Publications

3846

ABSTRACT In the past 10 years, the number of endosymbionts described within the bacterial order Rickettsiales has constantly grown. Since 2006, 18 novel Rickettsiales genera inhabiting protists, such as ciliates and amoebae, have been described. In this work, we characterize two novel bacterial endosymbionts from Paramecium collected near Bloomington, IN. Both endosymbiotic species inhabit the cytoplasm of the same host. The Gram-negative bacterium “ Candidatus Bealeia paramacronuclearis” occurs in clumps and is frequently associated with the host macronucleus. With its electron-dense cytoplasm and a distinct halo surrounding the cell, it is easily distinguishable from the second smaller symbiont, “ Candidatus Fokinia cryptica,” whose cytoplasm is electron lucid, lacks a halo, and is always surrounded by a symbiontophorous vacuole. For molecular characterization, the small-subunit rRNA genes were sequenced and used for taxonomic assignment as well as the design of species-specific oligonucleotide probes. Phylogenetic analyses revealed that “ Candidatus Bealeia paramacronuclearis” clusters with the so-called “basal” Rickettsiales , and “ Candidatus Fokinia cryptica” belongs to “ Candidatus Midichloriaceae.” We obtained tree topologies showing a separation of Rickettsiales into at least two groups: one represented by the families Rickettsiaceae , Anaplasmataceae , and “ Candidatus Midichloriaceae” (RAM clade), and the other represented by “basal Rickettsiales ,” including “ Candidatus Bealeia paramacronuclearis.” Therefore, and in accordance with recent publications, we propose to limit the order Rickettsiales to the RAM clade and to raise “basal Rickettsiales ” to an independent order, Holosporales ord. nov., inside Alphaproteobacteria , which presently includes four family-level clades. Additionally, we define the family “ Candidatus Hepatincolaceae” and redefine the family Holosporaceae . IMPORTANCE In this paper, we provide the characterization of two novel bacterial symbionts inhabiting the same Paramecium host (Ciliophora, Alveolata). Both symbionts belong to “traditional” Rickettsiales , one representing a new species of the genus “ Candidatus Fokinia” (“ Candidatus Midichloriaceae”), and the other representing a new genus of a “basal” Rickettsiales . According to newly characterized sequences and to a critical revision of recent literature, we propose a taxonomic reorganization of “traditional” Rickettsiales that we split into two orders: Rickettsiales sensu stricto and Holosporales ord. nov. This work represents a critical revision, including new records of a group of symbionts frequently occurring in protists and whose biodiversity is still largely underestimated.

AbstractGut-associated microbiota of ants include Rhizobiales bacteria with affiliation to the genus Bartonella. These bacteria may enable the ants to fix atmospheric nitrogen, but no genomes have been sequenced yet to test the hypothesis. Sequence reads from a member of the Rhizobiales were identified in the data collected in a genome project of the ant Harpegnathos saltator. We present an analysis of the closed 1.86 Mb genome of the ant-associated bacterium, for which we suggest the species name Candidatus Tokpelaia hoelldoblerii. A phylogenetic analysis reveals a relationship to Bartonella and Brucella, which infect mammals. Novel gene acquisitions include a gene for a putative extracellular protein of more than 6,000 amino acids secreted by the type I secretion system, which may be involved in attachment to the gut epithelium. No genes for nitrogen fixation could be identified, but genes for a multi-subunit urease protein complex are present in the genome. The urease genes are also present in Brucella, which has a fecal-oral transmission pathway, but not in Bartonella, which use blood-borne transmission pathways. We hypothesize that the gain and loss of the urease function is related to transmission strategies and lifestyle changes in the host-associated members of the Rhizobiales.

Cel.lu.lo.si.ly.ti.ca.ce'ae. N.L. neut. n.Cellulosilyticuma bacterial genus; N.L. fem. pl. n.CellulosilyticaceaetheCellulosilyticumfamily.Firmicutes / Clostridia / Clostridiales / CellulosilyticaceaeSlightly curved rod‐shaped cells. Gram‐positive cell‐wall structure but stain Gram‐negative. Terminal spores formed. Mesophilic, optimum growth temperature is approximately 40°C.Obligate anaerobic. No microaerophilic or aerobic growth occurs. Produce acetate, ethanol, and/or formate as the major end products, but not butyric or propionic acid. Hydrolyze cellulose and xylan and ferment cellobiose, but not ferment lactate and pyruvate. The predominant fatty acids are C16:0(27.1 %), C14:0(9.2 %), and iso‐C16:0(6.4%). Phylogenetically distant from the familyLachnospriaceae(Clostridum cluster XIVb), a member of the phylum Firmicutes, with less than 87.7% 16S rRNA similarity. Thus far, only one genusCellulosilyticumis described.DNA G + C content (mol%): 33–36.Type genus:CellulosilyticumCai and Dong 2010, 848VP.