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Cette contribution traite du développement de la jonction interpropositionnelle en français langue seconde. Elle ne considère pas uniquement ces cas où plusieurs propositions complètes sont reliées l’une à l’autre, mais se concentre également sur de différents types de relations interprédicationnelles , produits à des niveaux plus internes de la proposition (comme le prédicat, avec ou sans arguments). L’hypothèse de base postule que le processus d’appropriation des relations syntaxiques interpropositionnelles est déterminé par la complexité relative de celles-ci. Pour évaluer la validité de ce postulat, nous nous appuyerons sur la théorie fonctionnelle de la Grammaire du Rôle et de la Référence (Role and Reference Grammar ou RRG; Van Valin & LaPolla, 1997; Van Valin, 2005). Ce modèle avance un certain nombre d’hypothèses sur le plan du développement syntaxique de la jonction interpropositionnelle, dont nous essaierons de vérifier la validité pour l’acquisition d’une langue seconde, plus spécifiquement le français. Les données que nous analyserons sont des histoires de la grenouille (Mayer, 1969) produites à l’oral par des élèves de l’école secondaire en Flandre, ayant pour langue première le néerlandais. Les quarante apprenants impliqués dans cette recherche, qui fait partie d’un projet plus large, sont sous- divisés en quatre groupes selon leur niveau général de compétence en français. Nous comparerons également leurs productions à celles de dix locuteurs natifs du français. Cette première analyse syntaxique confirme une partie des hypothèses proposées par la RRG et en infirme d’autres; il faut remarquer, néanmoins, que cette déviation du principe général proposé est due en certains cas à la nature orale et quasi-spontanée des textes analysés. Dans le cadre du projet entier, ce facteur sera analysé en plus de détail.

La jonction propositionnelle, ou le processus visant à combiner plusieurs propositions dans un énoncé complexe, est un phénomène largement décrit en linguistique française (Bronckart & Schneuwly 1984, Clark 1998, Kern 2000). Dans un contexte d’acquisition du français comme langue seconde (FL2), en particulier, la complexification propositionnelle est un des objets de recherche centraux (Benazzo 2004, Kerr-Barnes 1998, Véronique 2005, Welcomme 2013). Certaines études axées sur le développement global d’apprenants du français exploitent en outre la jonction de propositions comme un indice essentiel pour évaluer la progression dans la maîtrise de la langue seconde (Bartning & Kirchmeyer 2003, Schlyter 2003, Bartning & Schlyter 2004). Le cadre d’analyse généralement adopté pour décrire le développement de la jonction propositionnelle est la bipartition traditionnelle entre la coordination d’une part (Georges est malade et il ne veut pas manger) et la subordination d’autre part (Georges dit qu’il est malade, complétive, ou George ne veut pas manger parce qu’il est malade, adverbiale). Pourtant, cette opposition binaire ne rend pas compte de l’écart très important entre le moment d’acquisition d’énoncés comme Quand il pleut, je prends un parapluie et celui de phrases composées comme Je connais l’homme dont tu me parles. De plus, elle ne permet pas la description adéquate de types de jonction propositionnelle tels que Il veut laver la voiture et J’essayerai de venir à ta fête. Pour permettre une approche plus affinée de la jonction propositionnelle, la Grammaire du Rôle et de la Référence (Role and Reference Grammar ou RRG ; Van Valin & LaPolla 1997, Van Valin 2005) avance un troisième type de relation syntaxique entre propositions, la cosubordination. Nous évaluerons l’apport de la tripartition proposée pour l’acquisition d’une L2 en analysant le développement progressif de la jonction propositionnelle chez des apprenants néerlandophones de FL2. La présente contribution se pose l’objectif de mettre en évidence les insuffisances de la saisie de la jonction propositionnelle au moyen de la bipartition traditionnelle corrdination/subordina-tion. Elle présente l’alternative avancée par la RRG, sur laquelle elle s’appuie pour décrire le processus d’appropriation de la connexion de propositions en FL2 par des apprenants néerlandophones. Elle discute finalement l’apport de la tripartition pour une meilleure appréhension du processus de jonction et sa pertinence pour la recherche en acquisition de langues étrangères.

