Human cytomegalovirus infection is associated with increased expression of the lissencephaly gene PAFAH1B1 encoding LIS1 in neural stem cells and congenitally infected brains

peer reviewedCongenital infection of the central nervous system by human cytomegalovirus (HCMV) is a leading cause of permanent sequelae, including mental retardation or neurodevelopmental abnormalities. The most severe complications include smooth brain or polymicrogyria, which are both indicative of abnormal migration of neural cells, although the underlying mechanisms remain to be determined. To gain better insight on the pathogenesis of such sequelae, we assessed the expression levels of a set of neurogenesis-related genes, using HCMV-infected human neural stem cells derived from embryonic stem cells (NSCs). Among the 84 genes tested, we found dramatically increased expression of the gene PAFAH1B1, encoding LIS1 (lissencephaly-1), in HCMV-infected versus uninfected NSCs. Consistent with these ndings, western blotting and immunouorescence analyses conrmed the increased levels of LIS1 in HCMV-infected NSCs at the protein level. We next assessed the migratory abilities of HCMV-infected NSCs and observed that infection strongly impaired the migration of NSCs, without detectable effect on their proliferation. Moreover, we observed increased immunostaining for LIS1 in brains of congenitally infected fetuses, but not in control samples, highlighting the clinical relevance of our ndings. Of note, PAFAH1B1 mutations (resulting in either haploinsufciency or gain of function) are primary causes of hereditary neurodevelopmental diseases. Notably, mutations resulting in PAFAH1B1 haploinsufciency cause classic lissencephaly. Taken together, our ndings suggest that PAFAH1B1 is a critical target of HCMV infection. They also shine a new light on the pathophysiological basis of the neurological outcomes of congenital HCMV infection, by suggesting that defective neural cell migration might contribute to the pathogenesis of the neurodevelopmental sequelae of infectio

Déterminants immuno-virologiques de l’infection congénitale à cytomégalovirus dans les prélèvements fœtaux périphériques et dans le tissu cérébral

CMV congenital infection has a worldwide incidence estimated at about 0.7% of all life births and represents the major cause of neurological handicap of infectious origin. The management of this infection remains highly debated. Several factors contribute to this and among them are the absence of recognized prognostic markers and gaps in the knowledge of its pathogenicity particularly that of the fetal brain. The first objective of this work was to describe and validate immune and virological predictive markers of vertical transmission and of neonatal sequelae. The second objective was to study in situ immune and virological correlates of the severity of fetal brain infection. We first validated a model of materno-fetal transmission based on maternal virological results (IgG avidity and blood CMV DNA). We then showed that the viral reservoir level, estimated by the viral load in the amniotic fluid and the fetal blood, was a predictive marker of neonatal sequelae. Prognosis models combining quantification of the viral reservoir to fetal imaging allow to reach positive and negative predictive values up to 80% and 100% respectively. We showed using immunohistochemistry and quantitative image analysis that viral multiplication as well as both innate immune responses (NK cells) and adaptive immune responses (CD8+ and plasma cells) were significantly higher in