The cell edit: Looking at and beyond non-structural proteins to understand membrane rearrangement in coronaviruses.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a positive-stranded RNA virus that sits at the centre of the recent global pandemic. As a member of the coronaviridae family of viruses, it shares features such as a very large genome (>30 kb) that is replicated in a purpose-built replication organelle. Biogenesis of the replication organelle requires significant and concerted rearrangement of the endoplasmic reticulum membrane, a job that is carried out by a group of integral membrane non-structural proteins (NSP3, 4 and 6) expressed by the virus along with a host of viral replication enzymes and other factors that support transcription and replication. The primary sites for RNA replication within the replication organelle are double membrane vesicles (DMVs). The small size of DMVs requires generation of high membrane curvature, as well as stabilization of a double-membrane arrangement, but the mechanisms that underlie DMV formation remain elusive. In this review, we discuss recent breakthroughs in our understanding of the molecular basis for membrane rearrangements by coronaviruses. We incorporate established models of NSP3-4 protein-protein interactions to drive double membrane formation, and recent data highlighting the roles of lipid composition and host factor proteins (e.g. reticulons) that influence membrane curvature, to propose a revised model for DMV formation in SARS-CoV-2.

A vaccine programme comprising GA08 (GI-27) and Mass (GI-1) strains prevents DMV1639 (GI-17) infectious bronchitis virus transmission among broiler chickens.

Effective control of infectious bronchitis is a challenge in commercial poultry operations due to the high transmissibility of the virus. Although multiple IBV lineages are circulating in the United States, the DMV1639-type IBV strain (GI-17) is currently the major circulating variant, creating production losses in the poultry industry. This study aimed to test whether the combination of a GA08 (GI-27) and a Mass-type (GI-1) IB vaccines could significantly reduce the transmission of a DMV1639-type (GI-17) field IBV strain in 4-week-old commercial broilers. Half of the birds were directly challenged, whereas the other half of the groupmates were put in contact 24 hours later. Two replicates of the same study setup, including 10 directly challenged and 10 contact birds per group, were run. Transmission of the challenge virus was significantly reduced in vaccinates (R = 0.0), whereas all unvaccinated birds became infected (R = 9.6). Reduced transmission of the DMV1639 IB challenge virus by the combined vaccination programme in broiler chickens was also accompanied by clinical protection. These data are important because prevention of IBV transmission by vaccination will result in overall reduced viral replication and consequently in reduced likelihood of genetic changes that can lead to new variants. This is the first published evidence of the successful transmission control of a DMV1639 IBV strain in chickens.

Submandibular Gland Pathogenesis Following SARS-CoV-2 Infection and Implications for Xerostomia.

Although SARS-CoV-2 induces mucin hypersecretion in the respiratory tract, hyposalivation/xerostomia has been reported by COVID-19 patients. We evaluate the submandibular gland (SMGs) pathogenesis in SARS-CoV-2-infected K18-hACE2 mice, focusing on the impact of infection on the mucin production and structural integrity of acini, ductal system, myoepithelial cells (MECs) and telocytes. The spike protein, the nucleocapsid protein, hACE2, actin, EGF, TNF-α and IL-1ß were detected by immunofluorescence, and the Egfr and Muc5b expression was evaluated. In the infected animals, significant acinar hypertrophy was observed in contrast to ductal atrophy. Nucleocapsid proteins and/or viral particles were detected in the SMG cells, mainly in the nuclear membrane-derived vesicles, confirming the nuclear role in the viral formation. The acinar cells showed intense TNF-α and IL-1ß immunoexpression, and the EGF-EGFR signaling increased, together with Muc5b upregulation. This finding explains mucin hypersecretion and acinar hypertrophy, which compress the ducts. Dying MECs and actin reduction were also observed, indicating failure of contraction and acinar support, favoring acinar hypertrophy. Viral assembly was found in the dying telocytes, pointing to these intercommunicating cells as viral transmitters in SMGs. Therefore, EGF-EGFR-induced mucin hypersecretion was triggered by SARS-CoV-2 in acinar cells, likely mediated by cytokines. The damage to telocytes and MECs may have favored the acinar hypertrophy, leading to ductal obstruction, explaining xerostomia in COVID-19 patients. Thus, acinar cells, telocytes and MECs may be viral targets, which favor replication and cell-to-cell viral transmission in the SMG, corroborating the high viral load in saliva of infected individuals.

Interactions between Brainstem Neurons That Regulate the Motility to the Stomach.

