Metagenome diversity illuminates the origins of pathogen effectors.

Recent metagenome-assembled genome (MAG) analyses have profoundly impacted Rickettsiology systematics. The discovery of basal lineages (novel families Mitibacteraceae and Athabascaceae) with predicted extracellular lifestyles exposed an evolutionary timepoint for the transition to host dependency, which seemingly occurred independent of mitochondrial evolution. Notably, these basal rickettsiae carry the Rickettsiales vir homolog (rvh) type IV secretion system and purportedly use rvh to kill congener microbes rather than parasitize host cells as described for later-evolving rickettsial pathogens. MAG analysis also substantially increased diversity for the genus Rickettsia and delineated a sister lineage (the novel genus Tisiphia) that stands to inform on the emergence of human pathogens from protist and invertebrate endosymbionts. Herein, we probed Rickettsiales MAG and genomic diversity for the distribution of Rickettsia rvh effectors to ascertain their origins. A sparse distribution of most Rickettsia rvh effectors outside of Rickettsiaceae lineages illuminates unique rvh evolution from basal extracellular species and other rickettsial families. Remarkably, nearly every effector was found in multiple divergent forms with variable architectures, indicating profound roles for gene duplication and recombination in shaping effector repertoires in Rickettsia pathogens. Lateral gene transfer plays a prominent role in shaping the rvh effector landscape, as evinced by the discovery of many effectors on plasmids and conjugative transposons, as well as pervasive effector gene exchange between Rickettsia and Legionella species. Our study exemplifies how MAGs can yield insight into pathogen effector origins, particularly how effector architectures might become tailored to the discrete host cell functions of different eukaryotic hosts.IMPORTANCEWhile rickettsioses are deadly vector-borne human diseases, factors distinguishing Rickettsia pathogens from the innumerable bevy of environmental rickettsial endosymbionts remain lacking. Recent metagenome-assembled genome (MAG) studies revealed evolutionary timepoints for rickettsial transitions to host dependency. The rvh type IV secretion system was likely repurposed from congener killing in basal extracellular species to parasitizing host cells in later-evolving pathogens. Our analysis of MAG diversity for over two dozen rvh effectors unearthed their presence in some non-pathogens. However, most effectors were found in multiple divergent forms with variable architectures, indicating gene duplication and recombination-fashioned effector repertoires of Rickettsia pathogens. Lateral gene transfer substantially shaped pathogen effector arsenals, evinced by the discovery of effectors on plasmids and conjugative transposons, as well as pervasive effector gene exchanges between Rickettsia and Legionella species. Our study exemplifies how MAGs yield insight into pathogen effector origins and evolutionary processes tailoring effectors to eukaryotic host cell biology.

Host association and intracellularity evolved multiple times independently in the Rickettsiales.

The order Rickettsiales (Alphaproteobacteria) encompasses multiple diverse lineages of host-associated bacteria, including pathogens, reproductive manipulators, and mutualists. Here, in order to understand how intracellularity and host association originated in this order, and whether they are ancestral or convergently evolved characteristics, we built a large and phylogenetically-balanced dataset that includes de novo sequenced genomes and a selection of published genomic and metagenomic assemblies. We perform detailed functional reconstructions that clearly indicates "late" and parallel evolution of obligate host-association in different Rickettsiales lineages. According to the depicted scenario, multiple independent horizontal acquisitions of transporters led to the progressive loss of biosynthesis of nucleotides, amino acids and other metabolites, producing distinct conditions of host-dependence. Each clade experienced a different pattern of evolution of the ancestral arsenal of interaction apparatuses, including development of specialised effectors involved in the lineage-specific mechanisms of host cell adhesion and/or invasion.

Microbial diversity of ticks and a novel typhus group Rickettsia species (Rickettsiales bacterium Ac37b) in Inner Mongolia, China.