Objectives: The development of External Quality Assessment Schemes (EQAS) for clinical flow cytometry (FCM) is challenging in the context of rare (immunological) diseases. Here, we introduce a novel EQAS monitoring the primary immunodeficiency Orientation Tube (PIDOT), developed by EuroFlow, in both a 'wet' and 'dry' format. This EQAS provides feedback on the quality of individual laboratories (i.e., accuracy, reproducibility and result interpretation), while eliminating the need for sample distribution., Methods: In the wet format, marker staining intensities (MedFIs) within landmark cell populations in PIDOT analysis performed on locally collected healthy control (HC) samples, were compared to EQAS targets. In the dry format, participants analyzed centrally distributed PIDOT flow cytometry data (n=10)., Results: We report the results of six EQAS rounds across 20 laboratories in 11 countries. The wet format (212 HC samples) demonstrated consistent technical performance among laboratories (median %rCV on MedFIs=34.5 %; average failure rate 17.3 %) and showed improvement upon repeated participation. The dry format demonstrated effective proficiency of participants in cell count enumeration (range %rCVs 3.1-7.1 % for the major lymphoid subsets), and in identifying lymphoid abnormalities (79.3 % alignment with reference)., Conclusions: The PIDOT-EQAS allows laboratories, adhering to the standardized EuroFlow approach, to monitor interlaboratory variations without the need for sample distribution, and provides them educational support to recognize rare clinically relevant immunophenotypic patterns of primary immunodeficiencies (PID). This EQAS contributes to quality improvement of PID diagnostics and can serve as an example for future flow cytometry EQAS in the context of rare diseases., (© 2024 Walter de Gruyter GmbH, Berlin/Boston.)

Common variable immunodeficiency (CVID) is a heterogeneous primary immunodeficiency (PID) characterized by an impaired immunoglobulin production, in association with an increased susceptibility to infections and a diversity of clinical manifestations. This narrative review summarizes immunophenotypic abnormalities in CVID patients and their relevance for diagnosis and disease classification. A comprehensive search across four databases - PubMED, Web of Science, EMBASE and Google Scholar - yielded 170 relevant studies published between 1988 and April 31, 2023. Over the past decades, the role of immunophenotyping in CVID diagnosis has become evident by identifying "hallmark" immunophenotypic aberrancies in patient subsets, with some now integrated in the consensus diagnostic criteria. Furthermore, the role of immunophenotyping in subclassifying CVID in relation to clinical presentation and prognosis has been extensively studied. Certain immunophenotypic patterns consistently correlate with clinical manifestations and/or subsets of CVID, particularly those associated with noninfectious complications (i.e. low switched memory B cells, shifts in follicular helper T cell subsets, low naïve CD4 + T cells, low regulatory T cells, and expansion of CD21low B cells, often associated with autoimmunity and/or splenomegaly). Also, efforts to associate subset levels of innate immune cells, such as Natural Killer (NK) cells, invariant (i)NKT cells, innate lymphoid cells (ILCs), and dendritic cells (DCs) to CVID complications are evident albeit in a lesser degree. However, inconsistencies regarding the role of flow cytometry in classification and prognosis persist, reflecting the disease complexity, but probably also cohort variations and methodological differences between published studies. This underscores the need for collaborative efforts to integrate emerging concepts, such as standardized flow cytometry and computational tools, for a more precise CVID classification approach. Additionally, recent studies suggest a potential value of (epi)genetic-based molecular assays to this effort.

Rickettsia, a genus of obligate intracellular bacteria, includes species that cause significant human diseases. This study challenges previous claims that the Leucine-973 residue in the RNA polymerase beta subunit is the primary determinant of rifampin resistance in Rickettsia. We investigated a previously untested Rickettsia species, R. lusitaniae, from the Transitional group and found it susceptible to rifampin, despite possessing the Leu-973 residue. Interestingly, we observed the conservation of this residue in several rifampin-susceptible species across most Rickettsia phylogenetic groups. Comparative genomics revealed potential alternative resistance mechanisms, including additional amino acid variants that could hinder rifampin binding and genes that could facilitate rifampin detoxification through efflux pumps. Importantly, the evolutionary history of Rickettsia genomes indicates that the emergence of natural rifampin resistance is phylogenetically constrained within the genus, originating from ancient genetic features shared among a unique set of closely related Rickettsia species. Phylogenetic patterns appear to be the most reliable predictors of natural rifampin resistance, which is confined to a distinct monophyletic subclade known as Massiliae. The distinctive features of the RNA polymerase beta subunit in certain untested Rickettsia species suggest that R. raoultii, R. amblyommatis, R. gravesii, and R. kotlanii may also be naturally rifampin-resistant species., (© 2024 The Author(s). MicrobiologyOpen published by John Wiley & Sons Ltd.)