Activity in the dorsal vagal complex (DVC) is essential to gastric motility regulation. We and others have previously shown that this activity is greatly influenced by local GABAergic signaling, primarily because of somatostatin (SST)-expressing GABAergic neurons. To further understand the network dynamics associated with gastric motility control in the DVC, we focused on another neuron prominently distributed in this complex, neuropeptide-Y (NPY) neurons. However, the effect of these neurons on gastric motility remains unknown. Here, we investigate the anatomic and functional characteristics of the NPY neurons in the nucleus tractus solitarius (NTS) and their interactions with SST neurons using transgenic mice of both sexes. We sought to determine whether NPY neurons influence the activity of gastric-projecting neurons, synaptically interact with SST neurons, and affect end-organ function. Our results using combined neuroanatomy and optogenetic in vitro and in vivo show that NPY neurons are part of the gastric vagal circuit as they are trans-synaptically labeled by a viral tracer from the gastric antrum, are primarily excitatory as optogenetic activation of these neurons evoke EPSCs in gastric-antrum-projecting neurons, are functionally coupled to each other and reciprocally connected to SST neurons, whose stimulation has a potent inhibitory effect on the action potential firing of the NPY neurons, and affect gastric tone and motility as reflected by their robust optogenetic response in vivo. These findings indicate that interacting NPY and SST neurons are integral to the network that controls vagal transmission to the stomach.SIGNIFICANCE STATEMENT The brainstem neurons in the dorsal nuclear complex are essential for regulating vagus nerve activity that affects the stomach via tone and motility. Two distinct nonoverlapping populations of predominantly excitatory NPY neurons and predominantly inhibitory SST neurons form reciprocal connections with each other in the NTS and with premotor neurons in the dorsal motor nucleus of the vagus to control gastric mechanics. Light activation and inhibition of NTS NPY neurons increased and decreased gastric motility, respectively, whereas both activation and inhibition of NTS SST neurons enhanced gastric motility.

IL-1ß/IL-1R1 signaling is involved in the propagation of α-synuclein pathology of the gastrointestinal tract to the brain.

The pathological hallmark of Parkinson's disease (PD) is the intraneuronal accumulation of misfolded alpha-synuclein (termed Lewy bodies) in dopaminergic neurons of substantia nigra par compacta (SNc). It is assumed that the α-syn pathology is induced by gastrointestinal inflammation and then transfers to the brain by the gut-brain axis. Therefore, the relationship between gastrointestinal inflammation and α-syn pathology leading to PD remains to be investigated. In our study, rotenone (ROT) oral administration induces gastrointestinal tract (GIT) inflammation in mice. In addition, we used pseudorabies virus (PRV) for tracing studies and performed behavioral testing. We observed that ROT treatments enhance macrophage activation, inflammatory mediator expression, and α-syn pathology in the GIT 6-week post-treatment (P6). Moreover, pathological α-syn was localized with IL-1R1 positive neural cells in GIT. In line with these findings, we also find pS129-α-syn signals in the dorsal motor nucleus of the vagus (DMV) and tyrosine hydroxylase in the nigral-striatum dynamically change from 3-week post-treatment (P3) to P6. Following that, pS129-α-syn was dominant in the enteric neural cell, DMV, and SNc, accompanied by microglial activation, and these phenotypes were absent in IL-1R1r/r mice. These data suggest that IL-1ß/IL-1R1-dependent inflammation of GIT can induce α-syn pathology, which then propagates to the DMV and SNc, resulting in PD.

The Inhaled Steroid Ciclesonide Blocks SARS-CoV-2 RNA Replication by Targeting the Viral Replication-Transcription Complex in Cultured Cells.