Ticks can carry multiple pathogens, and Inner Mongolia's animal husbandry provides excellent environmental conditions for ticks. This study characterized the microbiome of ticks from different geographical locations in Inner Mongolia; 905 Dermacentor nuttalli and 36 Ixodes persulcatus were collected from sheep in three main pasture areas and from bushes within the forested area. Mixed DNA samples were prepared from three specimens from each region and tick species. Microbial diversity was analyzed by 16S rRNA sequencing, and α and ß diversity were determined. The predominant bacterial genera were Rickettsia (54.60%), including Rickettsiales bacterium Ac37b (19.33%) and other Rickettsia (35.27%), Arsenophonus (11.21%), Candidatus Lariskella (10.84%), and Acinetobacter (7.17%). Rickettsia bellii was identified in I. persulcatus, while Rickettsiales bacterium Ac37b was found in D. nuttalli from Ordos and Chifeng. Potential Rickettsia and Anaplasma coinfections were observed in the Ordos region. Tick microbial diversity analysis in Inner Mongolia suggests that sheep at the sampling sites were exposed to multiple pathogens.

Title: Diversité microbienne des tiques et nouvelle espèce de Rickettsia du groupe du typhus (bactérie Rickettsiales Ac37b) en Mongolie intérieure, Chine. Abstract: Les tiques peuvent être porteuses de plusieurs agents pathogènes et l'élevage en Mongolie intérieure offre d'excellentes conditions environnementales pour les tiques. Cette étude a caractérisé le microbiome des tiques de différentes zones géographiques de Mongolie intérieure; 905 Dermacentor nuttalli et 36 Ixodes persulcatus ont été collectés sur des moutons dans trois principales zones de pâturage et dans des buissons de la zone forestière. Des échantillons d'ADN mixtes ont été préparés à partir de trois spécimens de chaque région et espèce de tique. La diversité microbienne a été analysée par séquençage de l'ARNr 16S et la diversité α et ß a été déterminée. Les genres bactériens prédominants étaient les Rickettsia (54,60 %), dont la bactérie Rickettsiales Ac37b (19,33 %) et d'autres Rickettsia (35,27 %), Arsenophonus (11,21 %), Candidatus Lariskella (10,84 %) et Acinetobacter (7,17 %). Rickettsia bellii a été identifiée chez I. persulcatus, tandis que la bactérie Rickettsiales Ac37b a été trouvée chez D. nuttalli d'Ordos et Chifeng. Des co-infections potentielles à Rickettsia et Anaplasma ont été observées dans la région d'Ordos. L'analyse de la diversité microbienne des tiques en Mongolie intérieure montre que les moutons présents sur les sites d'échantillonnage sont exposés à plusieurs agents pathogènes.

The evolutionary origin of host association in the Rickettsiales.

The evolution of obligate host-association of bacterial symbionts and pathogens remains poorly understood. The Rickettsiales are an alphaproteobacterial order of obligate endosymbionts and parasites that infect a wide variety of eukaryotic hosts, including humans, livestock, insects and protists. Induced by their host-associated lifestyle, Rickettsiales genomes have undergone reductive evolution, leading to small, AT-rich genomes with limited metabolic capacities. Here we uncover eleven deep-branching alphaproteobacterial metagenome assembled genomes from aquatic environments, including data from the Tara Oceans initiative and other publicly available datasets, distributed over three previously undescribed Rickettsiales-related clades. Phylogenomic analyses reveal that two of these clades, Mitibacteraceae and Athabascaceae, branch sister to all previously sampled Rickettsiales. The third clade, Gamibacteraceae, branch sister to the recently identified ectosymbiotic 'Candidatus Deianiraea vastatrix'. Comparative analyses indicate that the gene complement of Mitibacteraceae and Athabascaceae is reminiscent of that of free-living and biofilm-associated bacteria. Ancestral genome content reconstruction across the Rickettsiales species tree further suggests that the evolution of host association in Rickettsiales was a gradual process that may have involved the repurposing of a type IV secretion system.

Geographically driven differences in microbiomes of Acropora cervicornis originating from different regions of Florida's Coral Reef.

