Reconstitution of the SARS-CoV-2 ribonucleosome provides insights into genomic RNA packaging and regulation by phosphorylation

The nucleocapsid (N) protein of coronaviruses is responsible for compaction of the [~]30-kb RNA genome in the [~]100-nm virion. Cryo-electron tomography suggests that each virion contains 35-40 viral ribonucleoprotein (vRNP) complexes, or ribonucleosomes, arrayed along the genome. There is, however, little mechanistic understanding of the vRNP complex. Here, we show that N protein, when combined with viral RNA fragments in vitro, forms cylindrical 15-nm particles similar to the vRNP structures observed within coronavirus virions. These vRNPs form in the presence of stem-loop-containing RNA and depend on regions of N protein that promote protein-RNA and protein-protein interactions. Phosphorylation of N protein in its disordered serine/arginine (SR) region weakens these interactions and disrupts vRNP assembly. We propose that unmodified N binds stem-loop-rich regions in genomic RNA to form compact vRNP complexes within the nucleocapsid, while phosphorylated N maintains uncompacted viral RNA to promote the proteins transcriptional function.

CryoEM and AI reveal a structure of SARS-CoV-2 Nsp2, a multifunctional protein involved in key host processes.

The SARS-CoV-2 protein Nsp2 has been implicated in a wide range of viral processes, but its exact functions, and the structural basis of those functions, remain unknown. Here, we report an atomic model for full-length Nsp2 obtained by combining cryo-electron microscopy with deep learning-based structure prediction from AlphaFold2. The resulting structure reveals a highly-conserved zinc ion-binding site, suggesting a role for Nsp2 in RNA binding. Mapping emerging mutations from variants of SARS-CoV-2 on the resulting structure shows potential host-Nsp2 interaction regions. Using structural analysis together with affinity tagged purification mass spectrometry experiments, we identify Nsp2 mutants that are unable to interact with the actin-nucleation-promoting WASH protein complex or with GIGYF2, an inhibitor of translation initiation and modulator of ribosome-associated quality control. Our work suggests a potential role of Nsp2 in linking viral transcription within the viral replication-transcription complexes (RTC) to the translation initiation of the viral message. Collectively, the structure reported here, combined with mutant interaction mapping, provides a foundation for functional studies of this evolutionary conserved coronavirus protein and may assist future drug design.

Bivalent binding of a fully human IgG to the SARS-CoV-2 spike proteins reveals mechanisms of potent neutralization

In vitro antibody selection against pathogens from naive combinatorial libraries can yield various classes of antigen-specific binders that are distinct from those evolved from natural infection1-4. Also, rapid neutralizing antibody discovery can be made possible by a strategy that selects for those interfering with pathogen and host interaction5. Here we report the discovery of antibodies that neutralize SARS-CoV-2, the virus responsible for the COVID-19 pandemic, from a highly diverse naive human Fab library. Lead antibody 5A6 blocks the receptor binding domain (RBD) of the viral spike from binding to the host receptor angiotensin converting enzyme 2 (ACE2), neutralizes SARS-CoV-2 infection of Vero E6 cells, and reduces viral replication in reconstituted human nasal and bronchial epithelium models. 5A6 has a high occupancy on the viral surface and exerts its neutralization activity via a bivalent binding mode to the tip of two neighbouring RBDs at the ACE2 interaction interface, one in the "up" and the other in the "down" position, explaining its superior neutralization capacity. Furthermore, 5A6 is insensitive to several spike mutations identified in clinical isolates, including the D614G mutant that has become dominant worldwide. Our results suggest that 5A6 could be an effective prophylactic and therapeutic treatment of COVID-19.

Selection of homemade mask materials for preventing transmission of COVID-19: a laboratory study

