The essential genome of Streptococcus agalactiae

BMC Genomics

Research Article

Thomas A. Hooven¹  Leor H. Akabas¹  Tara M. Randis²  Andrew J. Catomeris²  Adam J. Ratner³  Sarah E. Peters⁴  Duncan J. Maskell⁴  Luke J. Tallon⁵  Sandra Ott⁵  Ivette Santana-Cruz⁵  Hervé Tettelin⁵ 

[1] Department of Pediatrics, Columbia University, New York, NY, USA;Department of Pediatrics, Division of Pediatric Infectious Diseases, New York University School of Medicine, 550 First Avenue (MSB 223), 10016, New York, NY, USA;Department of Pediatrics, Division of Pediatric Infectious Diseases, New York University School of Medicine, 550 First Avenue (MSB 223), 10016, New York, NY, USA;Department of Microbiology, New York University School of Medicine, New York, NY, USA;Department of Veterinary Medicine, University of Cambridge, Cambridge, UK;Institute for Genome Sciences, University of Maryland School of Medicine, Baltimore, MD, USA;

关键词: Mutant Library;    Streptococcus Agalactiae;    Quality Control Step;    Fitness Assignment;    A909 Genome;   

DOI  :  10.1186/s12864-016-2741-z

 received in 2016-01-21, accepted in 2016-05-14,  发布年份 2016

来源: Springer

【 摘 要 】

BackgroundNext-generation sequencing of transposon-genome junctions from a saturated bacterial mutant library (Tn-seq) is a powerful tool that permits genome-wide determination of the contribution of genes to fitness of the organism under a wide range of experimental conditions. We report development, testing, and results from a Tn-seq system for use in Streptococcus agalactiae (group B Streptococcus; GBS), an important cause of neonatal sepsis.MethodsOur method uses a Himar1 mini-transposon that inserts at genomic TA dinucleotide sites, delivered to GBS on a temperature-sensitive plasmid that is subsequently cured from the bacterial population. In order to establish the GBS essential genome, we performed Tn-seq on DNA collected from three independent mutant libraries—with at least 135,000 mutants per library—at serial 24 h time points after outgrowth in rich media.ResultsAfter statistical analysis of transposon insertion density and distribution, we identified 13.5 % of genes as essential and 1.2 % as critical, with high levels of reproducibility. Essential and critical genes are enriched for fundamental cellular housekeeping functions, such as acyl-tRNA biosynthesis, nucleotide metabolism, and glycolysis. We further validated our system by comparing fitness assignments of homologous genes in GBS and a close bacterial relative, Streptococcus pyogenes, which demonstrated 93 % concordance. Finally, we used our fitness assignments to identify signal transduction pathway components predicted to be essential or critical in GBS.ConclusionsWe believe that our baseline fitness assignments will be a valuable tool for GBS researchers and that our system has the potential to reveal key pathogenesis gene networks and potential therapeutic/preventative targets.

【 授权许可】

CC BY   
© The Author(s). 2016