Salt resistance genes revealed by functional metagenomics from brines and moderate-salinity rhizosphere within a hypersaline environment.

Salvador Mirete,María Lamprecht-Grandío,José E González-Pastor,Merit R Mora-Ruiz,Ramon Rosselló-Móra,Carolina G De Figueras

doi:10.3389/fmicb.2015.01121

Abstract

Hypersaline environments are considered one of the most extreme habitats on earth and microorganisms have developed diverse molecular mechanisms of adaptation to withstand these conditions. The present study was aimed at identifying novel genes from the microbial communities of a moderate-salinity rhizosphere and brine from the Es Trenc saltern (Mallorca, Spain), which could confer increased salt resistance to Escherichia coli. The microbial diversity assessed by pyrosequencing of 16S rRNA gene libraries revealed the presence of communities that are typical in such environments and the remarkable presence of three bacterial groups never revealed as major components of salt brines. Metagenomic libraries from brine and rhizosphere samples, were transferred to the osmosensitive strain E. coli MKH13, and screened for salt resistance. Eleven genes that conferred salt resistance were identified, some encoding for well-known proteins previously related to osmoadaptation such as a glycerol transporter and a proton pump, whereas others encoded proteins not previously related to this function in microorganisms such as DNA/RNA helicases, an endonuclease III (Nth) and hypothetical proteins of unknown function. Furthermore, four of the retrieved genes were cloned and expressed in Bacillus subtilis and they also conferred salt resistance to this bacterium, broadening the spectrum of bacterial species in which these genes can function. This is the first report of salt resistance genes recovered from metagenomes of a hypersaline environment.

Highlights

Life under extreme osmotic pressure in the environment represents a challenge for the vast majority of the microorganisms
DNA isolated from these samples was used to explore the bacterial and archaeal diversity. 16S rRNA gene sequences were clustered at an identity threshold 99%, resulting in a total of 970 operational taxonomic units (OTUs) (Supplementary Table S4) that after the phylogenetic inference produced a total of 226 operational phylogenetic units (OPUs), 200 for Bacteria and 26 for Archaea (Figure 1, Supplementary Table S5)
The bacterial diversity was dominated by three lineages never reported as major components of hypersaline habitats, and the expected major key player Salinibacter was in a noticeable minority

Summary

Introduction

Life under extreme osmotic pressure in the environment represents a challenge for the vast majority of the microorganisms Hypersaline habitats such as lakes, salt ponds, and sediments associated with marine ecosystems are considered extreme environments constituted by a discontinuous salinity gradient where salt can reach saturation by evaporation processes (Oren, 2002). The ‘saltin’ strategy is characterized by increasing the salt concentration inside the cell, leading to significant changes in the enzymatic machinery These include the over-representation of highly acidic amino acids such as aspartate (Asp), and a low proportion of hydrophobic residues that tend to form coil regions instead of helical structures when compared to nonhalophile proteins (Paul et al, 2008; Rhodes et al, 2010). Some of these solutes are found in specific phylogenetic groups while others are widely distributed in halophilic organisms (Oren, 2008)

Methods

Results

Conclusion