Interferon-stimulated gene (ISG)-expression screening reveals the specific antibunyaviral activity of ISG20

Bunyaviruses pose a significant threat to human health, prosperity and food security. In response to viral infections, interferons (IFNs) upregulate the expression of hundreds of interferon stimulated genes (ISGs) whose cumulative action can potently inhibit the replication of bunyaviruses. We used a flow cytometry-based method to screen the ability of ∼500 unique ISGs from humans and rhesus macaques to inhibit the replication of Bunyamwera orthobunyavirus (BUNV), the prototype of both the Peribunyaviridae family and Bunyavirales order. Candidates possessing antibunyaviral activity were further examined using a panel of divergent bunyaviruses. Interestingly, one candidate, ISG20, exhibited potent antibunyaviral activity against most viruses examined from the Peribunyaviridae, Hantaviridae and Nairoviridae families, whereas phleboviruses (Phenuiviridae) largely escaped inhibition. Similar to other viruses known to be targeted by ISG20, the antibunyaviral activity of ISG20 is dependent upon its functional ribonuclease activity. Through use of an infectious VLP assay (based on the BUNV minigenome system), we confirmed that gene expression from all 3 viral segments is strongly inhibited by ISG20. Using in vitro evolution, we generated a substantially ISG20-resistant BUNV and mapped the determinants of ISG20 sensitivity/resistance. Taken together, we report that ISG20 is a broad and potent antibunyaviral factor yet some bunyaviruses are remarkably ISG20 resistant. Thus, ISG20 sensitivity/resistance could influence the pathogenesis of bunyaviruses, many of which are emerging viruses of clinical or veterinary significance

Sensitivity to BST-2 restriction correlates with Orthobunyavirus host range

Orthobunyaviruses include several recently emerging viruses of significant medical and veterinary importance. There is currently very limited understanding on what determines the host species range of these pathogens. In this study we discovered that BST-2/tetherin restricts orthobunyavirus replication in a host-specific manner. We show that viruses with human tropism (Oropouche virus and La Crosse virus) are restricted by sheep BST-2 but not by the human orthologue, while viruses with ruminant tropism (Schmallenberg virus and others) are restricted by human BST-2 but not by the sheep orthologue. We also show that BST-2 blocks orthobunyaviruses replication by reducing the amount of envelope glycoprotein into viral particles egressing from infected cells. This is the first study identifying a restriction factor that correlates with species susceptibility to orthobunyavirus infection. This work provides insight to help us dissect the adaptive changes that bunyaviruses require to cross the species barrier and emerge into new species

Role of the cytosolic tails of Rift Valley fever virus envelope glycoproteins in viral morphogenesis

International audienceThe correct folding, heterodimerization and trafficking of Gn/Gc envelope glycoproteins of Rift Valley fever virus, RVFV (Bunyaviridae and Phlebovirus genus) are essential for Golgi assembly and budding of viral particles. The Gn and Gc carboxy-terminus contain a Golgi targeting and an ER-retrieval signal, respectively. We generated RVFV-like particles with mutations in the cytosolic tails of Gn or Gc and identified regions important for release of infectious particles. The role of specific amino-acids in these regions was further investigated by creating recombinant mutant viruses by reverse-genetics. Residues outside the suspected Golgi targeting motif, i.e. the di-lysine K29-K30 motif and the N43, R44 and I46 residues of the Gn cytosolic domain, appeared important for Golgi localization and RNP packaging. Concerning the Gc tail, replacement of K2 or K3 in the di-lysine motif, had a drastic impact on Gn trafficking and induced an important organelle redistribution and cell remodeling, greatly affecting particle formation and release

Fixation of Unstable Femoral Juvenile Osteochondritis Dissecans Lesions with Bioabsorbable Pins—Clinical and Radiographic Outcomes

Juvenile Osteochondritis Dissecans (JOCD) is a common reason for knee pain among children. The aim of this case study was to report on clinical and radiographic outcomes after fixation of an osteochondral fragment with bioabsorbable pins in children with open growth plates. We hypothesized that surgical treatment with this technique will result in good function, high rates of radiographic healing and high return to sport rates. A total of 13 knees in 12 patients (6 male, 6 female) with a median of 13 years (11, 17) were evaluated retrospectively at a minimum clinical follow-up of 24 months. Inclusion criteria were defined as open growth plates and an unstable osteochondral lesion grade III or IV. The clinical outcome was evaluated utilizing three standardized patient-reported outcome scores (Tegner Activity Scale [TAS], Knee Injury and Osteoarthritis Outcome Score [KOOS], Lysholm Score). All patients underwent magnetic resonance imaging 15 months (3, 34) after surgical treatment and defect healing was evaluated utilizing a modified version of the Magnetic Resonance Observation of Cartilage Repair Tissue (MOCART) score. Due to the small sample size, the data was reported descriptively. The interobserver variability was calculated with the Spearman rank correlation coefficient. Comparisons were made with Wilcoxon sign rank test (or sign test). At final follow-up the median KOOS Score was 98% (79.2%, 100%) and the median Lysholm Score was 94 (69, 100) points. The Tegner Activity Scale was 7 (4, 10) points preoperatively and 7 (4,10) points postoperatively (p = 0.5). Complete bony ingrowth occurred in 9 knees (69%), complete cartilage defect repair in 10 knees (77%) and integration to the border zone was found in 11 knees (85%) 15 (3, 34) months following surgical treatment. Fixation of osteochondral fragments with bioabsorbable pins resulted in good functional and radiographic outcomes, a high return to sport- and a low complication rate among children with open growth plates

Computational characterization and evaluation of deformation behavior of spherulite of high density polyethylene in mesoscale domain

We present a rigorously validated and highly sensitive confirmatory real-time RT-PCR assay (1A assay) that can be used in combination with the previously reported upE assay. Two additional RT-PCR assays for sequencing are described, targeting the RdRp gene (RdRpSeq assay) and N gene (NSeq assay), where an insertion/deletion polymorphism might exist among different hCoV-EMC strains. Finally, a simplified and biologically safe protocol for detection of antibody response by immunofluorescence microscopy was developed using convalescent patient serum