Ecology Of Microbial Communities Research Articles

Data-driven prediction of colonization outcomes for complex microbial communities

Microbial interactions can lead to different colonization outcomes of exogenous species, be they pathogenic or beneficial in nature. Predicting the colonization of exogenous species in complex communities remains a fundamental challenge in microbial ecology, mainly due to our limited knowledge of the diverse mechanisms governing microbial dynamics. Here, we propose a data-driven approach independent of any dynamics model to predict colonization outcomes of exogenous species from the baseline compositions of microbial communities. We systematically validate this approach using synthetic data, finding that machine learning models can predict not only the binary colonization outcome but also the post-invasion steady-state abundance of the invading species. Then we conduct colonization experiments for commensal gut bacteria species Enterococcus faecium and Akkermansia muciniphila in hundreds of human stool-derived in vitro microbial communities, confirming that the data-driven approaches can predict the colonization outcomes in experiments. Furthermore, we find that while most resident species are predicted to have a weak negative impact on the colonization of exogenous species, strongly interacting species could significantly alter the colonization outcomes, e.g., Enterococcus faecalis inhibits the invasion of E. faecium invasion. The presented results suggest that the data-driven approaches are powerful tools to inform the ecology and management of microbial communities.

Assessment of Endophytic Fungal Community in Palm Leaves and Their Biocontrol Potential

Microbial endophytes live asymptomatically within a host plant and contribute positively to its fitness. Endophytes associate ubiquitously with all plants examined so far and form an integral part of the host for processes such as nutrient uptake and stress response and traits like disease resistance. Well-characterized host–endophyte interaction models exist, but little is known about the microbiome of the monocot plant host, palm. In this study, we identified the endophytic fungal community associated with mule palm leaves with an objective to test their antagonist effects on lethal pathogens of palms. The leaflets of mule palms were sampled across five sites in Central and South Florida, and fungal endophytes were identified using culture-dependent (CD) and amplicon sequence-based culture-independent (CI) methods. Operational taxonomic units (OTUs) corresponding to phylum Ascomycota were identified by both approaches, whereas Basidiomycota were only detected using the CI method. Among the ascomycetes, a small (22%) overlap in the OTUs at the genus level was observed between the two methods, whereas the majority (64%) of OTUs were detected using the CI approach. In vitro cocultivation of endophytes with palm pathogens was observed to produce three distinct interaction phenotypes: inhibition zone, niche establishment, and overlapping growth. Eight endophytes capable of inhibiting fungal pathogen growth in vitro were also identified, suggesting a biocontrol potential for endophyte-derived secondary metabolites. A better understanding of the composition, assembly, and ecology of palm-associated microbial communities would eventually lead to healthy and resilient palms. [Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license .

Ubiquitous filter feeders shape open ocean microbial community structure and function.

The mechanism of mortality plays a large role in how microorganisms in the open ocean contribute to global energy and nutrient cycling. Salps are ubiquitous pelagic tunicates that are a well-known mortality source for large phototrophic microorganisms in coastal and high-latitude systems, but their impact on the immense populations of smaller prokaryotes in the tropical and subtropical open ocean gyres is not well quantified. We used robustly quantitative techniques to measure salp clearance and enrichment of specific microbial functional groups in the North Pacific Subtropical Gyre, one of the largest ecosystems on Earth. We discovered that salps are a previously unknown predator of the globally abundant nitrogen fixer Crocosphaera; thus, salps restrain new nitrogen delivery to the marine ecosystem. We show that the ocean's two numerically dominant cells, Prochlorococcus and SAR11, are not consumed by salps, which offers a new explanation for the dominance of small cells in open ocean systems. We also identified a double bonus for Prochlorococcus, wherein it not only escapes salp predation but the salps also remove one of its major mixotrophic predators, the prymnesiophyte Chrysochromulina. When we modeled the interaction between salp mesh and particles, we found that cell size alone could not account for these prey selection patterns. Instead, the results suggest that alternative mechanisms, such as surface property, shape, nutritional quality, or even prey behavior, determine which microbial cells are consumed by salps. Together, these results identify salps as a major factor in shaping the structure, function, and ecology of open ocean microbial communities.

