A Metagenomic Time-Series Approach to Assess the Ecological Stability of Microbial Mats in a Seasonally Fluctuating Environment.

Microbial mats are complex ecological assemblages that have been present in the rock record since the Precambrian and can still be found in extant marginalized environments. These structures are considered highly stable ecosystems. In this study, we evaluate the ecological stability of dome-forming microbial mats in a modern, water-level fluctuating, hypersaline pond located in the Cuatro Ciénegas Basin, Mexico. We conducted metagenomic sampling of the site from 2016 to 2019 and detected 2250 genera of Bacteria and Archaea, with only <20 belonging to the abundant taxa (>1%). The microbial community was dominated by Proteobacteria, Euryarchaeota, Bacteroidetes, Firmicutes, and Cyanobacteria, and was compositionally sensitive to disturbances, leading to high taxonomic replacement even at the phylum level, with a significant increase in Archaea from [Formula: see text]1-4% to [Formula: see text]33% throughout the 2016-2019 study period. Although a core community represented most of the microbial community (>75%), relative abundances shifted significantly between samples, as demonstrated by changes in the abundance of Coleofasciculus from 10.2% in 2017 to 0.05% in 2019. Although functional differences between seasons were subtle, co-occurrence networks suggest differential ecological interactions between the seasons, with the addition of a new module during the rainy season and the potential shift in hub taxa. Functional composition was slightly more similar between samples, but basic processes such as carbohydrate, amino acid, and nucleic acid metabolisms were widely distributed among samples. Major carbon fixation processes included sulfur oxidation, nitrogen fixation, and photosynthesis (both oxygenic and anoxygenic), as well as the Wood-Ljundgahl and Calvin cycles.

Fungal diversity in sediments of the eastern tropical Pacific oxygen minimum zone revealed by metabarcoding.

Oxygen minimum zones (OMZ) represent ~8% of the ocean, with the Pacific as the largest and top expanding area. These regions influence marine ecosystems, promoting anaerobic microbial communities. Nevertheless, only a fraction of microbial diversity has been studied, with fungi being the less explored component. So, herein we analyzed fungal diversity patterns in surface and subsurface sediments along a bathymetric transect using metabarcoding of the ITS1 region in the OMZ of the Mexican Pacific off Mazatlán. We identified 353 amplicon sequence variants (ASV), within the Ascomycota, Basidiomycota, and Rozellomycota. Spatial patterns evidenced higher alpha diversity in nearshore and subsurface subsamples, probably due to temporal fluctuations in organic matter inputs. Small-scale heterogeneity characterized the community with the majority of ASV (269 ASV) occurring in a single subsample, hinting at the influence of local biogeochemical conditions. This baseline data evidenced a remarkable fungal diversity presenting high variation along a bathymetric and vertical transects.

MicNet toolbox: Visualizing and unraveling a microbial network.

Applications of network theory to microbial ecology are an emerging and promising approach to understanding both global and local patterns in the structure and interplay of these microbial communities. In this paper, we present an open-source python toolbox which consists of two modules: on one hand, we introduce a visualization module that incorporates the use of UMAP, a dimensionality reduction technique that focuses on local patterns, and HDBSCAN, a clustering technique based on density; on the other hand, we have included a module that runs an enhanced version of the SparCC code, sustaining larger datasets than before, and we couple the resulting networks with network theory analyses to describe the resulting co-occurrence networks, including several novel analyses, such as structural balance metrics and a proposal to discover the underlying topology of a co-occurrence network. We validated the proposed toolbox on 1) a simple and well described biological network of kombucha, consisting of 48 ASVs, and 2) we validate the improvements of our new version of SparCC. Finally, we showcase the use of the MicNet toolbox on a large dataset from Archean Domes, consisting of more than 2,000 ASVs. Our toolbox is freely available as a github repository (https://github.com/Labevo/MicNetToolbox), and it is accompanied by a web dashboard (http://micnetapplb-1212130533.us-east-1.elb.amazonaws.com) that can be used in a simple and straightforward manner with relative abundance data. This easy-to-use implementation is aimed to microbial ecologists with little to no experience in programming, while the most experienced bioinformatics will also be able to manipulate the source code's functions with ease.