Effective coral restoration must include comprehensive investigations of the targeted coral community that consider all aspects of the coral holobiont-the coral host, symbiotic algae, and microbiome. For example, the richness and composition of microorganisms associated with corals may be indicative of the corals' health status and thus help guide restoration activities. Potential differences in microbiomes of restoration corals due to differences in host genetics, environmental condition, or geographic location, may then influence outplant success. The objective of the present study was to characterize and compare the microbiomes of apparently healthy Acropora cervicornis genotypes that were originally collected from environmentally distinct regions of Florida's Coral Reef and sampled after residing within Mote Marine Laboratory's in situ nursery near Looe Key, FL (USA) for multiple years. By using 16S rRNA high-throughput sequencing, we described the microbial communities of 74 A. cervicornis genotypes originating from the Lower Florida Keys (n = 40 genotypes), the Middle Florida Keys (n = 15 genotypes), and the Upper Florida Keys (n = 19 genotypes). Our findings demonstrated that the bacterial communities of A. cervicornis originating from the Lower Keys were significantly different from the bacterial communities of those originating from the Upper and Middle Keys even after these corals were held within the same common garden nursery for an average of 3.4 years. However, the bacterial communities of corals originating in the Upper Keys were not significantly different from those in the Middle Keys. The majority of the genotypes, regardless of collection region, were dominated by Alphaproteobacteria, namely an obligate intracellular parasite of the genus Ca. Aquarickettsia. Genotypes from the Upper and Middle Keys also had high relative abundances of Spirochaeta bacteria. Several genotypes originating from both the Lower and Upper Keys had lower abundances of Aquarickettsia, resulting in significantly higher species richness and diversity. Low abundance of Aquarickettsia has been previously identified as a signature of disease resistance. While the low-Aquarickettsia corals from both the Upper and Lower Keys had high abundances of an unclassified Proteobacteria, the genotypes in the Upper Keys were also dominated by Spirochaeta. The results of this study suggest that the abundance of Aquarickettsia and Spirochaeta may play an important role in distinguishing bacterial communities among A. cervicornis populations and compositional differences of these bacterial communities may be driven by regional processes that are influenced by both the environmental history and genetic relatedness of the host. Additionally, the high microbial diversity of low-Aquarickettsia genotypes may provide resilience to their hosts, and these genotypes may be a potential resource for restoration practices and management.

'Candidatus Mellornella promiscua' n. gen. n. sp. (Alphaproteobacteria: Rickettsiales: Anaplasmataceae): An intracytoplasmic, hepatopancreatic, pathogen of the flatback mud crab, Eurypanopeus depressus.

Bacterial pathogens are a long-standing threat to the longevity and survival of crustacean hosts. Their presence and continuing emergence require close monitoring to understand their impact on fished, cultured, and wild crustacean populations. We describe a new bacterial pathogen belonging to the Anaplasmataceae family (Alphaproteobacteria: Rickettsiales), providing pathological, ultrastructural, phylogenetic, and genomic evidence to determine a candidate genus and species ('Candidatus Mellornella promiscua'). This bacterium was found to infect the mud crab, Eurypanopeus depressus, on the North Carolina coastline (USA) at a prevalence of 10.8%. 'Candidatus Mellornella promiscua' was often observed in co-infection with the rhizocephalan barnacle, Loxothylacus panopaei. The bacterium was only found in the hepatopancreas of the mud crab host, causing cytoplasmic hypertrophy, tubule necrosis, large plaques within the cytoplasm of the host cell, and an abundance of sex-pili. The circular genome of the bacterium is 1,013,119 bp and encodes 939 genes in total. Phylogenetically, the new bacterium branches within the Anaplasmataceae. The genome is dissimilar from other described bacteria, with 16S gene similarity observed at a maximum of 85.3% to a Wolbachia endosymbiont. We explore this novel bacterial pathogen using genomic, phylogenetic, ultrastructural, and pathological methods, discussing these results in light of current bacterial taxonomy, similarity to other bacterial pathogens, and the potential impact upon the surrounding disease ecology of the host and benthic ecosystem.

A dual endosymbiosis supports nutritional adaptation to hematophagy in the invasive tick Hyalomma marginatum.

