Verni, Franco

AbstractProtists frequently host diverse bacterial symbionts, in particular those affiliated with the order Holosporales (Alphaproteobacteria). All characterised members of this bacterial lineage have been retrieved in obligate association with a wide range of eukaryotes, especially multiple protist lineages (e.g. amoebozoans, ciliates, cercozoans, euglenids, and nucleariids), as well as some metazoans (especially arthropods and related ecdysozoans). While the genus Paramecium and other ciliates have been deeply investigated for the presence of symbionts, known members of the family “Candidatus Paracaedibacteraceae” (Holosporales) are currently underrepresented in such hosts. Herein, we report the description of “Candidatus Intestinibacterium parameciiphilum” within the family “Candidatus Paracaedibacteraceae”, inhabiting the cytoplasm of Paramecium biaurelia. This novel bacterium is almost twice as big as its relative “Candidatus Intestinibacterium nucleariae” from the opisthokont Nuclearia and does not present a surrounding halo. Based on phylogenetic analyses of 16S rRNA gene sequences, we identified six further potential species-level lineages within the genus. Based on the provenance of the respective samples, we investigated the environmental distribution of the representatives of “Candidatus Intestinibacterium” species. Obtained results are consistent with an obligate endosymbiotic lifestyle, with protists, in particular freshwater ones, as hosts. Thus, available data suggest that association with freshwater protists could be the ancestral condition for the members of the “Candidatus Intestinibacterium” genus.

AbstractTaxonomy is the science of defining and naming groups of biological organisms based on shared characteristics and, more recently, on evolutionary relationships. With the birth of novel genomics/bioinformatics techniques and the increasing interest in microbiome studies, a further advance of taxonomic discipline appears not only possible but highly desirable. The present work proposes a new approach to modern taxonomy, consisting in the inclusion of novel descriptors in the organism characterization: (1) the presence of associated microorganisms (e.g.: symbionts, microbiome), (2) the mitochondrial genome of the host, (3) the symbiont genome. This approach aims to provide a deeper comprehension of the evolutionary/ecological dimensions of organisms since their very first description. Particularly interesting, are those complexes formed by the host plus associated microorganisms, that in the present study we refer to as “holobionts”. We illustrate this approach through the description of the ciliateEuplotes vanleeuwenhoekisp. nov. and its bacterial endosymbiont “CandidatusPinguicoccus supinus” gen. nov., sp. nov. The endosymbiont possesses an extremely reduced genome (~ 163 kbp); intriguingly, this suggests a high integration between host and symbiont.

Most of the microorganisms responsible for vector-borne diseases (VBD) have hematophagous arthropods as vector/reservoir. Recently, many new species of microorganisms phylogenetically related to agents of VBD were found in a variety of aquatic eukaryotic hosts; in particular, numerous new bacterial species related to the genus Rickettsia (Alphaproteobacteria, Rickettsiales) were discovered in protist ciliates and other unicellular eukaryotes. Although their pathogenicity for humans and terrestrial animals is not known, several indirect indications exist that these bacteria might act as etiological agents of possible VBD of aquatic organisms, with protists as vectors. In the present study, a novel strain of the Rickettsia-Like Organism (RLO) endosymbiont “Candidatus (Ca.) Trichorickettsia mobilis” was identified in the macronucleus of the ciliate Paramecium multimicronucleatum. We performed transfection experiments of this RLO to planarians (Dugesia japonica) per os. Indeed, the latter is a widely used model system for studying bacteria pathogenic to humans and other Metazoa. In transfection experiments, homogenized paramecia were added to food of antibiotic-treated planarians. Treated and non-treated (i.e. control) planarians were investigated at day 1, 3, and 7 after feeding for endosymbiont presence by means of PCR and ultrastructural analyses. Obtained results were fully concordant and suggest that this RLO endosymbiont can be transiently transferred from ciliates to metazoans, being detected up to day 7 in treated planarians’ enterocytes. Our findings might offer insights into the potential role of ciliates or other protists as putative vectors for diseases caused by Rickettsiales or other RLOs and occurring in fish farms or in the wild.

ABSTRACTThe orderRickettsiales(Alphaproteobacteria) is a well-known group containing obligate endocellular prokaryotes. The order encompasses three families (Rickettsiaceae,Anaplasmataceae, andHolosporaceae) and a fourth, family-level cluster, which includes only one candidate species, “CandidatusMidichloria mitochondrii,” as well as several unnamed bacterial symbionts. The broad host range exhibited by the members of the “CandidatusMidichloria” clade suggests their eventual relevance for a better understanding of the evolution of symbiosis and host specificity ofRickettsiales. In this paper, two new bacteria belonging to the “CandidatusMidichloria” clade, hosted by two different strains of the ciliate protistEuplotes harpa, are described on the basis of ultrastructural observations, comparative 16S rRNA gene sequence analysis, and an estimation of the percentage of infection. Ultrastructure of these bacteria shows some unusual features: one has an electron-dense cytoplasm, and the other one lacks a symbiosomal membrane. The latter was up to now considered an exclusive feature of bacteria belonging to the familyRickettsiaceae. 16S rRNA gene phylogenetic analysis unambiguously places the new bacteria in the “CandidatusMidichloria” clade, although their phylogenetic relationships with other members of the clade are not clearly resolved. This is the first report of a ciliate-borne bacterium belonging to the “CandidatusMidichloria” clade. On the basis of the data obtained, the two bacteria are proposed as two new candidate genera and species, “CandidatusAnadelfobacter veles” and “CandidatusCyrtobacter comes.”