The Coronavirus Disease 2019 (COVID-19) has swept the whole world with high mortality. Since droplet transmission is the main route of transmission, wearing a mask serves as a crucial preventive measure. However, the virus has spread quite quickly, causing severe mask shortage. Finding alternative materials for homemade masks while ensuring the significant performance indicators will help alleviate the shortage of masks. Referring to the national standard for the "Surgical Mask" of China, 17 materials to be selected for homemade masks were tested in four key indicators: pressure difference, particle filtration efficiency, bacterial filtration efficiency and resistance to surface wetting. Eleven single-layer materials met the standard of pressure difference ([≤]49 Pa), of which 3 met the standard of resistance to surface wetting ([≥]3), 1 met the standard of particle filtration efficiency ([≥]30%), but none met the standard of bacterial filtration efficiency ([≥]95%). Based on the testing results of single-layer materials, fifteen combinations of paired materials were tested. The results showed that three double-layer materials including double-layer medical non-woven fabric, medical non-woven fabric plus non-woven shopping bag, and medical non-woven fabric plus granular tea towel could meet all the standards of pressure difference, particle filtration efficiency, and resistance to surface wetting, and were close to the standard of the bacterial filtration efficiency. In conclusion, if resources are severely lacking and medical masks cannot be obtained, homemade masks using available materials, based on the results of this study, can minimize the chance of infection to the maximum extent.

Analysis of Risk Factors of SkinAdverse Reactions by Intravenous Injection of Iodine ContrastAgent / 中国药房

OBJECTIVE：To investiga te main risk factors for adverse drug reactions （ADR）of skin by intravenous injection of iodine contrast agent. METHODS ：From Jan. 2009 to Apr. 2020，the patients suffering from skin ADR after enhanced CT with iodine contrast agent were collected from our hospital. The basic information ，laboratory test results before using iodine contrast agent and ADR related information were collected through hospital information system （HIS）. The use of iodine contrast agent ，main manifestations of skin ADR and drug combination were analyzed statistically. Taking the sex ，age，body mass index （BMI），the dosage of iodine contrast agent ，length of stay ，laboratory examination ，tumor history ，basic disease ，allergy history ，drinking history as independent variables ，the incidence of skin ADR related to iodine contrast agent was analyzed by single factor analysis ，and the variables with statistically significant were selected for multivariate Logistic stepwise regression analysis. RESULTS ：There were 157 cases of skin ADR ，involving 79 males（50.3％）and 78 females（49.7％）. The age ranged from 19 to 86 years old ，being（52.68± 18.73）years old in average. BMI was 14.6-40.7 kg/m2，being（22.5±3.7） kg/m2. 67 cases（42.68％）were treated with iprodione ，34 cases（21.66％）with iodixanol ，31 cases（19.74％）with iohexol and 25 cases（15.92％）with iopamidol ；the dose of iodine contrast agent were 50-100 mL，being（73.06±13.29）mL in average. There was no significant difference among different dosage of 4 kinds of iodine contrast agents （P≤0.05）. Among 4 kinds of iodine contrast agents ，the incidence of skin ADR induced by iopromide was the highest（0.197％）. The skin ADR related to iodine contrast agent was mainly acute （89.2％），the severity was mild （75.2％），and urticaria（38.9％）was the most common. After symptomatic treatment ，135 cases were cured ，13 cases were improved and 9 cases were not improved. Among the patients with iodine contrast agent related skin ADR ，the incidence of ADR induced by combined use of anti infective drugs was the highest （33.1％）；however，the combined use of anti-tumor drugs was the main cause of severe skin ADR. The length of stay ｛11～20 d[OR＝1.21，95％CI（1.07，1.20），P＝0.042]、21～30 d[OR＝1.39，95％CI（1.12，1.52），P＝0.035]、31～40 d[OR＝1.15，95％CI（1.03，1.37），P＝0.008]、＞40 d[OR＝1.33，95％CI（1.28，1.53），P＝0.003]｝，respitatory and circulatory system tumor history[OR ＝1.51，95％CI（1.35，1.61），P＝0.037]，injection allergy history[OR ＝1.50，95％CI（1.37，1.59），P＝0.005] can significantly increase the incidence of iodine contrast agent related skin ADR. CONCLUSIONS ：The main manifestation of skin ADR related to iodine contrast agent was urticaria. The main risk factors of skin ADR related to iodine contrast agent were length of stay （＞ 10 d），respiratory and circulatory system tumor history and injection allergy history.