Uncultivated DPANN archaea are ubiquitous inhabitants of global oxygen-deficient zones with diverse metabolic potential.

Archaea from the DPANN (Diapherotrites, Parvarchaeota, Aenigmarchaeota, Nanoarchaeota, and Nanohaloarchaeota) superphylum have diverse metabolic capabilities and participate in multiple biogeochemical cycles. While metagenomics and enrichments have revealed that many DPANN are characterized by ultrasmall genomes, few biosynthetic genes, and episymbiotic lifestyles, much remains unknown about their biology. We report 33 new DPANN metagenome-assembled genomes originating from the three global marine oxygen-deficient zones (ODZs), the first from these regions. We survey DPANN abundance and distribution within the ODZ water column, investigate their biosynthetic capabilities, and report potential roles in the cycling of organic carbon, methane, and nitrogen. We test the hypothesis that nitrous oxide reductases found within several ODZ DPANN genomes may enable ultrasmall episymbionts to serve as nitrous oxide consumers when attached to a host nitrous oxide producer. Our results indicate DPANN archaea as ubiquitous residents within the anoxic core of ODZs with the potential to produce or consume key compounds.

Functional Swiss-type cheeses promote beneficial effects in mice gut microbiome during homeostasis and inflammation

The impact of Functional foods on the gut microbiota is a fundamental question since shifts in bacterial composition are associated with inflammation progression. Therefore, this study investigated the effects of two types of cheeses fermented by probiotic bacteria in C57BL6 mice either in healthy conditions or in dextran sulfate sodium (DSS)-induced colitis. One cheese was fermented by Propionibacterium freudenreichii CIRM-BIA129. A second kind of cheese was made in industrial conditions to reproduce Emmental but using the above-mentioned strain in combination with Lactobacillusdelbrueckii CNRZ327 and Streptococcus thermophilus LMD-9 as starters. The gut microbiota was investigated by shotgun metagenomic sequencing and the taxonomic and functional profiles were assessed through sequence alignment to NCBI taxonomy and KEGG pathways databases. Our results suggest that both treatments did not disturb the microbial community ecology. Emmental intake increased symbionts as Romboutsia and Akkermansia muciniphila. Furthermore, metabolic pathway analysis suggests that A. muciniphila may produce bioactive metabolites as acetate and cooperate with other commensal species to produce indole and gamma-aminobutyric acid. Mice with colitis have restored some of the impaired microbiome metabolic functions when treated with cheese fermented only by Propionibacterium, while the Emmental cheese promoted the increase of Ligilactobacillus murinus. This taxon presented several genes with immunomodulatory activity potential. This study provides insights for engineering functional cheeses to increase beneficial bacteria in the gut. However, further studies are required to investigate their role in regulating the gut-brain axis. Moreover, possible anti-inflammatory mechanisms based on the promotion of Lg. murinus interaction with the host was suggested.

Genome-Resolved Metagenomics and Metatranscriptomics Reveal Insights into the Ecology and Metabolism of Anaerobic Microbial Communities in PCB-Contaminated Sediments.

Growth of organohalide-respiring bacteria such as Dehalococcoides mccartyi on halogenated organics (e.g., polychlorinated biphenyls (PCBs)) at contaminated sites or in enrichment culture requires interaction and support from other microbial community members. To evaluate naturally occurring interactions between Dehalococcoides and key supporting microorganisms (e.g., production of H2, acetate, and corrinoids) in PCB-contaminated sediments, metagenomic and metatranscriptomic sequencing was conducted on DNA and RNA extracted from sediment microcosms, showing evidence of both Dehalococcoides growth and PCB dechlorination. Using a genome-resolved approach, 160 metagenome-assembled genomes (MAGs), including three Dehalococcoides MAGs, were recovered. A novel reductive dehalogenase gene, distantly related to the chlorophenol dehalogenase gene cprA (pairwise amino acid identity: 23.75%), was significantly expressed. Using MAG gene expression data, 112 MAGs were assigned functional roles (e.g., corrinoid producers, acetate/H2 producers, etc.). A network coexpression analysis of all 160 MAGs revealed correlations between 39 MAGs and the Dehalococcoides MAGs. The network analysis also showed that MAGs assigned with functional roles that support Dehalococcoides growth (e.g., corrinoid assembly, and production of intermediates required for corrinoid synthesis) displayed significant coexpression correlations with Dehalococcoides MAGs. This work demonstrates the power of genome-resolved metagenomic and metatranscriptomic analyses, which unify taxonomy and function, in investigating the ecology of dehalogenating microbial communities.

