Ebola virus VP35 interacts non-covalently with ubiquitin chains to promote viral replication.

Ebolavirus (EBOV) belongs to a family of highly pathogenic viruses that cause severe hemorrhagic fever in humans. EBOV replication requires the activity of the viral polymerase complex, which includes the cofactor and Interferon antagonist VP35. We previously showed that the covalent ubiquitination of VP35 promotes virus replication by regulating interactions with the polymerase complex. In addition, VP35 can also interact non-covalently with ubiquitin (Ub); however, the function of this interaction is unknown. Here, we report that VP35 interacts with free (unanchored) K63-linked polyUb chains. Ectopic expression of Isopeptidase T (USP5), which is known to degrade unanchored polyUb chains, reduced VP35 association with Ub and correlated with diminished polymerase activity in a minigenome assay. Using computational methods, we modeled the VP35-Ub non-covalent interacting complex, identified the VP35-Ub interacting surface, and tested mutations to validate the interface. Docking simulations identified chemical compounds that can block VP35-Ub interactions leading to reduced viral polymerase activity. Treatment with the compounds reduced replication of infectious EBOV in cells and in vivo in a mouse model. In conclusion, we identified a novel role of unanchored polyUb in regulating Ebola virus polymerase function and discovered compounds that have promising anti-Ebola virus activity.

Envelope protein ubiquitination drives entry and pathogenesis of Zika virus.

Zika virus (ZIKV) belongs to the family Flaviviridae, and is related to other viruses that cause human diseases. Unlike other flaviviruses, ZIKV infection can cause congenital neurological disorders and replicates efficiently in reproductive tissues1-3. Here we show that the envelope protein (E) of ZIKV is polyubiquitinated by the E3 ubiquitin ligase TRIM7 through Lys63 (K63)-linked polyubiquitination. Accordingly, ZIKV replicates less efficiently in the brain and reproductive tissues of Trim7-/- mice. Ubiquitinated E is present on infectious virions of ZIKV when they are released from specific cell types, and enhances virus attachment and entry into cells. Specifically, K63-linked polyubiquitin chains directly interact with the TIM1 (also known as HAVCR1) receptor of host cells, which enhances virus entry in cells as well as in brain tissue in vivo. Recombinant ZIKV mutants that lack ubiquitination are attenuated in human cells and in wild-type mice, but not in live mosquitoes. Monoclonal antibodies against K63-linked polyubiquitin specifically neutralize ZIKV and reduce viraemia in mice. Our results demonstrate that the ubiquitination of ZIKV E is an important determinant of virus entry, tropism and pathogenesis.

Dengue virus NS5 degrades ERC1 during infection to antagonize NF-kB activation.

Dengue virus (DENV) is the most important human virus transmitted by mosquitos. Dengue pathogenesis is characterized by a large induction of proinflammatory cytokines. This cytokine induction varies among the four DENV serotypes (DENV1 to 4) and poses a challenge for live DENV vaccine design. Here, we identify a viral mechanism to limit NF-κB activation and cytokine secretion by the DENV protein NS5. Using proteomics, we found that NS5 binds and degrades the host protein ERC1 to antagonize NF-κB activation, limit proinflammatory cytokine secretion, and reduce cell migration. We found that ERC1 degradation involves unique properties of the methyltransferase domain of NS5 that are not conserved among the four DENV serotypes. By obtaining chimeric DENV2 and DENV4 viruses, we map the residues in NS5 for ERC1 degradation, and generate recombinant DENVs exchanging serotype properties by single amino acid substitutions. This work uncovers a function of the viral protein NS5 to limit cytokine production, critical to dengue pathogenesis. Importantly, the information provided about the serotype-specific mechanism for counteracting the antiviral response can be applied to improve live attenuated vaccines.

Ubiquitination of Ebola virus VP35 at lysine 309 regulates viral transcription and assembly.

