Research Collection Journal Article Distinctive archaeal composition of an artisanal crystallizer pond and functional insights into salt-saturated hypersaline environment adaptation Author(s): Plominsky, Alvaro M.; Henríquez-Castillo, Carlos; Delherbe, Nathalie; Podell, Sheila; Ramirez-Flandes, Salvador; Ugalde, Juan A.; Santibañez, Juan F.; Van den Engh, Ger; Hanselmann, Kurt; Ulloa, Osvaldo; Allen, Eric E.; Trefault, Nicole Publication Date: 2018 Permanent Link: https://doi.org/10.3929/ethz-b-000284328 Originally published in: Frontiers in Microbiology 9, http://doi.org/10.3389/fmicb.2018.01800 Rights / License: Creative Commons Attribution 4.0 International This page was generated automatically upon download from the ETH Zurich Research Collection. For more information please consult the Terms of use. ETH Library fmicb-09-01800 August 13, 2018 Time: 15:20 # 1 ORIGINAL RESEARCH published: 14 August 2018 doi: 10.3389/fmicb.2018.01800 Distinctive Archaeal Composition of an Artisanal Crystallizer Pond and Functional Insights Into Salt-Saturated Hypersaline Environment Adaptation Edited by: Jesse G. Dillon, Alvaro M. Plominsky1,2†‡, Carlos Henríquez-Castillo1,2†, Nathalie Delherbe3, California State University, Sheila Podell4, Salvador Ramirez-Flandes2,5, Juan A. Ugalde6,7, Juan F. Santibañez1, Long Beach, United States Ger van den Engh8, Kurt Hanselmann9, Osvaldo Ulloa1,2, Rodrigo De la Iglesia10, Reviewed by: Eric E. Allen4,11* and Nicole Trefault12* Rohit Ghai, Biology Centre (ASCR), Czechia 1 Department of Oceanography, Faculty of Natural and Oceanographic Sciences, Universidad de Concepción, Concepción, Purificacion Lopez-Garcia, Chile, 2 Instituto Milenio de Oceanografía, Concepción, Chile, 3 Biology Department, Cell and Molecular Biology Joint Centre National de la Recherche Doctoral Program with UC San Diego, San Diego State University, San Diego, CA, United States, 4 Marine Biology Research Scientifique (CNRS), France Division, Scripps Institution of Oceanography, University of California, San Diego, La Jolla, CA, United States, 5 Programa de 6 *Correspondence: Doctorado en Ingeniería de Sistemas Complejos, Universidad Adolfo Ibáñez, Santiago, Chile, uBiome, Inc., San Francisco, 7 Eric E. Allen CA, United States, Center for Bioinformatics and Integrative Biology, Facultad de Ciencias Biológicas, Universidad Andrés 8 9 [email protected] Bello, Santiago, Chile, Center for Marine Cytometry, Concrete, WA, United States, Department of Earth Sciences, ETH 10 Nicole Trefault Zürich, Zurich, Switzerland, Department of Molecular Genetics and Microbiology, Faculty of Biological Sciences, Pontificia 11 [email protected] Universidad Católica de Chile, Santiago, Chile, Division of Biological Sciences, University of California, San Diego, La Jolla, CA, United States, 12 GEMA Center for Genomics, Ecology and Environment, Faculty of Sciences, Universidad Mayor, †These authors have contributed Santiago, Chile equally to this work ‡Present address: Alvaro M. Plominsky, Hypersaline environments represent some of the most challenging settings for life Marine Biology Research Division, on Earth. Extremely halophilic microorganisms have been selected to colonize and Scripps Institution of Oceanography, University of California, San Diego, thrive in these extreme environments by virtue of a broad spectrum of adaptations La Jolla, CA, United States to counter high salinity and osmotic stress. Although there is substantial data on microbial taxonomic diversity in these challenging ecosystems and their primary Specialty section: This article was submitted to osmoadaptation mechanisms, less is known about how hypersaline environments Extreme Microbiology, shape the genomes of microbial inhabitants at the functional level. In this study, we a section of the journal analyzed the microbial communities in five ponds along the discontinuous salinity Frontiers in Microbiology gradient from brackish to salt-saturated environments and sequenced the metagenome Received: 29 December 2017 Accepted: 17 July 2018 of the salt (halite) precipitation pond in the artisanal Cáhuil Solar Saltern system. Published: 14 August 2018 We combined field measurements with spectrophotometric pigment analysis and Citation: flow cytometry to characterize the microbial ecology of the pond ecosystems, Plominsky AM, Henríquez-Castillo C, Delherbe N, Podell S, including primary producers and applied metagenomic sequencing for analysis of Ramirez-Flandes S, Ugalde JA, archaeal and bacterial taxonomic diversity of the salt crystallizer harvest pond. Santibañez JF, van den Engh G, Comparative metagenomic analysis of the Cáhuil salt crystallizer pond against microbial Hanselmann K, Ulloa O, De la Iglesia R, Allen EE and Trefault N communities from other salt-saturated aquatic environments revealed a dominance (2018) Distinctive Archaeal of the archaeal genus Halorubrum and showed an unexpectedly low abundance Composition of an Artisanal Crystallizer Pond and Functional of Haloquadratum in the Cáhuil system. Functional comparison of 26 hypersaline Insights Into Salt-Saturated microbial metagenomes revealed a high proportion of sequences associated with Hypersaline Environment Adaptation. nucleotide excision repair, helicases, replication and restriction-methylation systems in Front. Microbiol. 