the most severely infected fetal brains. This paradox drove us to quantify PD-1 and its receptor PD-L1, PD-1expression was significantly higher in severely affected fetal brains. Cytometry flow analysis evidenced that PD-1 was expressed in 95% of CD8+ cells but also in at least 70% of NK cells and of B cells. These results demonstrate immune exhaustion of both adaptive and innate responses in fetal infected brains. Finally, viral replication was evidenced in stem cells, neurons and mature astrocytes after separation by flow cytometry of these neuronal cell types. In conclusion, the validation of immune-virological markers obtained within this work has usefully improved the algorithms for the clinical management of in utero CMV infection. Moreover, the demonstration that immune exhaustion and high viral multiplication are responsible of severe fetal brain affection is important to elaborate in utero treatment strategies.Avec une prévalence mondiale de 0,7%, l’infection in utero à cytomégalovirus (CMV) représente la première cause de handicap neurologique congénital d’origine infectieuse. Les modalités de prise en charge de cette infection restent débattues notamment en raison de l’absence de marqueurs pronostics fiables et d’inconnus sur sa physiopathologie notamment celle de l’atteinte du cerveau fœtal. Le premier objectif de notre travail était de décrire et valider des marqueurs immuno-virologiques prédictifs de la transmission verticale et de séquelles néonatales. Le deuxième objectif était d’étudier les corrélats immuno-virologiques in situ de la sévérité de l’atteinte du cerveau fœtal. Nous avons pu établir à partir du bilan virologique maternel (avidité des IgG et ADN CMV sanguin) un score de risque de transmission verticale du virus en cas de primo-infection maternelle. Nous avons montré que le niveau du réservoir viral fœtal, reflété par la charge virale dans le liquide amniotique et dans le sang fœtal, était un marqueur prédictif des séquelles néonatales. Ainsi, la combinaison de la mesure du réservoir viral fœtal avec l’imagerie fœtale a permis d’établir des scores pronostics avec des valeurs prédictives positives et négatives de 80 à 100% respectivement. Nous avons mis en évidence par immuno-histochimie couplée à une analyse quantitative d’images que la multiplication virale ainsi que la réponse immunitaire innée (cellules NK) et adaptative (CD8+ et plasmocytes) étaient significativement plus élevées dans les cerveaux fœtaux les plus sévèrement atteints. Ce résultat paradoxal nous a incité à quantifier la présence du marqueur PD-1 et celle de son récepteur PD-L1. PD-1 était significativement plus exprimé dans les cerveaux sévèrement atteints. L’analyse par cytométrie de flux montrait que PD-1 était exprimé par 96% des CD8+ mais aussi par plus de 70% des lymphocytes B et des cellules NK. Ces résultats témoignent de l’existence dans les cerveaux fœtaux infectés d’un épuisement immunitaire touchant la réponse adaptative mais aussi innée. Enfin, l’analyse par cytométrie de flux montrait la présence d’une réplication virale dans les différents types de cellules neuronales (cellules souches, neurones, astrocytes). En conclusion, les résultats de notre travail ont permis d’améliorer les algorithmes de prise en charge de l’infection à CMV in utero grâce à la validation de marqueurs prédictifs immuno-virologiques. Par ailleurs, le fait qu’un épuisement immunitaire et une forte multiplication virale soient associés à la sévérité de l’atteinte cérébrale est important pour l’élaboration de stratégies thérapeutiques in utero