Many animals are dependent on microbial partners that provide essential nutrients lacking from their diet. Ticks, whose diet consists exclusively on vertebrate blood, rely on maternally inherited bacterial symbionts to supply B vitamins. While previously studied tick species consistently harbor a single lineage of those nutritional symbionts, we evidence here that the invasive tick Hyalomma marginatum harbors a unique dual-partner nutritional system between an ancestral symbiont, Francisella, and a more recently acquired symbiont, Midichloria. Using metagenomics, we show that Francisella exhibits extensive genome erosion that endangers the nutritional symbiotic interactions. Its genome includes folate and riboflavin biosynthesis pathways but deprived functional biotin biosynthesis on account of massive pseudogenization. Co-symbiosis compensates this deficiency since the Midichloria genome encompasses an intact biotin operon, which was primarily acquired via lateral gene transfer from unrelated intracellular bacteria commonly infecting arthropods. Thus, in H. marginatum, a mosaic of co-evolved symbionts incorporating gene combinations of distant phylogenetic origins emerged to prevent the collapse of an ancestral nutritional symbiosis. Such dual endosymbiosis was never reported in other blood feeders but was recently documented in agricultural pests feeding on plant sap, suggesting that it may be a key mechanism for advanced adaptation of arthropods to specialized diets.

Dating Alphaproteobacteria evolution with eukaryotic fossils.

Elucidating the timescale of the evolution of Alphaproteobacteria, one of the most prevalent microbial lineages in marine and terrestrial ecosystems, is key to testing hypotheses on their co-evolution with eukaryotic hosts and Earth's systems, which, however, is largely limited by the scarcity of bacterial fossils. Here, we incorporate eukaryotic fossils to date the divergence times of Alphaproteobacteria, based on the mitochondrial endosymbiosis that mitochondria evolved from an alphaproteobacterial lineage. We estimate that Alphaproteobacteria arose ~1900 million years (Ma) ago, followed by rapid divergence of their major clades. We show that the origin of Rickettsiales, an order of obligate intracellular bacteria whose hosts are mostly animals, predates the emergence of animals for ~700 Ma but coincides with that of eukaryotes. This, together with reconstruction of ancestral hosts, strongly suggests that early Rickettsiales lineages had established previously underappreciated interactions with unicellular eukaryotes. Moreover, the mitochondria-based approach displays higher robustness to uncertainties in calibrations compared with the traditional strategy using cyanobacterial fossils. Further, our analyses imply the potential of dating the (bacterial) tree of life based on endosymbiosis events, and suggest that previous applications using divergence times of the modern hosts of symbiotic bacteria to date bacterial evolution might need to be revisited.

A human respiratory tract-associated bacterium with an extremely small genome.

Recent advances in culture-independent microbiological analyses have greatly expanded our understanding of the diversity of unculturable microbes. However, human pathogenic bacteria differing significantly from known taxa have rarely been discovered. Here, we present the complete genome sequence of an uncultured bacterium detected in human respiratory tract named IOLA, which was determined by developing a protocol to selectively amplify extremely AT-rich genomes. The IOLA genome is 303,838 bp in size with a 20.7% GC content, making it the smallest and most AT-rich genome among known human-associated bacterial genomes to our best knowledge and comparable to those of insect endosymbionts. While IOLA belongs to order Rickettsiales (mostly intracellular parasites), the gene content suggests an epicellular parasitic lifestyle. Surveillance of clinical samples provides evidence that IOLA can be predominantly detected in patients with respiratory bacterial infections and can persist for at least 15 months in the respiratory tract, suggesting that IOLA is a human respiratory tract-associated bacterium.

Cells within cells: Rickettsiales and the obligate intracellular bacterial lifestyle.

The Rickettsiales are a group of obligate intracellular vector-borne Gram-negative bacteria that include many organisms of clinical and agricultural importance, including Anaplasma spp., Ehrlichia chaffeensis, Wolbachia, Rickettsia spp. and Orientia tsutsugamushi. This Review provides an overview of the current state of knowledge of the biology of these bacteria and their interactions with host cells, with a focus on pathogenic species or those that are otherwise important for human health. This includes a description of rickettsial genomics, bacterial cell biology, the intracellular lifestyles of Rickettsiales and the mechanisms by which they induce and evade the innate immune response.

