorphic Variation ◽  
Sequencing Data ◽  
Extensive Evaluation ◽  
Culture Independent ◽  
Reference Sequences ◽  
Synthetic Microbial Communities

AbstractMotivationSynthetic microbial communities (SynComs) constitute an emergent and powerful tool in biological, biomedical, and biotechnological research. Despite recent advances in algorithms for analysis of culture-independent amplicon sequencing data from microbial communities, there is a lack of tools specifically designed for analysing SynCom data, where reference sequences for each strain are available.ResultsHere we present Rbec, a tool designed for analysing SynCom data that outperforms current methods by accurately correcting errors in amplicon sequences and identifying intra-strain polymorphic variation. Extensive evaluation using mock bacterial and fungal communities show that our tool performs robustly for samples of varying complexity, diversity, and sequencing depth. Further, Rbec also allows accurate detection of contaminations in SynCom experiments.AvailabilityRbec is freely available as an open-source R package and can be downloaded at: https://github.com/PengfanZhang/Microbiome.

scholarly journals The response of soil and phyllosphere microbial communities to repeated application of the fungicide iprodione: accelerated biodegradation or toxicity?

2020 ◽  
Vol 96 (6) ◽  
Author(s):  
A Katsoula ◽  
S Vasileiadis ◽  
M Sapountzi ◽  
Dimitrios G Karpouzas
Keyword(s):  
Microbial Communities ◽  
Agricultural Production ◽  
Soil Microorganisms ◽  
Amplicon Sequencing ◽  
Fungal Communities ◽  
Similar Response ◽  
Repeated Application ◽  
Plant Leaves ◽  
Soil System ◽  
Plant Soil

ABSTRACT Pesticides interact with microorganisms in various ways with the outcome being negative or positive for the soil microbiota. Pesticides' effects on soil microorganisms have been studied extensively in soil but not in other pesticides-exposed microbial habitats like the phyllosphere. We tested the hypothesis that soil and phyllosphere support distinct microbial communities, but exhibit a similar response (accelerated biodegradation or toxicity) to repeated exposure to the fungicide iprodione. Pepper plants received four repeated foliage or soil applications of iprodione, which accelerated its degradation in soil (DT50_1st = 1.23 and DT50_4th = 0.48 days) and on plant leaves (DT50_1st > 365 and DT50_4th = 5.95 days). The composition of the epiphytic and soil bacterial and fungal communities, determined by amplicon sequencing, was significantly altered by iprodione. The archaeal epiphytic and soil communities responded differently; the former showed no response to iprodione. Three iprodione-degrading Paenarthrobacter strains were isolated from soil and phyllosphere. They hydrolyzed iprodione to 3,5-dichloraniline via the formation of 3,5-dichlorophenyl-carboxiamide and 3,5-dichlorophenylurea-acetate, a pathway shared by other soil-derived arthrobacters implying a phylogenetic specialization in iprodione biotransformation. Our results suggest that iprodione-repeated application could affect soil and epiphytic microbial communities with implications for the homeostasis of the plant–soil system and agricultural production.

scholarly journals High-throughput amplicon sequencing reveals the spatiotemporal effects, abiotic and biotic shaping factors for the microbial communities in tropical mangrove sediments in Sanya, China

2020 ◽  
Author(s):  
Wu Qu ◽  
Boliang Gao ◽  
Jie Wu ◽  
Min Jin ◽  
Jianxin Wang ◽  
...  
Keyword(s):  
Community Structure ◽  
Microbial Community ◽  
Microbial Communities ◽  
Fungal Community ◽  
Amplicon Sequencing ◽  
Biotic Factors ◽  
Community Structures ◽  
Mangrove Sediments ◽  
Element Cycling ◽  
Microbial Community Structures