Ebola virus (EBOV) VP35 is a polyfunctional protein involved in viral genome packaging, viral polymerase function, and host immune antagonism. The mechanisms regulating VP35's engagement in different functions are not well-understood. We previously showed that the host E3 ubiquitin ligase TRIM6 ubiquitinates VP35 at lysine 309 (K309) to facilitate virus replication. However, how K309 ubiquitination regulates the function of VP35 as the viral polymerase co-factor and the precise stage(s) of the EBOV replication cycle that require VP35 ubiquitination are not known. Here, we generated recombinant EBOVs encoding glycine (G) or arginine (R) mutations at VP35/K309 (rEBOV-VP35/K309G/-R) and show that both mutations prohibit VP35/K309 ubiquitination. The K309R mutant retains dsRNA binding and efficient type-I Interferon (IFN-I) antagonism due to the basic residue conservation. The rEBOV-VP35/K309G mutant loses the ability to efficiently antagonize the IFN-I response, while the rEBOV-VP35/K309R mutant's suppression is enhanced. The replication of both mutants was significantly attenuated in both IFN-competent and -deficient cells due to impaired interactions with the viral polymerase. The lack of ubiquitination on VP35/K309 or TRIM6 deficiency disrupts viral transcription with increasing severity along the transcriptional gradient. This disruption of the transcriptional gradient results in unbalanced viral protein production, including reduced synthesis of the viral transcription factor VP30. In addition, lack of ubiquitination on K309 results in enhanced interactions with the viral nucleoprotein and premature nucleocapsid packaging, leading to dysregulation of virus assembly. Overall, we identified a novel role of VP35 ubiquitination in coordinating viral transcription and assembly.

The Host E3-Ubiquitin Ligase TRIM6 Ubiquitinates the Ebola Virus VP35 Protein and Promotes Virus Replication.

Ebola virus (EBOV), a member of the Filoviridae family, is a highly pathogenic virus that causes severe hemorrhagic fever in humans and is responsible for epidemics throughout sub-Saharan, central, and West Africa. The EBOV genome encodes VP35, an important viral protein involved in virus replication by acting as an essential cofactor of the viral polymerase as well as a potent antagonist of the host antiviral type I interferon (IFN-I) system. By using mass spectrometry analysis and coimmunoprecipitation assays, we show here that VP35 is ubiquitinated on lysine 309 (K309), a residue located on its IFN antagonist domain. We also found that VP35 interacts with TRIM6, a member of the E3-ubiquitin ligase tripartite motif (TRIM) family. We recently reported that TRIM6 promotes the synthesis of unanchored K48-linked polyubiquitin chains, which are not covalently attached to any protein, to induce efficient antiviral IFN-I-mediated responses. Consistent with this notion, VP35 also associated noncovalently with polyubiquitin chains and inhibited TRIM6-mediated IFN-I induction. Intriguingly, we also found that TRIM6 enhances EBOV polymerase activity in a minigenome assay and TRIM6 knockout cells have reduced replication of infectious EBOV, suggesting that VP35 hijacks TRIM6 to promote EBOV replication through ubiquitination. Our work provides evidence that TRIM6 is an important host cellular factor that promotes EBOV replication, and future studies will focus on whether TRIM6 could be targeted for therapeutic intervention against EBOV infection.IMPORTANCE EBOV belongs to a family of highly pathogenic viruses that cause severe hemorrhagic fever in humans and other mammals with high mortality rates (40 to 90%). Because of its high pathogenicity and lack of licensed antivirals and vaccines, EBOV is listed as a tier 1 select-agent risk group 4 pathogen. An important mechanism for the severity of EBOV infection is its suppression of innate immune responses. The EBOV VP35 protein contributes to pathogenesis, because it serves as an essential cofactor of the viral polymerase as well as a potent antagonist of innate immunity. However, how VP35 function is regulated by host cellular factors is poorly understood. Here, we report that the host E3-ubiquitin ligase TRIM6 promotes VP35 ubiquitination and is important for efficient virus replication. Therefore, our study identifies a new host factor, TRIM6, as a potential target in the development of antiviral drugs against EBOV.

Effect of oleic acid on store-operated calcium entry in immune-competent cells.