Here, we screened steroid compounds to obtain a drug expected to block host inflammatory responses and Middle East respiratory syndrome coronavirus (MERS-CoV) replication. Ciclesonide, an inhaled corticosteroid, suppressed the replication of MERS-CoV and other coronaviruses, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the cause of coronavirus disease 2019 (COVID-19), in cultured cells. The 90% effective concentration (EC90) of ciclesonide for SARS-CoV-2 in differentiated human bronchial tracheal epithelial cells was 0.55 µM. Eight consecutive passages of 43 SARS-CoV-2 isolates in the presence of ciclesonide generated 15 resistant mutants harboring single amino acid substitutions in nonstructural protein 3 (nsp3) or nsp4. Of note, ciclesonide suppressed the replication of all these mutants by 90% or more, suggesting that these mutants cannot completely overcome ciclesonide blockade. Under a microscope, the viral RNA replication-transcription complex in cells, which is thought to be detectable using antibodies specific for nsp3 and double-stranded RNA, was observed to fall in the presence of ciclesonide in a concentration-dependent manner. These observations indicate that the suppressive effect of ciclesonide on viral replication is specific to coronaviruses, highlighting it as a candidate drug for the treatment of COVID-19 patients.IMPORTANCE The outbreak of SARS-CoV-2, the cause of COVID-19, is ongoing. New and effective antiviral agents that combat the disease are needed urgently. Here, we found that an inhaled corticosteroid, ciclesonide, suppresses the replication of coronaviruses, including betacoronaviruses (murine hepatitis virus type 2 [MHV-2], MERS-CoV, SARS-CoV, and SARS-CoV-2) and an alphacoronavirus (human coronavirus 229E [HCoV-229E]), in cultured cells. Ciclesonide is safe; indeed, it can be administered to infants at high concentrations. Thus, ciclesonide is expected to be a broad-spectrum antiviral drug that is effective against many members of the coronavirus family. It could be prescribed for the treatment of MERS and COVID-19.

Hepatitis C Virus Replication Depends on Endosomal Cholesterol Homeostasis.

Similar to other positive-strand RNA viruses, hepatitis C virus (HCV) causes massive rearrangements of intracellular membranes, resulting in a membranous web (MW) composed of predominantly double-membrane vesicles (DMVs), the presumed sites of RNA replication. DMVs are enriched for cholesterol, but mechanistic details on the source and recruitment of cholesterol to the viral replication organelle are only partially known. Here we focused on selected lipid transfer proteins implicated in direct lipid transfer at various endoplasmic reticulum (ER)-membrane contact sites. RNA interference (RNAi)-mediated knockdown identified several hitherto unknown HCV dependency factors, such as steroidogenic acute regulatory protein-related lipid transfer domain protein 3 (STARD3), oxysterol-binding protein-related protein 1A and -B (OSBPL1A and -B), and Niemann-Pick-type C1 (NPC1), all residing at late endosome and lysosome membranes and required for efficient HCV RNA replication but not for replication of the closely related dengue virus. Focusing on NPC1, we found that knockdown or pharmacological inhibition caused cholesterol entrapment in lysosomal vesicles concomitant with decreased cholesterol abundance at sites containing the viral replicase factor NS5A. In untreated HCV-infected cells, unesterified cholesterol accumulated at the perinuclear region, partially colocalizing with NS5A at DMVs, arguing for NPC1-mediated endosomal cholesterol transport to the viral replication organelle. Consistent with cholesterol being an important structural component of DMVs, reducing NPC1-dependent endosomal cholesterol transport impaired MW integrity. This suggests that HCV usurps lipid transfer proteins, such as NPC1, at ER-late endosome/lysosome membrane contact sites to recruit cholesterol to the viral replication organelle, where it contributes to MW functionality.IMPORTANCE A key feature of the replication of positive-strand RNA viruses is the rearrangement of the host cell endomembrane system to produce a membranous replication organelle (RO). The underlying mechanisms are far from being elucidated fully. In this report, we provide evidence that HCV RNA replication depends on functional lipid transport along the endosomal-lysosomal pathway that is mediated by several lipid transfer proteins, such as the Niemann-Pick type C1 (NPC1) protein. Pharmacological inhibition of NPC1 function reduced viral replication, impaired the transport of cholesterol to the viral replication organelle, and altered organelle morphology. Besides NPC1, our study reports the importance of additional endosomal and lysosomal lipid transfer proteins required for viral replication, thus contributing to our understanding of how HCV manipulates their function in order to generate a membranous replication organelle. These results might have implications for the biogenesis of replication organelles of other positive-strand RNA viruses.

Survey of physician attitudes towards psychogenic nonepileptic seizures and driving.