Presence of minimal residual disease (MRD), detected by flow cytometry, is an important prognostic biomarker in the management of B-cell precursor acute lymphoblastic leukemia (BCP-ALL). However, data-analysis remains mainly expert-dependent. In this study, we designed and validated an Automated Gating & Identification (AGI) tool for MRD analysis in BCP-ALL patients using the two tubes of the EuroFlow 8-color MRD panel. The accuracy, repeatability, and reproducibility of the AGI tool was validated in a multicenter study using bone marrow follow-up samples from 174 BCP-ALL patients, stained with the EuroFlow BCP-ALL MRD panel. In these patients, MRD was assessed both by manual analysis and by AGI tool supported analysis. Comparison of MRD levels obtained between both approaches showed a concordance rate of 83%, with comparable concordances between MRD tubes (tube 1, 2 or both), treatment received (chemotherapy versus targeted therapy) and flow cytometers (FACSCanto versus FACSLyric). After review of discordant cases by additional experts, the concordance increased to 97%. Furthermore, the AGI tool showed excellent intra-expert concordance (100%) and good inter-expert concordance (90%). In addition to MRD levels, also percentages of normal cell populations showed excellent concordance between manual and AGI tool analysis. We conclude that the AGI tool may facilitate MRD analysis using the EuroFlow BCP-ALL MRD protocol and will contribute to a more standardized and objective MRD assessment. However, appropriate training is required for the correct analysis of MRD data., (© 2023 The Authors. Cytometry Part B: Clinical Cytometry published by Wiley Periodicals LLC on behalf of International Clinical Cytometry Society.)

Tick-borne bacteria of the genera Ehrlichia and Anaplasma cause several emerging human infectious diseases worldwide. In this study, we conduct an extensive survey for Ehrlichia and Anaplasma infections in the rainforests of the Amazon biome of French Guiana. Through molecular genetics and metagenomics reconstruction, we observe a high indigenous biodiversity of infections circulating among humans, wildlife, and ticks inhabiting these ecosystems. Molecular typing identifies these infections as highly endemic, with a majority of new strains and putative species specific to French Guiana. They are detected in unusual rainforest wild animals, suggesting they have distinctive sylvatic transmission cycles. They also present potential health hazards, as revealed by the detection of Candidatus Anaplasma sparouinense in human red blood cells and that of a new close relative of the human pathogen Ehrlichia ewingii, Candidatus Ehrlichia cajennense, in the tick species that most frequently bite humans in South America. The genome assembly of three new putative species obtained from human, sloth, and tick metagenomes further reveals the presence of major homologs of Ehrlichia and Anaplasma virulence factors. These observations converge to classify health hazards associated with Ehrlichia and Anaplasma infections in the Amazon biome as distinct from those in the Northern Hemisphere., (© 2024. The Author(s).)

Since the discovery of natural malaria vector populations infected by the endosymbiont bacterium Wolbachia , a renewed interest has arisen for using this bacterium as an alternative for malaria control. Among naturally infected mosquitoes, Anopheles moucheti , a major malaria mosquito in Central Africa, exhibits one of the highest prevalences of Wolbachia infection. To better understand whether this maternally inherited bacterium could be used for malaria control, we investigated Wolbachia influence in An. moucheti populations naturally infected by the malaria parasite Plasmodium falciparum . To this end, we collected mosquitoes in a village from Cameroon, Central Africa, where this mosquito is the main malaria vector. We found that the prevalence of Wolbachia bacterium was almost fixed in the studied mosquito population, and was higher than previously recorded. We also quantified Wolbachia in whole mosquitoes and dissected abdomens, confirming that the bacterium is also elsewhere than in the abdomen, but at lower density. Finally, we analyzed the association of Wolbachia presence and density on P. falciparum infection. Wolbachia density was slightly higher in mosquitoes infected with the malaria parasite than in uninfected mosquitoes. However, we observed no correlation between the P. falciparum and Wolbachia densities. In conclusion, our study indicates that naturally occurring Wolbachia infection is not associated to P. falciparum development within An. moucheti mosquitoes., Competing Interests: None declared., (© 2023 The Authors. Evolutionary Applications published by John Wiley & Sons Ltd.)

