| Summary: | <p>Abstract</p> <p>Background</p> <p><it>Porphyromonas gingivalis</it> is a Gram-negative anaerobic bacterium associated with periodontal disease onset and progression. Genetic tools for the manipulation of bacterial genomes allow for in-depth mechanistic studies of metabolism, physiology, interspecies and host-pathogen interactions. Analysis of the essential genes, protein-coding sequences necessary for survival of <it>P. gingivalis</it> by transposon mutagenesis has not previously been attempted due to the limitations of available transposon systems for the organism. We adapted a Mariner transposon system for mutagenesis of <it>P. gingivalis</it> and created an insertion mutant library. By analyzing the location of insertions using massively-parallel sequencing technology we used this mutant library to define genes essential for <it>P. gingivalis</it> survival under <it>in vitro</it> conditions.</p> <p>Results</p> <p>In mutagenesis experiments we identified 463 genes in <it>P. gingivalis</it> strain ATCC 33277 that are putatively essential for viability <it>in vitro</it>. Comparing the 463 <it>P. gingivalis</it> essential genes with previous essential gene studies, 364 of the 463 are homologues to essential genes in other species; 339 are shared with more than one other species. Twenty-five genes are known to be essential in <it>P. gingivalis</it> and <it>B. thetaiotaomicron</it> only. Significant enrichment of essential genes within Cluster of Orthologous Groups ‘D’ (cell division), ‘I’ (lipid transport and metabolism) and ‘J’ (translation/ribosome) were identified. Previously, the <it>P. gingivalis</it> core genome was shown to encode 1,476 proteins out of a possible 1,909; 434 of 463 essential genes are contained within the core genome. Thus, for the species <it>P. gingivalis</it> twenty-two, seventy-seven and twenty-three percent of the genome respectively are devoted to essential, core and accessory functions.</p> <p>Conclusions</p> <p>A Mariner transposon system can be adapted to create mutant libraries in <it>P. gingivalis</it> amenable to analysis by next-generation sequencing technologies. <it>In silico</it> analysis of genes essential for <it>in vitro</it> growth demonstrates that although the majority are homologous across bacterial species as a whole, species and strain-specific subsets are apparent. Understanding the putative essential genes of <it>P. gingivalis</it> will provide insights into metabolic pathways and niche adaptations as well as clinical therapeutic strategies.</p>
|
|---|