Immuno-virologic determinants of congenital cytomegalovirus infection in peripheral fetal samples and brain tissue

Avec une prévalence mondiale de 0,7%, l’infection in utero à cytomégalovirus (CMV) représente la première cause de handicap neurologique congénital d’origine infectieuse. Les modalités de prise en charge de cette infection restent débattues notamment en raison de l’absence de marqueurs pronostics fiables et d’inconnus sur sa physiopathologie notamment celle de l’atteinte du cerveau fœtal. Le premier objectif de notre travail était de décrire et valider des marqueurs immuno-virologiques prédictifs de la transmission verticale et de séquelles néonatales. Le deuxième objectif était d’étudier les corrélats immuno-virologiques in situ de la sévérité de l’atteinte du cerveau fœtal. Nous avons pu établir à partir du bilan virologique maternel (avidité des IgG et ADN CMV sanguin) un score de risque de transmission verticale du virus en cas de primo-infection maternelle. Nous avons montré que le niveau du réservoir viral fœtal, reflété par la charge virale dans le liquide amniotique et dans le sang fœtal, était un marqueur prédictif des séquelles néonatales. Ainsi, la combinaison de la mesure du réservoir viral fœtal avec l’imagerie fœtale a permis d’établir des scores pronostics avec des valeurs prédictives positives et négatives de 80 à 100% respectivement. Nous avons mis en évidence par immuno-histochimie couplée à une analyse quantitative d’images que la multiplication virale ainsi que la réponse immunitaire innée (cellules NK) et adaptative (CD8+ et plasmocytes) étaient significativement plus élevées dans les cerveaux fœtaux les plus sévèrement atteints. Ce résultat paradoxal nous a incité à quantifier la présence du marqueur PD-1 et celle de son récepteur PD-L1. PD-1 était significativement plus exprimé dans les cerveaux sévèrement atteints. L’analyse par cytométrie de flux montrait que PD-1 était exprimé par 96% des CD8+ mais aussi par plus de 70% des lymphocytes B et des cellules NK. Ces résultats témoignent de l’existence dans les cerveaux fœtaux infectés d’un épuisement immunitaire touchant la réponse adaptative mais aussi innée. Enfin, l’analyse par cytométrie de flux montrait la présence d’une réplication virale dans les différents types de cellules neuronales (cellules souches, neurones, astrocytes). En conclusion, les résultats de notre travail ont permis d’améliorer les algorithmes de prise en charge de l’infection à CMV in utero grâce à la validation de marqueurs prédictifs immuno-virologiques. Par ailleurs, le fait qu’un épuisement immunitaire et une forte multiplication virale soient associés à la sévérité de l’atteinte cérébrale est important pour l’élaboration de stratégies thérapeutiques in utero.CMV congenital infection has a worldwide incidence estimated at about 0.7% of all life births and represents the major cause of neurological handicap of infectious origin. The management of this infection remains highly debated. Several factors contribute to this and among them are the absence of recognized prognostic markers and gaps in the knowledge of its pathogenicity particularly that of the fetal brain. The first objective of this work was to describe and validate immune and virological predictive markers of vertical transmission and of neonatal sequelae. The second objective was to study in situ immune and virological correlates of the severity of fetal brain infection. We first validated a model of materno-fetal transmission based on maternal virological results (IgG avidity and blood CMV DNA). We then showed that the viral reservoir level, estimated by the viral load in the amniotic fluid and the fetal blood, was a predictive marker of neonatal sequelae. Prognosis models combining quantification of the viral reservoir to fetal imaging allow to reach positive and negative predictive values up to 80% and 100% respectively. We showed using immunohistochemistry and quantitative image analysis that viral multiplication as well as both innate immune responses (NK cells) and adaptive immune responses (CD8+ and plasma cells) were significantly higher in the most severely infected fetal brains. This paradox drove us to quantify PD-1 and its receptor PD-L1, PD-1expression was significantly higher in severely affected fetal brains. Cytometry flow analysis evidenced that PD-1 was expressed in 95% of CD8+ cells but also in at least 70% of NK cells and of B cells. These results demonstrate immune exhaustion of both adaptive and innate responses in fetal infected brains. Finally, viral replication was evidenced in stem cells, neurons and mature astrocytes after separation by flow cytometry of these neuronal cell types. In conclusion, the validation of immune-virological markers obtained within this work has usefully improved the algorithms for the clinical management of in utero CMV infection. Moreover, the demonstration that immune exhaustion and high viral multiplication are responsible of severe fetal brain affection is important to elaborate in utero treatment strategies

Stream restorations with meanders increase dragonfly and damselfly diversity and abundance, including an endangered species.

10 pagesInternational audienceThis study presents examples of successful restoration projects for biodiversity conservation. In West France, the Pinail National Nature Reserve is a protected wetland interspersed with more than 6000 ponds. This wetland is inhabited by 50 species of Odonata and thus is a key biodiversity area for damselflies and dragonflies conservation. In the past, when the limestone was exploited, the streams of the plateau were artificially channeled rectilinearly, running to the Vienne River. Eventually streams were blocked by biomass and sediments resulting in water flowing mainly underground. In 2011, two restoration projects dug and recreated lost habitats such as running streams and meanders by openly reconnecting bodies of standing water (two sites: Rivau (20 m) and Hutte (400 m) streams). The Odonata species diversity and abundance are annually monitored following transect inventories since 1995 and still ongoing. Diversity and abundance were compared before and after the restoration. The abundance and species diversity increased at both sites due to the addition of lotic habitats and consequently additional new species. The number of observed species almost doubled on the Rivau (from 5.4 observed species to 9.9 spp). By extrapolation the total species number on site increased from 15 to 18 spp to 29–37 spp. The abundance also greatly increased with 770 % more individuals on the Rivau. Similarly, on the 400 m Hutte stream, the extrapolated diversity increased from 31 to 38 spp to 35–43 spp; as well as the abundance with 475 % more individuals. These restoration projects created new habitats leading to local biodiversity enrichment and conservation success. More specifically, Coenagrion mercuriale (Odonata: Zygoptera), one of Europe's most threatened damselflies and listed in the European Habitats directive, successfully recolonized the Rivau stream and colonized the Hutte stream