PURPOSE: To study the mechanism by which oleic acid (OA) (C18:1) exerts its beneficial effects on immune-competent cells. Since store-operated Ca2+ entry (SOCE) is a Ca2+ influx pathway involved in the control of multiple physiological processes including cell proliferation, we studied the effect of OA in Ca2+ signals of Jurkat T cells and THP-1 monocytes, paying particular attention to SOCE. METHODS: Changes in [Ca2+]i were measured using the Fura-2 fluorescence dye. Mn2+ uptake was monitored as a rate of quenching of Fura-2 fluorescence measured at the Ca2+-insensitive wavelengths. Thapsigargin was used to induce SOCE in Fura-2-loaded cells. RESULTS: We showed a clear dose-dependent SOCE-inhibitory effect of OA in both cell lines. Such an inhibitory effect was PKC independent and totally restored by albumin, suggesting that OA exerts its effect somewhere in the membrane. We also demonstrated that OA induces increases in [Ca2+]i partly mediated by an extracellular Ca2+ influx through econazole-insensitive channels. Finally, we compared the effect of OA with stearic acid (C18:0), assuming the emerged evidence concerning the link between saturated fats and inflammation disorders. Stearic acid failed to inhibit SOCE, independently on the concentration tested, thus intensifying the physiological relevance of our findings. CONCLUSION: We suggest a physiological pathway for the beneficial effects of OA in inflammation.

The Dual Role of TRIM7 in Viral Infections.

The E3 ubiquitin ligase TRIM7 is known to have dual roles during viral infections. Like other TRIM proteins, TRIM7 can regulate the IFN pathway via the regulation of the cytosolic receptors RIG-I or MDA-5, which promote the production of type I interferons (IFN-I) and antiviral immune responses. Alternatively, under certain infectious conditions, TRIM7 can negatively regulate IFN-I signaling, resulting in increased virus replication. A growing body of evidence has also shown that TRIM7 can, in some cases, ubiquitinate viral proteins to promote viral replication and pathogenesis, while in other cases it can promote degradation of viral proteins through the proteasome, reducing virus infection. TRIM7 can also regulate the host inflammatory response and modulate the production of inflammatory cytokines, which can lead to detrimental inflammation. TRIM7 can also protect the host during infection by reducing cellular apoptosis. Here, we discuss the multiple functions of TRIM7 during viral infections and its potential as a therapeutic target.

TRIM7 ubiquitinates SARS-CoV-2 membrane protein to limit apoptosis and viral replication.

SARS-CoV-2 is a highly transmissible virus that causes COVID-19 disease. Mechanisms of viral pathogenesis include excessive inflammation and viral-induced cell death, resulting in tissue damage. We identified the host E3-ubiquitin ligase TRIM7 as an inhibitor of apoptosis and SARS-CoV-2 replication via ubiquitination of the viral membrane (M) protein. Trim7 -/- mice exhibited increased pathology and virus titers associated with epithelial apoptosis and dysregulated immune responses. Mechanistically, TRIM7 ubiquitinates M on K14, which protects cells from cell death. Longitudinal SARS-CoV-2 sequence analysis from infected patients revealed that mutations on M-K14 appeared in circulating variants during the pandemic. The relevance of these mutations was tested in a mouse model. A recombinant M-K14/K15R virus showed reduced viral replication, consistent with the role of K15 in virus assembly, and increased levels of apoptosis associated with the loss of ubiquitination on K14. TRIM7 antiviral activity requires caspase-6 inhibition, linking apoptosis with viral replication and pathology.

Pathogenesis of Zika Virus Infection.

Zika virus (ZIKV) is an emerging virus from the Flaviviridae family that is transmitted to humans by mosquito vectors and represents an important health problem. Infections in pregnant women are of major concern because of potential devastating consequences during pregnancy and have been associated with microcephaly in newborns. ZIKV has a unique ability to use the host machinery to promote viral replication in a tissue-specific manner, resulting in characteristic pathological disorders. Recent studies have proposed that the host ubiquitin system acts as a major determinant of ZIKV tropism by providing the virus with an enhanced ability to enter new cells. In addition, ZIKV has developed mechanisms to evade the host immune response, thereby allowing the establishment of viral persistence and enhancing viral pathogenesis. We discuss recent reports on the mechanisms used by ZIKV to replicate efficiently, and we highlight potential new areas of research for the development of therapeutic approaches.

Ebola Virus VP35 Interacts Non-Covalently with Ubiquitin Chains to Promote Viral Replication Creating New Therapeutic Opportunities.

