Resurgence of Ebola virus in 2021 in Guinea suggests a new paradigm for outbreaks.

Seven years after the declaration of the first epidemic of Ebola virus disease in Guinea, the country faced a new outbreak-between 14 February and 19 June 2021-near the epicentre of the previous epidemic1,2. Here we use next-generation sequencing to generate complete or near-complete genomes of Zaire ebolavirus from samples obtained from 12 different patients. These genomes form a well-supported phylogenetic cluster with genomes from the previous outbreak, which indicates that the new outbreak was not the result of a new spillover event from an animal reservoir. The 2021 lineage shows considerably lower divergence than would be expected during sustained human-to-human transmission, which suggests a persistent infection with reduced replication or a period of latency. The resurgence of Zaire ebolavirus from humans five years after the end of the previous outbreak of Ebola virus disease reinforces the need for long-term medical and social care for patients who survive the disease, to reduce the risk of re-emergence and to prevent further stigmatization.

Identification of oncolytic vaccinia restriction factors in canine high-grade mammary tumor cells using single-cell transcriptomics.

Mammary carcinoma, including triple-negative breast carcinomas (TNBC) are tumor-types for which human and canine pathologies are closely related at the molecular level. The efficacy of an oncolytic vaccinia virus (VV) was compared in low-passage primary carcinoma cells from TNBC versus non-TNBC. Non-TNBC cells were 28 fold more sensitive to VV than TNBC cells in which VV replication is impaired. Single-cell RNA-seq performed on two different TNBC cell samples, infected or not with VV, highlighted three distinct populations: naïve cells, bystander cells, defined as cells exposed to the virus but not infected and infected cells. The transcriptomes of these three populations showed striking variations in the modulation of pathways regulated by cytokines and growth factors. We hypothesized that the pool of genes expressed in the bystander populations was enriched in antiviral genes. Bioinformatic analysis suggested that the reduced activity of the virus was associated with a higher mesenchymal status of the cells. In addition, we demonstrated experimentally that high expression of one gene, DDIT4, is detrimental to VV production. Considering that DDIT4 is associated with a poor prognosis in various cancers including TNBC, our data highlight DDIT4 as a candidate resistance marker for oncolytic poxvirus therapy. This information could be used to design new generations of oncolytic poxviruses. Beyond the field of gene therapy, this study demonstrates that single-cell transcriptomics can be used to identify cellular factors influencing viral replication.

Synthesis, antiviral and antitumor activities investigations of a series of Ribavirin C-nucleoside analogue prodrugs.

Phosphoramidates obtained according to the ProTide strategy are known for their ability to increase the biological activity of various nucleosides. A series of such prodrugs of SRO-91, a non-natural ribofuranosyl-1,2,3-triazole C-nucleoside obtained by a synthetic sequence involving an indium mediated alkynylation and a Huisgen cycloaddition, was prepared and the antitumor activity on 3 strains of tumor cells was investigated. Two compounds 9a and 9c exhibited interesting cell proliferative inhibitions (IC50 = 2.5-12.1 µM) on two cell lines (pancreas and lung). Moreover, concerning the antiviral activity, another phosphoramidate 14 bearing a different aryl masking group exhibited an IC50 of 5 µM on Crimean-Congo Hemorrhagic Fever orthonairovirus. In both cases, free SRO-91 presented no activity on these cell lines.

Crimean-Congo hemorrhagic fever virus circulating among sheep of Portugal: a nationwide serosurvey assessment.

Crimean-Congo hemorrhagic fever virus (CCHFV) is a widespread zoonotic pathogen that can cause mild to severe hemorrhagic disease in humans. CCHFV may be transmitted through direct contact with tissue or blood of viremic animals; however, the primary transmission route is through infected tick bites. CCHFV RNA has been detected in ticks feeding on domestic and wild animals in western Spain, suggesting an established circulation of CCHFV in Western Europe. Ruminants have been recognized as important CCHFV reservoirs and have been linked to human cases in endemic regions. Given the emergence of CCHF in neighboring Spain, and a report of two CCHFV seropositive humans in southern Portugal in 1985, we investigated the potential circulation of this virus in the country by performing a nationwide anti-CCHFV IgG serosurvey in sentinel sheep of Portugal. Sera (n = 459) randomly selected from widely distributed farms (n = 20) of Portugal were tested using a commercial double-antigen enzyme-linked immunosorbent assay, yielding an overall seroprevalence of 0.4% (95% confidence interval [CI] 0.04-1.56%). Positive sheep were from the southern region of Portugal (Alentejo region), which raise the seroprevalence of this region to 0.74% (95% CI 0.09-2.66%). This is the first study reporting the presence of CCHFV antibodies in sheep of Portugal, thus suggesting a geographical expansion of CCHFV to this country. It seems likely that CCHFV may exist focally in southern Portugal.

