Role of Seaports and Imported Rats in Seoul Hantavirus Circulation, Africa.

Seoul orthohantavirus (SEOV) is not considered a major public health threat on the continent of Africa. However, Africa is exposed to rodentborne SEOV introduction events through maritime traffic after exponential growth of trade with the rest of the world. Serologic studies have already detected hantavirus antibodies in human populations, and recent investigations have confirmed circulation of hantavirus, including SEOV, in rat populations. Thus, SEOV is a possible emerging zoonotic risk in Africa. Moreover, the range of SEOV could rapidly expand, and transmission to humans could increase because of host switching from the usual brown rat (Rattus norvegicus) species, which is currently invading Africa, to the more widely installed black rat (R. rattus) species. Because of rapid economic development, environmental and climatic changes, and increased international trade, strengthened surveillance is urgently needed to prevent SEOV dissemination among humans in Africa.

Serologic Surveillance for SARS-CoV-2 Infection among Wild Rodents, Europe.

We report results from serologic surveillance for exposure to SARS-CoV-2 among 1,237 wild rodents and small mammals across Europe. All samples were negative, with the possible exception of 1. Despite suspected potential for human-to-rodent spillover, no evidence of widespread SARS-CoV-2 circulation in rodent populations has been reported to date.Esitämme tulokset serologisesta tutkimuksesta, jossa seulottiin SARS-CoV-2 tartuntojen varalta 1,237 luonnonvaraista jyrsijää ja piennisäkästä eri puolilta Eurooppaa. Kaikki näytteet olivat negatiivisia, yhtä näytettä lukuun ottamatta. SARS-CoV-2:n läikkymisen ihmisistä jyrsijöihin on arveltu olevan mahdollista, mutta todisteet viruksen laajamittaisesta leviämisestä jyrsijäpopulaatioissa puuttuvat.

Genetic Characterization of Seoul Virus in the Seaport of Cotonou, Benin.

Seoul virus is a zoonotic pathogen carried by the brown rat Rattus norvegicus. Information on its circulation in Africa is limited. In this study, the virus was detected in 37.5% of brown rats captured in the Autonomous Port of Cotonou, Benin. Phylogenetic analyses place this virus in Seoul virus lineage 7.

AQUAPONY: visualization and interpretation of phylogeographic information on phylogenetic trees.

MOTIVATION: The visualization and interpretation of evolutionary spatiotemporal scenarios is broadly and increasingly used in infectious disease research, ecology or agronomy. Using probabilistic frameworks, well-known tools can infer from molecular data ancestral traits for internal nodes in a phylogeny, and numerous phylogenetic rendering tools can display such evolutionary trees. However, visualizing such ancestral information and its uncertainty on the tree remains tedious. For instance, ancestral nodes can be associated to several geographical annotations with close probabilities and thus, several migration or transmission scenarios exist. RESULTS: We expose a web-based tool, named AQUAPONY, that facilitates such operations. Given an evolutionary tree with ancestral (e.g. geographical) annotations, the user can easily control the display of ancestral information on the entire tree or a subtree, and can view alternative phylogeographic scenarios along a branch according to a chosen uncertainty threshold. AQUAPONY interactively visualizes the tree and eases the objective interpretation of evolutionary scenarios. AQUAPONY's implementation makes it highly responsive to user interaction, and instantaneously updates the tree visualizations even for large trees (which can be exported as image files). AVAILABILITY AND IMPLEMENTATION: AQUAPONY is coded in JavaScript/HTML, available under Cecill license, and can be freely used at http://www.atgc-montpellier.fr/aquapony/.

PastView: a user-friendly interface to explore ancestral scenarios.

