title: Independent inhibition of the polymerase and deubiquitinase activities of the Crimean-Congo Hemorrhagic Fever Virus full-length L-protein

text: BACKGROUND: The Crimean-Congo hemorrhagic fever virus (CCHFV) is a segmented negative-sense RNA virus that can cause severe human disease. The World Health Organization (WHO) has listed CCHFVas a priority pathogen with an urgent need for enhanced research activities to develop effective countermeasures. Here we adopted a biochemical approach that targets the viral RNA-dependent RNA polymerase (RdRp). The CCHFV RdRp activity is part of a multifunctional L protein that is unusually large with a molecular weight of ~450 kDa. The CCHFV L-protein also contains an ovarian tumor (OTU) domain that exhibits deubiquitinating (DUB) activity, which was shown to interfere with innate immune responses and viral replication. We report on the expression, characterization and inhibition of the CCHFV full-length L-protein and studied both RNA synthesis and DUB activity. METHODOLOGY/PRINCIPLE FINDINGS: Recombinant full-length CCHFV L protein was expressed in insect cells and purified to near homogeneity using affinity chromatography. RdRp activity was monitored with model primer/templates during elongation in the presence of divalent metal ions. We observed a 14-mer full length RNA product as well as the expected shorter products when omitting certain nucleotides from the reaction mixture. The D2517N mutation of the putative active site rendered the enzyme inactive. Inhibition of RNA synthesis was studies with the broad-spectrum antivirals ribavirin and favipiravir that mimic nucleotide substrates. The triphosphate form of these compounds act like ATP or GTP; however, incorporation of ATP or GTP is markedly favored over the inhibitors. We also studied the effects of bona fide nucleotide analogues 2’-deoxy-2’-fluoro-CTP (FdC) and 2’-deoxy-2’-amino-CTP and demonstrate increased inhibitory effects due to higher rates of incorporation. We further show that the CCHFV L full-length protein and the isolated OTU domain cleave Lys48- and Lys63-linked polyubiqutin chains. Moreover, the ubiquitin analogue CC.4 inhibits the CCHFV-associated DUB activity of the full-length L protein and the isolated DUB domain to a similar extent. Inhibition of DUB activity does not affect elongation of RNA synthesis, and inhibition of RNA synthesis does not affect DUB activity. Both domains are functionally independent under these conditions. CONCLUSIONS/SIGNIFICANCE: The requirements for high biosafety measures hamper drug discovery and development efforts with infectious CCHFV. The availability of full-length CCHFV L-protein provides an important tool in this regard. High-throughput screening (HTS) campaigns are now feasible. The same enzyme preparations can be employed to identify novel polymerase and DUB inhibitors.


title: Levels of complexity in pathogen recognition by C-type lectins

text: In pathogen recognition by C-type lectins, several levels of complexity can be distinguished; these might modulate the immune response in different ways. Firstly, the pathogen-associated molecular pattern repertoire expressed at the microbial surface determines the interactions with specific receptors. Secondly, each immune cell type possesses a specific set of pathogen-recognition receptors. Thirdly, changes in the cell-surface distribution of C-type lectins regulate carbohydrate binding by modulating receptor affinity for different ligands. Crosstalk between these receptors results in a network of multimolecular complexes, adding a further level of complexity in pathogen recognition.


title: Targeting of Renin-Angiotensin System In COVID-19 Patients Affected by Stroke: Emerging Concerns About Detrimental vs. Benefit Effect

text: OBJECTIVE: The present short report summarizes some clinical characteristics of six patients affected by stroke while being on angiotensin-converting enzyme (ACE)2 inhibitors and angiotensin II receptor blockers (ARBs) before and during COVID-19. METHODS: Medical charts and images of six patients affected by stroke while being on ACE-Is and ARBs therapy before and during COVID-19 outbreak in Lombardy region, Italy, were reviewed. RESULTS: Three patients had a dural sinus thrombosis, whereas the remaining suffered by an arterial ischemia, which was a middle cerebral artery occlusion in one case, and a posterior-inferior cerebellar artery occlusion in the remaining two. All patients showed clinical features typical of SARS-CoV-2 infection and positive chest CT scan, and were treated with ACE-Is as needed. Hypercoagulability panel was negative in any case. A recovery was achieved in all cases, although in a variable manner. CONCLUSIONS: Whether or not and in which manner the pharmacomodulation of the renin-angiotensin system may had affect the clinical course of the reported six COVID-19 patients affected by stroke has to be still clarified. An urgent need of randomized clinical trials aimed to assess the safety profile and neuroprotective properties of ACE-Is and ARBs in COVID-19 patients diagnosed with stroke does exists.


title: Diagnostics-in-a-Suitcase: Development of a portable and rapid assay for the detection of the emerging avian influenza A (H7N9) virus

text: Abstract Background In developing countries, equipment necessary for diagnosis is only available in few central laboratories, which are less accessible and of limited capacity to test large numbers of incoming samples. Moreover, the transport conditions of samples are inadequate, therefore leading to unreliable results. Objectives The development of a rapid, inexpensive, and simple test would allow mobile detection of viruses. Study design A suitcase laboratory “Diagnostics-in-a-Suitcase” (56cm×45.5cm×26.5cm) containing all reagents and devices necessary for performing a reverse transcription recombinase polymerase amplification (RT-RPA) assay was developed. As an example, two RT-RPA assays were established for the detection of hemagglutinin (H) and neuraminidase (N) genes of the novel avian influenza (H7N9) virus. Results The sensitivities of the H7 and the N9 RT-RPA assays were 10 and 100 RNA molecules, respectively. The assays were performed at a single temperature (42°C). The results were obtained within 2–7min. The H7N9 RT-RPA assays did not show a cross-detection either of any other respiratory viruses affecting humans and/or birds or of the human or chicken genomes. All reagents were used, stored, and transported at ambient temperature, that is, cold chain independent. In addition, the Diagnostics-in-a-Suitcase was operated by a solar-powered battery. Conclusions The developed assay protocol and mobile setup performed well. Moreover, it can be easily implemented to perform diagnoses at airports, quarantine stations, or farms for rapid on-site viral nucleic acid detection.


title: Aberrant pathogenic GM-CSF+ T cells and inflammatory CD14+CD16+ monocytes in severe pulmonary syndrome patients of a new coronavirus

text: Pathogenic human coronavirus infections, such as severe acute respiratory syndrome CoV (SARS-CoV) and Middle East respiratory syndrome CoV (MERS-CoV), cause high morbidity and mortality 1,2. Recently, a severe pneumonia-associated respiratory syndrome caused by a new coronavirus was reported at December 2019 (2019-nCoV) in the city Wuhan, Hubei province, China3–5, which was also named as pneumonia-associated respiratory syndrome (PARS)6. Up to 9th of February 2020, at least 37, 251 cases have been reported with 812 fatal cases according to the report from China CDC. However, the immune mechanism that potential orchestrated acute mortality from patients of 2019-nCoV is still unknown. Here we show that after the 2019-nCoV infection, CD4+T lymphocytes are rapidly activated to become pathogenic T helper (Th) 1 cells and generate GM-CSF etc. The cytokines environment induces inflammatory CD14+CD16+ monocytes with high expression of IL-6 and accelerates the inflammation. These aberrant and excessive immune cells may enter the pulmonary circulation in huge numbers and play an immune damaging role to causing lung functional disability and quick mortality. Our results demonstrate that excessive non-effective host immune responses by pathogenic T cells and inflammatory monocytes may associate with severe lung pathology. Therefore, we suggest that monoclonal antibody that targets the GM-CSF or interleukin 6 receptor may potentially curb immunopathology caused by 2019-nCoV and consequently win more time for virus clearance.


title: Host cell proteins interacting with the 3′ end of TGEV coronavirus genome influence virus replication

text: Abstract Coronavirus RNA synthesis is performed by a multienzymatic replicase complex together with cellular factors. This process requires the specific recognition of RNA cis-acting signals located at the ends of the viral genome. To identify cellular proteins involved in coronavirus RNA synthesis, transmissible gastroenteritis coronavirus (TGEV) genome ends, harboring essential cis-acting signals for replication, were used as baits for RNA affinity protein purification. Ten proteins were preferentially pulled down with either the 5′ or 3′ ends of the genome and identified by proteomic analysis. Nine of them, including members of the heterogeneous ribonucleoprotein family of proteins (hnRNPs), the poly(A)-binding protein (PABP), the p100 transcriptional co-activator protein and two aminoacyl-tRNA synthetases, showed a preferential binding to the 3′ end of the genome, whereas only the polypyrimidine tract-binding protein (PTB) was preferentially pulled down with the 5′ end of the genome. The potential function of the 3′ end-interacting proteins in virus replication was studied by analyzing the effect of their silencing using a TGEV-derived replicon and the infectious virus. Gene silencing of PABP, hnRNP Q, and glutamyl-prolyl-tRNA synthetase (EPRS) caused a significant 2 to 3-fold reduction of viral RNA synthesis. Interestingly, the silencing of glyceraldehyde 3-phosphate dehydrogenase (GAPDH), initially used as a control gene, caused a 2 to 3-fold increase in viral RNA synthesis in both systems. These data suggest that PABP, hnRNP Q, and EPRS play a positive role in virus infection that could be mediated through their interaction with the viral 3′ end, and that GAPDH has a negative effect on viral infection.


title: Temporal relationship between antibiotic use and respiratory virus activities in the Republic of Korea: a time-series analysis

text: BACKGROUND: Inappropriate use of antibiotics increases resistance and reduces their effectiveness. Despite evidence-based guidelines, antibiotics are still commonly used to treat infections likely caused by respiratory viruses. In this study, we examined the temporal relationships between antibiotic usage and respiratory infections in the Republic of Korea. METHODS: The number of monthly antibiotic prescriptions and the incidence of acute respiratory tract infections between 2010 and 2015 at all primary care clinics were obtained from the Korean Health Insurance Review and Assessment Service. The monthly detection rates of respiratory viruses, including adenovirus, respiratory syncytial virus, influenza virus, human coronavirus, and human rhinovirus, were collected from Korea Centers for Disease Control and Prevention. Cross-correlation analysis was conducted to quantify the temporal relationship between antibiotic use and respiratory virus activities as well as respiratory infections in primary clinics. RESULTS: The monthly use of different classes of antibiotic, including penicillins, other beta-lactam antibacterials, macrolides and quinolones, was significantly correlated with influenza virus activity. These correlations peaked at the 0-month lag with cross-correlation coefficients of 0.45 (p < 0.01), 0.46 (p < 0.01), 0.40 (p < 0.01), and 0.35 (< 0.01), respectively. Furthermore, a significant correlation was found between acute bronchitis and antibiotics, including penicillin (0.73, p < 0.01), macrolides (0.74, p < 0.01), and quinolones (0.45, p < 0.01), at the 0-month lag. CONCLUSIONS: Our findings suggest that there is a significant temporal relationship between influenza virus activity and antibiotic use in primary clinics. This relationship indicates that interventions aimed at reducing influenza cases in addition to effort to discourage the prescription of antibiotics by physicians may help to decrease unnecessary antibiotic consumption.


