The hologenome concept: we need to incorporate function.

Are we in the midst of a paradigm change in biology and have animals and plants lost their individuality, i.e., are even so-called 'typical' organisms no longer organisms in their own right? Is the study of the holobiont-host plus its symbiotic microorganisms-no longer optional, but rather an obligatory path that must be taken for a comprehensive understanding of the ecology and evolution of the individual components that make up a holobiont? Or are associated microbes merely a component of their host's environment, and the holobiont concept is just a beautiful idea that does not add much or anything to our understanding of evolution? This article explores different aspects of the concept of the holobiont. We focus on the aspect of functional integration, a central holobiont property, which is only rarely considered thoroughly. We conclude that the holobiont comes in degrees, i.e., we regard the property of being a holobiont as a continuous trait that we term holobiontness, and that holobiontness is differentiated in several dimensions. Although the holobiont represents yet another level of selection (different from classical individual or group selection because it acts on a system that is composed of multiple species), it depends on the grade of functional integration whether or not the holobiont concept helps to cast light on the various degrees of interactions between symbiotic partners.

Flagellar movement in two bacteria of the family rickettsiaceae: a re-evaluation of motility in an evolutionary perspective.

Bacteria of the family Rickettsiaceae have always been largely studied not only for their importance in the medical field, but also as model systems in evolutionary biology. In fact, they share a recent common ancestor with mitochondria. The most studied species, belonging to genera Rickettsia and Orientia, are hosted by terrestrial arthropods and include many human pathogens. Nevertheless, recent findings show that a large part of Rickettsiaceae biodiversity actually resides outside the group of well-known pathogenic bacteria. Collecting data on these recently described non-conventional members of the family is crucial in order to gain information on ancestral features of the whole group. Although bacteria of the family Rickettsiaceae, and of the whole order Rickettsiales, are formally described as non-flagellated prokaryotes, some recent findings renewed the debate about this feature. In this paper we report the first finding of members of the family displaying numerous flagella and active movement inside their host cells. These two new taxa are hosted in aquatic environments by protist ciliates and are described here by means of ultrastructural and molecular characterization. Data here reported suggest that the ancestor of Rickettsiales displayed flagellar movement and re-evaluate the hypothesis that motility played a key-role in the origin of mitochondria. Moreover, our study highlights that the aquatic environment represents a well exploited habitat for bacteria of the family Rickettsiaceae. Our results encourage a deep re-consideration of ecological and morphological traits of the family and of the whole order.

'Candidatus Megaira polyxenophila' gen. nov., sp. nov.: considerations on evolutionary history, host range and shift of early divergent rickettsiae.

"Neglected Rickettsiaceae" (i.e. those harboured by non-hematophagous eukaryotic hosts) display greater phylogenetic variability and more widespread dispersal than pathogenic ones; yet, the knowledge about their actual host range and host shift mechanism is scarce. The present work reports the characterization following the full-cycle rRNA approach (SSU rRNA sequence, specific in situ hybridization, and ultrastructure) of a novel rickettsial bacterium, herewith proposed as 'Candidatus Megaira polyxenophila' gen. nov., sp. nov. We found it in association with four different free-living ciliates (Diophrys oligothrix, Euplotes octocarinatus, Paramecium caudatum, and Spirostomum sp., all belonging to Alveolata, Ciliophora); furthermore it was recently observed as intracellular occurring in Carteria cerasiformis and Pleodorina japonica (Chlorophyceae, Chlorophyta). Phylogenetic analyses demonstrated the belonging of the candidate new genus to the family Rickettsiaceae (Alphaproteobacteria, Rickettsiales) as a sister group of the genus Rickettsia. In situ observations revealed the ability of the candidate new species to colonize either nuclear or cytoplasmic compartments, depending on the host organism. The presence of the same bacterial species within different, evolutionary distant, hosts indicates that 'Candidatus Megaira polyxenophila' recently underwent several distinct host shifts, thus suggesting the existence of horizontal transmission pathways. We consider these findings as indicative of an unexpected spread of rickettsial infections in aquatic communities, possibly by means of trophic interactions, and hence propose a new interpretation of the origin and phylogenetic diversification of rickettsial bacteria.

Isolation of alveolar plates from Coleps hirtus.

