Optimised biomolecular extraction for metagenomic analysis of microbial biofilms from high-mountain streams

Busi, Susheel Bhanu; Pramateftaki, Paraskevi; Brandani, Jade; Fodelianakis, Stilianos; Peter, Hannes; Halder, Rashi; Wilmes, Paul; Battin, Tom J.

doi:10.7717/peerj.9973

Cited by 20 publications

(16 citation statements)

References 73 publications

(108 reference statements)

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“…A previously established protocol 85 was used to extract DNA from all samples. Briefly, 5 g of epipsammic and 0.05–0.1 g of epilithic biofilm were subjected to a phenol:chloroform-based extraction and purification method.…”

Section: Methodsmentioning

confidence: 99%

Genomic and metabolic adaptations of biofilms to ecological windows of opportunity in glacier-fed streams

et al. 2022

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In glacier-fed streams, ecological windows of opportunity allow complex microbial biofilms to develop and transiently form the basis of the food web, thereby controlling key ecosystem processes. Using metagenome-assembled genomes, we unravel strategies that allow biofilms to seize this opportunity in an ecosystem otherwise characterized by harsh environmental conditions. We observe a diverse microbiome spanning the entire tree of life including a rich virome. Various co-existing energy acquisition pathways point to diverse niches and the exploitation of available resources, likely fostering the establishment of complex biofilms during windows of opportunity. The wide occurrence of rhodopsins, besides chlorophyll, highlights the role of solar energy capture in these biofilms while internal carbon and nutrient cycling between photoautotrophs and heterotrophs may help overcome constraints imposed by oligotrophy in these habitats. Mechanisms potentially protecting bacteria against low temperatures and high UV-radiation are also revealed and the selective pressure of this environment is further highlighted by a phylogenomic analysis differentiating important components of the glacier-fed stream microbiome from other ecosystems. Our findings reveal key genomic underpinnings of adaptive traits contributing to the success of complex biofilms to exploit environmental opportunities in glacier-fed streams, which are now rapidly changing owing to global warming.

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Section: Methodsmentioning

confidence: 99%

Genomic and metabolic adaptations of biofilms to ecological windows of opportunity in glacier-fed streams

et al. 2022

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“…We extracted DNA from sediment samples using a phenol-chloroform method with certain modifications to address the nature of our samples [ 28 ]. We amplified the V3-V4 hypervariable regions of the bacterial 16 S rRNA gene using primers 341 f (5′-CCTACGGGNGGCWGCAG-3′) and 785r (5′-GACTACHVGGGTATCTAATCC-3′) that are general for amplifying Bacteria [ 29 ].…”

Section: Methodsmentioning

confidence: 99%

Microdiversity characterizes prevalent phylogenetic clades in the glacier-fed stream microbiome

et al. 2021

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Glacier-fed streams (GFSs) are extreme and rapidly vanishing ecosystems, and yet they harbor diverse microbial communities. Although our understanding of the GFS microbiome has recently increased, we do not know which microbial clades are ecologically successful in these ecosystems, nor do we understand potentially underlying mechanisms. Ecologically successful clades should be more prevalent across GFSs compared to other clades, which should be reflected as clade-wise distinctly low phylogenetic turnover. However, methods to assess such patterns are currently missing. Here we developed and applied a novel analytical framework, “phyloscore analysis”, to identify clades with lower spatial phylogenetic turnover than other clades in the sediment microbiome across twenty GFSs in New Zealand. These clades constituted up to 44% and 64% of community α-diversity and abundance, respectively. Furthermore, both their α-diversity and abundance increased as sediment chlorophyll a decreased, corroborating their ecological success in GFS habitats largely devoid of primary production. These clades also contained elevated levels of putative microdiversity than others, which could potentially explain their high prevalence in GFSs. This hitherto unknown microdiversity may be threatened as glaciers shrink, urging towards further genomic and functional exploration of the GFS microbiome.

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“…Glacier-fed stream sediment (GFS) [ 63 ] with 270,341 sequences (phenol-chloroform extraction batch No. 37);…”

Section: Discussionmentioning

confidence: 99%

Mantis: flexible and consensus-driven genome annotation

et al. 2021

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Background The rapid development of the (meta-)omics fields has produced an unprecedented amount of high-resolution and high-fidelity data. Through the use of these datasets we can infer the role of previously functionally unannotated proteins from single organisms and consortia. In this context, protein function annotation can be described as the identification of regions of interest (i.e., domains) in protein sequences and the assignment of biological functions. Despite the existence of numerous tools, challenges remain in terms of speed, flexibility, and reproducibility. In the big data era, it is also increasingly important to cease limiting our findings to a single reference, coalescing knowledge from different data sources, and thus overcoming some limitations in overly relying on computationally generated data from single sources. Results We implemented a protein annotation tool, Mantis, which uses database identifiers intersection and text mining to integrate knowledge from multiple reference data sources into a single consensus-driven output. Mantis is flexible, allowing for the customization of reference data and execution parameters, and is reproducible across different research goals and user environments. We implemented a depth-first search algorithm for domain-specific annotation, which significantly improved annotation performance compared to sequence-wide annotation. The parallelized implementation of Mantis results in short runtimes while also outputting high coverage and high-quality protein function annotations. Conclusions Mantis is a protein function annotation tool that produces high-quality consensus-driven protein annotations. It is easy to set up, customize, and use, scaling from single genomes to large metagenomes. Mantis is available under the MIT license at https://github.com/PedroMTQ/mantis.

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Optimised biomolecular extraction for metagenomic analysis of microbial biofilms from high-mountain streams

Cited by 20 publications

References 73 publications

Genomic and metabolic adaptations of biofilms to ecological windows of opportunity in glacier-fed streams

Genomic and metabolic adaptations of biofilms to ecological windows of opportunity in glacier-fed streams

Microdiversity characterizes prevalent phylogenetic clades in the glacier-fed stream microbiome

Mantis: flexible and consensus-driven genome annotation

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