Effects of hydrogeochemistry on the microbial ecology of terrestrial hot springs.

Temperature, pH, and hydrochemistry of terrestrial hot springs play a critical role in shaping thermal microbial communities. However, the interactions of biotic and abiotic factors at this terrestrial-aquatic interface are still not well understood on a global scale, and the question of how underground events influence microbial communities remains open. To answer this, 11 new samples obtained from the El Tatio geothermal field were analyzed by 16S rRNA amplicon sequencing (V4 region), along with 191 samples from previous publications obtained from the Taupo Volcanic Zone, the Yellowstone Plateau Volcanic Field, and the Eastern Tibetan Plateau, with their temperature, pH, and major ion concentration. Microbial alpha diversity was lower in acid-sulfate waters, and no significant correlations were found with temperature. However, moderate correlations were observed between chemical parameters such as pH (mostly constrained to temperatures below 70°C), SO4 2- and abundances of members of the phyla Armatimonadota, Deinococcota, Chloroflexota, Campilobacterota, and Thermoplasmatota. pH and SO4 2- gradients were explained by phase separation of sulfur-rich hydrothermal fluids and oxidation of reduced sulfur in the steam phase, which were identified as key processes shaping these communities. Ordination and permutational analysis of variance showed that temperature, pH, and major element hydrochemistry explain only 24% of the microbial community structure. Therefore, most of the variance remained unexplained, suggesting that other environmental or biotic factors are also involved and highlighting the environmental complexity of the ecosystem and its great potential to test niche theory ecological associated questions. IMPORTANCE This is the first approach to investigate whether geothermal processes could have an influence on the ecology of thermal microbial communities on a global scale. In addition to temperature and pH, microbial communities are structured by sulfate concentrations, which depends on the tectono-magmatic settings (such as the depth of magmatic chambers) and the local settings (such as the availability of a confining layer separating NaCl waters from steam after phase separation) and the possibility of mixing with more diluted fluids. Comparison of microbial communities from different geothermal areas by homogeneous sequence processing showed that no significant geographic distance decay was detected on the microbial communities according to Bray-Curtis, Jaccard, unweighted, and weighted Unifrac similarity/dissimilarity indices. Instead, an ancient potential divergence in the same taxonomic groups is suggested between globally distant thermal zones.

The Analysis of Microbial Community Characteristics Revealed that the Pathogens of Leaf Spot of Rosa roxburghii Originated from the Phyllosphere.

Members of the plant mycobiota are all associated to varying degrees with the development of plant diseases. Although many reports on the plant mycobiota are well documented, the relationships between mycobiota of Rosa roxburghii and plant diseases are poorly understood. Mutual interactions and extent of the roles of microbial communities associated with R. roxburghii and the source of pathogens are still unclear, and more research is needed on the health benefits of this ecologically important population. Using high-throughput sequencing, we analyzed the mycobiota composition and ecological guilds of the rhizosphere, root, and phyllosphere of healthy and diseased R. roxburghii from the Tianfu R. roxburghii Industrial Park in Panzhou city, Guizhou province. Analysis of community composition showed that the relative abundance of pathogens of leaf spot, including Alternaria, Pestalotiopsis and Neofusicoccum in the phyllosphere of diseased plant (LD), were 1.15%, 0.15% and 0.06%, and the relative abundance of Alternaria and Pestalotiopsis were 0.96% and 0.58% in healthy plant (LH). The alpha diversity indices indicated that fungal diversity was higher in healthy plants compared to diseased plants in each compartment. The alpha diversity index of fungi in the phyllosphere (LH) of healthy R. roxburghii, including Shannon, Chao-1, and Faith-pd indices, was 1.02, 81.50 and 10.42 higher than that of the diseased (LD), respectively. The fungi in the rhizosphere of healthy was 1.03, 59.00 and 5.56 higher than the diseased, respectively. The Shannon index of fungi in the root of healthy was 0.29 higher than that of diseased. Principal Coordinate analysis and ANOSIM results showed that there were significant differences in mycobiota composition between healthy and diseased phyllospheres (P < 0.05), as well as rhizosphere fungal community, while there was no significant difference between healthy and diseased roots (P > 0.05). Linear discriminant analysis effect size revealed that, at different taxonomic levels, there were significantly different taxa between the healthy and diseased plants in each compartment. The ecological guilds differed between healthy and diseased plants according to the FUNGuild analysis. For example, of healthy compared to diseased plants, the percentages of "lichenized-undefined saprotroph" were increased by 2.34%, 0.44%, and 1.54% in the phyllosphere, root, and rhizosphere, respectively. In addition, the plant pathogens existed in each compartment of R. roxburghii, but the percentages of "plant pathogen" were increased by 1.16% in the phyllosphere of diseased compared to healthy plants. Together, the ecological guild and co-occurrence network indicated that the potential pathogens of leaf spot were mainly found in the phyllosphere. This study explained one of pathogen origin of leaf spots of R. roxburghii by the microbial community ecology, which will provide the new insights for identification of plant pathogens.