In the ciliate Coleps hirtus, the alveoli contain rigid alveolar plates that are almost unstudied so far. Neither the exact composition nor the genesis and function are known. A necessary step to study the alveolar plates is to isolate these structures in an adequate amount. Therefore, culture conditions of C. hirtus were optimized to obtain an axenic and dense long-time culture. The protocol we developed to isolate C. hirtus alveolar plates is presented and clean alveolar plates were documented via scanning electron microscopy. The described procedure delivers alveolar plates of very good structure and integrity with preserved filigree details in sufficient amount. They can be analysed via a range of different material and biological characterisations. Since there are indications of a mineral phase within the alveolar plates, the presented results will allow to study C. hirtus alveolar plates also in the context of biomineralisation.

Tracing the role of R-bodies in the killer trait: absence of toxicity of R-body producing recombinant E. coli on paramecia.

R-bodies are coiled proteinaceous ribbons produced by Paramecium endosymbionts belonging to the genus Caedibacter. These intracellular bacteria confer upon their hosts a phenomenon called the killer trait. It is the ability to kill symbiont-free competitors called sensitives. The R-body is the crucial element of this process, but despite many efforts, the actual role of R-bodies in killing sensitive paramecia is still not satisfactory clarified. The open question is whether the R-body acts as transmitter for a yet unidentified toxin or whether it directly kills sensitive paramecia having intrinsic cytotoxic effects. In the present study, this problem is addressed by heterologous expression of Caedibacter taeniospiralis R-body in Escherichia coli followed by a detailed analysis of its potential intrinsic toxic effect on feeding sensitive Paramecium tetraurelia. Using this approach, we can exclude any eventual effects of additional, unidentified factors produced by C. taeniospiralis and thus observe the impact of the recombinant R-body itself. No cytotoxic effects of recombinant R-bodies were detected following this approach, strengthening the hypothesis that R-bodies act as releasing system for an unidentified C. taeniospiralis toxin.

The mitochondrial genomes of the ciliates Euplotes minuta and Euplotes crassus.

BACKGROUND: There are thousands of very diverse ciliate species from which only a handful mitochondrial genomes have been studied so far. These genomes are rather similar because the ciliates analysed (Tetrahymena spp. and Paramecium aurelia) are closely related. Here we study the mitochondrial genomes of the hypotrichous ciliates Euplotes minuta and Euplotes crassus. These ciliates are only distantly related to Tetrahymena spp. and Paramecium aurelia, but more closely related to Nyctotherus ovalis, which possesses a hydrogenosomal (mitochondrial) genome. RESULTS: The linear mitochondrial genomes of the hypotrichous ciliates Euplotes minuta and Euplotes crassus were sequenced and compared with the mitochondrial genomes of several Tetrahymena species, Paramecium aurelia and the partially sequenced mitochondrial genome of the anaerobic ciliate Nyctotherus ovalis. This study reports new features such as long 5'gene extensions of several mitochondrial genes, extremely long cox1 and cox2 open reading frames and a large repeat in the middle of the linear mitochondrial genome. The repeat separates the open reading frames into two blocks, each having a single direction of transcription, from the repeat towards the ends of the chromosome. Although the Euplotes mitochondrial gene content is almost identical to that of Paramecium and Tetrahymena, the order of the genes is completely different. In contrast, the 33273 bp (excluding the repeat region) piece of the mitochondrial genome that has been sequenced in both Euplotes species exhibits no difference in gene order. Unexpectedly, many of the mitochondrial genes of E. minuta encoding ribosomal proteins possess N-terminal extensions that are similar to mitochondrial targeting signals. CONCLUSION: The mitochondrial genomes of the hypotrichous ciliates Euplotes minuta and Euplotes crassus are rather different from the previously studied genomes. Many genes are extended in size compared to mitochondrial genes from other sources.

"Candidatus Paraholospora nucleivisitans", an intracellular bacterium in Paramecium sexaurelia shuttles between the cytoplasm and the nucleus of its host.

An intracellular bacterium was discovered in two isolates of Paramecium sexaurelia from an aquarium with tropical fish in Münster (Germany) and from a pond in the Wilhelma zoological-botanical garden, Stuttgart (Germany). The bacteria were regularly observed in the cytoplasm of the host, but on some occasions they were found in the macronucleus of the host cell. In these cases, only a few, if any, bacteria were observed remaining in the cytoplasm. The bacterium was not infectious to P. sexaurelia or other species of Paramecium and appeared to be an obligate intracellular bacterium, while bacteria-free host cells were completely viable. The fluorescence in situ hybridisation (FISH) and comparative 16SrDNA sequence analyses showed that the bacterium belonged to a new genus, and was most closely, yet quite distantly, related to Holospora obtusa. In spite of this relationship, the new bacteria differed from Holospora by at least two biological features. Whereas all Holospora species reside exclusively in the nuclei of various species of Paramecium and show a life cycle with a morphologically distinct infectious form, for the new bacterium no infectious form and no life cycle have been observed. For the new bacterium, the name Candidatus Paraholospora nucleivisitans is suggested. The host P. sexaurelia is usually known from tropical and subtropical areas and is not a species typically found in Germany and central Europe. Possibly, it had been taken to Germany with fish or plants from tropical or subtropical waters. Candidatus Paraholospora nucleivisitans may therefore be regarded as an intracellular neobacterium for Germany.

