Phylogenomics and comparative genomic studies delineate six main clades within the family Enterobacteriaceae and support the reclassification of several polyphyletic members of the family

Abstract

The family Enterobacteriaceae harbors many important pathogens, however it has proven difficult to reliably distinguish different members of this family or discern their interrelationships. To understand the interrelationships among the Enterobacteriaceae species, we have constructed two comprehensive phylogenetic trees for 78 genome-sequenced Enterobacteriaceae species based on 2487 core genome proteins, and another set of 118 conserved proteins. The genome sequences of Enterobacteriaceae species were also analyzed for genetic relatedness based on average amino acid identity and 16S rRNA sequence similarity. In parallel, comparative genomic studies on protein sequences from the Enterobacteriaceae have identified 88 molecular markers in the form of conserved signature indels (CSIs) that are uniquely shared by specific members of the family. All of these multiple lines of investigations provide consistent evidence that most of the species/genera within the family can be assigned to 6 different subfamily level clades which are designated as the "Escherichia clade", "Klebsiella clade", "Enterobacter clade", "Kosakonia clade", "Cronobacter clade" and "Cedecea clade". The members of the six described clades, in addition to their distinct branching in phylogenetic trees, can now be reliably demarcated in molecular terms on the basis of multiple identified CSIs that are exclusively shared by the group members. Several additional CSIs identified in this work that are either specific for individual genera (viz. Kosakonia, Kluyvera and Escherichia-Shigella), or are present at various taxonomic depths, offer information regarding the interrelationships among the different clades. The described molecular markers provide novel means for diagnostic as well as genetic and biochemical studies on the Enterobacteriaceae species and for resolving the polyphyly of its several genera viz. Escherichia, Enterobacter and Kluyvera. On the bases of our results, we are proposing the reclassification of Escherichia vulneris and Enterobacter massiliensis into two novel genera viz. Pseudescherichia gen. nov. and Metakosakonia gen. nov., respectively. Additionally, our results also support the transfer of "Enterobacter lignolyticus" and "Kluyvera intestini" to the genera Pluralibacter and Metakosakonia, respectively.

Keywords: Conserved signature indels; Enterobacteriaceae phylogeny; Enterobacteriaceae taxonomy; Molecular markers; Phylogenomics studies; Reclassification of Enterobacter and Escherichia species.