Fatal Cowpox Virus Infection in Human Fetus, France, 2017.

Cowpox virus (CPXV) has an animal reservoir and is typically transmitted to humans by contact with infected animals. In 2017, CPXV infection of a pregnant woman in France led to the death of her fetus. Fetal death after maternal orthopoxvirus (smallpox) vaccination has been reported; however, this patient had not been vaccinated. Investigation of the patient's domestic animals failed to demonstrate prevalence of CPXV infection among them. The patient's diagnosis was confirmed by identifying CPXV DNA in all fetal and maternal biopsy samples and infectious CPXV in biopsy but not plasma samples. This case of fetal death highlights the risk for complications of orthopoxvirus infection during pregnancy. Among orthopoxviruses, fetal infection has been reported for variola virus and vaccinia virus; our findings suggest that CPXV poses the same threats for infection complications as vaccinia virus.

Crimean-Congo Hemorrhagic Fever Virus Antibodies among Livestock on Corsica, France, 2014-2016.

We conducted a serologic survey for Crimean-Congo hemorrhagic fever virus antibodies in livestock (cattle, sheep, and goats; N = 3,890) on Corsica (island of France) during 2014-2016. Overall, 9.1% of animals were seropositive, suggesting this virus circulates on Corsica. However, virus identification is needed to confirm these results.

Correction to: Combination of ELISA screening and seroneutralisation tests to expedite Zika virus seroprevalence studies.

In the original publication of this article [1], Table 1 has some errors.

Description and characterization of a novel live-attenuated tri-segmented Machupo virus in Guinea pigs.

BACKGROUND: Machupo virus (MACV) is a member of the Mammarenavirus genus, Arenaviridae family and is the etiologic agent of Bolivian hemorrhagic fever, which causes small outbreaks or sporadic cases. Several other arenaviruses in South America Junín virus (JUNV) in Argentina, Guanarito in Venezuela, Sabiá in Brazil and Chapare in Bolivia, also are responsible for human hemorrhagic fevers. Among these arenaviruses, JUNV caused thousands of human cases until 1991, when the live attenuated Candid #1 vaccine, was used. Other than Candid #1 vaccine, few other therapeutic or prophylactic treatments exist. Therefore, new strategies for production of safe countermeasures with broad spectrum activity are needed. FINDINGS: We tested a tri-segmented MACV, a potential vaccine candidate with several mutations, (r3MACV). In cell culture, r3MACV showed a 2-log reduction in infectious virus particle production and the MACV inhibition of INF-1ß was removed from the construct and produced by infected cells. Furthermore, in an animal experiment, r3MACV was able to protect 50% of guinea pigs from a simultaneous lethal JUNV challenge. Protected animals didn't display clinical symptoms nor were virus particles found in peripheral blood (day 14) or in organs (day 28 post-inoculation). The r3MACV provided a higher protection than the Candid #1 vaccine. CONCLUSIONS: The r3MACV provides a potential countermeasure against two South America arenaviruses responsible of human hemorrhagic fever.

Combination of ELISA screening and seroneutralisation tests to expedite Zika virus seroprevalence studies.

Here we propose a strategy allowing implementing efficient and practicable large-scale seroepidemiological studies for Zika Virus (ZIKV). It combines screening by a commercial NS1 protein-based Zika IgG ELISA, and confirmation by a cytopathic effect-based virus neutralization test (CPE-based VNT). In post-epidemic samples from Martinique Island blood donors (a population with a dengue seroprevalence above 90%), this strategy allowed reaching specificity and sensitivity values over 98%. The CPE-based VNT consists of recording CPE directly under the optical microscope, which is easy to identify with ZIKV strain H/PF/2013 at day 5 pi. Overall, considered that CPE-based VNT is cost effective and widely automatable, the NS1 protein-based Zika IgG ELISA+CPE-based VNT combination strategy represents a convenient tool to expedite ZIKV seroprevalence studies.

Enhancement of Ebola Virus Infection via Ficolin-1 Interaction with the Mucin Domain of GP Glycoprotein.