title: Guidelines for Surgical Tracheostomy and Tracheostomy Tube Change During the COVID-19 Pandemic: A Review Article

text: The novel corona virus disease (COVID-19) has unfolded into a pandemic and is continuing to propagate at a frightening speed. The aim of this article is to share our protocol for performing a safe surgical tracheostomy in this COVID-19 era. Tracheostomy procedures have a high risk of aerosol generation. To standardize institutional safety measures with tracheostomy, we advocate using a dedicated tracheostomy protocol applicable to all patients including those suspected of having COVID-19. We also did explore the current literature and recommendations for tracheostomy in patients with COVID-19 and studied the previous data from severe acute respiratory syndrome coronavirus 1 (SARS-CoV-1), the virus responsible for the SARS outbreak of 2003. We have prepared a protocol for performing a safe surgical tracheotomy in patients affected by COVID-19. Surgeons who might be involved in performing the tracheostomies should become familiar with these guidelines.


title: Diabetes or endocrinopathy admitted in the COVID‐19 ward

text: The Covid‐19 pandemic confronted us with unknown clinical pictures, also in diabetology and endocrinology. Sharing clinical experiences is therefore of enormous importance. Actually, information about the care given in the Covid‐19 ward (in contrast to that provided in the Emergency Room/ICU) is still sparse. The last weeks we built experience and gathered knowledge while giving hospital care to patients who had a pre‐existent endocrine disease (and diabetes; most patients suffered from a type two diabetes). In our contribution we presented our insights obtained from this intensive period obtained in the Covid‐19 ward.


title: International travelers and genomics uncover a ‘hidden’ Zika outbreak

text: The ongoing Zika epidemic in the Americas has challenged public health surveillance, response, and control systems. Even as the epidemic appears to be near its end in the Americas, it is unclear whether substantial Zika virus transmission may still be ongoing. This issue is exacerbated by large discrepancies in local case reporting and significant delays in detecting outbreaks due to surveillance gaps. To uncover locations with lingering outbreaks in the Americas, we investigated travel-associated Zika cases diagnosed in the United States and Europe to identify signatures of transmission dynamics that were not captured by local reporting. We found that a large and unreported Zika outbreak occurred in Cuba during 2017, a year after peak transmission in neighboring countries, with cases still appearing in 2018. By sequencing Zika virus from infected travelers, we show that the 2017 outbreak in Cuba was sparked by long-lived lineages of Zika virus introduced from multiple places in the Americas a year prior. Our data suggest that while aggressive mosquito control in Cuba may initially have been effective at mitigating Zika virus transmission, in the absence of vaccines, herd immunity, or strong international coordination, such control measures may need to be maintained to be effective. Our study highlights how Zika virus may still be ‘silently’ spreading in the Americas and provides a framework for more accurately understanding outbreak dynamics.


Crisis del covid 19. El reto para dar el paso definitivo a la telerehabilitación


Preoperative radiotherapy for locally advanced rectal cancer during and after the COVID‐19 pandemic


Dysosmia and dysgeusia due to the 2019 Novel Coronavirus; a hypothesis that needs further investigation


title: Effect of integrated traditional Chinese medicine and western medicine on the treatment of severe acute respiratory syndrome: A meta-analysis

text: BACKGROUND: Data regarding the treatment efficacy of integrative treatment of Traditional Chinese Medicine (TCM) and Western Medicine (WM) in treating patients with (SARS) are conflicting. The effects of integrative TCM/WM treatment have not been fully quantified. OBJECTIVES: To systematically asses the treatment effects of integrated TCM with WM versus WM alone in patients with SARS, incorporating data from recently published studies. METHODS: A meta-analysis was conducted, using published randomized and nonrandomized controlled clinical studies that compared the treatment effects of integrative TCM/WM with WM alone from 2002 to 2006. The outcome measurements included mortality rate, cure rate, resolution of pulmonary infiltrate, use of corticosteroid, and time to defervescence. The effect sizes were presented as risk ratio (RR), rate difference (RD), and weighted mean difference (WMD). The pooled effect sizes were calculated by both fixed-effects and random-effects models. RESULTS: A total of 1,678 patients with a diagnosis of SARS were identified, including 866 patients from 16 randomized controlled studies and 812 patients from 8 nonrandomized controlled studies. There were no differences detected in mortality rate or cure rate between treatments. Compared with patients receiving WM treatment alone, patients receiving integrative treatment were more likely to have complete or partial resolution of pulmonary infiltrate (RD=0.18, 95%CI; 0.07 to 0.30), lower average daily dosage (mg) of corticosteroid (WMD=-60.27, 95% CI; -70.58 to -49.96), higher CD4+ counts (cells/uL) (WMD=167.96, 95% CI; 109.68 to 226.24), and shorter time to defervescence (days) (WMD= -1.06, 95%CI;-1.60 to -0.53). CONCLUSIONS: The experience of integrative TCM/WM in the treatment of SARS is encouraging. The use of TCM as an adjunctive therapy in the treatment of SARS should be further investigated.


title: Preliminary characterization of protein binding factor for porcine reproductive and respiratory syndrome virus on the surface of permissive and non-permissive cells

text: In its natural host, porcine reproductive and respiratory syndrome virus (PRRSV) has been reported to have a restricted tropism for cells of the monocyte/macrophage lineage. To date, cloned monkey kidney cell lines, such as MARC-145 and CL2621 cells which have been established from MA-104 cells, are the only non-porcine cells known to support PRRSV replication. In the present study, a binding assay was set up to follow by flow cytometry the attachment of PRRSV on the surface of porcine and non-porcine cells. PRRSV was found to be able to bind permissive cells like porcine alveolar macrophages and MARC-145. Further binding assays with porcine peripheral blood leukocytes showed that only monocytes can attach the virus. By their lack of binding factor, lymphocytes appeared to be refractory to PRRSV infection. Pre-incubation of MARC-145 cells with chymotrypsin and pronase E, but not neuraminidase, blocked their binding activity for PRRSV. The binding activity of the protease-treated cells was regenerated 8 hours after treatment, but cells remained unable to bind PRRSV if maintained in the presence of cycloheximide, thus confirming the proteinic nature of the specific binding factor(s). Experiments conduced with cells that have been previously characterized as non-permissive to PRRSV infection showed that many of them were able to bind the virus. Data obtained suggest that interaction of PRRSV with a specific binding factor on the surface of some cells is not sufficient to lead to a productive infection, and that a second putative receptor or other phenomena are probably required to pursue later events.


title: Control, Management, and Prevention of Bovine Respiratory Disease in Dairy Calves and Cows

text: Incidence rates for bovine respiratory disease (BRD) in dairy cattle have remained essentially unchanged over the last 20 years. Dairy calves are more commonly affected than adult animals, with BRD being the principal cause of death in weaned dairy calves. The lack of progress in controlling respiratory disease demonstrates that there continues to be significant room for improvement in controlling this multifactorial syndrome, and that dairy producers need assistance in applying evolving husbandry practices to improve the health of dairy cattle. Therefore, it seems prudent to focus the management strategies on preventing disease through sound management of the transition period, along with sound vaccination and biosecurity programs.


title: Kobuvirus (Aichivirus B) infection in Brazilian cattle herds

text: There are few studies involving the detection of Aichivirus B in cattle herds worldwide, and this virus has never been diagnosed in South America. This study evaluated 222 diarrhoeic faecal samples from four Brazilian geographical regions (South, Southeast, Midwest, and North), collected between February 2010 to May 2012. To evaluate the frequency of occurrence in different types of livestock, samples from beef (n = 105) and dairy (n = 117) cattle herds were evaluated. To determine the category of animals more susceptible to infection, the sampling included samples from calves (n = 182) and adults animals (n = 40). The 216 bp fragment of the Aichivirus RdRp gene was amplified by a RT-PCR assay in 18.2 % (40/222) of the samples evaluated in both beef and dairy cattle animals. The highest (P < 0.05) detection rate (20.9 %; 38/182) of the Aichivirus B was found in calves. The nucleotide sequencing analysis showed that the Brazilian Aichivirus B strains clustered in a distinct branch in the phylogenetic tree of the European and Asiatic strains. This is the first description of Aichivirus B infection in Brazilian cattle herds.


title: Tocilizumab for the treatment of severe COVID-19 pneumonia with hyperinflammatory syndrome and acute respiratory failure: A single center study of 100 patients in Brescia, Italy

text: A hyperinflammatory syndrome (HIS) may cause a life-threatening acute respiratory distress syndrome (ARDS) in patients with COVID-19 pneumonia. A prospective series of 100 consecutive patients admitted to the Spedali Civili University Hospital in Brescia (Italy) between March 9th and March 20th with confirmed COVID-19 pneumonia and ARDS requiring ventilatory support was analyzed to determine whether intravenous administration of tocilizumab (TCZ), a monoclonal antibody that targets the interleukin 6 (IL-6) receptor, was associated with improved outcome. Tocilizumab was administered at a dosage of 8 mg/kg by two consecutive intravenous infusions 12 h apart. A third infusion was optional based on clinical response. The outcome measure was an improvement in acute respiratory failure assessed by means of the Brescia COVID Respiratory Severity Score (BCRSS 0 to 8, with higher scores indicating higher severity) at 24–72 h and 10 days after tocilizumab administration. Out of 100 treated patients (88 M, 12 F; median age: 62 years), 43 received TCZ in the intensive care unit (ICU), while 57 in the general ward as no ICU beds were available. Of these 57 patients, 37 (65%) improved and suspended noninvasive ventilation (NIV) (median BCRSS: 1 [IQR 0–2]), 7 (12%) patients remained stable in NIV, and 13 (23%) patients worsened (10 died, 3 were admitted to ICU). Of the 43 patients treated in the ICU, 32 (74%) improved (17 of them were taken off the ventilator and were discharged to the ward), 1 (2%) remained stable (BCRSS: 5) and 10 (24%) died (all of them had BCRSS≥7 before TCZ). Overall at 10 days, the respiratory condition was improved or stabilized in 77 (77%) patients, of whom 61 showed a significant clearing of diffuse bilateral opacities on chest x-ray and 15 were discharged from the hospital. Respiratory condition worsened in 23 (23%) patients, of whom 20 (20%) died. All the patients presented with lymphopenia and high levels of C-reactive protein (CRP), fibrinogen, ferritin and IL-6 indicating a HIS. During the 10-day follow-up, three cases of severe adverse events were recorded: two patients developed septic shock and died, one had gastrointestinal perforation requiring urgent surgery and was alive at day 10. In conclusion, our series showed that COVID-19 pneumonia with ARDS was characterized by HIS. The response to TCZ was rapid, sustained, and associated with significant clinical improvement.