Predictive Ecology and Management of Phyllosphere Microbial Communities Through Cross-Scale Synthesis

In this article, we summarize the main takeaways from a symposium and hybrid virtual and in-person participatory discussion focused on the challenges of scale in understanding the ecology and management of phyllosphere microbial communities. We provide an overview of the confounding effects of spatial scale on inference in microbial ecology, the spatial organization of microbial interactions in the phyllosphere, advances and remaining gaps in measuring phyllosphere colonization across scales, and the epidemiology in the phyllosphere. We hope to motivate further discussion and the development and adoption of creative approaches to solving the challenges of scale to enhance fundamental understanding and practical management of the phyllosphere microbiomes.

Environmental heterogeneity and dispersal limitation simultaneously determine the spatial scaling of different microbial functional groups

Uncovering the mechanisms driving patterns of diversity across space and through time is of critical importance in microbial community ecology. Previous studies suggest that microorganisms also follow the same spatial scaling patterns as macro-organisms. However, it remains unclear whether different microbial functional groups differ in spatial scaling and how different ecological processes may contribute to such differences. In this study, two typical spatial scaling patterns, taxa-area (TAR) and distance-decay relationships (DDR), were investigated for the whole prokaryotic community and seven microbial functional groups using marker genes, including amoA (AOA), amoA (AOB), aprA, dsrB, mcrA, nifH and nirS. Different microbial functional groups harbored different spatial scaling patterns. Microbial functional groups had weaker TAR slope coefficients than the whole prokaryotic community. The archaeal ammonia-oxidizing group, however, displayed a stronger DDR pattern than the bacterial ammonia-oxidizing group. For both TAR and DDR, rare subcommunities were mainly responsible for the observed microbial spatial scaling patterns. Significant associations between environmental heterogeneity and spatial scaling metrics were observed for multiple microbial functional groups. Dispersal limitation, which positively correlated with phylogenetic breadth, was also strongly associated with the strength of microbial spatial scaling. The results demonstrated that environmental heterogeneity and dispersal limitation simultaneously contributed to microbial spatial scaling patterns. This study links microbial spatial scaling patterns with ecological processes, providing mechanistic insights into the typical diversity patterns followed by microbes.

Exploring the Bacterial Community in Aged Fecal Sources from Dairy Cows: Impacts on Fecal Source Tracking.

(1) Background: This paper discusses the impact of agricultural activities on stream health, particularly in relation to dairy cow fecal pollution. The study explores the fecal microbiome of cattle and the potential ecological implications of aging fecal pollution on waterways. (2) Methods: The study examines changes in the bacterial community available for mobilization from in-situ decomposing cowpats and the effects of simulated rainfall. The microbiome of individual cowpats was monitored over 5.5 months. We used 16S rRNA metagenomics and machine learning software, FEAST (Fast Expectation-mAximization for microbial Source Tracking), for bacterial and fecal source assignments. (3) Results: The phyla Bacillota and Bacteroidota are dominant in the fecal microbiota of fresh cow feces but shift to Pseudomonodota, Actinomycetota, and environmental Bacteroidota in aged cowpats. Potential impacts of these bacterial community shifts on inputs to local agricultural streams are discussed in relation to water quality monitoring and aging sources of fecal contamination. We identified taxon orders that are potential indicators of fresh cattle sources (Oscillospirales and Bacteroidales) and aged sources (Peptostreptococcales-Tissierellales) in water bodies. (4) The paper highlights that bacterial metagenomic profiling can inform our understanding of the ecology of microbial communities in aquatic environments and the potential impacts of agricultural activities on ecosystem health.