Hydras are freshwater cnidarians widely used as a biological model to study different questions such as senescence or phenotypic plasticity but also tumoral development. The spontaneous tumors found in these organisms have been so far described in two female lab strains domesticated years ago (Hydra oligactis and Pelmatohydra robusta) and the extent to which these tumors can be representative of tumors within the diversity of wild hydras is completely unknown. In this study, we examined individuals isolated from recently sampled wild strains of different sex and geographical origin, which have developed outgrowths looking like tumors. These tumefactions have common features with the tumors previously described in lab strains: are composed of an accumulation of abnormal cells, resulting in a similar enlargement of the tissue layers. However, we also found diversity within these new types of tumors. Indeed, not only females, but also males seem prone to form these tumors. Finally, the microbiota associated to these tumors is different from the one involved in the previous lineages exhibiting tumors. We found that tumorous individuals hosted yet undescribed Chlamydiales vacuoles. This study brings new insights into the understanding of tumor susceptibility and diversity in brown hydras from different origins., (© 2023. The Author(s).)

Background: Ticks and tick-borne pathogens significantly impact both human and animal health and therefore are of major concern to the scientific community. Knowledge of tick-borne pathogens is crucial for prescription of mitigation measures. In Africa, much research on ticks has focused on domestic animals. Little is known about ticks and their pathogens in wild habitats and wild animals like the endangered chimpanzee, our closest relative., Methods: In this study, we collected ticks in the forested habitat of a community of 100 chimpanzees living in Kibale National Park, Western Uganda, and assessed how their presence and abundance are influenced by environmental factors. We used non-invasive methods of flagging the vegetation and visual search of ticks both on human team members and in chimpanzee nests. We identified adult and nymph ticks through morphological features. Molecular techniques were used to detect and identify tick-borne piroplasmids and bacterial pathogens., Results: A total of 470 ticks were collected, which led to the identification of seven tick species: Haemaphysalis parmata (68.77%), Amblyomma tholloni (20.70%), Ixodes rasus sensu lato (7.37%), Rhipicephalus dux (1.40%), Haemaphysalis punctaleachi (0.70%), Ixodes muniensis (0.70%) and Amblyomma paulopunctatum (0.35%). The presence of ticks, irrespective of species, was influenced by temperature and type of vegetation but not by relative humidity. Molecular detection revealed the presence of at least six genera of tick-borne pathogens (Babesia, Theileria, Borrelia, Cryptoplasma, Ehrlichia and Rickettsia). The Afrotopical tick Amblyomma tholloni found in one chimpanzee nest was infected by Rickettsia sp., Conclusions: In conclusion, this study presented ticks and tick-borne pathogens in a Ugandan wildlife habitat whose potential effects on animal health remain to be elucidated., (© 2023. The Author(s).)

Background: Trypanosomes are protozoan parasites of vertebrates that are of medical and veterinary concern. A variety of blood-feeding invertebrates have been identified as vectors, but the role of ticks in trypanosome transmission remains unclear., Methods: In this study, we undertook extensive molecular screening for the presence and genetic diversity of trypanosomes in field ticks., Results: Examination of 1089 specimens belonging to 28 tick species from Europe and South America led to the identification of two new trypanosome strains. The prevalence may be as high as 4% in tick species such as the castor bean tick Ixodes ricinus, but we found no evidence of transovarial transmission. Further phylogenetic analyses based on 18S rRNA, EF1-α, hsp60 and hsp85 gene sequences revealed that different tick species, originating from different continents, often harbour phylogenetically related trypanosome strains and species. Most tick-associated trypanosomes cluster in a monophyletic clade, the Trypanosoma pestanai clade, distinct from clades of trypanosomes associated with transmission by other blood-feeding invertebrates., Conclusions: These observations suggest that ticks may be specific arthropod hosts for trypanosomes of the T. pestanai clade. Phylogenetic analyses provide further evidence that ticks may transmit these trypanosomes to a diversity of mammal species (including placental and marsupial species) on most continents., (© 2023. The Author(s).)