Abstract Background Microbial roles in element cycling and nutrient providing are crucial for mangrove ecosystems and serve as important regulators for climate change in Earth ecosystem. However, some key information about the spatiotemporal influences and abiotic and biotic shaping factors for the microbial communities in mangrove sediments remains lacking. Methods In this work, 22 sediment samples were collected from multiple spatiotemporal dimensions, including three locations, two depths, and four seasons, and the bacterial, archaeal, and fungal community structures in these samples were studied using amplicon sequencing. Results The microbial community structures were varied in the samples from different depths and locations based on the results of LDA effect size analysis, principal coordinate analysis, the analysis of similarities, and permutational multivariate ANOVA. However, these microbial community structures were stable among the seasonal samples. Linear fitting models and Mantel test showed that among the 13 environmental factors measured in this study, the sediment particle size (PS) was the key abiotic shaping factor for the bacterial, archaeal, or fungal community structure. Besides PS, salinity and humidity were also significant impact factors according to the canonical correlation analysis (p ≤ 0.05). Co-occurrence networks demonstrated that the bacteria assigned into phyla Ignavibacteriae, Proteobacteria, Bacteroidetes, Chloroflexi, and Acidobacteria were the key biotic factors for shaping the bacterial community in mangrove sediments. Conclusions This work showed the variability on spatial dimensions and the stability on temporal dimension for the bacterial, archaeal, or fungal microbial community structure, indicating that the tropical mangrove sediments are versatile but stable environments. PS served as the key abiotic factor could indirectly participate in material circulation in mangroves by influencing microbial community structures, along with salinity and humidity. The bacteria as key biotic factors were found with the abilities of photosynthesis, polysaccharide degradation, or nitrogen fixation, which were potential indicators for monitoring mangrove health, as well as crucial participants in the storage of mangrove blue carbons and mitigation of climate warming. This study expanded the knowledge of mangroves for the spatiotemporal variation, distribution, and regulation of the microbial community structures, thus further elucidating the microbial roles in mangrove management and climate regulation.

scholarly journals Copper affects composition and functions of microbial communities in marine biofilms at environmentally relevant concentrations

2018 ◽  
Author(s):  
Natàlia Corcoll ◽  
Jianghua Yang ◽  
Thomas Backhaus ◽  
Xiaowei Zhang ◽  
Martin Karl Eriksson ◽  
...  
Keyword(s):  
Community Structure ◽  
Primary Production ◽  
Microbial Communities ◽  
Algal Biomass ◽  
Amplicon Sequencing ◽  
Quality Standard ◽  
Aquatic Communities ◽  
Static Test ◽  
Community Function ◽  
Eukaryotic Organisms

Cu pollution in coastal areas is a worldwide threat for aquatic communities. This study assesses the effects of Cu exposure on microbial diversity, community structure and functions of microbial communities in marine periphyton biofilms at environmental relevant concentrations. Periphyton was exposed for 18 days to five Cu concentrations, between 0.01 and 10 μM, in a semi-static test. Diversity and community structure of prokaryotic and eukaryotic organisms were assessed by 16S and 18S amplicon sequencing, respectively. Community function was studied as impacts on algal biomass and primary production. Additionally, we studied Pollution-Induced Community Tolerance (PICT) using photosynthesis as the endpoint. Sequencing results detected an average of 9504 and 1242 OTUs for 16S and 18S, respectively, reflecting the huge biodiversity of marine periphytic biofilms. Eukaryotes represent the most Cu-sensitive kingdom, where effects were seen already at concentrations as low as 10 nM. The structure of the prokaryotic part of the community was impacted at slightly higher concentrations (60 nM), which is still in the range of the Cu concentrations observed in the area (80 nM).The current environmental quality standard for Cu of 70 nM therefore does not seem to be sufficiently protective for periphyton. Cu exposure resulted in a more Cu-tolerant community, which was accompanied by a reduced total algal biomass, increased relative abundance of diatoms and a reduction of primary production. Cu exposure changed the network of associations between taxa in the communities. A total of 23 taxa, including species within Proteobacteria, Bacteroidetes, Stramenopiles and Hacrobia, were identified as being particularly sensitive to Cu. DNA metabarcoding is presented as a sensitive tool for community-level ecotoxicological studies that allows to observe impacts simultaneously on a multitude of pro- and eukaryotic species, and therefore to identify particularly sensitive, non-cultivable species and taxa.