BACKGROUND: Ancestral character states computed from the combination of phylogenetic trees with extrinsic traits are used to decipher evolutionary scenarios in various research fields such as phylogeography, epidemiology, and ecology. Despite the existence of powerful methods and software in ancestral character state inference, difficulties may arise when interpreting the outputs of such inferences. The growing complexity of data (trees, annotations), the diversity of optimization criteria for computing trees and ancestral character states, the combinatorial explosion of potential evolutionary scenarios if some ancestral characters states do not stand out clearly from others, requires the design of new methods to explore associations of phylogenetic trees with extrinsic traits, to ease the visualization and interpretation of evolutionary scenarios. RESULT: We developed PastView, a user-friendly interface that includes numerical and graphical features to help users to import and/or compute ancestral character states from discrete variables and extract ancestral scenarios as sets of successive transitions of character states from the tree root to its leaves. PastView provides summarized views such as transition maps and integrates comparative tools to highlight agreements or discrepancies between methods of ancestral annotations inference. CONCLUSION: The main contribution of PastView is to assemble known numerical and graphical methods into a multi-maps graphical user interface dedicated to the computing, searching and viewing of evolutionary scenarios based on phylogenetic trees and ancestral character states. PastView is available publicly as a standalone software on www.pastview.org .

Puumala Hantavirus Genotypes in Humans, France, 2012-2016.

The analysis of the nucleoprotein gene of 77 Puumala hantavirus strains detected in human samples in France during 2012-2016 showed that all belonged to the Central European lineage. We observed 2 main clusters, geographically structured; one included strains with the Q64 signature and the other strains with the R64 signature.

Discovery of hantavirus circulating among Rattus rattus in French Mayotte island, Indian Ocean.

Hantaviruses are emerging zoonotic viruses that cause human diseases. In this study, sera from 642 mammals from La Réunion and Mayotte islands (Indian Ocean) were screened for the presence of hantaviruses by molecular analysis. None of the mammals from La Réunion island was positive, but hantavirus genomic RNA was discovered in 29/160 (18 %) Rattus rattus from Mayotte island. The nucleoprotein coding region was sequenced from the liver and spleen of all positive individuals allowing epidemiological and intra-strain variability analyses. Phylogenetic analysis based on complete coding genomic sequences showed that this Murinae-associated hantavirus is a new variant of Thailand virus. Further studies are needed to investigate hantaviruses in rodent hosts and in Haemorrhagic Fever with Renal Syndrome (HFRS) human cases.

Hybrid capture-based next-generation sequencing of new and old world Orthohantavirus strains and wild-type Puumala isolates from humans and bank voles.

Orthohantaviruses, transmitted primarily by rodents, cause hemorrhagic fever with renal syndrome (HFRS) in Eurasia and hantavirus pulmonary syndrome in the Americas. These viruses, with documented human-to-human transmission, exhibit a wide case-fatality rate, 0.5-40 %, depending on the virus species, and no vaccine or effective treatment for severe Orthohantavirus infections exists. In Europe, the Puumala virus (PUUV), carried by the bank vole Myodes glareolus, causes a milder form of HFRS. Despite the reliance on serology and PCR for diagnosis, the three genomic segments of Swedish wild-type PUUV have yet to be completely sequenced. We have developed a targeted hybrid-capture method aimed at comprehensive genomic sequencing of wild-type PUUV isolates and the identification of other Orthohantaviruses. Our custom-designed panel includes >11,200 probes covering the entire Orthohantavirus genus. Using this panel, we sequenced complete viral genomes from bank vole lung tissue, human plasma samples, and cell-cultured reference strains. Analysis revealed that Swedish PUUV isolates belong to the Northern Scandinavian lineage, with nucleotide diversity ranging from 2.8 % to 3.7 % among them. Notably, no significant genotypic differences were observed between the viral sequences from reservoirs and human cases except in the nonstructural protein. Despite the high endemicity of PUUV in Northern Sweden, these are the first complete Swedish wild-type PUUV genomes and substantially increase our understanding of PUUV evolution and epidemiology. The panel's sensitivity enables genomic sequencing of human samples with viral RNA levels reflecting the natural progression of infection and underscores our panel's diagnostic value, and could help to uncover novel Orthohantavirus transmission routes.

Evolution and genetic characterization of Seoul virus in wild rats Rattus norvegicus from an urban park in Lyon, France 2020-2022.

