Novel acid resistance genes from the metagenome of the Tinto River, an extremely acidic environment

Abstract

Microorganisms that thrive in acidic environments are endowed with specialized molecular mechanisms to survive under this extremely harsh condition. In this work, we performed functional screening of six metagenomic libraries from planktonic and rhizosphere microbial communities of the Tinto River, an extremely acidic environment, to identify genes involved in acid resistance. This approach has revealed 15 different genes conferring acid resistance to Escherichia coli, most of which encoding putative proteins of unknown function or previously described proteins not known to be related to acid resistance. Moreover, we were able to assign function to one unknown and three hypothetical proteins. Among the recovered genes were the ClpXP protease, the transcriptional repressor LexA and nucleic acid-binding proteins such as an RNA-binding protein, HU and Dps. Furthermore, nine of the retrieved genes were cloned and expressed in Pseudomonas putida and Bacillus subtilis and, remarkably, most of them were able to expand the capability of these bacteria to survive under severe acid stress. From this set of genes, four presented a broad-host range as they enhance the acid resistance of the three different organisms tested. These results expand our knowledge about the different strategies used by microorganisms to survive under extremely acid conditions.

Acid-resistant genes of oral plaque microbiome from the functional metagenomics

“…The DnaK molecular chaperone (U-57) is a ubiquitously distributed class I stress protein, which tolerates the low pH potentially through regulating the biogenesis or stabilization of the F-ATPase compound [13,24]. The functions of Clp protease (U-38) and methyltransferase (U-41) have been corroborated by a previous study [17]. Only SMU_1725 (U-23), mreD and gbpB (U-8) were newly identified functional genes from S. mutans UA159.…”

“…Both the inserts orf1 and ffh are known aciduric-related genes. The former encodes RNA-binding protein and a separate functional metagenomics study of the environmental sample confirmed its function [17], the latter comes from the S. mutans strain [11]. The purpose of construction of the strain E. coli DH10B/pSL-ffh and its functional verification was to detect whether the acid-resistant genes from oral bacteria could be heterologously expressed in E. coli .…”

“…In addition, we used two known acid-resistant genes, orf1 and ffh , to construct two positive controls, E. coli DH10B/pSL-orf1 and pSL-ffh, the former from the environmental microbes [17], the latter from S. mutans [11]. The detailed protocol is described in the Supplementary File.…”

“…Currently, function-based screening of metagenomic libraries has been widely deployed in different fields, including the screening of environmental libraries [17], the investigation of mouse gut colonization [18], of human faecal samples [19] and others. However, acid-resistance screening studies of oral plaque samples have not yet been reported.…”

Plant and microbe genomics and beyond: potential for developing a novel molecular plant nutrition approach

“…Functional analyses of environmental genomic libraries are not designed to identify genes involved in uptake, transport, and nutrient metabolism processes (Mirete et al 2007;Guazzaroni et al 2013). While these functions could be studied using microarrays, the nature of the samples and the limited availability of universal controls impose further complications for understanding the intensity of the detected signals and their correlation with specific species.…”