9:1800. doi: 10.3389/fmicb.2018.01800 all of them. Moreover, we found distinctive functional signatures between the microbial Frontiers in Microbiology| www.frontiersin.org 1 August 2018| Volume 9| Article 1800 fmicb-09-01800 August 13, 2018 Time: 15:20 # 2 Plominsky et al. Functional Insights Into Salt-Saturated Environments communities from salt-saturated (>30% [w/v] total salinity) compared to sub-saturated hypersaline environments mainly due to a higher representation of sequences related to replication, recombination and DNA repair in the former. The current study expands our understanding of the diversity and distribution of halophilic microbial populations inhabiting salt-saturated habitats and the functional attributes that sustain them. Keywords: hypersaline environments, solar salterns, metagenomics, microbial ecology, environmental adaptation, functional metagenomics, artisanal crystallizer pond INTRODUCTION from diverse phyla, including Proteobacteria, Firmicutes, Cyanobacteria, Bacteroidetes, Spirochaetes, and methanogenic Solar salterns provide tractable model ecosystems for Euryarchaeota (Ventosa et al., 2015). understanding microbial selection, successions, and habitat The Cáhuil Solar Saltern consists of a series of artisanal ponds resilience, and are thus one of the most studied hypersaline at the shores of the Cáhuil Lagoon that forms at the mouth environments. These are engineered pond systems for the of the Nilahue creek, in central Chile. During the summer, it commercial production of halite (NaCl) that can be found in is separated from the ocean by a sandbar that forms due to a many regions throughout the world, particularly in tropical and decrease in the water flux from the creek and by high littoral subtropical areas (Oren, 2002, 2013a; Ventosa et al., 2014). As the sediment transport from the coast. During winter, the higher water passes through a series of evaporation ponds, it generates a water flux from the creek transforms the Cáhuil Lagoon into discontinuous gradient of increasing salinities, beginning close to a seasonally stratified estuary by breaking through the sandbar that of the surrounding input water and ending at the crystallizer and connecting it with the South Pacific Ocean. This seasonal pond, where salinities reach the crystallization point of halite dynamic drastically transforms the hydrological properties of the (Oren, 2002, 2013a; Ventosa et al., 2014). Cáhuil Lagoon with regards to its surface temperature (22.1◦C The extreme osmotic stress associated with hypersaline in summer and 10.5◦C in winter) and salinity (ranging from 2.4 habitats elicits strong selective pressures for adaptations that to 2.2% [w/v] in summer and from 3.1 to 0.01% in winter, in sustain life. Cellular life in hypersaline habitats is dominated by locations nearest to the sea and the creek, respectively) (Andrade halophilic archaea and bacteria, with a few microbial eukaryotes, and Grau, 2005). such as photosynthetic microalgae, heterotrophic protists and This study describes the physico-chemical properties and fungi, and the crustacean Artemia salina (Ventosa et al., microbial communities inhabiting the Cáhuil Solar Saltern 2014). Microbial inhabitants of hypersaline environments have ponds. Comparative metagenomic analysis of the Cáhuil developed specialized adaptations to live under the high ionic microbial ecosystem against 25 other hypersaline metagenomes strength of these systems (Oren, 2002, 2013a,b). “Low-salt- collected worldwide provides insight into the abiotic-biotic in” halophilic microorganisms maintain lower intracellular salt coupling, functional convergence, and the distinctive metabolic concentrations than that of the external environment (especially adaptations that unite or distinguish the microbial communities NaC). In contrast, “high-salt-in” halophiles accumulate high that define extreme hypersaline habitats. concentrations of inorganic ions in the cytoplasm, usually KC, and Cl−. While “high-salt-in” halophiles are physiologically constrained to environments with a constant presence of high salt concentrations, “low-salt-in” halophiles can regulate MATERIALS AND METHODS the intracellular concentrations of their compatible solutes accordingly to approximate