"Candidatus Cryptoprodotis polytropus," a novel Rickettsia-like organism in the ciliated protist Pseudomicrothorax dubius (Ciliophora, Nassophorea).

Rickettsia-like organisms (RLO) are obligate, often highly fastidious, intracellular bacterial parasites associated with a variety of vertebrate and invertebrate hosts. Despite their importance as causative agents of severe mortality outbreaks in farmed aquatic species, little is known about their life cycle and their host range. The present work reports the characterization of "Candidatus Cryptoprodotis polytropus," a novel Rickettsia-like bacterium associated with the common ciliate species Pseudomicrothorax dubius by means of the "Full-Cycle rRNA Approach" and ultrastructural observations. The morphological description by in vivo and scanning electron microscopy and the 18S rRNA gene sequence of the host species is provided as well. Phylogenetic analysis based on the 16S rRNA gene supports the inclusion of "Candidatus Cryptoprodotis polytropus" within the family Rickettsiaceae (cl. Alphaproteobacteria) together with the genera Rickettsia and Orientia. Observations on natural ciliate populations account for the occasional nature of this likely parasitic association. The presence of a previously unknown RLO in ciliates sheds a new light on the possible role of protists as transient hosts, vectors or natural reservoir for some economically important pathogens.

The genus Caedibacter comprises endosymbionts of Paramecium spp. related to the Rickettsiales (Alphaproteobacteria) and to Francisella tularensis (Gammaproteobacteria).

Obligate bacterial endosymbionts of paramecia able to form refractile inclusion bodies (R bodies), thereby conferring a killer trait upon their ciliate hosts, have traditionally been grouped into the genus CAEDIBACTER: Of the six species described to date, only the Paramecium caudatum symbiont Caedibacter caryophilus has been phylogenetically characterized by its 16S rRNA gene sequence, and it was found to be a member of the Alphaproteobacteria related to the RICKETTSIALES: In this study, the Caedibacter taeniospiralis type strain, an R-body-producing cytoplasmatic symbiont of Paramecium tetraurelia strain 51k, was investigated by comparative 16S rRNA sequence analysis and fluorescence in situ hybridization with specific oligonucleotide probes. C. taeniospiralis is not closely related to C. caryophilus (80% 16S rRNA sequence similarity) but forms a novel evolutionary lineage within the Gammaproteobacteria with the family Francisellaceae as a sister group (87% 16S rRNA sequence similarity). These findings demonstrate that the genus Caedibacter is polyphyletic and comprises at least two phylogenetically different bacterial species belonging to two different classes of the PROTEOBACTERIA: Comparative phylogenetic analysis of C. caryophilus, five closely related Acanthamoeba endosymbionts (including one previously uncharacterized amoebal symbiont identified in this study), and their hosts suggests that the progenitor of the alphaproteobacterial C. caryophilus lived within acanthamoebae prior to the infection of paramecia.

Intracellular bacteria in ciliates

Ciliates are frequently colonized by other micro-organisms. The large size of ciliate cells offers habitats for hundreds to thousands of bacteria in different compartments, such as cytoplasm, nuclei and even perinuclear spaces. Size, phagocytic feeding habit and other features appear to be favorable pre-adaptations of ciliates for symbiosis with bacteria. Certain intracellular bacteria are permanent symbionts that are not infectious, whereas others are highly infectious. Both types show specific adaptations. With their wide spectrum of phylogenetic positions, intracellular bacteria in ciliates show relationships to different taxa of free-living bacteria and even archaea. Certain symbionts may be deleterious for their host ciliates, whereas others may provide a selective advantage under appropriate conditions or even be essential for the host cells. Depending on the nature of a symbiont, its prevalence in a host population may be low or high. Symbionts that express a killer toxin affecting non-infected ciliates achieve high infection rates in a host population, whereas certain infectious bacteria may only show a low prevalence (AU)

No disponible