'Candidatus Sarmatiella mevalonica' endosymbiont of the ciliate Paramecium provides insights on evolutionary plasticity among Rickettsiales.

Members of the bacterial order Rickettsiales are obligatorily associated with a wide range of eukaryotic hosts. Their evolutionary trajectories, in particular concerning the origin of shared or differential traits among distant sub-lineages, are still poorly understood. Here, we characterized a novel Rickettsiales bacterium associated with the ciliate Paramecium tredecaurelia and phylogenetically related to the Rickettsia genus. Its genome encodes significant lineage-specific features, chiefly the mevalonate pathway gene repertoire, involved in isoprenoid precursor biosynthesis. Not only this pathway has never been described in Rickettsiales, it also is very rare among bacteria, though typical in eukaryotes, thus likely representing a horizontally acquired trait. The presence of these genes could enable an efficient exploitation of host-derived intermediates for isoprenoid synthesis. Moreover, we hypothesize the reversed reactions could have replaced canonical pathways for producing acetyl-CoA, essential for phospholipid biosynthesis. Additionally, we detected phylogenetically unrelated mevalonate pathway genes in metagenome-derived Rickettsiales sequences, likely indicating evolutionary convergent effects of independent horizontal gene transfer events. Accordingly, convergence, involving both gene acquisitions and losses, is highlighted as a relevant evolutionary phenomenon in Rickettsiales, possibly favoured by plasticity and comparable lifestyles, representing a potentially hidden origin of other more nuanced similarities among sub-lineages.

Molecular detection of pathogens from ectoparasites recovered from peri-domestic animals, and the first description of a Candidatus Midichloria sp. from Haemaphysalis wellingtoni from rural communities in Malaysia.

Rural communities in Malaysia have been shown to be exposed to Coxiella, Borrelia and rickettsial infections in previous seroprevalence studies. Further research is necessary to identify the actual causative agents and the potential vectors of these infections. The arthropods parasitizing peri-domestic animals in these communities may serve as the vector in transmitting arthropod-borne and zoonotic agents to the humans. Molecular screening of bacterial and zoonotic pathogens from ticks and fleas collected from dogs, cats and chickens from six rural communities in Malaysia was undertaken. These communities were made up of mainly the indigenous people of Malaysia, known as the Orang Asli, as well as settlers in oil palm plantations. The presence of Coxiella burnetii, Borrelia, and rickettsial agents, including Rickettsia and Anaplasma, was investigated by performing polymerase chain reaction (PCR) and DNA sequencing. Candidatus Rickettsia senegalensis was detected in one out of eight pools of Ctenocephalides felis fleas. A relapsing fever group Borrelia sp. was identified from one of seven Haemaphysalis hystricis ticks tested. The results from the PCR screening for Anaplasma unexpectedly revealed the presence of Candidatus Midichloria sp., a potential tick endosymbiont, in two out of fourteen Haemaphysalis wellingtoni ticks tested. C. burnetii was not detected in any of the samples tested. The findings here provide evidence for the presence of potentially novel strains of rickettsial and borrelial agents in which their impact on public health risks among the rural communities in Malaysia merit further investigation. The detection of a potential endosymbiont of ticks also suggest that the presence of tick endosymbionts in the region is not fully explored.

Abalone Withering Syndrome Disease Dynamics: Infectious Dose and Temporal Stability in Seawater.