title: Case series of six kidney transplanted patients with COVID‐19 pneumonia treated with tocilizumab

text: Few reports described the outcome of kidney transplanted patients (KTs) affected by COVID‐19 treated with interleukin‐6 receptor inhibitor tocilizumab (TCZ). We report our case series of 6 KTs with COVID‐19 pneumonia who received TCZ: All were of male gender, with a mean age of 55.5 ± 8.4 years, a median time from transplantation of 3611 days (1465‐5757); 5/6 had cardiovascular comorbidities, 1/6 had diabetes, and 3/6 have one or more previous KTs. Four out of six patients died, at an average time of 9.75 ± 2.4 days after tocilizumab administration, 3/6 due to a coexistent septic shock. Two patients improved after TCZ and were discharged at 20 and 21 days, respectively; in both patient, a significant increase of total lymphocyte count was observed. In conclusion, KTs, where the role of peculiar factors such as chronic immunosuppression is still undetermined, represent a high‐risk group with significant COVID‐19‐associated mortality. The evaluation of the TCZ effect in COVID‐19 pneumonia requires controlled studies (ideally RCTs) in this specific population.


Internal clock, sleep and desynchronization of confined children and adolescents


title: Human organoids: model systems for human biology and medicine

text: The historical reliance of biological research on the use of animal models has sometimes made it challenging to address questions that are specific to the understanding of human biology and disease. But with the advent of human organoids — which are stem cell-derived 3D culture systems — it is now possible to re-create the architecture and physiology of human organs in remarkable detail. Human organoids provide unique opportunities for the study of human disease and complement animal models. Human organoids have been used to study infectious diseases, genetic disorders and cancers through the genetic engineering of human stem cells, as well as directly when organoids are generated from patient biopsy samples. This Review discusses the applications, advantages and disadvantages of human organoids as models of development and disease and outlines the challenges that have to be overcome for organoids to be able to substantially reduce the need for animal experiments.


title: Expanding roles for CD4(+) T cells in immunity to viruses

text: Viral pathogens often generate strong CD4(+) T cell responses that are best known for their ability to help B cell and CD8(+) T cell responses. However, recent studies reveal additional roles for CD4(+) T cells, some of which are independent of other lymphocytes, and indicate that memory cells are more effective in most functions than naïve CD4 T cells. Here, we review the full spectrum of antiviral functions of CD4(+) T cells, including their helper activities, innate immune induction, and direct anti-viral roles, and suggest how these functions are integrated to provide highly protective responses against viral pathogens.


title: The burden and clinical manifestation of hospitalized influenza among different pediatric age‐groups in the tropics

text: INTRODUCTION: In tropical Singapore, influenza occurs all year‐round. This study of influenza‐confirmed hospitalized pediatric patients compared clinical characteristics and complications by age‐group and differences between influenza A and B. METHODS: This was a retrospective study of pediatric inpatients from January 2013 to December 2014. Patients were grouped into: <6 months, 6 months to <5 years, 5‐ to <10‐year and ≥10 years. Complications were classified into neurologic, pulmonary, and other. We also calculated the incidence of hospitalized influenza cases per 100 000 age‐related population. RESULTS: There were a total of 1272 patients with a median age of 37 months. The highest hospitalization rates were in the <6 months age‐group. Majority (75.2%) had no comorbidity; 25.6% had complications: neurologic 11.9%, pulmonary 9.6%, other 4.1%. Patients with other complications were older, male, and had the highest influenza B rates and the longest length of stay. Influenza A comprised 76.9% of cases and had higher complication rates especially neurologic, compared to influenza B. Influenza B patients were older and were more likely to develop other complications. The 6‐month to <5‐year‐age‐group had the highest complication rate (30.6%), especially neurologic. However, ≥10 years old had the highest other complications, ICU/ high‐dependency admissions and influenza B Victoria rates. CONCLUSIONS: Infants <6 months had the highest hospitalization rates for influenza. The 6‐month to <5‐year‐age‐group had the highest complication rate especially neurologic. Influenza A patients were younger, had higher seizure rates and complications compared to influenza B.


title: Chapter 4 Alphavirus Nonstructural Proteases and Their Inhibitors

text: Abstract Alphaviruses, such as Chikungunya virus, O’Nyong–Nyong virus, Ross River virus, have been widely known to cause fever, rash, and rheumatic diseases. In addition, several other alphaviruses, for instance Eastern equine encephalitis virus, Venezuelan equine encephalitis virus, and Western equine encephalitis virus, potentially cause fatal encephalitis in humans. These diseases are considered as neglected tropical diseases for which there are no current antiviral therapies or vaccines available. The replication process in alphaviruses depends on four nonstructural proteins, NSP1–NSP4, which are produced as a single polyprotein. Therefore, the Alphavirus-mediated diseases in humans remain challenging among the virologists worldwide. Thus researchers are trying to find out proficient approaches, including the discovery of novel chemotherapeutic agents for the possible management and treatment of infected patients. Attempts were also made to identify an active compound against alphaviruses from natural sources. The genomes of various alphaviruses have already been revealed, and the function of proteins may be predicted by homology modeling, with the known proteins of closely related viruses. With the help of this information of protein modeling and subsequent virtual screening approach, the research teams will be able to identify few potential leads. The drug discovery against various alphaviruses is still in its early stages. Moreover, consolidating the available information and making it available for the scientific community are urgent requirements to expedite the research of potential drug discovery. The current chapter describes the techniques available to prevent Alphavirus infection and to treat Alphavirus-associated malignancies. In addition, we also discuss the recent outcomes in the fields of synthetic and natural medicinal chemistry research that were solely aimed to fight against Alphavirus infection. Thus the present chapter may also help and expedite the drug discovery and development of inhibitors against nonstructural proteins of various alphaviruses.


title: Quantitative Proteomics Reveal Peroxiredoxin Perturbation Upon Persistent Lymphocytic Choriomeningitis Virus Infection in Human Cells

text: Experimental data indicate that during persistent infection, lymphocytic choriomeningitis virus (LCMV) may both directly or indirectly modulate regulatory cellular processes and alter cellular functions that are not critical for survival, but are essential for cell homeostasis. In order to shed more light on these processes, two-dimensional differential in-gel electrophoresis (2D-DIGE) and MALDI-TOF tandem mass spectrometry were used to determine the proteome response of the HeLa cell line to persistent LCMV infection. Quantitative analysis revealed 24 differentially abundant proteins. Functional analysis showed that LCMV-responsive proteins were primarily involved in metabolism, stress, and the defense response. Among identified proteins, we discovered significant changes for peroxiredoxins, a family of antioxidant enzymes. Decreased amount of these antioxidant proteins correlated with elevation of reactive oxygen species (ROS) in infected cells. Increased levels of ROS were accompanied by changes in the pattern of telomere restriction fragments (TRFs) in infected cells and mediated activation of hypoxia-inducible transcription factor-1 (HIF-1) and phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathways. Moreover, treatment with antioxidants resulted in reduced levels of viral nucleoprotein, indicating a connection between ROS-dependent signaling and viral replication.


Annulation du congrès annuel de la SIFUD-PP et nouvelles dates des futurs événements


title: Data as evidence

text: In an earlier article, Gordon Drummond summarized ongoing changes in how statistics are being used in experimental physiology (Drummond, 2020). He described the near ubiquitous use of the P value, cautioning against declaring an analysis “statistically significant”. He mentioned alternative approaches including Bayesian and likelihood approaches. This article focusses on the latter approach although we will first have another look at that P value. Then the likelihood approach will be introduced with a very artificial example that allows us to easily grasp the concept. A more realistic example is then described with associated calculations. A further example using real categorical data is explained, and how it relates to and is superior to the oft used χ(2) test. A final discussion reveals that the likelihood approach, although mathematically and statistically accurate, is poorly supported by literature and training. This article is protected by copyright. All rights reserved


title: Immediate Psychological Responses and Associated Factors during the Initial Stage of the 2019 Coronavirus Disease (COVID-19) Epidemic among the General Population in China

text: Background: The 2019 coronavirus disease (COVID-19) epidemic is a public health emergency of international concern and poses a challenge to psychological resilience. Research data are needed to develop evidence-driven strategies to reduce adverse psychological impacts and psychiatric symptoms during the epidemic. The aim of this study was to survey the general public in China to better understand their levels of psychological impact, anxiety, depression, and stress during the initial stage of the COVID-19 outbreak. The data will be used for future reference. Methods: From 31 January to 2 February 2020, we conducted an online survey using snowball sampling techniques. The online survey collected information on demographic data, physical symptoms in the past 14 days, contact history with COVID-19, knowledge and concerns about COVID-19, precautionary measures against COVID-19, and additional information required with respect to COVID-19. Psychological impact was assessed by the Impact of Event Scale-Revised (IES-R), and mental health status was assessed by the Depression, Anxiety and Stress Scale (DASS-21). Results: This study included 1210 respondents from 194 cities in China. In total, 53.8% of respondents rated the psychological impact of the outbreak as moderate or severe; 16.5% reported moderate to severe depressive symptoms; 28.8% reported moderate to severe anxiety symptoms; and 8.1% reported moderate to severe stress levels. Most respondents spent 20–24 h per day at home (84.7%); were worried about their family members contracting COVID-19 (75.2%); and were satisfied with the amount of health information available (75.1%). Female gender, student status, specific physical symptoms (e.g., myalgia, dizziness, coryza), and poor self-rated health status were significantly associated with a greater psychological impact of the outbreak and higher levels of stress, anxiety, and depression (p < 0.05). Specific up-to-date and accurate health information (e.g., treatment, local outbreak situation) and particular precautionary measures (e.g., hand hygiene, wearing a mask) were associated with a lower psychological impact of the outbreak and lower levels of stress, anxiety, and depression (p < 0.05). Conclusions: During the initial phase of the COVID-19 outbreak in China, more than half of the respondents rated the psychological impact as moderate-to-severe, and about one-third reported moderate-to-severe anxiety. Our findings identify factors associated with a lower level of psychological impact and better mental health status that can be used to formulate psychological interventions to improve the mental health of vulnerable groups during the COVID-19 epidemic.