BACKGROUND: Seoul virus (SEOV) is an orthohantavirus primarily carried by rats. In humans, it may cause hemorrhagic fever with renal syndrome (HFRS). Its incidence is likely underestimated and given the expansion of urban areas, a better knowledge of SEOV circulation in rat populations is called for. Beyond the need to improve human case detection, we need to deepen our comprehension of the ecological, epidemiological, and evolutionary processes involved in the transmission of SEOV. METHODOLOGY / PRINCIPAL FINDINGS: We performed a comprehensive serological and molecular characterization of SEOV in Rattus norvegicus in a popular urban park within a large city (Lyon, France) to provide essential information to design surveillance strategies regarding SEOV. We sampled rats within the urban park of 'La Tête d'Or' in Lyon city from 2020 to 2022. We combined rat population genetics, immunofluorescence assays, SEOV high-throughput sequencing (S, M, and L segments), and phylogenetic analyses. We found low structuring of wild rat populations within Lyon city. Only one sampling site within the park (building created in 2021) showed high genetic differentiation and deserves further attention. We confirmed the circulation of SEOV in rats from the park with high seroprevalence (17.2%) and high genetic similarity with the strain previously described in 2011 in Lyon city. CONCLUSION/SIGNIFICANCE: This study confirms the continuous circulation of SEOV in a popular urban park where the risk for SEOV transmission to humans is present. Implementing a surveillance of this virus could provide an efficient early warning system and help prepare risk-based interventions. As we reveal high gene flow between rat populations from the park and the rest of the city, we advocate for SEOV surveillance to be conducted at the scale of the entire city.

Sequence, assembly and count datasets of viruses associated to the pine processionary moth Thaumetopoea pityocampa (Denis & Schiffermüller) (Lepidoptera, Notodontidae) identified from transcriptomic high-throughput sequencing.

The pine processionary moth Thaumetopoea pityocampa is a Lepidopteran pest species occurring in the Western Mediterranean. It causes heavy pine defoliations and it is a public and animal health concern because of its urticating caterpillars. Very little is known about the viruses associated to this species, as only two viruses were described so far. We here present a dataset corresponding to 34 viral transcripts, among which 27 could be confidently assigned to 9 RNA and DNA viral families (Iflaviridae, Reoviridae, Partitiviridae, Permutotetraviridae, Flaviviridae, Rhabdoviridae, Parvoviridae, Baculoviridae and PolyDNAviridae). These transcripts were identified from an original transcriptome assembled for the insect host, using both blast search and phylogenetic approaches. The data were acquired from 2 populations in Portugal and 2 populations in Italy. The transcripts were de novo assembled and used to identify viral sequences by homology searches. We also provide information about the populations and life stages in which each virus was identified. The data produced will allow to enrich the virus taxonomy in Lepidopteran hosts, and to develop PCR-based diagnostic tools to screen colonies across the range and determine the distribution and prevalence of the identified viral species.

Puumala orthohantavirus circulation in its wild reservoir, the bank vole, during the 2021 outbreak of hemorrhagic fever with renal syndrome in Jura, France.

OBJECTIVE: A large and unprecedented outbreak of an attenuated form of hemorrhagic fever with renal syndrome called nephropathia epidemica (NE) and caused by Puumala virus (PUUV) occurred in 2021 in the southern Jura Mountains (France) leading to numerous hospitalizations. The aim of this study was to investigate the circulation of PUUV in its animal reservoir at the time of this outbreak. METHODS: We conjointly surveyed bank vole relative abundance, small mammal community composition, and PUUV circulation in bank voles (seroprevalence and genetic diversity) in the Jura NE epidemic area, between 2020 and 2022. RESULTS: Trapping results showed a higher relative abundance of bank voles in 2021 compared to 2020 and 2022. Extremely high levels of PUUV seroprevalence in bank voles were found at the time of the human NE epidemic with seropositive animals trapped in almost all trap lines as of spring 2021. Genetic analyses of PUUV (S segment) gathered in 2021 at two sampling sites revealed a strong clustering of these strains within the "Jura" clade. No significant genetic variation was detected compared to what was already known to be circulating in the Jura region. CONCLUSION: These results underline a need for enhanced monitoring of PUUV circulation in host reservoir populations in NE endemic areas. This would enable the relevant actors to better inform and sensitize the public on this zoonotic risk, and to implement prevention strategies in collaboration with physicians.