Urban land uses shape soil microbial abundance and diversity

Soil microbial biodiversity provides many useful services in cities. However, the ecology of microbial communities in urban soils remains poorly documented, and studies are required to better predict the impact of urban land use. We characterized microbial communities (archea/bacteria and fungi) in urban soils in Dijon (Burgundy, France). Three main land uses were considered – public leisure, traffic, and urban agriculture – sub-categorized in sub-land uses according to urban indexes and management practices. Microbial biomass and diversity were determined by quantifying and high-throughput sequencing of soil DNA. Variation partitioning analysis was used to rank soil physicochemical characteristics and land uses according to their relative contribution to the variation of soil microbial communities. Urban soils in Dijon harbored high levels of microbial biomass and diversity that varied according to land uses. Microbial biomass was 1.8 times higher in public leisure and traffic sites than in urban agriculture sites. Fungal richness increased by 25 % in urban agriculture soils, and bacterial richness was lower (by 20 %) in public leisure soils. Partitioning models explained 25.7 %, 46.2 % and 75.6 % of the variance of fungal richness, bacterial richness and microbial biomass, respectively. The organic carbon content and the C/N ratio were the best predictors of microbial biomass, whereas soil bacterial diversity was mainly explained by soil texture and land use. Neither metal trace elements nor polycyclic aromatic hydrocarbons contents explained variations of microbial communities, probably due to their very low concentration in the soils. The microbial composition results highlighted that leisure sites represented a stabilized habitat favoring specialized microbial groups and microbial plant symbionts, as opposed to urban agriculture sites that stimulated opportunistic populations able to face the impact of agricultural practices. Altogether, our results provide evidence that there is scope for urban planners to drive soil microbial diversity through sustainable urban land use and associated management practices.

The Robogut: A Bioreactor Model of the Human Colon for Evaluation of Gut Microbial Community Ecology and Function

The human colon is inhabited by a complex community of microbes. These microbes are integral to host health and physiology. Understanding how and when the microbiome causally influences host health will require microbiome models that can be tightly controlled and manipulated. While in vivo models are unrivalled in their ability to study host-microbial interplay, in vitro models are gaining in popularity as methods to study the ecology and function of the gut microbiota, and benefit from tight controllability and reproducibility, as well as reduced ethical constraints. In this set of protocols, we describe the Robogut, a single-stage bioreactor system designed to replicate the conditions of the distal human colon, to culture whole microbial communities derived from stool and/or colonic biopsy samples, with consideration of methods to create culture medium formulations and to build, run, and sample the bioreactor apparatus. Cleaning and maintenance of the bioreactor system are also described. © 2023 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Growth medium preparation Support Protocol 1: Preparing medium supplements Basic Protocol 2: Preparing the bioreactor vessels Support Protocol 2: Making acid and base bottles Support Protocol 3: Preparing the effluent bottles Support Protocol 4: Making acid solution Support Protocol 5: Making base solution Basic Protocol 3: Preparing inoculum and inoculating bioreactors Alternate Protocol 1: Preparing inoculum less than 0.5% (w/v) of vessel volume Alternate Protocol 2: Preparing synthetic community aliquots and inoculation via the septum Alternate Protocol 3: Preparing inoculum from a tissue sample Basic Protocol 4: Sampling the bioreactor vessel Basic Protocol 5: Harvesting bioreactor vessel contents at end of experiment Support Protocol 6: Cleaning and sterilizing sampling needles Basic Protocol 6: Cleaning the bioreactor vessel Support Protocol 7: Cleaning bioreactor support bottles.