Recent Differentiation of Aquatic Bacterial Communities in a Hydrological System in the Cuatro Ciénegas Basin, After a Natural Perturbation.

Pozas Rojas is a hydrological system comprising nine isolated shallow ponds and a deep lagoon, which were temporally merged in 2010 by increased rainfall due to a tropical cyclone. In this work, we assess which components, biotic interactions, or environment filtering effects, drive the assembly of microbial communities after a natural perturbation. Arsenic, pH, and temperature are among the most significant environmental variables between each pond, clustering the samples in two main groups, whereas microbial composition is diverse and unique to each site, with no core at the operational taxonomic unit level and only 150 core genera when studied at the genus level. Los Hundidos lagoon has the most differentiated community, which is highly similar to the epipelagic Mediterranean Sea communities. On the other hand, the shallow ponds at the Pozas Rojas system resemble more to epicontinental hydrological systems, such as some cold rivers of the world and the phreatic mantle from Iowa. Overall, despite being a sole of water body 2 years prior to the sampling, interspecific interactions, rather than environmental selection, seem to play a more important role in Pozas Rojas, bolstered by founder effects on each poza and subsequent isolation of each water body.

Diversity of an uncommon elastic hypersaline microbial mat along a small-scale transect.

We evaluated the microbial diversity and metabolome profile of an uncommon hypersaline elastic microbial mat from Cuatro Ciénegas Basin (CCB) in the Chihuahuan Desert of Coahuila, México. We collected ten samples on a small scale transect (1.5-m) and described its microbial diversity through NGS-based ITS and 16S rDNA gene sequencing. A very low number of taxa comprised a considerable proportion of the mat and were shared across all sampling points, whereas the rare biosphere was more phylogenetically diverse (Faith's Phylogenetic Diversity (FPD) index) and phylogenetically disperse (using a null model distribution of Phylogenetic Species Clustering (nmdPSC)) than the abundant (high read count) taxa for both analyzed libraries. We also found a distinctive metabolome profile for each sample and were able to tentatively annotate several classes of compounds with relevant biological properties.

Diversity of aquatic prokaryotic communities in the Cuatro Cienegas basin.

The Cuatro Cienegas basin (Coahuila, México) is a composite of different water systems in the middle of the desert with unusually high levels of endemism and diversity in different taxa. Although the diversity of macrobiota has been well described, little is known about the diversity and distribution of microorganisms in the oligotrophic ponds. Here we describe the extent and distribution of diversity found in aquatic prokaryotic communities by analysis of terminal restriction fragment length polymorphisms (T-RFLP) of 16S rRNA genes and phylogenetic analysis of cloned genes. Twelve locations within the basin were sampled. Among all the samples, we found a total of 117 operational taxonomic units (OTUs) using T-RFLPs, which ranged in any single sample from four to 49. OTU richness and Shannon diversity indices for different sites varied, but none were particularly high. 16S rRNA gene sequence data showed 68 different phylotypes among 198 clones. The most abundant phylotypes were Gamma- and Betaproteobacteria, and extreme halophiles. The differences among sites were significant; 45 TRFs were found only once, and 37% of the total diversity was represented by differences between sites, suggesting high beta-diversity. Further studies are needed to test whether this is a direct consequence of environmental heterogeneity in the basin.

Experimental and molecular approximation to microbial niche: trophic interactions between oribatid mites and microfungi in an oligotrophic freshwater system.