title: Jetzt kommt es auf uns Hausärzte an!: Coronavirus in Deutschland

text: Ausgangssperren, steigende Infektionszahlen, Angst in der Bevölkerung — die Coronavirus-Pandemie hält das ganze Land in Atem. Hausärzte müssen in dieser Situation das hohe Vertrauen nutzen, das sie genießen: Durch klar strukturierte, qualitativ hochwertige Versorgung, aber auch durch die Verbreitung von Ruhe und Zuversicht.


title: Loop mediated isothermal amplification of 5.8S rDNA for specific detection of Tritrichomonas foetus

text: Abstract Tritrichomonas foetus is the causative agent of bovine trichomonosis, a sexually transmitted disease leading to infertility and abortion. A test based on loop mediated isothermal amplification (LAMP) targeting the 5.8S rDNA subunit was designed for the specific identification of T. foetus. The LAMP assay was validated using 28 T. foetus and 35 non-T. foetus trichomonads strains. It did not exhibit cross-reaction with closely related parasites commonly found in smegma cultures like Tetratrichomonas spp. and Pentatrichomonas hominis. Bovine smegma did not show interferences for the detection of the parasite and, the sensitivity of the method (4×103 CFU/mL, approximately 10cells/reaction) was slightly higher than that found for PCR amplification with TFR3 and TFR4 primers. The LAMP approach has potential applications for diagnosis and control of T. foetus and, practical use for low skill operators in rural areas.


title: Sci-athon: Promoting Interdisciplinary Science and Peer Learning with Adrenaline and Pizza

text: Sci-athon is a collaborative event for graduate students and postdoctoral scholars designed to harness diverse scientific backgrounds toward solving global issues in chemistry. The last 2 years have yielded surprisingly different versions of this annual competition.


Welcome to Viruses: A New Open-Access, Multidisciplinary Forum for Virology


title: Temporal dynamics of Middle East respiratory syndrome coronavirus in the Arabian Peninsula, 2012–2017

text: Middle East respiratory syndrome coronavirus (MERS-CoV) remains a notable disease and poses a significant threat to global public health. The Arabian Peninsula is considered a major global epicentre for the disease and the virus has crossed regional and continental boundaries since 2012. In this study, we focused on exploring the temporal dynamics of MERS-CoV in human populations in the Arabian Peninsula between 2012 and 2017, using publicly available data on case counts and combining two analytical methods. Disease progression was assessed by quantifying the time-dependent reproductive number (TD-Rs), while case series temporal pattern was modelled using the AutoRegressive Integrated Moving Average (ARIMA). We accounted for geographical variability between three major affected regions in Saudi Arabia including Eastern Province, Riyadh and Makkah. In Saudi Arabia, the epidemic size was large with TD-Rs >1, indicating significant spread until 2017. In both Makkah and Riyadh regions, the epidemic progression reached its peak in April 2014 (TD-Rs > 7), during the highest incidence period of MERS-CoV cases. In Eastern Province, one unique super-spreading event (TD-R > 10) was identified in May 2013, which comprised of the most notable cases of human-to-human transmission. Best-fitting ARIMA model inferred statistically significant biannual seasonality in Riyadh region, a region characterised by heavy seasonal camel-related activities. However, no statistical evidence of seasonality was identified in Eastern Province and Makkah. Instead, both areas were marked by an endemic pattern of cases with sporadic outbreaks. Our study suggested new insights into the epidemiology of the virus, including inferences about epidemic progression and evidence for seasonality. Despite the inherent limitations of the available data, our conclusions provide further guidance to currently implement risk-based surveillance in high-risk populations and, subsequently, improve related interventions strategies against the epidemic at country and regional levels.


title: Virus infections of the gastrointestinal tract of poultry

text: Abstract Several different viruses have been identified as causes of gastrointestinal tract infections in poultry. These include rotaviruses, coronaviruses, enteroviruses, adenoviruses, astroviruses, and reoviruses. In addition, a number of other viruses of unknown importance have been associated with gastrointestinal diseases in poultry based on electron microscopic examination of feces and intestinal contents. Viral infections of the gastrointestinal tract of poultry are known to negatively impact poultry production, and they likely contribute to the development of other, extragastrointestinal diseases. Our current understanding of the viruses that cause gastrointestinal tract infections in poultry is reviewed, with emphasis given to those of greatest importance.


Emerging viruses


title: Master sensors of pathogenic RNA – RIG-I like receptors

text: Abstract Initiating the immune response to invading pathogens, the innate immune system is constituted of immune receptors (pattern recognition receptors, PRR) that sense microbe-associated molecular patterns (MAMPs). Detection of pathogens triggers intracellular defense mechanisms, such as the secretion of cytokines or chemokines to alarm neighboring cells and attract or activate immune cells. The innate immune response to viruses is mostly based on PRRs that detect the unusual structure, modification or location of viral nucleic acids. Most of the highly pathogenic and emerging viruses are RNA genome-based viruses, which can give rise to zoonotic and epidemic diseases or cause viral hemorrhagic fever. As viral RNA is located in the same compartment as host RNA, PRRs in the cytosol have to discriminate between viral and endogenous RNA by virtue of their structure or modification. This challenging task is taken on by the homologous cytosolic DExD/H-box family helicases RIG-I and MDA5, which control the innate immune response to most RNA viruses. This review focuses on the molecular basis for RIG-I like receptor (RLR) activation by synthetic and natural ligands and will discuss controversial ligand definitions.


title: Decline in child vaccination coverage during the COVID‐19 pandemic — Michigan Care Improvement Registry, May 2016‐May 2020

text: This article describes a reduction in childhood vaccinations during the COVID‐19 pandemic, which may leave communities more vulnerable to vaccine‐preventable diseases.


title: Isolation of a coronavirus during studies on puffinosis, a disease of the Manx shearwater(Puffinus puffinus)

text: A virus was isolated from 2 day-old mice inoculated with homogenates of either the lungs or blood of 2 different shearwaters affected by puffinosis. Examination of infected suckling mouse brain and infected NCTC-1469 (mouse liver) cell cultures, by electron microscopy, revealed virus particles and inclusion bodies characteristic of a coronavirus. Neutralization, complement fixation and fluorescent antibody tests showed that the virus was related to mouse hepatitis virus. The virus was not isolated from 445 control, uninfected mice. Neutralising antibodies were not detected in 39 sera from diseased shearwaters and 2 sera from apparently healthy birds. Two shearwaters inoculated with the virus did not develop clinical signs of infection. The question of whether the virus was isolated from shearwaters or from laboratory mice is discussed.


title: An evaluation of co-use of chloroquine or hydroxychloroquine plus azithromycin on cardiac outcomes: A pharmacoepidemiological study to inform use during the COVID19 pandemic

text: Abstract Background Chloroquine or hydroxychloroquine (chloroquine) plus azithromycin is considered as therapy for COVID19. With benefit evaluations underway, safety concerns due to potential additive effects on QTc prolongation should be addressed. Objective We compared risk of cardiac adverse events between combinations of chloroquine and azithromycin and chloroquine and amoxicillin. Methods We conducted a retrospective cohort study using the IBM MarketScan Commercial Claims and Medicare Supplemental Databases, 2005–2018. We included autoimmune disease patients aged ≥18 years initiating azithromycin or amoxicillin for ≥5 days during chloroquine treatment. Patients had continuous insurance coverage ≥6 months before combination use until 5 days thereafter or inpatient death. Two outcomes were sudden cardiac arrest/ventricular arrhythmias (SCA/VA) and cardiac symptoms. We followed patients for up to 5 days to estimate hazard ratios (HR). Covariates were adjusted using stabilized inverse probability treatment weighting. Results We identified two SVC/VA events among >145,000 combination users. The adjusted incidence of cardiac symptoms among azithromycin and amoxicillin users was 276 vs 254 per 10,000 person-years with an adjusted HR of 1.10 (95%CI, 0.62–1.95). Conclusion Combination use of chloroquine and azithromycin at routine doses did not show pronounced increases in arrhythmias in this real-world population, though small sample size and outcome rates limit conclusions.


title: Incidentally discovered COVID-19 in low-suspicion patients—a threat to front line health care workers

text: PURPOSE: The COVID-19 pandemic has been responsible for thousands of deaths worldwide. Testing remains at a premium, and criteria for testing remains reserved for those with lower respiratory infection symptoms and/or a known high-risk exposure. The role of imaging in COVID-19 is rapidly evolving; however, few algorithms include imaging criteria, and it is unclear what should be done in low-suspicion patients with positive imaging findings. METHODS: From 03/01/2020–03/20/2020, a retrospective review of all patients with suspected COVID-19 on imaging was performed. Imaging was interpreted by a board-certified, fellowship-trained radiologist. Patients were excluded if COVID-19 infection was suspected at the time of presentation, was the reason for imaging, or if any lower respiratory symptoms were present. RESULTS: Eight patients with suspected COVID-19 infection on imaging were encountered. Seven patients received testing due to suspicious imaging findings with subsequent lab-confirmed COVID-19. No patients endorsed prior exposure to COVID-19 or recent international travel. COVID-19 was suggested in six patients incidentally on abdominal CT and two on chest radiography. At the time of presentation, no patients were febrile, and seven endorsed gastrointestinal symptoms. Five COVID-19 patients eventually developed respiratory symptoms and required intubation. Two patients expired during the admission. CONCLUSIONS: Patients with imaging findings suspicious for COVID-19 warrant prompt reverse transcription polymerase chain reaction (RT-PCR) testing even in low clinical suspicion cases. The prevalence of disease in the population may be underestimated by the current paradigm of RT-PCR testing with the current clinical criteria of lower respiratory symptoms and exposure risk.


title: Severe Acute Respiratory Syndrome

text: The first cases of severe acute respiratory syndrome (SARS) occurred in China in November 2002. The agent causing this illness has been identified as a novel coronavirus, SARS-coronavirus. Since its introduction <1 year ago, this virus has infected 8098 people in 26 countries, killing 774 of them. We present an overview of the epidemiology, clinical presentation, diagnosis, and treatment of SARS based on the current state of knowledge derived from published studies and our own personal experience.


title: The virome hunters

text: Ambitious efforts to catalog viruses across the globe may facilitate our understanding of viral communities and ecology, boost infectious disease diagnostics and surveillance, and spur new therapeutics. Charles Schmidt investigates.