Small terrestrial mammals (Rodentia and Soricomorpha) along a gradient of forest anthropisation (reserves, managed forests, urban parks) in France.

Background: Understanding the relationships between wildlife biodiversity and zoonotic infectious diseases in a changing climate is a challenging issue that scientists must address to support further policy actions. We aim at tackling this challenge by focusing on small mammal-borne diseases in temperate forests and large urban green spaces. Small mammals are important reservoirs of zoonotic agents, with a high transmission potential for humans and domestic animals. Forests and large urban green spaces are ecosystems where efforts are undertaken to preserve biodiversity. They are put forward for their contribution to human well-being in addition to other ecosystem services (e.g. provisioning and regulating services). Moreover, forests and large urban green spaces are environments where small mammals are abundant and human/domestic-wildlife interactions are plausible to occur. These environments are, therefore, focal points for conservation management and public health issues. New information: The European Biodiversa BioRodDis project (https://www6.inrae.fr/biodiversa-bioroddis) aims at better understanding the relationships between small terrestrial mammal biodiversity and health in the context of global change and, in particular, of forest anthropisation and urbanisation. Here, we present the data gathered in France. The dataset will enable us to describe the diversity of small terrestrial mammal communities in forested areas corresponding to different levels of anthropisation and to evaluate the variability of this diversity over time, between seasons and years.The dataset contains occurrences of small terrestrial mammals (Rodentia and Soricomorpha) trapped in forested areas in eastern France (administrative Departments: Rhône, Ain, Jura). The sampling sites correspond to different degrees of anthropisation. Forests included in biological reserves are the least anthropised sites. Then, public forests and urban parks experience increasing levels of anthropisation. Data were collected during spring and autumn 2020 (three to four sampling sites), 2021 (six sampling sites) and 2022 (four sampling sites). These variations in the number of sites between years were due to lockdown restrictions in 2020 or to the legal authorisation to trap around biological reserves granted in 2021 only. The capture of animals was carried out in various types of forests (pine, deciduous, mixed) and in different habitats within urban parks (wooded areas, buildings, hay storage yards, riverside vegetation, restaurants, playground for kids, botanical garden, landfills). Animals were captured using live traps that were set on the ground for one to 11 nights. During this study period, 1593 small mammals were trapped and identified. They belong to 15 species, amongst which were nine species of rodents (Muridae, Cricetidae, Gliridae) and six species of shrews (Soricidae). They were weighted (gram) and measured (cm): head-body length, tail length and hind-foot length. Sexual characteristics were also recorded.

Phage display of combinatorial peptide libraries: application to antiviral research.

Given the growing number of diseases caused by emerging or endemic viruses, original strategies are urgently required: (1) for the identification of new drugs active against new viruses and (2) to deal with viral mutants in which resistance to existing antiviral molecules has been selected. In this context, antiviral peptides constitute a promising area for disease prevention and treatment. The identification and development of these inhibitory peptides require the high-throughput screening of combinatorial libraries. Phage-display is a powerful technique for selecting unique molecules with selective affinity for a specific target from highly diverse combinatorial libraries. In the last 15 years, the use of this technique for antiviral purposes and for the isolation of candidate inhibitory peptides in drug discovery has been explored. We present here a review of the use of phage display in antiviral research and drug discovery, with a discussion of optimized strategies combining the strong screening potential of this technique with complementary rational approaches for identification of the best target. By combining such approaches, it should be possible to maximize the selection of molecules with strong antiviral potential.

Impact of Landscape on Host-Parasite Genetic Diversity and Distribution Using the Puumala orthohantavirus-Bank Vole System.