UNLABELLED: Ebola virus infection requires the surface viral glycoprotein to initiate entry into the target cells. The trimeric glycoprotein is a highly glycosylated viral protein which has been shown to interact with host C-type lectin receptors and the soluble complement recognition protein mannose-binding lectin, thereby enhancing viral infection. Similarly to mannose-binding lectin, ficolins are soluble effectors of the innate immune system that recognize particular glycans at the pathogen surface. In this study, we demonstrate that ficolin-1 interacts with the Zaire Ebola virus (EBOV) glycoprotein, and we characterized this interaction by surface plasmon resonance spectroscopy. Ficolin-1 was shown to bind to the viral glycoprotein with a high affinity. This interaction was mediated by the fibrinogen-like recognition domain of ficolin-1 and the mucin-like domain of the viral glycoprotein. Using a ficolin-1 control mutant devoid of sialic acid-binding capacity, we identified sialylated moieties of the mucin domain to be potential ligands on the glycoprotein. In cell culture, using both pseudotyped viruses and EBOV, ficolin-1 was shown to enhance EBOV infection independently of the serum complement. We also observed that ficolin-1 enhanced EBOV infection on human monocyte-derived macrophages, described to be major viral target cells,. Competition experiments suggested that although ficolin-1 and mannose-binding lectin recognized different carbohydrate moieties on the EBOV glycoprotein, the observed enhancement of the infection likely depended on a common cellular receptor/partner. In conclusion, ficolin-1 could provide an alternative receptor-mediated mechanism for enhancing EBOV infection, thereby contributing to viral subversion of the host innate immune system. IMPORTANCE: A specific interaction involving ficolin-1 (M-ficolin), a soluble effector of the innate immune response, and the glycoprotein (GP) of EBOV was identified. Ficolin-1 enhanced virus infection instead of tipping the balance toward its elimination. An interaction between the fibrinogen-like recognition domain of ficolin-1 and the mucin-like domain of Ebola virus GP occurred. In this model, the enhancement of infection was shown to be independent of the serum complement. The facilitation of EBOV entry into target host cells by the interaction with ficolin-1 and other host lectins shunts virus elimination, which likely facilitates the survival of the virus in infected host cells and contributes to the virus strategy to subvert the innate immune response.

Past, present and future therapeutic and prophylactic strategies against arenaviruses responsible of human hemorrhagic fever.

For most viral hemorrhagic fevers caused by arenaviruses, no prophylactic vaccine is available yet. Only one therapeutic treatment is currently available and should be administered at the early stages of the infection. This is particularly problematic as these diseases are difficult to diagnose and cure. Lassa fever is the most important pathology caused by arenaviruses, including millions of people at risk in West Africa. For decades, promising studies focusing on the development of vaccine candidates targeting Lassa virus have been published, but no vaccine candidate had reached the clinical phase. The second arenavirus in terms of number of human infections is the Junín virus in Argentina. The Junín infected case number has drastically decreased since the use of the Candid #1 vaccine. This review summarizes past and present experimental studies regarding treatments against arenaviruses responsible for human hemorrhagic fevers from a prophylactic and therapeutic point of view. It also discusses future breakthroughs to get available and effective treatments.

Crystal Structure of the Vaccinia Virus Uracil-DNA Glycosylase in Complex with DNA.

Vaccinia virus polymerase holoenzyme is composed of the DNA polymerase catalytic subunit E9 associated with its heterodimeric co-factor A20·D4 required for processive genome synthesis. Although A20 has no known enzymatic activity, D4 is an active uracil-DNA glycosylase (UNG). The presence of a repair enzyme as a component of the viral replication machinery suggests that, for poxviruses, DNA synthesis and base excision repair is coupled. We present the 2.7 Å crystal structure of the complex formed by D4 and the first 50 amino acids of A20 (D4·A201-50) bound to a 10-mer DNA duplex containing an abasic site resulting from the cleavage of a uracil base. Comparison of the viral complex with its human counterpart revealed major divergences in the contacts between protein and DNA and in the enzyme orientation on the DNA. However, the conformation of the dsDNA within both structures is very similar, suggesting a dominant role of the DNA conformation for UNG function. In contrast to human UNG, D4 appears rigid, and we do not observe a conformational change upon DNA binding. We also studied the interaction of D4·A201-50 with different DNA oligomers by surface plasmon resonance. D4 binds weakly to nonspecific DNA and to uracil-containing substrates but binds abasic sites with a Kd of <1.4 µm. This second DNA complex structure of a family I UNG gives new insight into the role of D4 as a co-factor of vaccinia virus DNA polymerase and allows a better understanding of the structural determinants required for UNG action.