Introduction: Multiparameter flow cytometry (FCM) immunophenotyping is an important tool in the diagnostic screening and classification of primary immunodeficiencies (PIDs). The EuroFlow Consortium recently developed the PID Orientation Tube (PIDOT) as a universal screening tool to identify lymphoid-PID in suspicious patients. Although PIDOT can identify different lymphoid-PIDs with high sensitivity, clinical validation in a broad spectrum of patients with suspicion of PID is missing. In this study, we investigated the diagnostic performance of PIDOT, as part of the EuroFlow diagnostic screening algorithm for lymphoid-PID, in a daily practice at a tertiary reference center for PID., Methods: PIDOT was tested in 887 consecutive patients suspicious of PID at the Ghent University Hospital, Belgium. Patients were classified into distinct subgroups of lymphoid-PID vs. non-PID disease controls (non-PID DCs), according to the IUIS and ESID criteria. For the clinical validation of PIDOT, comprehensive characterization of the lymphoid defects was performed, together with the identification of the most discriminative cell subsets to distinguish lymphoid-PID from non-PID DCs. Next, a decision-tree algorithm was designed to guide subsequent FCM analyses., Results: The mean number of lymphoid defects detected by PIDOT in blood was 2.87 times higher in lymphoid-PID patients vs. non-PID DCs (p < 0.001), resulting in an overall sensitivity and specificity of 87% and 62% to detect severe combined immunodeficiency (SCID), combined immunodeficiency with associated or syndromic features (CID), immune dysregulation disorder (ID), and common variable immunodeficiency (CVID). The most discriminative populations were total memory and switched memory B cells, total T cells, TCD4+cells, and naive TCD4+cells, together with serum immunoglobulin levels. Based on these findings, a decision-tree algorithm was designed to guide further FCM analyses, which resulted in an overall sensitivity and specificity for all lymphoid-PIDs of 86% and 82%, respectively., Conclusion: Altogether, our findings confirm that PIDOT is a powerful tool for the diagnostic screening of lymphoid-PID, particularly to discriminate (S)CID, ID, and CVID patients from other patients suspicious of PID. The combination of PIDOT and serum immunoglobulin levels provides an efficient guide for further immunophenotypic FCM analyses, complementary to functional and genetic assays, for accurate PID diagnostics., Competing Interests: JD and AO each report being two of the inventors on the EuroFlow-owned patent PCT/NL 2015/050762 (diagnosis of primary immunodeficiencies). The Infinicyt software is based on intellectual property (IP) of the EuroFlow laboratories (University of Salamanca in Spain and Federal University of Rio de Janeiro in Brazil) and the scientific input of other EuroFlow members. All abovementioned intellectual property and related patents are licensed to Cytognos (Salamanca, ES) and BD Biosciences (San José, CA), for which companies pay royalties to the EuroFlow Consortium. These royalties are exclusively used for continuation of the EuroFlow collaboration and sustainability of the EuroFlow Consortium. JD and AO report an Educational Services Agreement from BD Biosciences and a Scientific Advisory Agreement from Cytognos; all related fees and honoraria are for the involved university departments at Leiden University Medical Center and University of Salamanca. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Neirinck, Emmaneel, Buysse, Philippé, Van Gassen, Saeys, Bossuyt, De Buyser, Burg, Pérez-Andrés, Orfao, van Dongen, Lambrecht, Kerre, Hofmans, Haerynck and Bonroy.)

We report a case of unusual human anaplasmosis in the Amazon rainforest of French Guiana. Molecular typing demonstrated that the pathogen is a novel Anaplasma species, distinct to all known species, and more genetically related to recently described Anaplasma spp. causing infections in rainforest wild fauna of Brazil.

Symbiosis with vitamin-provisioning microbes is essential for the nutrition of animals with some specialized feeding habits. While coevolution favors the interdependence between symbiotic partners, their associations are not necessarily stable: Recently acquired symbionts can replace ancestral symbionts. In this study, we demonstrate successful replacement by Francisella-like endosymbionts (-LE), a group of B-vitamin-provisioning endosymbionts, across tick communities driven by horizontal transfers. Using a broad collection of Francisella-LE-infected tick species, we determined the diversity of Francisella-LE haplotypes through a multi-locus strain typing approach and further characterized their phylogenetic relationships and their association with biological traits of their tick hosts. The patterns observed showed that Francisella-LE commonly transfer through similar ecological networks and geographic distributions shared among different tick species and, in certain cases, through preferential shuffling across congeneric tick species. Altogether, these findings reveal the importance of geographic, ecological, and phylogenetic proximity in shaping the replacement pattern in which new nutritional symbioses are initiated., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

The symbiont-associated (SA) environmental package is a new extension to the minimum information about any (x) sequence (MIxS) standards, established by the Parasite Microbiome Project (PMP) consortium, in collaboration with the Genomics Standard Consortium. The SA was built upon the host-associated MIxS standard, but reflects the nestedness of symbiont-associated microbiota within and across host-symbiont-microbe interactions. This package is designed to facilitate the collection and reporting of a broad range of metadata information that apply to symbionts such as life history traits, association with one or multiple host organisms, or the nature of host-symbiont interactions along the mutualism-parasitism continuum. To better reflect the inherent nestedness of all biological systems, we present a novel feature that allows users to co-localize samples, to nest a package within another package, and to identify replicates. Adoption of the MIxS-SA and of the new terms will facilitate reports of complex sampling design from a myriad of environments., (© 2022. The Author(s).)