Withering syndrome (WS) is a chronic bacterial disease that affects numerous northeastern Pacific abalone Haliotis spp. The causative agent of WS is an obligate intracellular Rickettsiales-like bacterium (WS-RLO) that remains unculturable, thereby limiting our understanding of WS disease dynamics. The objectives of our study were to (1) determine the temporal stability of WS-RLO DNA outside of its abalone host in 14°C and 18°C seawater, (2) develop a standardized protocol for exposing abalones to known concentrations of WS-RLO DNA, and (3) calculate the dose of WS-RLO DNA required to generate 50% infection prevalence (ID50) in the highly cultured red abalone Haliotis rufescens. The WS-RLO stability trials were conducted in October 2016, February 2017, and June 2017. A quantitative PCR (qPCR) analysis was used to quantify bacterial DNA for 7 d in seawater collected at an abalone farm in southern California, where the pathogen is now endemic. For all trials and temperature treatments, WS-RLO DNA was unstable in seawater for longer than 2 d. To determine an ID50, groups of uninfected juvenile red abalone were subjected to 3-h bath exposures with four concentrations of WS-RLO at 0, 103 , 104 , and 105 DNA copies/mL. Abalone feces were tested biweekly for the presence of WS-RLO DNA, and abalone tissues were sampled 9 weeks postinfection for histological and qPCR analyses. The ID50 results indicated that our protocol was successful in generating WS-RLO infections; a pathogen dose of 2.3 × 103 DNA copies/mL was required to generate a 50% infection prevalence in red abalone tissue. These findings are critical components of disease dynamics that will help assess WS transmission risk within and among abalone populations and facilitate appropriate management and restoration strategies for both wild and cultured abalone species in WS-endemic areas.

Molecular identification of protozoal and bacterial organisms in domestic animals and their infesting ticks from north-eastern Algeria.

A molecular survey was undertaken to determine the presence of protozoal and bacterial organisms in 120 ticks and 87 blood samples collected from mammals in north-eastern Algeria. Eight tick species were morphologically identified including 70 Rhipicephalus (Boophilus) annulatus, 23 Rhipicephalus bursa, five Rhipicephalus sanguineus sensu lato, 11 Hyalomma impeltatum, five Hyalomma scupense, two Hyalommma marginatum, one Hyalomma anatolicum and three Ixodes ricinus. Quantitative PCR screening of the ticks showed that Theileria annulata, "Candidatus Ehrlichia urmitei", Theileria buffeli and Anaplasma platys were detected in Rh. annulatus. Rickettsia massiliae and Anaplasma ovis were detected in Rh. sanguineus s.l. and Rh. bursa. Rickettsia aeschlimannii was detected in Hy. marginatum, Hy. scupense and Hy. impeltatum. Finally, "Candidatus Rickettsia barbariae" was detected in Rh. bursa. In the screening blood samples, Theileria equi, T.annulata, T. buffeli, Babesia bovis, Anaplasma marginale, A. ovis and Borrelia spp. were detected in cattle. Theileria ovis, T. annulata, and A. ovis were detected in sheep. In addition, A. ovis and T. equi were detected in goats and equidea respectively. In this study, T. equi and "Candidatus Rickettsia barbariae" were identified for the first time in Algeria as well as potential new species of Ehrlichia and Anaplasma. Although molecular detection does not indicate vector/reservoir competence when investigating ticks removed from animals, this study expands the knowledge of the microorganisms detected in ticks in north-east of Algeria.

Epidemiology and Diversity of Rickettsiales Bacteria in Humans and Animals in Jiangsu and Jiangxi provinces, China.

Diseases caused by Rickettsiales bacteria are a global public health problem. To better understand the diversity and origins of Rickettsiales infection in humans and animals, we sampled 134 febrile patients, 173 rodents and 43 shrews, as well as 358 ticks, from two cities in Jiangsu and Jiangxi provinces, China. Our data revealed a relatively high prevalence of scrub typhus cases in both localities. In addition, both serological tests and genetic analysis identified three patients infected with Anaplasma bovis, Rickettsia monacensis, and Orientia tsutsugamushi bacteria. Molecular epidemiological investigation revealed the co-circulation of multiple species of Rickettsiales bacteria in small mammals and ticks in both provinces, potentially including novel bacterial species. In sum, these data demonstrate the ongoing importance of Rickettsiales infection in China and highlight the need for the regular surveillance of local arthropods, mammals and humans.

Phylogenetic, genomic, and biogeographic characterization of a novel and ubiquitous marine invertebrate-associated Rickettsiales parasite, Candidatus Aquarickettsia rohweri, gen. nov., sp. nov.