title: Chapter 7 Camelidae

text: Abstract New world (NW) camelids, alpaca, llama, vicuña, and guanaco, and old world (OW) camelids, Bactrian and dromedary camels are related and have many of the same anatomical features and disease susceptibilities though they are also very different. Only the free-ranging population of wild Bactrian camel is endangered. Bactrian camels held in zoos are generally of domestic origin. Vicuña are listed as vulnerable. In addition to those camelids held in captivity, there are domesticated populations of camelids (except vicuña and guanaco) maintained throughout the world. Most are fairly hardy animals, but there are some specific disease concerns. Domesticated llamas and alpacas have become hobby pets and thus management and genetic issues are an increasing source of disease. These include obesity, vitamin and mineral deficiencies and intoxications, metabolic derangements, and congenital malformations (particularly in the young). Domesticated animals are also more prone to degenerative arthropathy and dental disease as they age than wild camelids. Ovarian hydrobursitis is an important source of infertility in dromedaries. Important infectious diseases include coccidiosis, bovine viral diarrhea virus, alpaca fever, and meningeal worm of NW camelids, trypanosomiasis and camelpox in OW camels, and foot and mouth disease in Bactrian camels and NW camelids. These and other disease processes are discussed in this chapter.


title: Vaccine Efficacy in Senescent Mice Challenged with Recombinant SARS-CoV Bearing Epidemic and Zoonotic Spike Variants

text: BACKGROUND: In 2003, severe acute respiratory syndrome coronavirus (SARS-CoV) was identified as the etiological agent of severe acute respiratory syndrome, a disease characterized by severe pneumonia that sometimes results in death. SARS-CoV is a zoonotic virus that crossed the species barrier, most likely originating from bats or from other species including civets, raccoon dogs, domestic cats, swine, and rodents. A SARS-CoV vaccine should confer long-term protection, especially in vulnerable senescent populations, against both the 2003 epidemic strains and zoonotic strains that may yet emerge from animal reservoirs. We report the comprehensive investigation of SARS vaccine efficacy in young and senescent mice following homologous and heterologous challenge. METHODS AND FINDINGS: Using Venezuelan equine encephalitis virus replicon particles (VRP) expressing the 2003 epidemic Urbani SARS-CoV strain spike (S) glycoprotein (VRP-S) or the nucleocapsid (N) protein from the same strain (VRP-N), we demonstrate that VRP-S, but not VRP-N vaccines provide complete short- and long-term protection against homologous strain challenge in young and senescent mice. To test VRP vaccine efficacy against a heterologous SARS-CoV, we used phylogenetic analyses, synthetic biology, and reverse genetics to construct a chimeric virus (icGDO3-S) encoding a synthetic S glycoprotein gene of the most genetically divergent human strain, GDO3, which clusters among the zoonotic SARS-CoV. icGD03-S replicated efficiently in human airway epithelial cells and in the lungs of young and senescent mice, and was highly resistant to neutralization with antisera directed against the Urbani strain. Although VRP-S vaccines provided complete short-term protection against heterologous icGD03-S challenge in young mice, only limited protection was seen in vaccinated senescent animals. VRP-N vaccines not only failed to protect from homologous or heterologous challenge, but resulted in enhanced immunopathology with eosinophilic infiltrates within the lungs of SARS-CoV–challenged mice. VRP-N–induced pathology presented at day 4, peaked around day 7, and persisted through day 14, and was likely mediated by cellular immune responses. CONCLUSIONS: This study identifies gaps and challenges in vaccine design for controlling future SARS-CoV zoonosis, especially in vulnerable elderly populations. The availability of a SARS-CoV virus bearing heterologous S glycoproteins provides a robust challenge inoculum for evaluating vaccine efficacy against zoonotic strains, the most likely source of future outbreaks.


title: Animal Models for Target Diseases in Gene Therapy — using DNA and siRNA Delivery Strategies

text: Nanoparticles, including lipopolyamines leading to lipoplexes, liposomes, and polyplexes are targeted drug carrier systems in the current search for a successful delivery system for polynucleic acids. This review is focused on the impact of gene and siRNA delivery for studies of efficacy, pharmacodynamics, and pharmacokinetics within the setting of the wide variety of in vivo animal models now used. This critical appraisal of the recent literature sets out the different models that are currently being investigated to bridge from studies in cell lines through towards clinical reality. Whilst many scientists will be familiar with rodent (murine, fecine, cricetine, and musteline) models, few probably think of fish as a clinically relevant animal model, but zebrafish, madake, and rainbow trout are all being used. Larger animal models include rabbit, cat, dog, and cow. Pig is used both for the prevention of foot-and-mouth disease and human diseases, sheep is a model for corneal transplantation, and the horse naturally develops arthritis. Non-human primate models (macaque, common marmoset, owl monkey) are used for preclinical gene vector safety and efficacy trials to bridge the gap prior to clinical studies. We aim for the safe development of clinically effective delivery systems for DNA and RNAi technologies.


title: A single-tube approach for in vitro diagnostics using diatomaceous earth and optical sensor

text: Versatile, simple and efficient sample preparation is desirable for point-of-care testing of emerging diseases such as zoonoses, but current sample preparation assays are insensitive, labour-intensive and time-consuming and require multiple instruments. We developed a single-tube sample preparation approach involving direct pathogen enrichment and extraction from human specimens using diatomaceous earth (DE). Amine-modified DE was used to directly enrich a zoonotic pathogen, Brucella, in a large sample volume. Next, a complex of amine-modified DE and dimethyl suberimidate was used for nucleic acid extraction from the enriched pathogen. Using our single-tube approach, the pathogen can be enriched and extracted within 60 min at a level of 1 colony formation unit (CFU) from a 1 ml sample volume in the same tube. The performance of this approach is 10–100 times better than that of a commercial kit (10(2) to 10(3) CFU/ml) but does not require a large centrifuge. Finally, we combined the single-tube approach with a bio-optical sensor for rapid and accurate zoonotic pathogen detection in human urine samples. Using the combination system, Brucella in human urine can be efficiently enriched (~ 8-fold) and the detection limit is enhanced by up to 100 times (1 CFU/ml bacteria in urine) compared with the commercial kit. This combined system is fast and highly sensitive and thus represents a promising approach for disease diagnosis in the clinical setting.


title: Respiratory epithelial cells as master communicators during viral infections

text: PURPOSE OF REVIEW: Communication by epithelial cells during respiratory viral infections is critical in orchestrating effective anti-viral responses but also can lead to excessive inflammation. This review will evaluate studies that investigate how respiratory epithelial cells influence the behavior of immune cells and how epithelial cell/immune cell interactions contribute to antiviral responses and immunopathology outcomes. RECENT FINDINGS: Previous studies have characterized cytokine responses of virus-infected epithelial cells. More recent studies have carefully demonstrated the effects of these cytokines on cellular behaviors within the infected lung. Infected epithelial cells release exosomes that specifically regulate responses of monocytes and neighboring epithelial cells without promoting spread of virus. In contrast, rhinovirus-infected cells induce monocytes to upregulate expression of the viral receptor, promoting spread of the virus to alternate cell types. The precise alteration of PDL expression on infected epithelial cells has been shown to switch between inhibition and activation of antiviral responses. SUMMARY: These studies have more precisely defined the interactions between epithelial and immune cells during viral infections. This level of understanding is critical for the development of novel therapeutic strategies that promote effective antiviral responses or epithelial repair, or inhibit damaging inflammatory responses during severe respiratory viral infections.


title: Seroepidemiology of enterovirus D68 infection in China

text: Human enterovirus 68 (EV-D68) is a rarely reported virus that has been linked to respiratory disease. In recent years, reports about EV-D68 infection have markedly increased worldwide. However, the epidemiological features of this emerging infection are not well understood. To evaluate the emerging EV-D68 epidemic, we isolated the circulating viral strain and investigated the seroprevalence of neutralizing antibodies (NAbs) in Beijing between 2004 and 2011. We found that the titers of EV-D68 NAbs were generally low in all age groups in sampled populations in 2004 but significantly higher in 2009. From 2007 to 2011, the NAbs against EV-D68 significantly increased over time. These findings indicate that EV-D68 has spread widely in the Chinese population in recent years, although only a limited number of cases were reported.


title: Clinical impact of COVID-19 on patients with cancer (CCC19): a cohort study

text: BACKGROUND: Data on patients with COVID-19 who have cancer are lacking. Here we characterise the outcomes of a cohort of patients with cancer and COVID-19 and identify potential prognostic factors for mortality and severe illness. METHODS: In this cohort study, we collected de-identified data on patients with active or previous malignancy, aged 18 years and older, with confirmed severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection from the USA, Canada, and Spain from the COVID-19 and Cancer Consortium (CCC19) database for whom baseline data were added between March 17 and April 16, 2020. We collected data on baseline clinical conditions, medications, cancer diagnosis and treatment, and COVID-19 disease course. The primary endpoint was all-cause mortality within 30 days of diagnosis of COVID-19. We assessed the association between the outcome and potential prognostic variables using logistic regression analyses, partially adjusted for age, sex, smoking status, and obesity. This study is registered with ClinicalTrials.gov, NCT04354701, and is ongoing. FINDINGS: Of 1035 records entered into the CCC19 database during the study period, 928 patients met inclusion criteria for our analysis. Median age was 66 years (IQR 57–76), 279 (30%) were aged 75 years or older, and 468 (50%) patients were male. The most prevalent malignancies were breast (191 [21%]) and prostate (152 [16%]). 366 (39%) patients were on active anticancer treatment, and 396 (43%) had active (measurable) cancer. At analysis (May 7, 2020), 121 (13%) patients had died. In logistic regression analysis, independent factors associated with increased 30-day mortality, after partial adjustment, were: increased age (per 10 years; partially adjusted odds ratio 1·84, 95% CI 1·53–2·21), male sex (1·63, 1·07–2·48), smoking status (former smoker vs never smoked: 1·60, 1·03–2·47), number of comorbidities (two vs none: 4·50, 1·33–15·28), Eastern Cooperative Oncology Group performance status of 2 or higher (status of 2 vs 0 or 1: 3·89, 2·11–7·18), active cancer (progressing vs remission: 5·20, 2·77–9·77), and receipt of azithromycin plus hydroxychloroquine (vs treatment with neither: 2·93, 1·79–4·79; confounding by indication cannot be excluded). Compared with residence in the US-Northeast, residence in Canada (0·24, 0·07–0·84) or the US-Midwest (0·50, 0·28–0·90) were associated with decreased 30-day all-cause mortality. Race and ethnicity, obesity status, cancer type, type of anticancer therapy, and recent surgery were not associated with mortality. INTERPRETATION: Among patients with cancer and COVID-19, 30-day all-cause mortality was high and associated with general risk factors and risk factors unique to patients with cancer. Longer follow-up is needed to better understand the effect of COVID-19 on outcomes in patients with cancer, including the ability to continue specific cancer treatments. FUNDING: American Cancer Society, National Institutes of Health, and Hope Foundation for Cancer Research.