Ebolavirus (EBOV) belongs to a family of highly pathogenic viruses that cause severe hemorrhagic fever in humans. EBOV replication requires the activity of the viral polymerase complex, which includes the co-factor and Interferon antagonist VP35. We previously showed that the covalent ubiquitination of VP35 promotes virus replication by regulating interactions with the polymerase complex. In addition, VP35 can also interact non-covalently with ubiquitin (Ub); however, the function of this interaction is unknown. Here, we report that VP35 interacts with free (unanchored) K63-linked polyUb chains. Ectopic expression of Isopeptidase T (USP5), which is known to degrade unanchored polyUb chains, reduced VP35 association with Ub and correlated with diminished polymerase activity in a minigenome assay. Using computational methods, we modeled the VP35-Ub non-covalent interacting complex, identified the VP35-Ub interacting surface and tested mutations to validate the interface. Docking simulations identified chemical compounds that can block VP35-Ub interactions leading to reduced viral polymerase activity that correlated with reduced replication of infectious EBOV. In conclusion, we identified a novel role of unanchored polyUb in regulating Ebola virus polymerase function and discovered compounds that have promising anti-Ebola virus activity.

Impact of prior dengue virus infection on Zika virus infection during pregnancy in marmosets.

Zika virus (ZIKV) infection during pregnancy causes severe developmental defects in newborns, termed congenital Zika syndrome (CZS). Factors contributing to a surge in ZIKV-associated CZS are poorly understood. One possibility is that ZIKV may exploit the antibody-dependent enhancement of infection mechanism, mediated by cross-reactive antibodies from prior dengue virus (DENV) infection, which may exacerbate ZIKV infection during pregnancy. In this study, we investigated the impact of prior DENV infection or no DENV infection on ZIKV pathogenesis during pregnancy in a total of four female common marmosets with five or six fetuses per group. The results showed that negative-sense viral RNA copies increased in the placental and fetal tissues of DENV-immune dams but not in DENV-naïve dams. In addition, viral proteins were prevalent in endothelial cells, macrophages, and neonatal Fc receptor-expressing cells in the placental trabeculae and in neuronal cells in the brains of fetuses from DENV-immune dams. DENV-immune marmosets maintained high titers of cross-reactive ZIKV-binding antibodies that were poorly neutralizing, raising the possibility that these antibodies might be involved in the exacerbation of ZIKV infection. These findings need to be verified in a larger study, and the mechanism involved in the exacerbation of ZIKV infection in DENV-immune marmosets needs further investigation. However, the results suggest a potential negative impact of preexisting DENV immunity on subsequent ZIKV infection during pregnancy in vivo.

Validation of the TRI-SCORE in patients undergoing surgery for isolated tricuspid regurgitation.

INTRODUCTION: Estimation of peri-procedural risk in patients with tricuspid regurgitation (TR) undergoing isolated tricuspid valve surgery (ITVS) is of paramount importance. The TRI-SCORE is a new surgical risk scale specifically developed for this purpose, which ranged from 0 to 12 points and included eight parameters: right-sided heart failure signs, daily dose of furosemide ≥125 mg, glomerular filtration rate <30 mL/min, elevated bilirubin (with a value of 2 points), age ≥70 years, New York Heart Association Class III-IV, left ventricular ejection fraction <60% and moderate/severe right ventricular dysfunction (with a value of 1 point). The objective of the study was to evaluate the performance of the TRI-SCORE in an independent cohort of patients undergoing ITVS. METHODS: A retrospective observational study was performed in four centres, including consecutive adult patients undergoing ITVS for TR between 2005 and 2022. The TRI-SCORE and the traditional risk scores used in cardiac surgery (Logistic EuroScore (Log-ES) and EuroScore-II (ES-II)) were applied for each patient, and discrimination and calibration of the three scores were evaluated in the entire cohort. RESULTS: A total of 252 patients were included. The mean age was 61.5±11.2 years, 164 (65.1%) patients were female, and TR mechanism was functional in 160 (63.5%) patients. The observed in-hospital mortality was 10.3%. The estimated mortality by the Log-ES, ES-II and TRI-SCORE was 8.7±7.3%, 4.7±5.3% and 11.0±16.6%, respectively. Patients with a TRI-SCORE ≤4 and >4 had an in-hospital mortality of 1.3% and 25.0%, p=0.001, respectively. The discriminatory capacity of the TRI-SCORE had a C-statistic of 0.87 (0.81-0.92), which was significantly higher than both the Log-ES (0.65 (0.54-0.75)) and ES-II (0.67 (0.58-0.79)), p=0.001 (for both comparisons). CONCLUSION: This external validation of the TRI-SCORE demonstrated good performance to predict in-hospital mortality in patients undergoing ITVS, which was significantly better than the Log-ES and ES-II, which underestimated the observed mortality. These results support the widespread use of this score as a clinical tool.