Spatial profiling of microbial communities by sequential FISH with error-robust encoding

Spatial analysis of microbiomes at single cell resolution with high multiplexity and accuracy has remained challenging. Here we present spatial profiling of a microbiome using sequential error-robust fluorescence in situ hybridization (SEER-FISH), a highly multiplexed and accurate imaging method that allows mapping of microbial communities at micron-scale. We show that multiplexity of RNA profiling in microbiomes can be increased significantly by sequential rounds of probe hybridization and dissociation. Combined with error-correction strategies, we demonstrate that SEER-FISH enables accurate taxonomic identification in complex microbial communities. Using microbial communities composed of diverse bacterial taxa isolated from plant rhizospheres, we apply SEER-FISH to quantify the abundance of each taxon and map microbial biogeography on roots. At micron-scale, we identify clustering of microbial cells from multiple species on the rhizoplane. Under treatment of plant metabolites, we find spatial re-organization of microbial colonization along the root and alterations in spatial association among microbial taxa. Taken together, SEER-FISH provides a useful method for profiling the spatial ecology of complex microbial communities in situ.

Determinants of synergistic cell-cell interactions in bacteria.

Bacteria are ubiquitous and colonize virtually every conceivable habitat on earth. To achieve this, bacteria require different metabolites and biochemical capabilities. Rather than trying to produce all of the needed materials by themselves, bacteria have evolved a range of synergistic interactions, in which they exchange different commodities with other members of their local community. While it is widely acknowledged that synergistic interactions are keytothe ecology of both individual bacteria and entire microbial communities, the factors determining their establishment remain poorly understood. Here we provide acomprehensive overview over our current knowledge onthedeterminants of positive cell-cell interactions amongbacteria. Taking a holistic approach, we review the literature on the molecular mechanisms bacteria use to transfer commodities between bacterial cells and discuss towhichextent these mechanisms favour or constrain the successful establishment of synergistic cell-cell interactions. In addition, we analyse how these different processes affect the specificity among interaction partners. By drawing together evidence from different disciplines that study the focal question on different levels of organisation, this work not only summarizes the state of the art in this exciting field of research, but also identifies new avenues for future research.

Ecological successions throughout the desiccation of Tirez lagoon (Spain) as an astrobiological time-analog for wet-to-dry transitions on Mars

Tirez was a small and seasonal endorheic athalassohaline lagoon that was located in central Spain. In recent years, the lagoon has totally dried out, offering for the first time the opportunity to analyze its desiccation process as a “time-analog” to similar events occurred in paleolakes with varying salinity during the wet-to-dry transition on early Mars. On the martian cratered highlands, an early period of water ponding within enclosed basins evolved to a complete desiccation of the lakes, leading to deposition of evaporitic sequences during the Noachian and into the Late Hesperian. As Tirez also underwent a process of desiccation, here we describe (i) the microbial ecology of Tirez when the lagoon was still active 20 years ago, with prokaryotes adapted to extreme saline conditions; (ii) the composition of the microbial community in the dried lake sediments today, in many case groups that thrive in sediments of extreme environments; and (iii) the molecular and isotopic analysis of the lipid biomarkers that can be recovered from the sediments today. We discuss the implications of these results to better understanding the ecology of possible Martian microbial communities during the wet-to-dry transition at the end of the Hesperian, and how they may inform about research strategies to search for possible biomarkers in Mars after all the water was lost.

Spatial self-organization of metabolism in microbial systems: A matter of enzymes and chemicals.

Most bacteria live in dense, spatially structured communities such as biofilms. The high density allows cells to alter the local microenvironment, whereas the limited mobility can cause species to become spatially organized. Together, these factors can spatially organize metabolic processes within microbial communities so that cells in different locations perform different metabolic reactions. The overall metabolic activity of a community depends both on how metabolic reactions are arranged in space and on how they are coupled, i.e., how cells in different regions exchange metabolites. Here, we review mechanisms that lead to the spatial organization of metabolic processes in microbial systems. We discuss factors that determine the length scales over which metabolic activities are arranged in space and highlight how the spatial organization of metabolic processes affects the ecology and evolution of microbial communities. Finally, we define key open questions that we believe should be the main focus of future research.

Pitfalls of genotyping microbial communities with rapidly growing genome collections.