title: Transmission of Panton-Valentine Leukocidin–Producing Staphylococcus aureus to a Physician during Resuscitation of a Child

text: We report the first case of transmission of Panton-Valentine leukocidin–producing Staphylococcus aureus to a physician during the resuscitation of an infant with fatal pneumonia. The physician exhibited numerous furuncles. This case highlights the necessity for health care workers to protect themselves against transmission of infectious diseases from patient to care giver.


title: Phylogenetic analysis of avian infectious bronchitis virus S1 glycoprotein regions reveals emergence of a new genotype in Moroccan broiler chicken flocks

text: BACKGROUND: Infectious bronchitis virus (IBV), a major pathogen of commercial poultry flocks, circulates in the form of several serotypes/genotypes. Only a few amino-acid changes in the S1 subunit of wild-type IBVS proteins may result in mutants unaffected by current vaccines. METHODS: Partial S1 gene sequences of 3 IBV isolates of the Moroccan Italy 02 genotype from vaccinated and unvaccinated broiler chicken flocks, located in southern and central regions of Morocco, were amplified by RT-PCR, sequenced, and aligned for phylogenetic and amino-acid similarity analyses. RESULTS: The three isolates were found genetically highly distant from known avian IBV based on partial sequences of their S1 genes: gammaCoV/chicken/Morocco/I01/2011(IBV/Morocco/01), gammaCoV/chicken/Morocco/I30/2010 (IBV/Morocco/30), and gammaCoV/chicken/Morocco/I38/2013 (IBV/Morocco/38), nucleotide sequence identities reached 89.5 % to 90.9 % among the three isolates. The deduced protein sequence identities ranged from 29.7 % (between IBV/Morocco/38 and Egypt SCU-14/2013-1) to 78.2 % (between IBV/Morocco/01 and Spain/05/866). Amino acid sequence comparison and phylogenetic analysis indicated the emergence of a new Moroccan genotype, clustering with regionally related isolates from Spain (Spain/05/866) and belonging to a new sub-genotype. CONCLUSION: Our sequencing results demonstrate a co-circulation of wild-type infectious bronchitis viruses in broiler chickens. These results justify permanent monitoring of circulating strains in order to rationally modify vaccination strategies to make them appropriate to the evolving field situation.


title: Universal face masking in community: Material, design, and considerations—reply

text: See related letter See related letter


title: On the Host Side of the Hepatitis E Virus Life Cycle

text: Hepatitis E virus (HEV) infection is one of the most common causes of acute hepatitis in the world. HEV is an enterically transmitted positive-strand RNA virus found as a non-enveloped particle in bile as well as stool and as a quasi-enveloped particle in blood. Current understanding of the molecular mechanisms and host factors involved in productive HEV infection is incomplete, but recently developed model systems have facilitated rapid progress in this area. Here, we provide an overview of the HEV life cycle with a focus on the host factors required for viral entry, RNA replication, assembly and release. Further developments of HEV model systems and novel technologies should yield a broader picture in the future.


title: CovidCounties - an interactive, real-time tracker of the COVID-19 pandemic at the level of US counties

text: Management of the COVID-19 pandemic has proven to be a significant challenge to policy makers. This is in large part due to uneven reporting and the absence of open-access visualization tools to present local trends and infer healthcare needs. Here we report the development of CovidCounties.org, an interactive web application that depicts daily disease trends at the level of US counties using time series plots and maps. This application is accompanied by a manually curated dataset that catalogs all major public policy actions made at the state-level, as well as technical validation of the primary data. Finally, the underlying code for the site is also provided as open source, enabling others to validate and learn from this work.


title: Analysis of intrapatient heterogeneity uncovers the microevolution of Middle East respiratory syndrome coronavirus

text: Genome sequence analysis of Middle East respiratory syndrome coronavirus (MERS-CoV) variants from patient specimens has revealed the evolutionary dynamics and mechanisms of pathogenesis of the virus. However, most studies have analyzed the consensus sequences of MERS-CoVs, precluding an investigation of intrapatient heterogeneity. Here, we analyzed non–consensus sequences to characterize intrapatient heterogeneity in cases associated with the 2015 outbreak of MERS in South Korea. Deep-sequencing analysis of MERS-CoV genomes performed on specimens from eight patients revealed significant intrapatient variation; therefore, sequence heterogeneity was further analyzed using targeted deep sequencing. A total of 35 specimens from 24 patients (including a super-spreader) were sequenced to detect and analyze variants displaying intrapatient heterogeneity. Based on the analysis of non–consensus sequences, we demonstrated the intrapatient heterogeneity of MERS-CoVs, with the highest level in the super-spreader specimen. The heterogeneity could be transmitted in a close association with variation in the consensus sequences, suggesting the occurrence of multiple MERS-CoV infections. Analysis of intrapatient heterogeneity revealed a relationship between D510G and I529T mutations in the receptor-binding domain (RBD) of the viral spike glycoprotein. These two mutations have been reported to reduce the affinity of the RBD for human CD26. Notably, although the frequency of both D510G and I529T varied greatly among specimens, the combined frequency of the single mutants was consistently high (87.7% ± 1.9% on average). Concurrently, the frequency of occurrence of the wild type at the two positions was only 6.5% ± 1.7% on average, supporting the hypothesis that selection pressure exerted by the host immune response played a critical role in shaping genetic variants and their interaction in human MERS-CoVs during the outbreak.


COVID-19 pandemic: Impact on children, families and the future


title: Acute phase response in bovine coronavirus positive post-weaned calves with diarrhea

text: Bovine coronavirus (BCoV) is associated with severe diarrhea in calves, winter dysentery in adult cattle, and respiratory diseases in cattle of all ages. This study aimed to investigate the relationship between white blood cell counts and haptoglobin (Hp) and serum amyloid A (SAA) levels in post-weaned calves with diarrhea caused by BCoV and those that recovered from diarrhea. Blood and fecal samples were collected twice from the same animals; 17 post-weaned calves with diarrhea (first) and 15 post-weaned calves that recovered from diarrhea (second). Real-time polymerase chain reaction revealed that all 17 fecal samples from post-weaned calves with diarrhea and one out of 15 from diarrhea-recovered calves were positive for BCoV and negative for Cryptosporidium spp., Escherichia coli K99, Salmonella spp., bovine rotavirus, and bovine viral diarrhea virus. No Eimeria oocysts were detected using the flotation method. In comparison with post-weaned calves with diarrhea, in diarrhea-recovered calves, the lymphocyte count was significantly higher (P = 0.018), and the monocyte count was significantly lower (P = 0.001); however, the number of monocytes was still high. Post-weaned calves with diarrhea had a significantly higher Hp concentration (P < 0.001) compared with diarrhea-recovered calves. The results indicated that increased Hp concentration and monocytosis but not SAA may be associated with diarrhea caused by BCoV. The present study suggests that the monitoring of Hp concentration and monocyte count is useful in the diagnosis of post-weaned calves with diarrhea caused by BCoV in this field.


title: Drug combination therapy in control of cryptosporidiosis in Ludhiana district of Punjab

text: The present report describes outbreak of cryptosporidiosis in neonatal cross bred cattle calves ageing 1–2 months in an organized dairy farm. The protozoan infection was confirmed by identifying bright red oocysts of Cryptosporidium spp. in the faecal samples after staining with modified acid Fast Zeihl–Neelsen stain. Metronidazole and furazolidone combination was able to induce clinically and parasitological recovery. This is believed to be the first report on the successful use of this drug combination against cryptosporidiosis.


title: A comparative analysis of control measures on-board ship against COVID-19 and similar novel viral respiratory disease outbreak: Quarantine ship or disembark suspects?

text: BACKGROUND: Management of novel viral respiratory disease outbreak on-board a ship with person-to-person transmission can be a public health challenge because of close proximity of inhabitants due to confined space and air-conditioned environment. It has a potential to be explosive, with high secondary attack rate (SAR) and cause significant morbidity and mortality. This study compares control measures instituted on-board two ships with similar outbreaks and recommends a standardized evidence-based outbreak response against them. METHODS: This is a descriptive study, showing comparative analysis of control measures instituted on-board two ships, a cruise ship in case of COVID-19 and a warship in case of H1N1 influenza, with novel viral respiratory disease outbreak, at different span of time. Data of the date of onset, clinical details, laboratory results, history of travel, history of contact with positive case and control measures initiated were collected, analysed and compared. RESULTS: Of the two ships compared, one was a cruise ship with 712 COVID-19 cases, with an attack rate (AR) of 19.2% and 13 deaths, and other a warship with 14 cases of H1N1 influenza and an AR of 4.83%. The epidemic curve for both the outbreaks was plotted to study time distribution. CONCLUSION: Active surveillance, early self-reporting and immediate disembarkation of the suspects, along with strict compliance of hand hygiene, cough etiquettes and disinfection enhancement, will help in early mitigation of the outbreak. Health education should be undertaken to impart evidence-based knowledge and alleviate fear of the unknown. Vaccination may not be present but if available should only be administered after strict risk–benefit, cost–benefit and effectiveness analysis.


title: Skin layer in cyclic loading-cleaning of a nanofiber filter in filtering nano-aerosols

text: Abstract Nano-aerosols from viruses to virgin pollutant particulates from combustion, 100nm or smaller, are harmful to our health as they penetrate readily into our body causing various diseases. Nanofiber filter can capture effectively these nano-aerosols. However, over time the pressure drop increases dramatically and cleaning of the filter by backpulse/backblow is essential for filter reuse. The cyclic loading-and-cleaning of a nanofiber filter has been investigated for the first time experimentally and theoretically. The “skin” layer, a thin region upstream of the nanofiber filter, plays a pivoting role in controlling the pressure drop excursion of the filter. We model the skin layer to be made up of numerous fine capillaries and examine how continuous aerosols deposited in the capillaries affect rapid rise in pressure drop followed by bridging of aerosols across the capillary openings leading to more bridging and ultimately formation of cake on top of the bridges and filter surface. We have been able to describe the deposition of aerosols in the capillary pores for depth filtration, the deposition of aerosols in the cake (surface filtration), and the intermediate bridging regime between these two. We can depict the complete pressure drop excursion including the S-shaped curve behavior from depth filtration transiting to surface filtration for a filter with severe skin effect. Our prediction matches extremely well with the 6 cycles of loading/cleaning on a 280-nm nanofiber filter subject to challenging nano-aerosols, 50–400nm. During cyclic loading and cleaning, the porosity and permeability in the skin layer for our experiment drop to 68% and 11–21% of their original values, respectively, and the effective pore diameter also drops from 1.2 to 0.6μm.


