Diplorickettsia massiliensis species is an obligate intracellular, gram negative bacterium isolated from Ixodes ricinus ticks collected in Slovak republic forest geographically from southeastern part of Rovinka in 2006. They belong to the gammaproteobacteria class and are non endospore forming, small rods usually grouped in pairs. The bacteria are non-motile, and 16S rRNA, rpoB, parC and ftsY gene sequencing indicate that this bacterium is clearly different from all other recognized species. An initial phylogenetic analysis based on 16S rRNA, clustered D. massiliensis with Rickettsiella grylli. Because of its low 16S rDNA similarity (94%) with R. grylli, it was classified as a new genus Diplorickettsia into the family Coxiellaceae and the order Legionellales. D. massiliensis strain 20B was identified in three patients with suspected tick-borne infections that exhibited a specific seroconversion. The evidence of infection was further reconfirmed by using PCR-assay, thus established its role as a human pathogen and later whole genome sequencing was performed.
Description
Diplorickettsia massiliensis (mas.si' li.en.sis. L. gen. adj. massiliensis, from Massilia, the Latin name of Marseille, France, where the organism was first grown, identified and characterized). The description is for that of genus type strain 20B. The known geographical distribution of this bacterium is Slovakia. This isolate has been deposited in the collection of the two World Health Organization Collaborative Centers for Rickettsial Reference and Research in Bratislava, Slovak Republic and the Faculté de Médecine, University of the Mediterranean in Marseille, France, as well as in the German Collection of Microorganisms and Cell Cultures (Deutsche Sammlung von Mikroorganismen und Zellkulturen, DSMZ) under the reference DSM 233381.
The GenBank/EMBL/DDBJ accession numbers for the 16S rRNA gene, rpoB, parC and ftsY sequences of strain B20 are GQ857549, GQ983049, GQ983050, and GU289825, respectively.
Phylogenetic analysis and Molecular Signatures
Comparative 16S rRNA gene sequence analysis showed that this strain belongs to the family Coxiellaceae, order Legionellales of Gamma-proteobacteria, and the closest relatives are different Rickettsiella spp. A 1476-bp fragment of the rrs gene encoding the 16S ribosomal RNA was amplified and sequenced. Surprisingly, a BLAST search did not result in a close similarity with any published 16S RNA gene sequences from other bacteria. The level of 16S rRNA gene sequence similarity between strain 20B and other recognized species of the family was below 94.5%. Partial sequences of the rpoB, parC and ftsY genes confirmed the phylogenetic position of the new isolate. The G+C content estimated on the basis of whole genome analysis of strain 20B was 37.88%. On the basis of its phenotypic and genotypic properties, together with phylogenetic distinctiveness, Mediannikov et al. proposed that strain 20B to be classified in the new genus Diplorickettsia as the type strain of a novel species named Diplorickettsia massiliensis sp. nov.
Diplorickettsia massiliensis and members of the genus Rickettsiella were observed to form a reliable clade in phylogenetic trees, suggesting that they might belong to a single genus. 12 conserved signature indels (CSIs) were also identified for this clade in the proteins inositol monophosphatase, lysyl-tRNA synthetase, elongation factor P-(R)-beta-lysine ligase, tol-Pal system beta propeller repeat protein TolB, FKBP-type peptidyl-prolyl cis-trans isomerase, response regulator transcription factor, 30S ribosomal protein S2, glycine cleavage system aminomethyltransferase GcvT, M3 family metallopeptidase, lysm peptidoglycan-binding domain-containing protein and oligopeptide transporter. The presence of these CSIs molecularly distinguishes this clade from other members of the family Coxiellaceae, further supporting the combination of the genera Rickettsiella and Diplorickettsia.
Additionally, three CSIs were identified as uniquely shared between Diplorickettsia, Rickettsiella and Coxiella, another genus belonging to the family Coxiellaceae. The absence of Aquicella species (the remaining genus within the family Coxiellaceae) from the shared CSIs suggests that the genera Coxiella, Diplorickettsia and Rickettsiella share a common ancestor exclusive of Aquicella.