Bacterial symbionts are integral to the health and homeostasis of invertebrate hosts. Notably, members of the Rickettsiales genus Wolbachia influence several aspects of the fitness and evolution of their terrestrial hosts, but few analogous partnerships have been found in marine systems. We report here the genome, phylogenetics, and biogeography of a ubiquitous and novel Rickettsiales species that primarily associates with marine organisms. We previously showed that this bacterium was found in scleractinian corals, responds to nutrient exposure, and is associated with reduced host growth and increased mortality. This bacterium, like other Rickettsiales, has a reduced genome indicative of a parasitic lifestyle. Phylogenetic analysis places this Rickettsiales within a new genus we define as "Candidatus Aquarickettsia." Using data from the Earth Microbiome Project and SRA databases, we also demonstrate that members of "Ca. Aquarickettsia" are found globally in dozens of invertebrate lineages. The coral-associated "Candidatus A. rohweri" is the first finished genome in this new clade. "Ca. A. rohweri" lacks genes to synthesize most sugars and amino acids but possesses several genes linked to pathogenicity including Tlc, an antiporter that exchanges host ATP for ADP, and a complete Type IV secretion system. Despite its inability to metabolize nitrogen, "Ca. A. rohweri" possesses the NtrY-NtrX two-component system involved in sensing and responding to extracellular nitrogen. Given these data, along with visualization of the parasite in host tissues, we hypothesize that "Ca. A. rohweri" reduces coral health by consuming host nutrients and energy, thus weakening and eventually killing host cells. Last, we hypothesize that nutrient enrichment, which is increasingly common on coral reefs, encourages unrestricted growth of "Ca. A. rohweri" in its host by providing abundant N-rich metabolites to be scavenged.

Molecular detection and genetic characterization of the potentially pathogenic Coxiella burnetii and the endosymbiotic Candidatus Midichloria mitochondrii in ticks infesting camels (Camelus dromedarius) from Tunisia.

Tick-borne bacteria are considered to be emerging in camels, but data about their occurrence in Tunisian dromedaries and their infesting ticks remain scarce. In this study, 412 camel blood samples and 327 partially engorged ticks were collected and tested for the presence of Coxiella burnetii and/or related strains, and Rickettsiales bacteria. Coxiella burnetii was detected in 9 Hyalomma impeltatum and 3 H. dromedarii with an overall prevalence rate of 3.6% (12/327). Candidatus Midichloria mitochondrii DNA was identified in 16 H. impeltatum and 10 H. dromedarii with an overall prevalence rate of 8% (26/327). Six ticks (2%) were found to be co-infected with these two bacteria. No positive DNA camel blood sample was observed for both bacteria. Genotyping and phylogenetic analysis of obtained C. burnetii partial sequences based on the IS1111 and htpB genes revealed 99-100% similarity to the pathogenic C. burnetii strains isolated from humans, ruminants and ticks, and that were genetically distant to those isolated from all endosymbiotic related strains (Coxiella-like bacteria). The analysis of the rickettsial partial sequences of the 16S rRNA gene showed 100% similarity to Ca. M. mitochondrii strains infecting Ixodid ticks and dogs. This is the first time that C. burnetii and Ca. M. mitochondrii have been detected in ticks from Tunisia, which raises the possibility of the involvement of Hyalomma tick species in the active diffusion of these bacteria among camels, other domestic animals and humans.

Tick-borne pathogens Bartonella spp., Borrelia burgdorferi sensu lato, Coxiella burnetii and Rickettsia spp. may trigger endocarditis.