Detecting genetic variants in metagenomic data is a priority for understanding the evolution, ecology, and functional characteristics of microbial communities. Many tools that perform this metagenotyping rely on aligning reads of unknown origin to a database of sequences from many species before calling variants. In this synthesis, we investigate how databases of increasingly diverse and closely related species have pushed the limits of current alignment algorithms, thereby degrading the performance of metagenotyping tools. We identify multi-mapping reads as a prevalent source of errors and illustrate a trade-off between retaining correct alignments versus limiting incorrect alignments, many of which map reads to the wrong species. Then we evaluate several actionable mitigation strategies and review emerging methods showing promise to further improve metagenotyping in response to the rapid growth in genome collections. Our results have implications beyond metagenotyping to the many tools in microbial genomics that depend upon accurate read mapping.

Sediment Microbiota as a Proxy of Environmental Health: Discovering Inter- and Intrakingdom Dynamics along the Eastern Mediterranean Continental Shelf.

Sedimentary marine habitats are the largest ecosystem on our planet in terms of area. Marine sediment microbiota govern most of the benthic biological processes and therefore are responsible for much of the global biogeochemical activity. Sediment microbiota respond, even rapidly, to natural change in environmental conditions as well as disturbances of anthropogenic sources. The latter greatly impact the continental shelf. Characterization and monitoring of the sediment microbiota may serve as an important tool for assessing environmental health and indicate changes in the marine ecosystem. This study examined the suitability of marine sediment microbiota as a bioindicator for environmental health in the eastern Mediterranean Sea. Integration of information from Bacteria, Archaea, and Eukaryota enabled robust assessment of environmental factors controlling sediment microbiota composition: seafloor-depth (here representing sediment grain size and total organic carbon), core depth, and season (11%, 4.2%, and 2.5% of the variance, respectively). Furthermore, inter- and intrakingdom cooccurrence patterns indicate that ecological filtration as well as stochastic processes may control sediment microbiota assembly. The results show that the sediment microbiota was robust over 3 years of sampling, in terms of both representation of region (outside the model sites) and robustness of microbial markers. Furthermore, anthropogenic disturbance was reflected by significant transformations in sediment microbiota. We therefore propose sediment microbiota analysis as a sensitive approach to detect disturbances, which is applicable for long-term monitoring of marine environmental health. IMPORTANCE Analysis of data, curated over 3 years of sediment sampling, improves our understanding of microbiota assembly in marine sediment. Furthermore, we demonstrate the importance of cross-kingdom integration of information in the study of microbial community ecology. Finally, the urgent need to propose an applicable approach for environmental health monitoring is addressed here by establishment of sediment microbiota as a robust and sensitive model.

Genotyping Microbial Communities with MIDAS2: From Metagenomic Reads to Allele Tables.

The Metagenomic Intra-Species Diversity Analysis System 2 (MIDAS2) is a scalable pipeline that identifies single nucleotide variants and gene copy number variants in metagenomes using comprehensive reference databases built from public microbial genome collections (metagenotyping). MIDAS2 is the first metagenotyping tool with functionality to control metagenomic read mapping filters and to customize the reference database to the microbial community, features that improve the precision and recall of detected variants. In this article we present four basic protocols for the most common use cases of MIDAS2, along with supporting protocols for installation and use. In addition, we provide in-depth guidance on adjusting command line parameters, editing the reference database, optimizing hardware utilization, and understanding the metagenotyping results. All the steps of metagenotyping, from raw sequencing reads to population genetic analysis, are demonstrated with example data in two downloadable sequencing libraries of single-end metagenomic reads representing a mixture of multiple bacterial species. This set of protocols empowers users to accurately genotype hundreds of species in thousands of samples, providing rich genetic data for studying the evolution and strain-level ecology of microbial communities. © 2022 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Species prescreening Basic Protocol 2: Download MIDAS reference database Basic Protocol 3: Population single nucleotide variant calling Basic Protocol 4: Pan-genome copy number variant calling Support Protocol 1: Installing MIDAS2 Support Protocol 2: Command line inputs Support Protocol 3: Metagenotyping with a custom collection of genomes Support Protocol 4: Metagenotyping with advanced parameters.