Intrapartum fetal heart rate characteristics of nonanomalous fetuses with trisomy 21.

OBJECTIVE: To compare the intrapartum fetal heart rate (FHR) characteristics and selected birth outcomes of nonanomalous trisomy 21 (T21) fetuses to matched controls. STUDY DESIGN: Nonanomalous, non-growth-restricted T21 infants were identified and matched by gestational age to presumed euploid controls. We excluded women with scheduled cesarean deliveries and multiple gestations. The incidence of abnormal FHR patterns within 3 hours of delivery and birth outcomes were compared between T21 fetuses and controls. The presence of any abnormal FHR pattern was the primary outcome. Birth outcomes included 5-minute Apgar <7, neonatal intensive care unit admission, and cesarean delivery for fetal indications. RESULTS: Forty-four T21 infants and 44 controls were compared. Of the T21 infants, 83% were postnatally diagnosed. No significant differences were noted in the primary outcome (68% versus 59%, p = 0.37) or birth outcomes. CONCLUSION: T21 is not associated with an increased incidence of abnormal FHR patterns or adverse birth outcomes compared with presumed euploid fetuses.

PD1 CD44 antiviral peptide as an inhibitor of the protein-protein interaction in dengue virus invasion.

Dengue virus (DENV) infection is mediated by the interaction between the virus envelope protein and cellular receptors of the host cells. In this study, we designed peptides to inhibit protein-protein interaction between dengue virus and CD44 receptor, which is one of the receptors used by DENV for entry. In silico model complexes were designed between domain III of the viral envelope protein of dengue virus 2 and the domain of human CD44 receptor using ClusPro 2.0, (https://cluspro.bu.edu/login.php), and inhibition peptides were designed with Rosetta Online-Server(http://rosie.rosettacommons.org/peptiderive). We identified one linear antiviral peptide of 18 amino acids derived from the human CD44 receptor, PD1 CD44. It did not show hemolysis or toxicity in HepG2 or BHK cell lines, nor did it stimulate the release of IL-1ß, IL-6, TNF-α, and IFN-γ, below 100 µM. It had an IC50 of 13.8 µM and maximum effective dose of 54.9 µM evaluated in BHK cells. The decrease in plaque-forming units/mL for DENV1, DENV2, DENV3, and DENV4 was 99.60%, 99.40%, 97.80%, and 70.50%, respectively, and similar results were obtained by RT-qPCR. Non-structural protein 1 release was decreased in pre- and co-treatment but not in post-treatment. Competition assays between the DN59 peptide, envelope protein, and the fragment of domain III "MDKLQLKGMSYSMCTGKF" of the viral envelope of DENV2 and PD1 CD44 showed that our peptide lost its antiviral activity. We demonstrated that our peptide decreased endosome formation, and we propose that it binds to the envelope protein of DENV, inhibiting viral invasion/fusion.

In Vitro Inhibition of Replication of Dengue Virus Serotypes 1-4 by siRNAs Bound to Non-Toxic Liposomes.

Dengue virus is a ssRNA+ flavivirus, which produces the dengue disease in humans. Currently, no specific treatment exists. siRNAs regulate gene expression and have been used systematically to silence viral genomes; however, they require controlled release. Liposomes show favorable results encapsulating siRNA for gene silencing. The objective herein was to design and evaluate in vitro siRNAs bound to liposomes that inhibit DENV replication. siRNAs were designed against DENV1-4 from conserved regions using siDirect2.0 and Web-BLOCK-iT™ RNAiDesigner; the initial in vitro evaluation was carried out through transfection into HepG2 cells. siRNA with silencing capacity was encapsulated in liposomes composed of D-Lin-MC3-DMA, DSPC, Chol. Cytotoxicity, hemolysis, pro-inflammatory cytokine release and antiviral activity were evaluated using plaque assay and RT-qPCR. A working concentration of siRNA was established at 40 nM. siRNA1, siRNA2, siRNA3.1, and siRNA4 were encapsulated in liposomes, and their siRNA delivery through liposomes led to a statistically significant decrease in viral titers, yielded no cytotoxicity or hemolysis and did not stimulate release of pro-inflammatory cytokines. Finally, liposomes were designed with siRNA against DENV, which proved to be safe in vitro.