Mite-fungal interactions play a key role in structuring core ecosystem processes such as nutrient dynamics. Despite their ecological relevance, these cross-kingdom interactions remain poorly understood particularly in extreme environments. Herein, we investigated feeding preferences of a novel genetic lineage of aquatic oribatids obtained from an oligotrophic freshwater system in the Cuatro Ciénegas Basin (CCB) within the Chihuahuan Desert, Mexico. During in vitro diet preference bioassays, transient aquatic microfungi (Aspergillus niger, Talaromyces sp., and Pleosporales sp.) recovered from the same mesocosm samples were offered individually and simultaneously to mites. Gut content was analyzed using classic plating and culture-independent direct PCR (focusing on the fungal barcoding region) methods. Our results indicated that oribatids fed on all tested fungal isolates, yet the profusely developing A. niger was preferentially consumed with all fungal components being digested. This feeding habit is particularly interesting since A. niger has been reported as an unsuitable dietary element for population growth, being consistently avoided by mites in previous laboratory experiments. It is possible that our mites from the CCB have adapted to exploit available resources within this oligotrophic site. This work confirms the trophic relationship between microfungi and mites, two rarely investigated major components of the microbial community, shedding light on the niche dynamics under low-nutrient conditions.

Nutrient Dependent Cross-Kingdom Interactions: Fungi and Bacteria From an Oligotrophic Desert Oasis.

Microbial interactions play a key role in ecosystem functioning, with nutrient availability as an important determinant. Although phylogenetically distant bacteria and fungi commonly co-occur in nature, information on their cross-kingdom interactions under unstable, extreme environments remains poor. Hence, the aims of this work were to evaluate potential in vitro interactions among fungi and bacteria isolated from a phosphorous oligotrophic aquatic system in the Cuatro Ciénegas Basin, Mexico, and to test the nutrients-based shifts. We assessed growth changes in bacteria (Aeromonas and Vibrio) and fungi (Coprinellus micaceus, Cladosporium sp., and Aspergillus niger) on co-cultures in relation to monocultures under diverse nutrient scenarios on Petri dishes. Interactions were explored using a network analysis, and a metabolome profiling for specific taxa. We identified nutrient-dependent patterns, as beneficial interactions dominated in low-nutrients media and antagonistic interactions dominated in rich media. This suggests that cross-kingdom synergistic interactions might favor microbial colonization and growth under low nutrient conditions, representing an adaptive trait to oligotrophic environments. Moreover, our findings agree with the stress-gradient hypothesis, since microbial interactions shifted from competition to cooperation as environmental stress (expressed as low nutrients) increased. At a functional level consistent differences were detected in the production of secondary metabolites, agreeing with plate bioassays. Our results based on culture experiments, provides evidence to understand the complexity of microbial dynamics and survival in phosphorous-depleted environments.

Microfungal oasis in an oligotrophic desert: diversity patterns and community structure in three freshwater systems of Cuatro Ciénegas, Mexico.

The Cuatro Ciénegas Basin (CCB) comprises several oligotrophic aquatic ecosystems limited by phosphorus. These aquatic systems are dominated by a high prokaryotic diversity, shaped by the stress of low nutrient supplies and interspecific competition. Although fungi constitute a diverse and important component of microbial diversity, the microfungal diversity in the CCB remains to be unveiled. With the aim to explore microfungal diversity and ecological patterns in this area, we present the first investigation analyzing cultivable taxa from sediment and water, as well as lignocellulolytic taxa obtained from incubated submerged plant debris, and wood panels in three contrasting freshwater systems in the CCB: Churince, Becerra and Pozas Rojas. We chose a culture-based approach to analyze sediment and water samples in order to obtain fungal cultures, providing opportunities for a posteriori studies, and the possibility of ex situ preservation of the diversity. We evaluated sequence data from the nuclear ribosomal internal transcribed spacer including the 5.8 rDNA region for 126 isolates, revealing 37 OTUs. These OTUs were phylogenetically affiliated to several genera in the fungal phyla: Zygomycota, Basidiomycota, and Ascomycota. We recorded two OTUs with saline affinity, agreeing with previous findings on the prokaryotic communities with ancestral marine resemblances. All the studied systems showed moderate diversity levels, however discrepancies among the diversity indexes were observed, due to the occurrence of abundant taxa in the samples. Our results indicated that lignocellulolytic microfungal communities are dominated by transient fungal taxa, as resident species were not recorded perhaps as a result of the long-term strong competition with the highly adapted prokaryotic community. Moreover, the obtained microfungal taxa occurred mostly on the resident plant debris, rather than submerged wood panels, perhaps as a result of the high adaptation to specific environmental conditions. In conclusion, the CCB possess a moderate taxonomical diversity compared to other arid environments, probably as a result of high selective pressures. Nonetheless, due to high spatial and temporal heterogeneity, the functional fungal diversity was considerable as predicted by the intermediate disturbance hypothesis. Decisively, the assessment of microfungal diversity freshwater systems is relevant, since this ecological group of microorganisms represents an important indicator of trophic complexity and biotic interactions among microbial communities, having important implications for understanding eukaryotic survival at the oligotrophic limit for life.

