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      Longitudinal metabarcode analysis of karst bacterioplankton microbiomes provide evidence of epikarst to cave transport and community succession

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          Abstract

          Caves are often assumed to be static environments separated from weather changes experienced on the surface. The high humidity and stability of these subterranean environments make them attractive to many different organisms including microbes such as bacteria and protists. Cave waters generally originate from the surface, may be filtered by overlying soils, can accumulate in interstitial epikarst zones underground, and emerge in caves as streams, pools and droplets on speleothems. Water movement is the primary architect of karst caves, and depending on the hydrologic connectivity between surface and subsurface, is the most likely medium for the introduction of microbes to caves. Recently published metabarcoding surveys of karst cave soils and speleothems have suggested that the vast majority of bacteria residing in these habitats do not occur on the surface, calling into question the role of microbial transport by surface waters. The purpose of this study was to use metabarcoding to monitor the aquatic prokaryotic microbiome of a cave for 1 year, conduct longitudinal analyses of the cave’s aquatic bacterioplankton, and compare it to nearby surface water. Water samples were collected from two locations inside Panel Cave in Natural Tunnel State Park in Duffield, VA and two locations outside of the cave. Of the two cave locations, one was fed by groundwater and drip water and the other by infiltrating surface water. A total of 1,854 distinct prokaryotic ASVs were detected from cave samples and 245 (13.1%) were not found in surface samples. PCo analysis demonstrated a marginal delineation between two cave sample sites and between cave and surface microbiomes suggesting the aquatic bacterioplankton in a karst cave is much more similar to surface microbes than reported from speleothems and soils. Most surprisingly, there was a cave microbe population and diversity bloom in the fall months whereas biodiversity remained relatively steady on the surface. The cave microbiome was more similar to the surface before the bloom than during and afterwards. This event demonstrates that large influxes of bacteria and particulate organic matter can enter the cave from either the surface or interstitial zones and the divergence of the cave microbiome from the surface demonstrates movement of microbes from the epikarst zones into the cave.

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          DADA2: High resolution sample inference from Illumina amplicon data

          We present DADA2, a software package that models and corrects Illumina-sequenced amplicon errors. DADA2 infers sample sequences exactly, without coarse-graining into OTUs, and resolves differences of as little as one nucleotide. In several mock communities DADA2 identified more real variants and output fewer spurious sequences than other methods. We applied DADA2 to vaginal samples from a cohort of pregnant women, revealing a diversity of previously undetected Lactobacillus crispatus variants.
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            Reproducible, interactive, scalable and extensible microbiome data science using QIIME 2

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              Metagenomics: application of genomics to uncultured microorganisms.

               Jo Handelsman (2004)
              Metagenomics (also referred to as environmental and community genomics) is the genomic analysis of microorganisms by direct extraction and cloning of DNA from an assemblage of microorganisms. The development of metagenomics stemmed from the ineluctable evidence that as-yet-uncultured microorganisms represent the vast majority of organisms in most environments on earth. This evidence was derived from analyses of 16S rRNA gene sequences amplified directly from the environment, an approach that avoided the bias imposed by culturing and led to the discovery of vast new lineages of microbial life. Although the portrait of the microbial world was revolutionized by analysis of 16S rRNA genes, such studies yielded only a phylogenetic description of community membership, providing little insight into the genetics, physiology, and biochemistry of the members. Metagenomics provides a second tier of technical innovation that facilitates study of the physiology and ecology of environmental microorganisms. Novel genes and gene products discovered through metagenomics include the first bacteriorhodopsin of bacterial origin; novel small molecules with antimicrobial activity; and new members of families of known proteins, such as an Na(+)(Li(+))/H(+) antiporter, RecA, DNA polymerase, and antibiotic resistance determinants. Reassembly of multiple genomes has provided insight into energy and nutrient cycling within the community, genome structure, gene function, population genetics and microheterogeneity, and lateral gene transfer among members of an uncultured community. The application of metagenomic sequence information will facilitate the design of better culturing strategies to link genomic analysis with pure culture studies.
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                Author and article information

                Contributors
                Journal
                PeerJ
                PeerJ
                PeerJ
                PeerJ
                PeerJ
                PeerJ Inc. (San Diego, USA )
                2167-8359
                8 March 2021
                2021
                : 9
                Affiliations
                University of Virginia’s College at Wise , Wise, VA, USA
                Article
                10757
                10.7717/peerj.10757
                7950216
                © 2021 Morse et al.

                This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, reproduction and adaptation in any medium and for any purpose provided that it is properly attributed. For attribution, the original author(s), title, publication source (PeerJ) and either DOI or URL of the article must be cited.

                Funding
                Funded by: John C. Buchanan Chair in Biology endowment
                Funded by: UVa-Wise Fellowship in Natural Sciences
                Funded by: UVa-Wise’s Chancellor Donna Henry and the UVa-Wise Foundation
                This project was funded by the John C. Buchanan Chair in Biology endowment. Kendall V. Morse and Dylan R. Richardson were supported by the UVa-Wise Fellowship in Natural Sciences. Meteorology equipment was purchased with funds provided by UVa-Wise’s Chancellor Donna Henry and the UVa-Wise foundation. There was no additional external funding received for this study. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
                Categories
                Biodiversity
                Ecosystem Science
                Microbiology
                Freshwater Biology
                Ecohydrology

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