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., Competing Interests: MB, AF, YG, TN, FC, EO, AG, AP, BM, CB, AC, DS, OD No competing interests declared, (© 2021, Buysse et al.)

Q fever is a widespread zoonotic disease caused by the intracellular bacterium Coxiella burnetii. While transmission is primarily but not exclusively airborne, ticks are usually thought to act as vectors on the basis of early microscopy studies. However, recent observations revealed that endosymbionts of ticks have been commonly misidentified as C. burnetii, calling the importance of tick-borne transmission into question. In this study, we re-evaluated the vector competence of the African soft tick Ornithodoros moubata for an avirulent strain of C. burnetii. To this end, we used an artificial feeding system to initiate infection of ticks, specific molecular tools to monitor further infections, and culture assays in axenic and cell media to check for the viability of C. burnetii excreted by ticks. We observed typical traits associated with vector competence: The exposure to an infected blood meal resulted in viable and persistent infections in ticks, trans-stadial transmissions of infection from nymphs to adults and the ability of adult ticks to transmit infectious C. burnetii. However, in contrast to early studies, we found that infection differed substantially between tick organs. In addition, while adult female ticks were infected, we did not observe C. burnetii in eggs, suggesting that transovarial transmission is not effective. Finally, we detected only a sporadic presence of C. burnetii DNA in tick faeces, but no living bacterium was further isolated in culture assays, suggesting that excretion in faeces is not a common mode of transmission in O. moubata., Competing Interests: The authors have declared that no competing interests exist.

Bone marrow-derived multipotent mesenchymal stromal cells (BM-MSCs) are non-haematopoietic cells present in the bone marrow stroma. They have the potential to modulate immune responses and exhibit a capacity to promote immune tolerance. Although the efficacy of immunosuppressive drugs has improved significantly, thereby ameliorating renal graft outcome, the use of these drugs still carries an increased risk of malignancies and opportunistic infections, and sometimes fail to prevent chronic allograft rejection or recurrence of the original kidney disease. As such, there is strong interest in ways to induce immune tolerance and thereby tempering or avoiding conventional immunosuppressive drugs. Cellular immunomodulation by MSCs can create a new way to induce transplant tolerance. This review will give a critical overview of the use of BM-MSCs as a cell-based immunosuppressive therapy in kidney transplant recipients. In vitro studies revealed several mechanisms that can clarify the immunomodulatory potential of BM-MSCs. Several clinical studies showed that BM-MSCs can modulate T-cell proliferation and can alter the ratio of T-cell subsets, favoring immune tolerance. However, this immunomodulation was often not associated with better clinical outcome during follow-up when compared to control groups. Some clinical studies found that BM-MSCs allow a reduction in dose of conventional immunosuppressive drugs and prevent acute graft dysfunction. Most clinical studies emphasized that BM-MSC infusion was safe. This review suggests that the use of BM-MSCs as cell-based immunosuppression therapy in kidney transplant recipients has potential, however some caution regarding their clinical use is appropriate. Mechanisms by which BM-MSCs induce transplant tolerance and factors that can alter their functionality need to be analyzed in more detail before clinical use., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Lyme disease (LD) and relapsing fevers (RF) are vector-borne diseases caused by bacteria of the Borrelia genus. Here, we report on the widespread infection by a non-described Borrelia species in passerine-associated ticks in tropical rainforests of French Guiana, South America. This novel Borrelia species is common in two tick species, Amblyomma longirostre and A. geayi, which feed on a broad variety of neotropical mammal and bird species, including migratory species moving to North America. The novel Borrelia species is divergent from the LD and RF species, and is more closely related to the reptile- and echidna-associated Borrelia group that was recently described. Genome sequencing showed that this novel Borrelia sp. has a relatively small genome consisting of a 0.9-Mb-large chromosome and an additional 0.3 Mb dispersed on plasmids. It harbors an RF-like genomic organization but with a unique mixture of LD- and RF-specific genes, including genes used by RF Borrelia for the multiphasic antigen-switching system and a number of immune-reactive protein genes used for the diagnosis of LD. Overall, our data indicate that this novel Borrelia is an intermediate taxon between the LD and RF species that may impact a large host spectrum, including American mammals. The designation "Candidatus Borrelia mahuryensis" is proposed for this species.