Development and Evaluation from Laboratory to Field Trial of a Dual-Purpose Fracturing Nanofluid: Inhibition of Associated Formation Damage and Increasing Heavy Crude Oil Mobility.

This study aims to develop and evaluate fracturing nanofluids from the laboratory to the field trial with the dual purpose of increasing heavy crude oil mobility and reducing formation damage caused by the remaining fracturing fluid (FF). Two fumed silica nanoparticles of different sizes, and alumina nanoparticles were modified on the surface through basic and acidic treatments. The nanoparticles were characterized by transmission electron microscopy, dynamic light scattering, zeta potential and total acidity. The rheological behavior of the linear gel and the heavy crude oil after adding different chemical nature nanoparticles were measured at two concentrations of 100 and 1000 mg/L. Also, the contact angle assessed the alteration of the rock wettability. The nanoparticle with better performance was the raw fumed silica of 7 nm at 1000 mg/L. These were employed to prepare a fracturing nanofluid from a commercial FF. Both fluids were evaluated through their rheological behavior as a function of time at high pressure following the API RP39 test, and spontaneous imbibition tests were carried out to assess the FF's capacity to modify the wettability of the porous media. It was possible to conclude that the inclusion of 7 nm commercial silica nanoparticles allowed obtaining a reduction of 10 and 20% in the two breakers used in the commercial fracture fluid formulation without altering the rheological properties of the system. Displacement tests were also performed on proppant and rock samples at reservoir conditions of overburden and pore pressures of 3200 and 1200 psi, respectively, while the temperature was set at 77 °C and the flow rate at 0.3 cm3/min. According to the effective oil permeability, a decrease of 31% in the damage was obtained. Based on these results, the fracturing nanofluid was selected and used in the first worldwide field application in a Colombian oil field with a basic sediment and water (BSW%) of 100 and without oil production. After two weeks of the hydraulic fracture operation, crude oil was produced. Finally, one year after this work, crude oil viscosity and BSW% kept showing reductions near 75% and 33%, respectively; and having passed two years, the cumulative incremental oil production is around 120,000 barrels.

Co-Management Reduces Mortality in Post-Sternotomy Mediastinitis.

Background: Post-sternotomy mediastinitis (PSM) is one of the most feared complications of cardiac surgery. The impact of a multidisciplinary management approach on this pathology is yet unknown. Patients and Methods: A multidisciplinary approach based on a co-management model (CMM) of care was initiated in January 2018 because of the incorporation of a hospitalist unit on a cardiac surgery department. An observational retrospective cohort study was designed to evaluate the impact of the CMM of care compared to the standard model (SM) of care in patients diagnosed with PSM. Our primary and secondary outcomes were survival time and treatment failure rate (two or more surgical procedures needed to solve PSM or PSM-related death), respectively. Data related to patient death date were collected from the Spanish National Death Index. A multivariable Cox regression model was created using those variables believed to be clinically relevant. Results: Ninety-one patients developed PSM from January 2010 to June 2020. Regarding the pre-operative clinical status, surgical procedure, and PSM severity, both groups had similar baseline characteristics. Patients were followed for a mean of 27.54 ± 30.5 months. A total of 60.3% of the SM group and 11.1% of the CMM group (p < 0.001) died. Treatment failure occurred in 53 patients (72.6%) in the SM group versus 7 (38.6%) in the CMM group (p = 0.007). The CMM independently reduced overall mortality (hazard ratio [HR], 0.11; 95% confidence interval [CI]. 0.01-0.83) and treatment failure rate (HR, 0.01; 95% CI, 0.001-0.183). Gram-positive bacterial infection (HR, 3.73; 95% CI, .6-8.3), and complete osteosynthesis material removal (HR, 0.47; 95% CI, 0.24-0.91) also influenced mortality in our model. Conclusions: A co-management care model reduced overall mortality in patients diagnosed with post-sternotomy mediastinitis.

The RNA helicase DHX16 recognizes specific viral RNA to trigger RIG-I-dependent innate antiviral immunity.

