Abstract
Gallbladder metagenome involves a wide range of unidentified sequences comprising the so-called metagenomic dark matter. Therefore, this study aimed to characterise three gallbladder metagenomes and a fosmid library with an emphasis on metagenomic dark matter fraction. For this purpose, a novel data analysis strategy based on the combination of remote homology and molecular modelling has been proposed. According to the results obtained, several protein functional domains were annotated in the metagenomic dark matter fraction including acetyltransferases, outer membrane transporter proteins, membrane assembly factors, DNA repair and recombination proteins and response regulator phosphatases. In addition, one deacetylase involved in mycothiol biosynthesis was found in the metagenomic dark matter fraction of the fosmid library. This enzyme may exert a protective effect in Actinobacteria against bile components exposure, in agreement with the presence of multiple antibiotic and multidrug resistance genes. Potential mechanisms of action of this novel deacetylase were elucidated by molecular simulations, highlighting the role of histidine and aspartic acid residues. Computational pipelines presented in this work may be of special interest to discover novel microbial enzymes which had not been previously characterised.
Highlights
The human gallbladder comprises a scarcely studied microbial ecological niche compared to the human faecal microbiome
A comprehensive data analysis strategy involving two different steps was followed: (1) identification of antibiotic resistance genes (ARGs) and multiple drug resistance (MDR) and (2) tentative assignment of biological functions to metagenomic dark matter domains through remote homology and computational chemistry techniques
Main families present in fosmid library include Caulobacteraceae, Chitinophagaceae, Lachnospiraceae, Gammaproteobacteria, Sphingomonadaceae, Ruminococcaceae, Muribaculaceae, Rikenellaceae, Burkholderiaceae, Bacteroidaceae, Streptomycetaceae, Enterobacteriaceae, Barnesiellaceae and Prevotellaceae in agreement with 16S sequencing data of bile samples used to prepare fosmid library reported by Molinero et al [1]
Summary
The human gallbladder comprises a scarcely studied microbial ecological niche compared to the human faecal microbiome. Few studies characterised the human biliary metagenome. Song et al [2], performed metagenomic sequencing of the biliary microbiota of individuals who had undergone laparoscopic cholecystectomy or radical cholecystectomy. Shen et al [3], reported metagenomic characterisation of bile samples from patients with gallstone disease. Previous works in our group were performed to characterise, by 16S rDNA and shotgun metagenomic sequencing, bile samples from the gallbladder of individuals suffering from lithiasis as well as subjects without any record of the hepatobiliary disorder [1]. We observed that bile metagenomes contain a larger number of unclassified sequences and protein functional domains than faecal samples from healthy
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