title: Amino acid substitutions within the heptad repeat domain 1 of murine coronavirus spike protein restrict viral antigen spread in the central nervous system

text: Abstract Targeted recombination was carried out to select mouse hepatitis viruses (MHVs) in a defined genetic background, containing an MHV-JHM spike gene encoding either three heptad repeat 1 (HR1) substitutions (Q1067H, Q1094H, and L1114R) or L1114R alone. The recombinant virus, which expresses spike with the three substitutions, was nonfusogenic at neutral pH. Its replication was significantly inhibited by lysosomotropic agents, and it was highly neuroattenuated in vivo. In contrast, the recombinant expressing spike with L1114R alone mediated cell-to-cell fusion at neutral pH and replicated efficiently despite the presence of lysosomotropic agents; however, it still caused only subclinical morbidity and no mortality in animals. Thus, both recombinant viruses were highly attenuated and expressed viral antigen which was restricted to the olfactory bulbs and was markedly absent from other regions of the brains at 5 days postinfection. These data demonstrate that amino acid substitutions, in particular L1114R, within HR1 of the JHM spike reduced the ability of MHV to spread in the central nervous system. Furthermore, the requirements for low pH for fusion and viral entry are not prerequisites for the highly attenuated phenotype.


title: Impact of infection preventionists on Centers for Medicare and Medicaid quality measures in Maryland nursing homes

text: BACKGROUND: Health care-associated infections are the leading cause of morbidity and mortality in US nursing homes (NHs). The objective of the research is to assess the impact of Maryland NH infection preventionists (IPs) on NH quality measures. METHODS: Two hundred thirty-four NHs were queried through mailed survey. These survey data were then linked with 2008 quality data from Nursing Home Compare and the On-line Survey Certification of Automated Records. RESULTS: Three of the 8 quality measures examined—influenza vaccination for both short- and long-stay residents and pressure ulcer prevention in high-risk residents—were significantly associated with the number of IPs. None of the quality measures were shown to be significant with IPs who received specialized training on infection prevention and management compared with those who did not receive specialized training. CONCLUSION: IPs play a critical role in preventing and managing health care-associated infections in nursing homes, especially in the areas of influenza vaccination and pressure ulcer prevention among high-risk nursing home residents. Quality measures that reflect the effects of IP training may not have been elucidated yet. Further research is needed to support the IP role in order for policy to advocate for increased IP funding.


title: Dangerous infectious diseases: Bad news for Main Street, good news for Wall Street?

text: Abstract We examine whether investor mood, driven by World Health Organization (WHO) alerts and media news on dangerous infectious diseases, is priced in pharmaceutical companies' stocks in the United States. We argue that disease-related news (DRNs) should not trigger rational trading. We find that DRNs have a positive and significant sentiment effect among investors (on Wall Street). The effect is stronger (weaker) for small (large) companies, who are less (more) likely to engage in the development of new vaccines. A potential negative investor climate (on Main Street) – induced by disease-related fear – does not alter the positive sentiment effect.


title: Porcine alveolar macrophage polarization is involved in inhibition of porcine reproductive and respiratory syndrome virus (PRRSV) replication

text: Macrophage polarization is a process by which macrophages acquire a distinct phenotypic and functional profile in response to microenvironmental signals. The classically and alternatively activated (M1 and M2, respectively) macrophage phenotypes are defined by the specific molecular characteristics induced in response to prototypic pro- and anti-inflammatory cues. In this study, we used LPS/IFN-γ and IL-4 to stimulate porcine alveolar macrophages (PAMs) in vitro and investigated the expression changes of several novel markers during macrophage polarization. Notably, we found that LPS/IFN-γ-stimulated PAMs express prototypical M1 molecules, whereas IL-4-stimulated PAMs express M2 molecules. We also demonstrated that replication of the highly pathogenic porcine reproductive and respiratory syndrome virus (PRRSV) strain HuN4 was effectively suppressed in LPS/IFN-γ-stimulated M1 PAMs (M1 type), but not IL-4 stimulated M2 PAMs. However, this was not observed with the classic, less pathogenic CH-1a strain. Moreover, we found that M2 marker expression gradually increased after PAM infection with PRRSV, whereas no significant changes were found with M1 marker expression, suggesting that PRRSV infection may skew macrophage polarization towards an M2 phenotype. Finally, we found that anti-viral cytokine expression was significantly higher in M1 macrophages than in M2 macrophages or nonpolarized controls. In summary, our results show that PRRSV replication was significantly impaired in M1 PAMs, which may serve as a foundation for further understanding of the dynamic phenotypic changes during macrophage polarization and their effects on viral infection.


Clarification on mask use during droplet precautions


title: Cytoplasmic Viral Replication Complexes

text: Many viruses that replicate in the cytoplasm compartmentalize their genome replication and transcription in organelle-like structures that enhance replication efficiency and protection from host defenses. In particular, recent studies with diverse positive-strand RNA viruses have further elucidated the ultrastructure of membrane-bounded RNA replication complexes and their close coordination with virion assembly and budding. The structure, function and assembly of some positive-strand RNA virus replication complexes have parallels and potential evolutionary links with the replicative cores of double-strand RNA virus and retrovirus virions, and more general similarities with the replication factories of cytoplasmic DNA viruses.


April 26–May 2, 2014


title: Developmentally regulated O-acetylated sialoglycans in the central nervous system revealed by a new monoclonal antibody 493D4 recognizing a wide range of O-acetylated glycoconjugates

text: We have previously detected an alkali-labile and developmentally regulated antigen in rat embryonic cerebral cortex, which may be 9-O-acetylsialylated GT3 ganglioside (Hirabayashi Y, Hirota M, Suzuki Y, Matsumoto M, Obata K, Ando S (1989) Neurosci Lett 106:193-98). In this study we established a mouse monoclonal antibody, 493D4, that recognizes 9-O-acetyl GT3 ganglioside, but not non-O-acetyl gangliosides. This antibody also reacted with 9-O-acetyl GD3 to a much lesser extent. By using this antibody, we found that O-acetyl GT3 as well as O-acetyl GD3 were expressed strongly in fetal murine cerebral cortex and decreased to an undetectable level after birth. With the assistance of TLC-immunostaining using 493D4 together with Q-Sepharose column chromatography, O-acetyl gangliosides of bovine brain were purified and the structural analysis showed the presence of O-acetyl GD3, O-acetyl LD1, O-acetyl GD2 and O-acetyl GD1b in the adult brain as extremely minor components. Interestingly, the antibody 493D4 could detect O-acetyl sialoglycoproteins in rat brain tissues. One of the major immunoreactive proteins was shown to be synaptophysin, an integral membrane protein specifically present in synaptic vesicles. This monoclonal antibody was therefore useful for sensitive detection of both O-acetylated gangliosides and glycoproteins with O-acetylated sialic acids.


title: Chapter 4 Infrared Thermography and Soft Computing for Diabetic Foot Assessment

text: Abstract Recent advancements in digital image processing and soft computing techniques have widened the scope of computer aided diagnosis of medical conditions. Many imaging modalities like MRI, CT, PET, Ultrasound combined with soft computing techniques is already contributing to this trend. With the recent inclusion of infrared thermal imaging, the capability of computer aided diagnosis has increased and has become more safe and convenient. Research in this noncontact and noninvasive imaging technology has steadily increased over the last 50 years. Disease diagnosis based on the correlation of surface temperature distribution of skin is being studied at large and has shown promising results. This chapter will give the reader a solid understanding of the theory behind infrared thermography and the use of soft computing techniques applied to medical image analysis, particularly for diabetic foot complication assessment. The issues and challenges to be addressed in using infrared thermography for diagnostic purposes are also discussed. The reader will get a complete overview of building an intelligent diagnostic system using the two sensational topics of research in machine learning and medical imaging—infrared thermography and soft computing.


title: Sequence–function analysis of the Sendai virus L protein domain VI

text: Abstract The large (about 2200 amino acids) L polymerase protein of nonsegmented negative-strand RNA viruses (order Mononegavirales) has six conserved sequence regions (“domains”) postulated to constitute the specific enzymatic activities involved in viral mRNA synthesis, 5′-end capping, cap methylation, 3′ polyadenylation, and genomic RNA replication. Previous studies with vesicular stomatitis virus identified amino acid residues within the L protein domain VI required for mRNA cap methylation. In our recent study we analyzed four amino acid residues within domain VI of the Sendai virus L protein and our data indicated that there could be differences in L protein sequence requirements for cap methylation in two different families of Mononegavirales — rhabdoviruses and paramyxoviruses. In this study, we conducted a more comprehensive mutational analysis by targeting the entire SeV L protein domain VI, creating twenty-four L mutants, and testing these mutations for their effects on viral mRNA synthesis, cap methylation, viral genome replication and virus growth kinetics. Our analysis identified several residues required for successful cap methylation and virus replication and clearly showed the importance of the K-D-K-E tetrad and glycine-rich motif in the SeV cap methylation. This study is the first extensive sequence analysis of the L protein domain VI in the family Paramyxoviridae, and it confirms structural and functional similarity of this domain across different families of the order Mononegavirales.


title: Identification of an ideal adjuvant for receptor-binding domain-based subunit vaccines against Middle East respiratory syndrome coronavirus

text: Middle East respiratory syndrome (MERS), an emerging infectious disease caused by MERS coronavirus (MERS-CoV), has garnered worldwide attention as a consequence of its continuous spread and pandemic potential, making the development of effective vaccines a high priority. We previously demonstrated that residues 377–588 of MERS-CoV spike (S) protein receptor-binding domain (RBD) is a very promising MERS subunit vaccine candidate, capable of inducing potent neutralization antibody responses. In this study, we sought to identify an adjuvant that optimally enhanced the immunogenicity of S377–588 protein fused with Fc of human IgG (S377–588-Fc). Specifically, we compared several commercially available adjuvants, including Freund's adjuvant, aluminum, Monophosphoryl lipid A, Montanide ISA51 and MF59 with regard to their capacity to enhance the immunogenicity of this subunit vaccine. In the absence of adjuvant, S377–588-Fc alone induced readily detectable neutralizing antibody and T-cell responses in immunized mice. However, incorporating an adjuvant improved its immunogenicity. Particularly, among the aforementioned adjuvants evaluated, MF59 is the most potent as judged by its superior ability to induce the highest titers of IgG, IgG1 and IgG2a subtypes, and neutralizing antibodies. The addition of MF59 significantly augmented the immunogenicity of S377–588-Fc to induce strong IgG and neutralizing antibody responses as well as protection against MERS-CoV infection in mice, suggesting that MF59 is an optimal adjuvant for MERS-CoV RBD-based subunit vaccines.