Type I interferons (IFN-I) are essential to establish antiviral innate immunity. Unanchored (or free) polyubiquitin (poly-Ub) has been shown to regulate IFN-I responses. However, few unanchored poly-Ub interactors are known. To identify factors regulated by unanchored poly-Ub in a physiological setting, we developed an approach to isolate unanchored poly-Ub from lung tissue. We identified the RNA helicase DHX16 as a potential pattern recognition receptor (PRR). Silencing of DHX16 in cells and in vivo diminished IFN-I responses against influenza virus. These effects extended to members of other virus families, including Zika and SARS-CoV-2. DHX16-dependent IFN-I production requires RIG-I and unanchored K48-poly-Ub synthesized by the E3-Ub ligase TRIM6. DHX16 recognizes a signal in influenza RNA segments that undergo splicing and requires its RNA helicase motif for direct, high-affinity interactions with specific viral RNAs. Our study establishes DHX16 as a PRR that partners with RIG-I for optimal activation of antiviral immunity requiring unanchored poly-Ub.

Breastfeeding experiences during the COVID-19 pandemic in Spain:a qualitative study.

BACKGROUND: The pandemic caused by COVID-19 has affected reproductive and perinatal health both through the infection itself and, indirectly, as a consequence of changes in medical care, social policy or social and economic circumstances. The objective of this study is to explore the impact of the pandemic and of the measures adopted on breastfeeding initiation and maintenance. METHODS: A qualitative descriptive study was conducted by means in-depth semi-structured interviews, until reaching data saturation. The study was conducted between the months of January to May 2021. Participants were recruited by midwives from the Primary Care Centres of the Andalusian provinces provinces of Seville, Cádiz, Huelva, Granada, and Jaén. The interviews were conducted via phone call and were subsequently transcribed and analysed by means of reflexive inductive thematic analysis, using Braun and Clarke's thematic analysis. RESULTS: A total of 30 interviews were conducted. Five main themes and ten subthemes were developed, namely: Information received (access to the information, figure who provided the information), unequal support from the professionals during the pandemic (support to postpartum hospitalization, support received from Primary Health Care during the postpartum period), social and family support about breastfeeding (support groups, family support), impact of confinement and of social restriction measures (positive influence on breastfeeding, influence on bonding with the newborn), emotional effect of the pandemic (insecurity and fear related to contagion by coronavirus, feelings of loneliness). CONCLUSION: The use of online breastfeeding support groups through applications such as WhatsApp®, Facebook® or Instagram® has provided important breastfeeding information and support sources. The main figure identified that has provided formal breastfeeding support during this period was that of the midwife. In addition, the social restrictions inherent to the pandemic have exerted a positive effect for women in bonding and breastfeeding, as a consequence of the increase in the time spent at their homes and in the family nucleus co-living.

Use of antibiotics in patients who were attacked by animals that can transmit rabies.

Animal bites have a high probability of becoming infected. In high-risk wounds, the use of antibiotics that kill the associated microorganisms is recommended. The aim of this study was to determine the use of inappropriate antibiotics in patients in different regions of Colombia who were attacked in 2020 by animals that can transmit rabies. This was a retrospective follow-up study of a cohort of patients with wounds caused by animals; the patients were affiliated with the Colombian Health System. Sociodemographic, clinical and pharmacological data were collected. A total of 280 patients were analysed. The median age was 28.0 years, and 52.1% were men. The attacks were attributed to dogs (85.0%) and cats (15.0%), and 9.6% were classified as severe exposure. Antibiotics were used in 71.1% of cases for prophylaxis and in 4.3% of the cases for the treatment of superinfection; cephalexin (37.5%), amoxicillin/clavulanic acid (15.7%) and dicloxacillin (10.4%) were the predominant antibiotics used. A total of 72.0% of patients who received antibiotics received inadequate prescriptions, and 49.2% had no indication to receive prophylaxis. Being treated in the Caribbean region (OR: 4.09; 95% CI: 1.79-9.30) and receiving analgesics (OR: 3.15; 95% CI: 1.25-7.94) were associated with a greater probability of being prescribed antibiotics inadequately; attacks resulting in severe exposure were associated with a lower probability (OR: 0.40; 95% CI: 0.16-0.98). A significant proportion of patients had no indication for prophylactic antibiotics or was prescribed antibiotics not recommended by clinical practice guidelines. There was a low prevalence of prescriptions for amoxicillin/clavulanic acid, the most recommended antibiotic.