In nature, host specificity has a strong impact on the parasite's distribution, prevalence, and genetic diversity. The host's population dynamics is expected to shape the distribution of host-specific parasites. In turn, the parasite's genetic structure is predicted to mirror that of the host. Here, we study the tandem Puumala orthohantavirus (PUUV)-bank vole system. The genetic diversity of 310 bank voles and 33 PUUV isolates from 10 characterized localities of Northeast France was assessed. Our findings show that the genetic diversity of both PUUV and voles, was positively correlated with forest coverage and contiguity of habitats. While the genetic diversity of voles was weakly structured in space, that of PUUV was found to be strongly structured, suggesting that the dispersion of voles was not sufficient to ensure a broad PUUV dissemination. Genetic diversity of PUUV was mainly shaped by purifying selection. Genetic drift and extinction events were better reflected than local adaptation of PUUV. These contrasting patterns of microevolution have important consequences for the understanding of PUUV distribution and epidemiology.

Isolation and Genetic Characterization of Puumala Orthohantavirus Strains from France.

Puumala orthohantavirus (PUUV) causes a mild form of haemorrhagic fever with renal syndrome (HFRS) called nephropathia epidemica (NE), regularly diagnosed in Europe. France represents the western frontier of the expansion of NE in Europe with two distinct areas: an endemic area (north-eastern France) where PUUV circulates in rodent populations, with the detection of many human NE cases, and a non-endemic area (south-western France) where the virus is not detected, with only a few human cases being reported. In this study, we describe the different stages of the isolation of two PUUV strains from two distinct French geographical areas: Ardennes (endemic area) and Loiret (non-endemic area). To isolate PUUV efficiently, we selected wild bank voles (Myodes glareolus, the specific reservoir of PUUV) captured in these areas and that were seronegative for anti-PUUV IgG (ELISA) but showed a non-negligible viral RNA load in their lung tissue (qRT-PCR). With this study design, we were able to cultivate and maintain these two strains in Vero E6 cells and also propagate both strains in immunologically neutral bank voles efficiently and rapidly. High-throughput and Sanger sequencing results provided a better assessment of the impact of isolation methods on viral diversity.

Puumala Virus Variants Circulating in Forests of Ardennes, France: Ten Years of Genetic Evolution.

In Europe, Puumala virus (PUUV) transmitted by the bank vole (Myodes glareolus) is the causative agent of nephropathia epidemica (NE), a mild form of haemorrhagic fever with renal syndrome. In France, very little is known about the spatial and temporal variability of the virus circulating within bank vole populations. The present study involved monitoring of bank vole population dynamics and PUUV microdiversity over a ten-year period (2000-2009) in two forests of the Ardennes region: Elan and Croix-Scaille. Ardennes region is characterised by different environmental conditions associated with different NE epidemiology. Bank vole density and population parameters were estimated using the capture/marking/recapture method, and blood samples were collected to monitor the overall seroprevalence of PUUV in rodent populations. Phylogenetic analyses of fifty-five sequences were performed to illustrate the genetic diversity of PUUV variants between forests. The pattern of the two forests differed clearly. In the Elan forest, the rodent survival was higher, and this limited turn-over resulted in a lower seroprevalence and diversity of PUUV sequences than in the Croix-Scaille forest. Uncovering the links between host dynamics and virus microevolution is improving our understanding of PUUV distribution in rodents and the NE risk.

Peptides that mimic the amino-terminal end of the rabies virus phosphoprotein have antiviral activity.