Aquatic bacterial assemblage structure in Pozas Azules, Cuatro Cienegas Basin, Mexico: Deterministic vs. stochastic processes.

The aim of this study was to determine the contributions of stochastic vs. deterministic processes in the distribution of microbial diversity in four ponds (Pozas Azules) within a temporally stable aquatic system in the Cuatro Cienegas Basin, State of Coahuila, Mexico. A sampling strategy for sites that were geographically delimited and had low environmental variation was applied to avoid obscuring distance effects. Aquatic bacterial diversity was characterized following a culture-independent approach (16S sequencing of clone libraries). The results showed a correlation between bacterial beta diversity (1-Sorensen) and geographic distance (distance decay of similarity), which indicated the influence of stochastic processes related to dispersion in the assembly of the ponds' bacterial communities. Our findings are the first to show the influence of dispersal limitation in the prokaryotic diversity distribution of Cuatro Cienegas Basin.

Evidence of biogeography in surface ocean bacterioplankton assemblages.

Regardless of the importance of bacterial assemblages as essential components of ecosystems, little is known about how their populations are structured. We analyzed the composition and turnover rates, based on 16S rDNA sequences, of surface water oceanic bacterial assemblages of the fraction between 0.1 and 0.8 µm along a latitudinal gradient (45°6'42''N in the North Atlantic to 15°8'37''S in the South Pacific) including geographic distance, temperature, chlorophyll a and salinity. Here we show that oceanic bacterial assemblages between 0.1 and 0.8 µm, can be structured by a variety of environmental interactions that include separation by distance and chlorophyll a concentration in temperate North Atlantic coastal samples and temperature in tropical Atlantic and Pacific coastal and open ocean samples. Bacterial phyla composition diverged between temperate and tropical regions. This study suggests that some bacterial assemblages could be structured both by environmental and spatial factors, while others by environmental factors alone.

An endangered oasis of aquatic microbial biodiversity in the Chihuahuan desert.

The Cuatro Cienegas basin in the Chihuahuan desert is a system of springs, streams, and pools. These ecosystems support >70 endemic species and abundant living stromatolites and other microbial communities, representing a desert oasis of high biodiversity. Here, we combine data from molecular microbiology and geology to document the microbial biodiversity of this unique environment. Ten water samples from locations within the Cuatro Cienegas basin and two neighboring valleys as well as three samples of wet sediments were analyzed. The phylogeny of prokaryotic populations in the samples was determined by characterizing cultured organisms and by PCR amplification and sequencing of 16S rRNA genes from total community DNA. The composition of microbial communities was also assessed by determining profiles of terminal restriction site polymorphisms of 16S rRNA genes in total community DNA. There were 250 different phylotypes among the 350 cultivated strains. Ninety-eight partial 16S rRNA gene sequences were obtained and classified. The clones represented 38 unique phylotypes from ten major lineages of Bacteria and one of Archaea. Unexpectedly, 50% of the phylotypes were most closely related to marine taxa, even though these environments have not been in contact with the ocean for tens of millions of years. Furthermore, terminal restriction site polymorphism profiles and geological data suggest that the aquatic ecosystems of Cuatro Cienegas are hydrologically interconnected with adjacent valleys recently targeted for agricultural intensification. The findings underscore the conservation value of desert aquatic ecosystems and the urgent need for study and preservation of freshwater microbial communities.