Impact of visceral obesity on the short-term outcomes after radical operation for mid-low rectal cancers / 中国肿瘤临床

Objective: To investigate the effect of visceral obesity on the short-term outcomes after radical operation for mid-low rectal cancers. Methods: We conducted a prospective study on patients who underwent selective rectal cancer resection at The Second Affili-ated Hospital of Wenzhou Medical University between April 2017 and October 2018. The cutoff visceral fat area (VFA) for visceral obe-sity was≥134.6 cm2 for men and≥91.1 cm2 for women. Results: A total of 127 patients were included in the study, of whom 64 were diagnosed as having visceral obesity and 63 as having non-visceral obesity. The patients with visceral obesity had a higher body mass index (BMI) (P<0.001) than those without visceral obesity. The proportions of female patients and those who had a laparoscopy-assist-ed surgery were higher in the visceral obesity group than in the non-visceral obesity group. We found no significant differences in age, albumin level, hemoglobin count, American Society of Anesthesiologists (ASA) stage, Nutritional Risk Screening (NRS) 2002 score, Charlson comorbidity index, tumor location, TNM stage, lymphatic invasion, and laparoscopy-assisted surgery between the two groups. The postoperative complication rate was significantly higher in the visceral obesity group than in the non-visceral obesity group (35.9% vs . 19% , P=0.033). A multivariate Logistic regression analysis revealed that visceral obesity [odds ratio (OR)=2.732, P=0.019] and NRS 2002 scores of≥3 (OR=2.574, P=0.042) were independent risk factors for postoperative complications. Conclusions:Visceral obesity was an independent risk factor for complications after surgery for mid-low rectal cancers.

Study on the Migration of Residual Styrene to Moxifloxacin Hydrochloride and Sodium Chloride Injection from Five-layer Co-extrusion Bags / 中国药师

Objective:To establish a monitoring method for the detection of styrene monomer migration to moxifloxacin hydrochlo-ride and sodium chloride injection. Methods:A GC-MS method with a DB-5MS (30 m×0.25 mm,0.25 μm) column was used to determine styrene by the manner of large volume headspace purge-trap. Results:The linearity of styrene was good within the range of 0.009 1-0.364 μg·ml-1(r=0.999 9),and the average recovery of styrene in moxifloxacin hydrochloride and sodium chloride injec-tion was 99.4%(RSD=1.1%,n=9). After the detection of several batches of samples,the amount of styrene migration was all be-low the detection limit.Conclusion:The method can be used to measure the migration of styrene remained in five-layer co-extrusion in-fusion bags to moxifloxacin hydrochloride and sodium chloride injection.

Application of Microwave Technology in the Preparation of Plastic Beads / 中国药房

OBJECTIVE:To study the advantage of microwave technology in the preparation of plastic beads. METHODS:Three kinds of representative processing methods were applied to prepare Equus asinus,Colla Cornus Cervi and Colla Carapax et Plastri Testudinis. The processing methods and the quality of plastic beads were compared. RESULTS:Study results showed mic-rowave technology was superior to other processing methods in the preparation of plastic beads and quality. CONCLUSION:The microwave technology has its specific advantages in the preparation of plastic beads.

Orthogonal optimization of plastic beads microwave processing of Colla Corii Asini,Colla Carapacis et Plastri Testudinis,and Colla Cornus Cervi / 中草药

Objective Colla Corii Asini,Colla Carapacis et Plastri Testudinis,Colla Cornus Cervi,three types of plastic bead microwave process parameters were optimized in order to standardize the production of the types of plastic beads and to enhance the stability of the quality of plastic beads.Methods Using a number of quality indicators to evaluate the quality of plastic beads and orthogonal test to optimize the plastic bead process parameters.Results The microwave process parameters of Colla Corii Asini,Colla Carapacis et Plastri Testudinis,Colla Cornus Cervi,three types of plastic beads were obtained through orthogonal optimization.Conclusion Different types of plastic type drugs,different production lot, and different manufacturers of similar plastic herbs have their own optimum process parameters.L4(23) orthogonal optimization can be used to screen out the plastic type drugs rapidly.

Interference test in bacterial endotoxins detection of Shenmai Freeze-dried Powder Injection / 中草药

Object To prove the feasibility of bacterial endotoxin test (BET) in Shenmai Freeze-dried Powder Injection by interference test. Methods Interference primary screening test and interference test were conducted in BET of Shenmai Freeze-dried Powder Injection to detect whether the interference existed or not and to explore the method of removing the interference. Results There was inhibition on BET with 0.5 EU/mL tachypleus amebocyte lysate (TAL). Interference could be excluded by using 0.25 EU/mL TAL and diluting the samples. Conclusion It is feasible to use 0.25 EU/mL TAL or TAL with more sensitivity in BET of Shenmai Freeze-dried Powder Injection.