We wanted to develop a therapeutic approach against rabies disease by targeting the lyssavirus transcription/replication complex. Because this complex (nucleoprotein N-RNA template processed by the L polymerase and its cofactor, the phosphoprotein P) is similar to that of other negative-strand RNA viruses, we aimed to design broad-spectrum antiviral drugs that could be used as a complement to postexposure vaccination and immunotherapy. Recent progress in understanding the structure/function of the rabies virus P, N, and L proteins predicts that the amino-terminal end of P is an excellent target for destabilizing the replication complex because it interacts with both L (for positioning onto the N-RNA template) and N (for keeping N soluble, as needed for viral RNA encapsidation). Thus, peptides mimicking various lengths of the amino-terminal end of P have been evaluated, as follows: (i) for binding properties to the N-P-L partners by the two-hybrid method; (ii) for their capacity to inhibit the transcription/replication of a rabies virus minigenome encoding luciferase in BHK-21-T7 cells; and (iii) for their capacity to inhibit rabies virus infection of BHK-21-T7 cells and of two derivatives of the neuronal SK-N-SH cell line. Peptides P60 and P57 (the first 60 and first 57 NH2 residues of P, respectively) exhibited a rapid, strong, and long-lasting inhibitory potential on luciferase expression (>95% from 24 h to 55 h). P42 was less efficient in its inhibition level (75% for 18 to 30 h) and duration (40% after 48 h). The most promising peptides were synthesized in tandem with the Tat sequence, allowing cell penetration. Their inhibitory effects were observed on BHK-21-T7 cells infected with rabies virus and Lagos bat virus but not with vesicular stomatitis virus. In neuronal cells, a significant inhibition of both nucleocapsid inclusions and rabies virus release was observed.

How Bank Vole-PUUV Interactions Influence the Eco-Evolutionary Processes Driving Nephropathia Epidemica Epidemiology-An Experimental and Genomic Approach.

In Europe, Puumala virus (PUUV) is responsible for nephropathia epidemica (NE), a mild form of hemorrhagic fever with renal syndrome (HFRS). Despite the presence of its reservoir, the bank vole, on most of French territory, the geographic distribution of NE cases is heterogeneous and NE endemic and non-endemic areas have been reported. In this study we analyzed whether bank vole-PUUV interactions could partly shape these epidemiological differences. We performed crossed-experimental infections using wild bank voles from French endemic (Ardennes) and non-endemic (Loiret) areas and two French PUUV strains isolated from these areas. The serological response and dynamics of PUUV infection were compared between the four cross-infection combinations. Due to logistical constraints, this study was based on a small number of animals. Based on this experimental design, we saw a stronger serological response and presence of PUUV in excretory organs (bladder) in bank voles infected with the PUUV endemic strain. Moreover, the within-host viral diversity in excretory organs seemed to be higher than in other non-excretory organs for the NE endemic cross-infection but not for the NE non-endemic cross-infection. Despite the small number of rodents included, our results showed that genetically different PUUV strains and in a lesser extent their interaction with sympatric bank voles, could affect virus replication and diversity. This could impact PUUV excretion/transmission between rodents and to humans and in turn at least partly shape NE epidemiology in France.

Detection and Genetic Characterization of Puumala Orthohantavirus S-Segment in Areas of France Non-Endemic for Nephropathia Epidemica.

Puumala virus (PUUV) in Europe causes nephropathia epidemica (NE), a mild form of hemorrhagic fever with renal syndrome (HFRS). The incidence of NE is highly heterogeneous spatially, whereas the geographic distribution of the wild reservoir of PUUV, the bank vole, is essentially homogeneous. Our understanding of the processes driving this heterogeneity remains incomplete due to gaps in knowledge. Little is known about the current distribution and genetic variation of PUUV in the areas outside the well-identified zones of NE endemicity. We trapped bank voles in four forests in French regions in which NE is considered non-endemic, but sporadic NE cases have been reported recently. We tested bank voles for anti-PUUV IgG and characterized the S segment sequences of PUUV from seropositive animals. Phylogenetic analyses revealed specific amino-acid signatures and genetic differences between PUUV circulating in non-endemic and nearby NE-endemic areas. We also showed, in temporal surveys, that the amino-acid sequences of PUUV had undergone fewer recent changes in areas non-endemic for NE than in endemic areas. The evolutionary history of the current French PUUV clusters was investigated by phylogeographic approaches, and the results were considered in the context of the history of French forests. Our findings highlight the need to monitor the circulation and genetics of PUUV in a larger array of bank vole populations, to improve our understanding of the risk of NE.