Abstract
BackgroundBacteroides fragilis is a Gram-negative anaerobe that is normally a human gut commensal; it comprises a small percentage of the gut Bacteroides but is the most frequently isolated Bacteroides from human infections. Identification of the essential genes necessary for the survival of B. fragilis provides novel information which can be exploited for the treatment of bacterial infections.ResultsMassive parallel sequencing of saturated transposon mutant libraries (two mutant pools of approximately 50,000 mutants each) was used to determine the essential genes for the growth of B. fragilis 638R on nutrient rich medium. Among the 4326 protein coding genes, 550 genes (12.7%) were found to be essential for the survival of B. fragilis 638R. Of the 550 essential genes, only 367 genes were assigned to a Cluster of Orthologous Genes, and about 290 genes had Kyoto Encyclopedia of Genes and Genomes orthologous members. Interestingly, genes with hypothetical functions accounted for 41.3% of essential genes (227 genes), indicating that the functions of a significant percentage of the genes used by B. fragilis 638R are still unknown. Global transcriptome analysis using RNA-Seq indicated that most of the essential genes (92%) are, in fact, transcribed in B. fragilis 638R including most of those coding for hypothetical proteins. Three hundred fifty of the 550 essential genes of B. fragilis 638R are present in Database of Essential Genes. 10.02 and 31% of those are genes included as essential genes for nine species (including Gram-positive pathogenic bacteria).ConclusionsThe essential gene data described in this investigation provides a valuable resource to study gene function and pathways involved in B. fragilis survival. Thorough examination of the B. fragilis-specific essential genes and genes that are shared between divergent organisms opens new research avenues that will lead to enhanced understanding of survival strategies used by bacteria in different microniches and under different stress situations.Electronic supplementary materialThe online version of this article (doi: 10.1186/1471-2164-15-429) contains supplementary material, which is available to authorized users.
Highlights
Bacteroides fragilis is a Gram-negative anaerobe that is normally a human gut commensal; it comprises a small percentage of the gut Bacteroides but is the most frequently isolated Bacteroides from human infections
After the genomic DNA preparation, each of these samples was split into two technical replicates (TR) to minimize any changes due to technical variation introduced by downstream manipulations
We identified the essential genes required for the survival of B. fragilis 638R in brain heart infusion (BHI) medium using a transposon delivery vector and Illumina sequencing technology
Summary
Bacteroides fragilis is a Gram-negative anaerobe that is normally a human gut commensal; it comprises a small percentage of the gut Bacteroides but is the most frequently isolated Bacteroides from human infections. As a commensal it provides many benefits to the host, including digestion of complex polysaccharides, While B. fragilis accounts for only a small percentage of the gut Bacteroides, it is the major Bacteroides species isolated from human infections [3]. While the scope of the factors that account for the particular virulence of B. fragilis are not fully known, several virulence factors have been described including the ability to withstand low concentrations of oxygen [6], release of degradative enzymes such as fibrinogenolysin [7], enterotoxin production, evasion of complement-mediated killing and phagocytosis, induction of abscess formation, and extensive within-strain variation of surface proteins and polysaccharides (PSs). Abscesses may rupture and result in bacteremia [3,4]
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