scholarly journals Depth and location influence prokaryotic and eukaryotic microbial community structure in New Zealand fjords

2019 ◽  
Author(s):  
Sven P. Tobias-Hünefeldt ◽  
Stephen R. Wing ◽  
Nadjejda Espinel-Velasco ◽  
Federico Baltar ◽  
Sergio E. Morales
Keyword(s):  
Community Structure ◽  
New Zealand ◽  
Microbial Communities ◽  
National Park ◽  
Amplicon Sequencing ◽  
Forested Catchments ◽  
Environmental Forcing ◽  
Low Salinity ◽  
The Impact ◽  
Depth View

SummarySystems with strong horizontal and vertical gradients, such as fjords, are useful models for studying environmental forcing. Here we examine microbial (prokaryotic and eukaryotic) community changes associated with the surface low salinity layer (LSL) and underlying seawater in multiple fjords in Fiordland National Park (New Zealand). High rainfall (1200-8000 mm annually) and linked runoff from native forested catchments results in surface LSLs with high tannin concentrations within each fjord. These gradients are expected to drive changes in microbial communities. We used amplicon sequencing (16S and 18S) to assess the impact of these gradients on microbial communities and identified depth linked changes in diversity and community structure. With increasing depth we observed significant increases in Proteobacteria (15%) and SAR (37%), decreases in Opisthokonta (35%), and transiently increased Bacteroidetes (3% increase from 0 to 40 m, decreasing by 8% at 200 m). Community structure differences were observed along a transect from inner to outer regions, specifically 25% mean relative abundance decreases in Opisthokonta and Bacteroidetes, and increases in SAR (25%) and Proteobacteria (>5%) at the surface, indicating changes based on distance from the ocean. This provides the first in-depth view into the ecological drivers of microbial communities within New Zealand fjords.

scholarly journals Microbial Communities on Plastic Polymers in the Mediterranean Sea

2021 ◽  
Vol 12 ◽  
Author(s):  
Annika Vaksmaa ◽  
Katrin Knittel ◽  
Alejandro Abdala Asbun ◽  
Maaike Goudriaan ◽  
Andreas Ellrott ◽  
...  
Keyword(s):  
Microbial Communities ◽  
Mediterranean Sea ◽  
Amplicon Sequencing ◽  
Polymer Surfaces ◽  
Rrna Gene ◽  
Sequencing Data ◽  
Microbial Biofilms ◽  
Degrading Bacteria ◽  
The Mediterranean

Plastic particles in the ocean are typically covered with microbial biofilms, but it remains unclear whether distinct microbial communities colonize different polymer types. In this study, we analyzed microbial communities forming biofilms on floating microplastics in a bay of the island of Elba in the Mediterranean Sea. Raman spectroscopy revealed that the plastic particles mainly comprised polyethylene (PE), polypropylene (PP), and polystyrene (PS) of which polyethylene and polypropylene particles were typically brittle and featured cracks. Fluorescence in situ hybridization and imaging by high-resolution microscopy revealed dense microbial biofilms on the polymer surfaces. Amplicon sequencing of the 16S rRNA gene showed that the bacterial communities on all plastic types consisted mainly of the orders Flavobacteriales, Rhodobacterales, Cytophagales, Rickettsiales, Alteromonadales, Chitinophagales, and Oceanospirillales. We found significant differences in the biofilm community composition on PE compared with PP and PS (on OTU and order level), which shows that different microbial communities colonize specific polymer types. Furthermore, the sequencing data also revealed a higher relative abundance of archaeal sequences on PS in comparison with PE or PP. We furthermore found a high occurrence, up to 17% of all sequences, of different hydrocarbon-degrading bacteria on all investigated plastic types. However, their functioning in the plastic-associated biofilm and potential role in plastic degradation needs further assessment.