BACKGROUND: Physicians from various disciplines encounter patients presenting with psychogenic nonepileptic seizures (PNES) as part of their routine clinical practice. Recommendations towards assessing fitness to drive and reporting are clearer for conditions such as neurocognitive disorders and epilepsy, but such guidelines do not exist for patients with PNES. Here, we assess physicians' attitudes towards driving for patients diagnosed with PNES. METHODS: Electronic questionnaires were sent to Neurology and Family Medicine physicians practicing at Creighton University Medical Center and Psychiatry physicians practicing at Creighton-Nebraska Psychiatry Residency Program to assess their opinion regarding driving risk when encountering PNES. RESULTS: The survey request was sent to 125 physicians, of which close to 60% completed the survey. Eighty-eight percent of participants encountered PNES in their clinical practice, and 69.1% agreed it was a difficult problem to assess, with only 8.3% endorsing a belief that these patients should drive without restrictions. Ninety-three percent felt having guidelines would help them assess the driving risk in this population. CONCLUSION: Psychogenic nonepileptic seizures are common across neurology, psychiatry, and primary care, and most physicians find assessing driving risk in such individuals highly warranted yet difficult. Developing such assessment guidelines and recommendations is of great need for clinicians.

A Department of Motor Vehicle-Based Intervention to Promote Organ Donor Registrations in New York State.

CONTEXT: Examination of efficacy of motor vehicle representative educational training and dissemination of promotional materials as a means to promote organ donation enrollments in New York State. OBJECTIVE: To increase the number of New York State residents who consent to donation through the department of motor vehicle transactions during project period. SETTING: County-run motor vehicle offices across New York State. PARTICIPANTS: Customers who present to New York Department of Motor Vehicle offices and the representative who work at designated bureaus. INTERVENTIONS: point-of-decision materials including promotional posters, brochures, website, and the motor vehicle representative training sessions. MAIN OUTCOME MEASURES: Reasons for enrollment decision, knowledge/experience with donation, monthly consent rates, enrollment in state organ, and tissue registry. RESULTS: Customers who elected not to register reported no reason or uncertainty surrounding enrollment. The representatives reported experience with donation, discussion with customers, and need for additional education on organ donation. Enrollment cards were mailed to 799 project staff; counties where offices participated in intervention did not indicate significantly higher monthly enrollments when comparing pre- to postenrollment rates. CONCLUSIONS: Use of point-of-decision materials and enrollment cards proved inexpensive method to register customers with a 3.6% return rate. Customers report low (27%) enrollment rate and reticence to consent to donation. Educational training sessions with representatives did not yield significant enrollment increases when evaluating data at county-level enrollment.

Mechanism, structural and functional insights into nidovirus-induced double-membrane vesicles.

During infection, positive-stranded RNA causes a rearrangement of the host cell membrane, resulting in specialized membrane structure formation aiding viral genome replication. Double-membrane vesicles (DMVs), typical structures produced by virus-induced membrane rearrangements, are platforms for viral replication. Nidoviruses, one of the most complex positive-strand RNA viruses, have the ability to infect not only mammals and a few birds but also invertebrates. Nidoviruses possess a distinctive replication mechanism, wherein their nonstructural proteins (nsps) play a crucial role in DMV biogenesis. With the participation of host factors related to autophagy and lipid synthesis pathways, several viral nsps hijack the membrane rearrangement process of host endoplasmic reticulum (ER), Golgi apparatus, and other organelles to induce DMV formation. An understanding of the mechanisms of DMV formation and its structure and function in the infectious cycle of nidovirus may be essential for the development of new and effective antiviral strategies in the future.

Protection of laying chickens against the Canadian DMV/1639 infectious bronchitis virus infection through priming with heterologous live vaccine and boosting with heterologous or homologous inactivated vaccine.

The emergence of the Canadian Delmarva (DMV)/1639 infectious bronchitis virus (IBV) type strains was associated with egg production disorders in Eastern Canadian layer operations. While developing vaccines for novel IBV variants is not typically a reasonable approach, the consideration of an autogenous vaccine becomes more appealing, particularly when the new variant presents significant economic challenges. The current study aimed to compare the efficacies of two vaccination programs that included heterologous live priming by Massachusetts (Mass) and Connecticut (Conn) type vaccines followed by either a commercial inactivated Mass type vaccine or a locally prepared autogenous inactivated DMV/1639 type vaccine against DMV/1639 IBV challenge. The protection parameters evaluated were egg production, viral shedding, dissemination of the virus in tissues, gross and microscopic lesions, and immunological responses. The challenge with the DMV/1639 caused severe consequences in the non-vaccinated laying hens including significant drop in egg production, production of low-quality eggs, serious damage to the reproductive organs, and yolk peritonitis. The two vaccination programs protected the layers from the poor egg-laying performance and the pathology. The vaccination program incorporating the autogenous inactivated DMV/1639 type vaccine was more effective in reducing vial loads in renal and reproductive tissues. This was associated with a higher virus neutralization titer compared to the group that received the commercial inactivated Mass type vaccine. Additionally, the autogenous vaccine boost led to a significant reduction in the viral shedding compared to the non-vaccinated laying hens. However, both vaccination programs induced significant level of protection considering all parameters examined. Overall, the findings from this study underscore the significance of IBV vaccination for protecting laying hens.

