The essential genome of Streptococcus agalactiae
In: BMC Genomics, Jg. 17, Heft 1
Online
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Zugriff:
Background Next-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. Methods Our 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. Results After 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. Conclusions We 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. Electronic supplementary material The online version of this article (doi:10.1186/s12864-016-2741-z) contains supplementary material, which is available to authorized users.
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The essential genome of Streptococcus agalactiae
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Autor/in / Beteiligte Person: | Tallon, Luke J. ; Maskell, Duncan J. ; Tettelin, Hervé ; Ott, Sandra ; Catomeris, Andrew J. ; Randis, Tara M. ; Ratner, Adam J. ; Akabas, Leor H. ; Hooven, Thomas A. ; Santana-Cruz, Ivette ; Peters, Sarah ; Maskell, Duncan [0000-0002-5065-653X] ; Apollo - University of Cambridge Repository |
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Zeitschrift: | BMC Genomics, Jg. 17, Heft 1 |
Veröffentlichung: | Springer Nature |
Medientyp: | unknown |
ISSN: | 1471-2164 (print) |
DOI: | 10.1186/s12864-016-2741-z |
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