Parapoxviruses, little-known zoonotic pathogens.

Parapoxviruses, double-stranded DNA viruses of the Poxviridæ family, are etiologic agents of cutaneaous infectious diseases among farm animals. These highly contagious viruses are responsible for wide outbreaks among livestock. The clinical manifestations are generally mild and consist of cutaneous or mucosal lesions, which resolve spontaneously within a few weeks. However, secondary bacterial or fungal infections on the lesion sites can aggravate the symptoms. Sore lesions located within the oral cavity and on the udders can impair feeding or nursing, thus leading to death. Livestock parapoxviruses can infect humans by direct or indirect transmission and affect mainly farmers, slaughters and veterinarians. Human symptoms generally consist of small cutaneous lesions located at the inoculation points but more severe forms can occur, peculiarly in immunocompromised persons. The parapoxvirus epidemiology is poorly understood: their respective host range and ecology among wild animals are to be clarified. The identification of parapoxviruses among marine mammals suggests that the genetic diversity within the genus is still underestimated.

Concomitant human infections with 2 cowpox virus strains in related cases, France, 2011.

We investigated 4 related human cases of cowpox virus infection reported in France during 2011. Three patients were infected by the same strain, probably transmitted by imported pet rats, and the fourth patient was infected by another strain. The 2 strains were genetically related to viruses previously isolated from humans with cowpox infection in Europe.

Special Issue "Endemic Arboviruses".

Arthropod-borne viruses (Arbovirus) is an ecological term defining viruses that are maintained in nature through biological transmission between a susceptible vertebrate host and a hematophagous arthropod such as a mosquito [...].

The Relationship between DUGBE Virus Infection and Autophagy in Epithelial Cells.

Dugbe orthonairovirus (DUGV) is a tick-borne arbovirus within the order Bunyavirales. Although displaying mild pathogenic potential, DUGV is genetically related to the Crimean-Congo hemorrhagic fever virus (CCHFV), another orthonairovirus that causes severe liver dysfunction and hemorrhagic fever with a high mortality rate in humans. As we previously observed that CCHFV infection could massively recruit and lipidate MAP1LC3 (LC3), a core factor involved in the autophagic degradation of cytosolic components, we asked whether DUGV infection also substantially impacts the autophagy machinery in epithelial cells. We observed that DUGV infection does impose LC3 lipidation in cultured hepatocytes. DUGV infection also caused an upregulation of the MAP1LC3 and SQSTM1/p62 transcript levels, which were, however, more moderate than those seen during CCHFV infection. In contrast, unlike during CCHFV infection, the modulation of core autophagy factors could influence both LC3 lipidation and viral particle production: the silencing of ATG5 and/or ATG7 diminished the induction of LC3 lipidation and slightly upregulated the level of infectious DUGV particle production. Overall, the results are compatible with the notion that in epithelial cells infected with DUGV in vitro, the autophagy machinery may be recruited to exert a certain level of restriction on viral replication. Thus, the relationship between DUGV infection and autophagy in epithelial cells appears to present both similarities and distinctions with that seen during CCHFV infection.

Reduction in SARS-CoV-2 Virus Infectivity in Human and Hamster Feces.

OBJECTIVE: There is extensive evidence that SARS-CoV-2 replicates in the gastrointestinal tract. However, the infectivity of virions in feces is poorly documented. Although the primary mode of transmission is airborne, the risk of transmission from contaminated feces remains to be assessed. DESIGN: The persistence of SARS-CoV-2 (infectivity and RNA) in human and animal feces was evaluated by virus isolation on cell culture and RT-qPCR, respectively. The exposure of golden Syrian hamsters to experimentally contaminated feces through intranasal inoculation has also been tested to assess the fecal-oral transmission route. RESULTS: For periods that are compatible with average intestinal transit, the SARS-CoV-2 genome was noticeably stable in human and animal feces, contrary to the virus infectivity that was reduced in a time- and temperature-dependent manner. In human stools, this reduction was variable depending on the donors. Viral RNA was excreted in the feces of infected hamsters, but exposure of naïve hamsters to feces of infected animals did not lead to any productive infection. Conversely, hamsters could be experimentally infected following exposure to spiked fresh feces. CONCLUSION: Infection following exposure to naturally contaminated feces has been suspected but has not been established so far. The present work demonstrates that SARS-CoV-2 rapidly lost infectivity in spiked or naturally infected feces. Although the possibility of persistent viral particles in human or animal feces cannot be fully ruled out, SARS-CoV-2 transmission after exposure to contaminated feces is unlikely.