BACKGROUND: Infections caused by tick-borne pathogens such as Bartonella spp., Borrelia burgdorferi s.l., Coxiella burnetii, and Rickettsia spp. are capable of causing serious lesions of the mitral and aortic valves, leading to a need for valve replacement. OBJECTIVES: The aim of the study was to determine whether such cases are sporadic or frequent. An additional goal was to establish effective diagnostic methods to detect these infections. MATERIAL AND METHODS: The study involved 148 patients undergoing valve replacement. Blood samples were drawn for serological testing. Samples of the removed mitral and aortic valves were tested with polymerase chain reaction and immunohistochemical staining. RESULTS: Specific antibodies to Bartonella spp. were detected in 47 patients (31.7%) and in 1 of the healthy controls (1%) (p < 0.05). Antibodies to B. burgdorferi spirochetes were found in 18 of the patients (12.2%) and in 6 blood donors from the control group (5.8%) (p < 0.1). Antibodies to Rickettsia spp. were detected in 12 (8.1%) and to C. burnetii phase I and II antigens in the serum of 1 patient. All the participants in the control group were seronegative to C. burnetii and Rickettsia spp. antigens. Polymerase chain reaction (PCR) tests for detection of Bartonella spp., B. burgdorferi s.l., C. burnetii and Rickettsia spp. DNA in the valve samples were all negative. Inflammation foci with mononuclear lymphoid cells in the aortic and mitral valves were seen in sections stained with hematoxiline and eozine. In sections dyed using the indirect immunofluorescence method with hyperimmune sera, Bartonella spp. and Rickettsia spp. were found. CONCLUSIONS: The results obtained indicate that laboratory diagnostics for patients with heart disorders should be expanded to include tests detecting tick-borne zoonoses such as bartonelloses, Lyme borreliosis, rickettsioses and Q fever.

Tissue tropism and metabolic pathways of Midichloria mitochondrii suggest tissue-specific functions in the symbiosis with Ixodes ricinus.

A wide range of arthropod species harbour bacterial endosymbionts in various tissues, many of them playing important roles in the fitness and biology of their hosts. In several cases, many different symbionts have been reported to coexist simultaneously within the same host and synergistic or antagonistic interactions can occur between them. While the associations with endosymbiotic bacteria have been widely studied in many insect species, in ticks such interactions are less investigated. The females and immatures of Ixodes ricinus (Ixodidae), the most common hard tick in Europe, harbour the intracellular endosymbiont "Candidatus Midichloria mitochondrii" with a prevalence up to 100%, suggesting a mutualistic relationship. Considering that the tissue distribution of a symbiont might be indicative of its functional role in the physiology of the host, we investigated M. mitochondrii specific localization pattern and the corresponding abundance in selected organs of I. ricinus females. We paired these experiments with in silico analysis of the metabolic pathways of M. mitochondrii, inferred from the available genome sequence, and additionally compared the presence of these pathways in seven other symbionts commonly harboured by ticks to try to obtain a comparative understanding of their biological effects on the tick hosts. M. mitochondrii was found to be abundant in ovaries and tracheae of unfed I. ricinus, and in ovaries, Malpighian tubules and salivary glands of semi-engorged females. These results, together with the in silico metabolic reconstruction allow to hypothesize that the bacterium could play multiple tissue-specific roles in the host, both enhancing the host fitness (supplying essential nutrients, enhancing the reproductive fitness, helping in the anti-oxidative defence, in the energy production and in the maintenance of homeostasis and water balance) and/or for ensuring its presence in the host population (nutrients acquisition, vertical and horizontal transmission). The ability of M. mitochondrii to colonize different tissues allows to speculate that distinctive sub-populations may display different specializations in accordance with tissue tropism. Our hypotheses should be corroborated with future nutritional and physiological experiments for a better understanding of the mechanisms underlying this symbiotic interaction.

Deianiraea, an extracellular bacterium associated with the ciliate Paramecium, suggests an alternative scenario for the evolution of Rickettsiales.

Rickettsiales are a lineage of obligate intracellular Alphaproteobacteria, encompassing important human pathogens, manipulators of host reproduction, and mutualists. Here we report the discovery of a novel Rickettsiales bacterium associated with Paramecium, displaying a unique extracellular lifestyle, including the ability to replicate outside host cells. Genomic analyses show that the bacterium possesses a higher capability to synthesise amino acids, compared to all investigated Rickettsiales. Considering these observations, phylogenetic and phylogenomic reconstructions, and re-evaluating the different means of interaction of Rickettsiales bacteria with eukaryotic cells, we propose an alternative scenario for the evolution of intracellularity in Rickettsiales. According to our reconstruction, the Rickettsiales ancestor would have been an extracellular and metabolically versatile bacterium, while obligate intracellularity would have evolved later, in parallel and independently, in different sub-lineages. The proposed new scenario could impact on the open debate on the lifestyle of the last common ancestor of mitochondria within Alphaproteobacteria.