Freshwater acidification: an example of an endangered crayfish species sensitive to pH

10 pagesInternational audienceCarbon dioxide released in the atmosphere and dissolved in water leads to acidification. Relatively few studies have focused on fresh waters, where biocalcifying species are more readily impacted by changes in pH. Sensitivity to pH of an endangered calcium-demanding organism, the crayfish Austropotamobius pallipes, was investigated in the Pinail nature reserve, a natural system with 3000 permanent ponds, some inhabited by the crayfish and others not, originally due to human introduction. From the 14 chemical parameters measured in this study, the main limiting factor preventing crayfish establishment appears to be water acidity (pH 7)

Evaluation of midwives’ practises on herpetic infections during pregnancy ::a french vignette-based study

Background: One out of two pregnant women has a history of herpes infection. Initial infections have a high risk of neonatal transmission. Our objective was to analyse the professional practises of midwives regarding the management of herpes infections during pregnancy in France; Methods: A national survey conducted via an online self-questionnaire, including clinical vignettes for which the midwives proposed a diagnosis, a drug treatment, a mode of birth, and a prognosis. These responses were used to evaluate the conformity of the responses to the guidelines, as well as the influence of certain criteria, such as mode of practise and experience; Results: Of 728 responses, only 26.1% of the midwives reported being aware of the 2017 clinical practise guidelines. The midwives proposed taking the appropriate actions in 56.1% of the responses in the case of a recurrence, and in 95.1% of the responses in the case of a primary infection. For the specific, high-risk case of a nonprimary initial infection at 38 weeks of gestation, reporting knowledge of the recommendations improved the compliance of the proposed care by 40% (p = 0.02). However, 33.8% of the midwives underestimated the neonatal risk at term after a primary initial infection, and 43% underestimated the risk after a primary initial infection at term; Conclusions: The majority of reported practises were compliant despite a low level of knowledge of the guidelines. The dissemination of guidelines may be important to improve information and adherence to appropriate therapeutic practise

Native top-predator cannot eradicate an invasive fish from small pond ecosystems.

5 pagesInternational audiencePumpkinseed (Lepomis gibbosus) originates from North America and has been widely introduced in Europe where invasive populations have established. We tested the effectiveness of a biomanipulation approach based on the stocking of a native top-predatory species, the northern pike (Esox lucius), in 23 small and oligotrophic ponds at the Pinail Nature Reserve (Vienne, France) among which 10 ponds were stocked twice. In addition, 16 ponds with similar environmental characteristics were used as control with no pike stocking. Our study revealed that, even with limited space and limited alternative prey species, northern pike did not eradicate pumpkinseed populations. Instead, we found that pumpkinseed were younger and larger when reaching sexual maturity in the stocked ponds, suggesting an increased growth rate in ponds with the predator. These results suggest that invasion populations might adapt and respond to management practices. These changes were likely driven by an adaptation to predation pressure and/or changes in food availability with reduced intraspecific competition. Importantly, such changes might actually modify the level of invasiveness potential of non-native populations and lead to counterproductive results for managers

Climate change would prevail over land use change in shaping the future distribution of Triturus marmoratus in France

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Predicting suitable habitats of four range margin amphibians under climate and land-use changes in southwestern France

International audienceWhereas the effects of climate change on the potential range shifts of amphibians have been modeled at various scales, exploring the future impacts of land-use changes has rarely been considered. We modeled the potential distribution of Bombina variegata, Hyla arborea, Hyla meridionalis, and Triturus cristatus using ecological niche modeling, in the French administrative region Nouvelle-Aquitaine. The species are all at range margin in the study area which makes them interesting model species to study range shifts because of their potential local adaptations to their marginal habitat. We used Dyna-CLUE modeling framework to produce land-use change scenarios and combined them with RCP 2.6 and RCP 8.5 climate change scenarios to predict their impact on local amphibian distributions. B. variegata, H. arborea, and T. cristatus were predicted to shift eastward and northward. H. meridionalis was predicted to increase its probability of presence in the study area. The effect of climate determined the general pattern of distribution and was modulated depending on the scenarios of land-use change

Habitat patches for newts in the face of climate change: local scale assessment combining niche modelling and graph theory

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