Functional metagenomics reveals novel salt tolerance loci from the human gut microbiome

Abstract

Metagenomics is a powerful tool that allows for the culture-independent analysis of complex microbial communities. One of the most complex and dense microbial ecosystems known is that of the human distal colon, with cell densities reaching up to 10 12 per gram of faeces. With the majority of species as yet uncultured, there are an enormous number of novel genes awaiting discovery. In the current study, we conducted a functional screen of a metagenomic library of the human gut microbiota for potential salt-tolerant clones. Using transposon mutagenesis, three genes were identified from a single clone exhibiting high levels of identity to a species from the genus Collinsella (closest relative being Collinsella aerofaciens) (COLAER_01955, COLAER_01957 and COLAER_01981), a high G þ C, Gram-positive member of the Actinobacteria commonly found in the human gut. The encoded proteins exhibit a strong similarity to GalE, MurB and MazG. Furthermore, pyrosequencing and bioinformatic analysis of two additional fosmid clones revealed the presence of an additional galE and mazG gene, with the highest level of genetic identity to Akkermansia muciniphila and Eggerthella sp. YY7918, respectively. Cloning and heterologous expression of the genes in the osmosensitive strain, Escherichia coli MKH13, resulted in increased salt tolerance of the transformed cells. It is hoped that the identification of atypical salt tolerance genes will help to further elucidate novel salt tolerance mechanisms, and will assist our increased understanding how resident bacteria cope with the osmolarity of the gastrointestinal tract.

Effects of transportation stress and addition of salt to transport water on the skin mucosal homeostasis of rainbow trout (Oncorhynchus mykiss)

“…It is known that different bacterial species, including symbionts of the human microbiome have different tolerance to NaCl (Culligan et al, 2012). Thus, it is likely that addition of NaCl to transport water favors the growth of certain bacterial groups whereas limits the growth of others.…”

Functional metagenomic discovery of bacterial effectors in the human microbiome and isolation of commendamide, a GPCR G2A/132 agonist

“…In addition, it couples each observed phenotype to a single fragment of cloned metagenomic DNA, making it possible to simultaneously identify specific effector molecules and the specific effector genes that encode these molecules. Functional metagenomics has been used to isolate small molecules and proteins from soil metagenomes (15)(16)(17)(18)(19)(20)(21)(22)(23)(24); however, its application to the human microbiome has to date been very limited (16,25,26). In this study, a set of arrayed, large-insert cosmid libraries hosted in Escherichia coli was created from DNA isolated from the stool of three patients who, based on their phenotype, were predicted to have different commensal bacteria cohorts.…”

Adaptation in Natural Microbial Populations

“…Hemme et al 2010) and specific phenotypes can be discovered 30 through functional metagenomics, where random sequence fragments isolated from a 8 community sample are cloned into a vector, allowing host bacteria to be screened for specific traits encoded by these sequences (e.g. Culligan et al 2012). These correlation patterns are important to establish, but causation can be more directly addressed through controlled manipulation of the environment, such as the artificial warming of soil (Rousk et al 2012).…”