Microbial community of the deep-sea brine Lake Kryos seawater-brine interface is active below the chaotropicity limit of life as revealed by recovery of mRNA Yakimov, M. M., La Cono, V., Spada, G. L., Bortoluzzi, G., Messina, E., Smedile, F., ... Giuliano, L. (2015). Microbial community of the deep-sea brine Lake Kryos seawater-brine interface is active below the chaotropicity limit of life as revealed by recovery of mRNA. Environmental Microbiology, 17 (2) This is the accepted version of the following article: Microbial community of the deep-sea brine Lake Kryos seawater-brine interface is active below the chaotropicity limit of life as revealed by recovery of mRNA, Michail M. Yakimov, Violetta La Cono, Gina La Spada, Giovanni Bortoluzzi, Enzo Messina, Francesco Smedile, Erika Arcadi, Mireno Borghini, Manuel Ferrer, Phillippe Schmitt-Kopplin, Norbert Hertkorn, Jonathan A. Cray, John E. Hallsworth, Peter N. Golyshin andLaura Giuliano, which has been published in final form at ttp://onlinelibrary.wiley.com/doi/10.1111/1462-2920.12587/abstract. General rightsCopyright for the publications made accessible via the Queen's University Belfast Research Portal is retained by the author(s) and / or other copyright owners and it is a condition of accessing these publications that users recognise and abide by the legal requirements associated with these rights.Take down policy The Research Portal is Queen's institutional repository that provides access to Queen's research output. Every effort has been made to ensure that content in the Research Portal does not infringe any person's rights, or applicable UK laws. If you discover content in the Research Portal that you believe breaches copyright or violates any law, please contact openaccess@qub.ac.uk. 25This article has been accepted for publication and undergone full peer review but has not been through the copyediting, typesetting, pagination and proofreading process, which may lead to differences between this version and the Version of Record. Please cite this article as doi: 10.1111/1462-2920.12587 Accepted ArticleThis article is protected by copyright. All rights reserved. the microbiology of the seawater-Kryos brine interface and managed to recover mRNA from the 10 2.27-3.03 M MgCl 2 layer (equivalent to 0.747-0.631 water-activity) thereby expanding the established 11 chaotropicity window-for-life. The primary bacterial taxa present there were KB1 candidate division 12 and DHAL-specific group of organisms, distantly related to Desulfohalobium. Two euryarchaeal 13 candidate divisions MSBL1 and HC1, detected in minority in the overlaying layers, accounted for 14 more than 85% of the rRNA-containing archaeal clones analyzed in 2.27-3.03 M MgCl 2 layer. These 15 findings shed light on the plausibility of life in highly chaotropic environments, geochemical 16 windows for microbial extremophiles, and have implications for habitability elsewhere in the Solar 17 System.

Nano-sized archaeota, with their small genomes and limited metabolic capabilities, are known to associate with other microbes, thereby compensating for their own auxotrophies. These diminutive and yet ubiquitous organisms thrive in hypersaline habitats that they share with haloarchaea. Here, we reveal the genetic and physiological nature of a nanohaloarchaeon–haloarchaeon association, with both microbes obtained from a solar saltern and reproducibly cultivated together in vitro. The nanohaloarchaeon Candidatus Nanohalobium constans LC1Nh is an aerotolerant, sugar-fermenting anaerobe, lacking key anabolic machinery and respiratory complexes. The nanohaloarchaeon cells are found physically connected to the chitinolytic haloarchaeon Halomicrobium sp. LC1Hm. Our experiments revealed that this haloarchaeon can hydrolyze chitin outside the cell (to produce the monosaccharide N-acetylglucosamine), using this beta-glucan to obtain carbon and energy for growth. However, LC1Hm could not metabolize either glycogen or starch (both alpha-glucans) or other polysaccharides tested. Remarkably, the nanohaloarchaeon’s ability to hydrolyze glycogen and starch to glucose enabled growth of Halomicrobium sp. LC1Hm in the absence of a chitin. These findings indicated that the nanohaloarchaeon–haloarchaeon association is both mutualistic and symbiotic; in this case, each microbe relies on its partner’s ability to degrade different polysaccharides. This suggests, in turn, that other nano-sized archaeota may also be beneficial for their hosts. Given that availability of carbon substrates can vary both spatially and temporarily, the susceptibility of Halomicrobium to colonization by Ca. Nanohalobium can be interpreted as a strategy to maximize the long-term fitness of the host.

Deep-sea hypersaline anoxic lakes (DHALs) of the Eastern Mediterranean represent some of the most hostile environments on our planet. We investigated microbial life in the recently discovered Lake Medee, the largest DHAL found to-date. Medee has two unique features: a complex geobiochemical stratification and an absence of chemolithoautotrophic Epsilonproteobacteria, which usually play the primary role in dark bicarbonate assimilation in DHALs interfaces. Presumably because of these features, Medee is less productive and exhibits reduced diversity of autochthonous prokaryotes in its interior. Indeed, the brine community almost exclusively consists of the members of euryarchaeal MSBL1 and bacterial KB1 candidate divisions. Our experiments utilizing cultivation and [14C]-assimilation, showed that these organisms at least partially rely on reductive cleavage of osmoprotectant glycine betaine and are engaged in trophic cooperation. These findings provide novel insights into how prokaryotic communities can adapt to salt-saturated conditions and sustain active metabolism at the thermodynamic edge of life.

Hydrated, magnesium-rich minerals and subglacial brines exist on the martian surface, so the habitability of high-Mg2+ environments on Earth has extraterrestrial (as well as terrestrial) implications. Here, we report the discovery of a MgCl2-dominated (4.72 M) brine lake on the floor of the Mediterranean Ridge that underlies a 3500-m water column, and name it Lake Hephaestus. Stable isotope analyses indicated that the Hephaestus brine is derived from interactions between ancient bishofite-enriched evaporites and subsurface fluids. Analyses of sediment pore waters indicated that the Hephaestus depression had contained the MgCl2 brine for a remarkably short period; only 700 years. Lake Hephaestus is, therefore, the youngest among currently known submarine athalassohaline brine lakes on Earth. Due to its biologically hostile properties (low water-activity and extreme chaotropicity), the Hephaestus brine is devoid of life. By contrast, the seawater-Hephaestus brine interface has been shown to act as refuge for extremely halophilic and magnesium-adapted stratified communities of microbes, even at MgCl2 concentrations that approach the water-activity limit for life (0.653).

We used a combination of molecular and microbiological approaches to determine the activity, abundance and diversity of archaeal populations inhabiting meromictic saline Lake Faro (Messina, Italy). Analysis of archaeal 16S rRNA, amoA, accA and hbd genes and transcripts revealed that sub- and anoxic layers of Lake Faro are primarily inhabited by the organisms related to the clusters of Marine Group I.1a of Thaumarchaeota frequently recovered from oxygen-depleted marine ecosystems. These organisms dominated the metabolically active archaea down to the bottom of the lake, indicating their adaptation to recurrent changes in the levels of water column hypoxia. The upper microaerobic layer of Lake Faro redoxcline has the maximal rates of dark primary production much lower than those of other previously studied pelagic redoxclines, but comparable to the values of meso- and bathypelagic areas of Mediterranean Sea. Application of bacterial inhibitors, especially azide, significantly declined the CO2 fixation rates in the low interface and monimolimnion, whereas archaea-specific inhibitor had effect only in upper part of the redoxcline. Based on these findings, we hypothesize that dark bicarbonate fixation in suboxic zone of Lake Faro results mainly from archaeal activity which is affected by the predicted lack in oxygen in lower layers.