title: Direct rhizogenesis, in vitro stolon proliferation and high-throughput regeneration of plantlets in Glycyrrhiza glabra

text: Direct rhizogenesis from leaf explants and establishment of an in vitro stolon culture system and subsequent plant regeneration for Glycyrrhiza glabra have been described. MS liquid medium supplemented with 0.01 mg l(−1) of NAA was most effective for stolon proliferation. Extensive proliferation of stolon and shoot regeneration was achieved on medium containing 3 % sucrose with 0.01 mg l(−1) NAA. Stolons with nodes showing growth was transferred under light for plantlet regeneration in the same medium. This paper is the first report in G. glabra describing a complete regeneration procedure via in vitro stolon proliferation along with quantitative data for glycyrrhizin and genetic fidelity of plant regenerated in vitro there from. In vitro stolon proliferation described here would be an efficient way for regeneration of plants for functional genomics studies and better understanding of glycyrrhizin (GA) metabolism.


title: Chapter 8 Zoonoses in Wildlife Integrating Ecology into Management

text: Abstract Zoonoses in wildlife not only play an important ecological role, but pose significant threats to the health of humans, domestic animals and some endangered species. More than two‐thirds of emerging, or re‐emerging, infectious diseases are thought to originate in wildlife. Despite this, co‐ordinated surveillance schemes are rare, and most efforts at disease control operate at the level of crisis management. This review examines the pathways linking zoonoses in wildlife with infection in other hosts, using examples from a range of key zoonoses, including European bat lyssaviruses and bovine tuberculosis. Ecologically based control, including the management of conditions leading to spill‐overs into target host populations, is likely to be more effective and sustainable than simple reductions in wildlife populations alone.


title: Molecular Evidence of Adenosine Deaminase Linking Adenosine A(2A) Receptor and CD26 Proteins

text: Adenosine is an endogenous purine nucleoside that acts in all living systems as a homeostatic network regulator through many pathways, which are adenosine receptor (AR)-dependent and -independent. From a metabolic point of view, adenosine deaminase (ADA) is an essential protein in the regulation of the total intracellular and extracellular adenosine in a tissue. In addition to its cytosolic localization, ADA is also expressed as an ecto-enzyme on the surface of different cells. Dipeptidyl peptidase IV (CD26) and some ARs act as binding proteins for extracellular ADA in humans. Since CD26 and ARs interact with ADA at opposite sites, we have investigated if ADA can function as a cell-to-cell communication molecule by bridging the anchoring molecules CD26 and A(2A)R present on the surfaces of the interacting cells. By combining site-directed mutagenesis of ADA amino acids involved in binding to A(2A)R and a modification of the bioluminescence resonance energy transfer (BRET) technique that allows detection of interactions between two proteins expressed in different cell populations with low steric hindrance (NanoBRET), we show direct evidence of the specific formation of trimeric complexes CD26-ADA-A(2A)R involving two cells. By dynamic mass redistribution assays and ligand binding experiments, we also demonstrate that A(2A)R-NanoLuc fusion proteins are functional. The existence of this ternary complex is in good agreement with the hypothesis that ADA could bridge T-cells (expressing CD26) and dendritic cells (expressing A(2A)R). This is a new metabolic function for ecto-ADA that, being a single chain protein, it has been considered as an example of moonlighting protein, because it performs more than one functional role (as a catalyst, a costimulator, an allosteric modulator and a cell-to-cell connector) without partitioning these functions in different subunits.


title: Tackling dengue fever: Current status and challenges

text: According to recent statistics, 96 million apparent dengue infections were estimated worldwide in 2010. This figure is by far greater than the WHO prediction which indicates the rapid spread of this disease posing a growing threat to the economy and a major challenge to clinicians and health care services across the globe particularly in the affected areas. This article aims at bringing to light the current epidemiological and clinical status of the dengue fever. The relationship between genetic mutations, single nucleotide polymorphism (SNP) and the pathophysiology of disease progression will be put into perspective. It will also highlight the recent advances in dengue vaccine development. Thus far, a significant progress has been made in unraveling the risk factors and understanding the molecular pathogenesis associated with the disease. However, further insights in molecular features of the disease and the development of animal models will enormously help improving the therapeutic interventions and potentially contribute to finding new preventive measures for population at risk.


The development and application of Laboratory Animal Science in China


title: Chapter 10 Genetic Basis of Salmonella Enteritidis Pathogenesis in Chickens

text: Abstract Salmonella Enteritidis (SE) is the predominant cause of the food-borne salmonellosis in humans, in part because this serotype has the unique ability to contaminate chicken eggs without causing discernible illness in the infected birds. Attempts to develop effective vaccines and eradicate SE from chickens are undermined by significant limitations in our current understanding of the genetic basis of pathogenesis of SE in this reservoir host. In this chapter, we summarize the infection kinetics and provide an overview of the current understanding of genetic factors underlying SE infection in the chicken host. We also discuss the important knowledge gaps that, if addressed, will improve our understanding of the complex biology of SE in young chickens and in egg laying hens.


title: Construction of a food-grade cell surface display system for Lactobacillus casei

text: Abstract In this study, a food-grade cell surface display host/vector system for Lactobacillus casei was constructed. The food-grade host L. casei Q-5 was a lactose-deficient derivative of L. casei ATCC 334 obtained by plasmid elimination. The food-grade cell surface display vector was constructed based on safe DNA elements from lactic acid bacteria containing the following: pSH71 replicon from Lactococcus lactis, lactose metabolism genes from L. casei ATCC 334 as complementation markers, and surface layer protein gene from Lactobacillus acidophilus ATCC 4356 for cell surface display. The feasibility of the new host/vector system was verified by the expression of green fluorescent protein (GFP) on L. casei. Laser scanning confocal microscopy and immunofluorescence analysis using anti-GFP antibody confirmed that GFP was anchored on the surface of the recombinant cells. The stability of recombinant L. casei cells in artificial gastrointestinal conditions was verified, which is beneficial for oral vaccination applications. These results indicate that the food-grade host/vector system can be an excellent antigen delivery vehicle in oral vaccine construction.


title: Human coronavirus OC43 3CL protease and the potential of ML188 as a broad-spectrum lead compound: Homology modelling and molecular dynamic studies

text: BACKGROUND: The coronavirus 3 chymotrypsin-like protease (3CL(pro)) is a validated target in the design of potential anticoronavirus inhibitors. The high degree of homology within the protease’s active site and substrate conservation supports the identification of broad spectrum lead compounds. A previous study identified the compound ML188, also termed 16R, as an inhibitor of the Severe Acute Respiratory Syndrome coronavirus (SARS-CoV) 3CL(pro). This study will detail the generation of a homology model of the 3CL(pro) of the human coronavirus OC43 and determine the potential of 16R to form a broad-spectrum lead compound. MODELLER was used to generate a suitable three-dimensional model of the OC43 3CL(pro) and the Prime module of Schrӧdinger predicted the binding conformation and free energy of binding of 16R within the 3CL(pro) active site. Molecular dynamics further confirmed ligand stability and hydrogen bonding networks. RESULTS: A high quality homology model of the OC43 3CL(pro) was successfully generated in an active conformation. Further studies reproduced the binding pose of 16R within the active site of the generated model, where its free energy of binding was shown to equal that of the 3CL(pro) of SARS-CoV, a receptor it is experimentally proven to inhibit. The stability of the ligand was subsequently confirmed by molecular dynamics. CONCLUSION: The lead compound 16R may represent a broad-spectrum inhibitor of the 3CL(pro) of OC43 and potentially other coronaviruses. This study provides an atomistic structure of the 3CL(pro) of OC43 and supports further experimental validation of the inhibitory effects of 16R. These findings further confirm that the 3CL(pro) of coronaviruses can be inhibited by broad spectrum lead compounds.


Part 2: Adult basic life support


title: Special Features of Bat Microbiota Differ From Those of Terrestrial Mammals

text: Bats (order Chiroptera) are one of the most diverse and widely distributed group of mammals with a close relationship to humans. Over the past few decades, a number of studies have been performed on bat viruses; in contrast, bacterial pathogens carried by bats were largely neglected. As more bacterial pathogens are being identified from bats, the need to study their natural microbiota is becoming urgent. In the current study, fecal samples of four bat species from different locations of China were analyzed for their microbiota composition. Together with the results of others, we concluded that bat microbiota is most commonly dominated by Firmicutes and Proteobacteria; the strict anaerobic phylum Bacteroidetes, which is dominant in other terrestrial mammals, especially humans and mice, is relatively rare in bats. This phenomenon was interpreted as a result of a highly specified gastrointestinal tract in adaptation to the flying lifestyle of bats. Further comparative study implied that bat microbiota resemble those of the order Carnivora. To discover potential bacterial pathogens, a database was generated containing the 16S rRNA gene sequences of known bacterial pathogens. Potential bacterial pathogens belonging to 12 genera were detected such as Salmonella, Shigella, and Yersinia, among which some have been previously reported in bats. This study demonstrated high resolution and repeatability in detecting organisms of rare existence, and the results could be used as guidance for future bacterial pathogen isolation.


title: Prehabilitation May Help Mitigate an Increase in COVID-19 Peripandemic Surgical Morbidity and Mortality

text: As physicians specializing in rehabilitation medicine consider sequelae from the novel coronavirus pandemic that began in 2019, one issue that should be top of mind is the physiologic effect that large-scale social distancing had on the health of patients in general but, more specifically, on preoperative patients who had their surgeries delayed or will have newly scheduled procedures during the peripandemic period. Predictably, as the virus becomes less prevalent, there will be a tremendous motivation to move forward with scheduling operations from both patient care and institutional perspectives. However, one can anticipate a pandemic-related increase in surgical morbidity and mortality above prepandemic levels, particularly in older or medically frail patients even if they did not have a novel coronavirus (i.e., COVID-19) infection. Therefore, now is the time to consider for patients awaiting surgery a wider adoption of prehabilitation—physical and psychological assessments that establish a baseline functional level, identify impairments, and provide interventions that promote physical and psychological health to reduce the incidence and/or severity of future impairments.