Community Structure and Co-Occurrence Network Analysis of Bacteria and Fungi in Wheat Fields vs Fruit Orchards

2021 ◽  
Author(s):  
Xinyu Cui ◽  
Huan He ◽  
Fengxiao Zhu ◽  
Xiaobo Liu ◽  
You Ma ◽  
...  
Keyword(s):  
Community Structure ◽  
Amplicon Sequencing ◽  
Yangtze River Delta ◽  
Vital Role ◽  
Fungal Communities ◽  
Rrna Gene ◽  
Network Analyses ◽  
Planting Methods ◽  
Bacteria And Fungi ◽  
Occurrence Patterns

Abstract Soil microorganisms play a vital role in biogeochemical processes and nutrient turnover in agricultural ecosystems. However, the information on how the structure and co-occurrence patterns of microbial communities respond to the change of planting methods is still limited. In this study, a total of 34 soil samples were collected from 17 different fields of two planting types (wheat and orchards) along the Taige Canal in Yangtze River Delta. The distribution and diversity of bacterial and fungal communities in soil were determined using amplicon sequencing targeting the 16S rRNA gene and ITS gene, respectively. The dominated bacteria were Proteobacteria, Acidobacteriota, Actinobacteriota, Chloroflexi, Bacteroidota, and Firmicutes. The relative abundance of Actinobacteriota and Firmicutes was higher in the orchards, while Chloroflexi and Nitrospirota were more abundant in wheat fields. Ascomycota, Mortierellomycota, and Basidiomycota were the predominant fungi in both types of soils. The diversity of bacterial and fungal communities was greater in the wheat fields than in the orchards. The statistical analyses showed that pH was the main factor shaping the community structure. Moreover, high co-occurrence patterns of bacteria and fungi were confirmed in both wheat fields and orchards. Network analyses showed that both the wheat fields and orchards occurred modular structure, which mainly contained nodes of Acidobacteriota, Chloroflexi, Gemmatimonadota, Nitrospirota and Ascomycota. In summary, our work showed the co-occurrence network and the convergence/divergence of microbial community structure in wheat fields and orchards, giving a comprehensive understanding of the microbe-microbe interaction during planting methods changes.

Microbiome variation during culture growth of the European house dust mite, Dermatophagoides pteronyssinus

2021 ◽  
Vol 97 (4) ◽  
Author(s):  
Marta Nesvorna ◽  
Stano Pekar ◽  
Elena Shcherbachenko ◽  
Vit Molva ◽  
Tomas Erban ◽  
...  
Keyword(s):  
Community Structure ◽  
Microbial Community ◽  
Microbial Communities ◽  
Relative Abundance ◽  
Environmental Samples ◽  
Dermatophagoides Pteronyssinus ◽  
Dust Mite ◽  
Mite Density ◽  
Culture Environment ◽  
Culture Growth

ABSTRACT In culture, the house dust mite, Dermatophagoides pteronyssinus, shows different growth patterns, but the composition and changes in the associated microbial community during mite culture growth are poorly known. In this study, we analyzed temporal changes in microbial communities including ‘internal’ communities (inside mites, ingested) and ‘environmental’ communities (from culture environment). Microbial community structure was correlated with guanine content (a nitrogenous waste product of mites) and mite population density. Both internal and environmental microbial communities were remarkably consistent between biological replicates from the same culture age group and were composed of relatively few dominant taxa–11 bacterial and 3 fungal operational taxonomic units (OTUs). Significant changes over time in microbial community structure in the bulk culture environment and in internal mite samples were observed. The yeast, Saccharomyces cerevisiae, a main component of the mite diet, gradually disappeared during mite culture growth and was replaced by fungi from the genera Aspergillus and Candida in both ‘internal’ and ‘environmental’ samples. In environmental samples, bacteria from the genus Lactobacillus and S. cerevisiae were negatively correlated, and Aspergillus and Candida positively correlated, with guanine content. The relative abundance of bacteria from the genus Kocuria increased with mite density but declined with increasing guanine content. The relative abundance of bacteria from the genus Virgibacillus was negatively correlated with mite density in ‘internal’ samples. Gram-positive bacteria dominated bacterial microbiomes at all time points in our experiments, indicating a more limited possibility for vaccine contamination by bacterial endotoxins (heat-stable lipopolysaccharides produced mostly by Gram-negative bacteria) in our experimental cultures.

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