Rickettsia are obligate intracellular bacteria often reported from hard ticks but more rarely from soft ticks. In this study, we detected in Northern Africa two putatively novel Rickettsia species in soft tick species of the Ornithodoros erraticus complex: Ornithodoros occidentalis from Morocco, Ornithodoros erraticus from Algeria and Ornithodoros normandi from Tunisia. We characterized these two novel Rickettsia species on the basis of comparative DNA sequence analyses and phylogenetics of four genes (gltA, 16S rRNA, coxA and ompB). These Rickettsia, provisionally named 'Candidatus Rickettsia africaseptentrionalis' and 'Candidatus Rickettsia mauretanica', differed in nucleotide sequence from those of other Rickettsia species by 0.38-21.43 % depending on the gene examined. Phylogenetics further showed that the two novel Rickettsia species are closely related to each other and represent sister taxa to R. hoogstraalii, R. felis and R. asembonensis within the transitional Rickettsia group. While Ornithodoros host species of 'Candidatus Rickettsia africaseptentrionalis' and 'Candidatus Rickettsia mauretanica' are among the most common soft ticks to bite humans, their pathogenicity remains to be investigated., Competing Interests: Declaration of Competing Interest The authors declare that they have no competing interests., (Copyright © 2020 Elsevier GmbH. All rights reserved.)

Mutualistic interactions with microbes have facilitated the adaptation of major eukaryotic lineages to restricted diet niches. Hence, ticks with their strictly blood-feeding lifestyle are associated with intracellular bacterial symbionts through an essential B vitamin supplementation. In this study, examination of bacterial diversity in 25 tick species of the genus Amblyomma showed that three intracellular bacteria, Coxiella-like endosymbionts (LE), Francisella-LE and Rickettsia, are remarkably common. No other bacterium is as uniformly present in Amblyomma ticks. Almost all Amblyomma species were found to harbour a nutritive obligate symbiont, Coxiella-LE or Francisella-LE, that is able to synthesize B vitamins. However, despite the co-evolved and obligate nature of these mutualistic interactions, the structure of microbiomes does not mirror the Amblyomma phylogeny, with a clear exclusion pattern between Coxiella-LE and Francisella-LE across tick species. Coxiella-LE, but not Francisella-LE, form evolutionarily stable associations with ticks, commonly leading to co-cladogenesis. We further found evidence for symbiont replacements during the radiation of Amblyomma, with recent, and probably ongoing, invasions by Francisella-LE and subsequent replacements of ancestral Coxiella-LE through transient co-infections. Nutritional symbiosis in Amblyomma ticks is thus not a stable evolutionary state, but instead arises from conflicting origins between unrelated but competing symbionts with similar metabolic capabilities., (© 2020 John Wiley & Sons Ltd.)

Rickettsia are obligate intracellular bacteria often associated with ticks and best known for causing human diseases (rickettsiosis), including typhus fever and sporadic cases of serious infection. In this study, we conducted a large survey of ticks in French Guiana to understand the overall diversity of Rickettsia in this remote area largely covered by dense rainforests. Out of 819 individuals (22 tick species in six genera), 252 (30.8%) samples were positive for Rickettsia infection. Multilocus typing and phylogenetic analysis identified 19 Rickettsia genotypes, but none was 100% identical to already known Rickettsia species or strains. Among these 19 genotypes, we identified two validated Rickettsia species, Rickettsia amblyommatis (spotted fever group) and Rickettsia bellii (bellii group), and characterized a novel and divergent Rickettsia phylogenetic group, the guiana group. While some tick hosts of these Rickettsia genotypes are among the most common ticks to bite humans in French Guiana, their potential pathogenicity remains entirely unknown. However, we found a strong association between Rickettsia genotypes and their host tick species, suggesting that most of these Rickettsia genotypes may be nonpathogenic forms maintained through transovarial transmission.

Ticks are commonly infected by Coxiella-like endosymbionts (Coxiella-LE) which are thought to supply missing B vitamin nutrients required for blood digestion.While this nutritional symbiosis is essential for the survival and reproduction of infected tick species, our knowledge of where Coxiella-LE is localized in tick tissues is partial at best since previous studies have focused on a limited number of Asian or American tick species. To fill this gap, we investigated the tissue localization of Coxiella-LE in three European tick species, Ornithodoros maritimus, Dermacentor marginatus and Ixodes hexagonus, using a diagnostic fluorescence in situ hybridization (FISH) assay, combined with PCR-based detection. Specific fluorescent foci were observed in several tick tissues. We visualized a pronounced tissue tropism of Coxiella-LE for tick ovaries and Malpighian tubules, a pattern suggestive of a high degree of lifestyle specialization toward mutualism: infection of the ovaries is indicative of transovarial transmission, whereas infection of the Malpighian tubules suggests a nutritional function. We postulate that Malpighian tubules are key organs for the nutritional symbiosis, notably the synthesis of B vitamins by Coxiella-LE, whereas the infection of the ovaries ensures vertical transmission of the symbionts to future generations. We also detected occasional infections in other organs, such as salivary glands and the midgut. Finally, we discuss the potential significance of the different tissue tropism for tick biology., (Copyright © 2019 Elsevier GmbH. All rights reserved.)

Background: Ticks are obligate blood feeders transmitting major pathogens worldwide. Over the past few years, considerable research efforts have focused on the diversity, distribution and impact of gut and intracellular bacterial symbionts on tick development and tick-borne pathogen transmission. The study of this internal microbiome requires the use of a sterilization method to remove external (i.e. cuticular) microbes present on the tick's surface and to avoid any further contamination. Several sterilization methods exist, including ethanol- or bleach-based treatments that are both effective in killing microbes but with different potential effects on DNA denaturation., Methods: We examined how these different sterilization methods impact the measure of internal microbial diversity hosted by the Cayenne tick Amblyomma cajennense (sensu stricto). Bacterial barcoding investigations based on 16S rRNA gene sequences were conducted on two batches of 50 individuals each: Ticks of the first batch were sterilized with bleach diluted at 1% and the second batch with 70% ethanol. Tick external microbiome was also determined from cuticle smearing and water samples used for tick washing., Results: Bacterial barcoding investigations showed major differences between ethanol- and bleach-treated specimens. Both methods led to the detection of major intracellular bacteria associated with A. cajennense (s.s.) but ethanol-treated ticks always harbored a higher bacterial diversity than bleach-treated ticks. Further examinations of tick gut and tick external microbiome revealed that ethanol-based surface sterilization method is inefficient to eliminate the DNA of external bacteria., Conclusions: We herein provide evidence that studies investigating the internal microbiome of ticks should consider bleach as the gold standard to efficiently remove cuticular bacterial DNA. Indeed, this method does not impact the internal bacterial diversity hosted by ticks and is thus a better method than the ethanol-based one for studying the internal microbiome.

Mutualistic interactions with microbes have facilitated the radiation of major eukaryotic lineages [1, 2]. Microbes can notably provide biochemical abilities, allowing eukaryotes to adapt to novel habitats or to specialize on particular feeding niches [2-4]. To investigate the importance of mutualisms for the exclusive blood feeding habits of ticks, we focused on a bacterial genus of medical interest, Francisella, which is known to include both virulent intracellular pathogens of vertebrates [5, 6] and maternally inherited symbionts of ticks [7-9]. Through a series of physiological experiments, we identified a Francisella type, F-Om, as an obligate nutritional mutualist in the life cycle of the African soft tick Ornithodoros moubata. Francisella F-Om mutualism synthesizes B vitamins that are deficient in the blood meal of ticks. Indeed, experimental elimination of Francisella F-Om resulted in alteration of tick life history traits and physical abnormalities, deficiencies which were fully restored with an oral supplement of B vitamins. We also show that Francisella F-Om is maternally transmitted to all maturing tick oocytes, suggesting that this heritable symbiont is an essential adaptive element in the life cycle of O. moubata. The Francisella F-Om genome further revealed a recent origin from a Francisella pathogenic life style, as observed in other Francisella symbionts [6, 7, 10]. Though half of its protein-coding sequences are now pseudogenized or lost, Francisella F-Om has kept several B vitamin synthesis pathways intact, confirming the importance of these genes in evolution of its nutritional mutualism with ticks., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

The UvrB protein is a subunit of the UvrABC endonuclease which is involved in the repair of a large variety of DNA lesions. We have 91 isolated random uvrB mutants which are impaired in the repair of UV-damage in vivo. These mutants were classified on the basis of the ability to form normal levels of protein and the position of the mutations in the gene. The amino acid substitutions in the N-terminal part or in the C-terminal part of the UvrB protein are exclusively found in the conserved boxes of the so-called "helicase motifs" present in these parts of the protein, indicating that these motifs are essential for UvrB function. The proteins of four C-terminal mutants were purified: two mutants in motif V (E514K and G509S), one mutant in motif VI (R544H) and a double mutant in both motifs (E514K + R541H). In vitro experiments with these mutant proteins show that the helicase motifs V and VI are involved in the induction of ATP hydrolysis in the presence of (damaged) DNA and in the strand-displacement activity of the UvrA2B complex as is observed in a helicase assay. Furthermore, our results suggest that this strand-displacement activity is correlated to a local unwinding, which seems to be used to form the UvrB-DNA preincision complex.