Association of ADAMTS13 activity with cerebral deep medullary vein: A community-based cross-sectional study.

Objectives: This study aims to investigate whether circulating ADAMTS13 activity can offer insights into the mechanism of pathophysiological changes in deep medullary veins (DMVs). Methods: This study was conducted on a community cohort of elderly individuals in Shanghai. Plasma von Willebrand factor (VWF) levels and ADAMTS13 activity were measured. A validated DMV score described the overall burden of DMV on the brain. Through ordinal regression models, we investigated the correlation between VWF levels, ADAMTS13 activity, and increasing severity of DMV score while adjusting for demographics and cardiovascular risk factors. Results: The study enrolled 262 subjects according to the inclusion criteria. The mean VWF level (1.35 ± 0.25) was higher in the DMV group than in the group without DMV (1.25 ± 0.30) (p = 0.025), and ADAMTS13 activity (83.76 ± 7.96) was relatively lower. After adjusting for age, sex, alcohol consumption, smoking, hypertension, and diabetes, reduced ADAMTS13 activity [ß = -7.78; 95 % CI (-10.21, -5.35) p < 0.01] was associated with DMV. Moreover, correlation analysis indicated that ADAMTS13 activity was negatively correlated with the DMV score (Kendall's tau-b = -0.53, p < 0.001). Discussion: In summary, there was an inverse correlation observed between ADAMTS13 activity and the DMV score, which may provide some clinical clues for exploring the potential pathogenesis of DMV.

Heterologous maternal antibodies derived from infectious bronchitis vaccines prevent the development of lesions associated with false layer syndrome.

Infectious bronchitis virus (IBV) strains of the Delmarva (DMV)/1639 genotype have been causing false layer syndrome (FLS) in the Eastern Canadian layer operations since the end of 2015. FLS is characterized by the development of cystic oviducts in layer pullets infected at an early age. Currently, there are no homologous vaccines for the control of this IBV genotype. Our previous research showed that a heterologous vaccination regimen incorporating Massachusetts (Mass) and Connecticut (Conn) IBV types protects layers against DMV/1639 genotype IBV. The aim of this study was to investigate the role of maternal antibodies conferred by breeders received the same vaccination regimen in the protection against the development of DMV/1639-induced FLS in pullets. Maternal antibody-positive (MA+) and maternal antibody-negative (MA-) female progeny chicks were challenged at 1 day of age and kept under observation for 16 weeks. Oviductal cystic formations were observed in 3 of 14 birds (21.4 %) in the MA- pullets, while the lesions were notably absent in the MA+ pullets. Milder histopathological lesions were observed in the examined tissues of the MA+ pullets. However, the maternal derived immunity failed to demonstrate protection against the damage to the tracheal ciliary activity, viral shedding, and viral tissue distribution. Overall, this study underscores the limitations of maternal derived immunity in preventing certain aspects of viral pathogenesis, emphasizing the need for comprehensive strategies to address different aspects of IBV infection.

Deep vein diameters and perivascular space scores are associated with deep medullary vein hypo-visibility in patients with white matter hyperintensity.

Background: Deep medullary vein (DMV) hypo-visibility is correlated with white matter hyperintensity (WMH), but the underlying causes remain unclear. This study aimed to explore the relationship between deep vein diameters and perivascular space (PVS) scores, and DMV hypo-visibility in the presence of WMH. Methods: This cross-sectional study prospectively analyzed the clinical and imaging data of 190 cerebral small vessel disease patients with WMH and 40 healthy controls from the Lishui Hospital of Traditional Chinese Medicine affiliated with Zhejiang Chinese Medical University. PVS scores ranging from 0 to 4 were determined according to the PVS counts in the basal ganglia area on T2-weighted magnetic resonance images; high-grade PVS was defined as a PVS score >1. The diameters of the deep cerebral veins, including the bilateral septal veins (SVs), thalamostriate veins (TSVs), lateral ventricular veins (LVVs), and internal cerebral veins, were measured using susceptibility weighted imaging (SWI). Left and right DMV scores, ranging from 0 to 9, were calculated based on the visibility of the DMV on SWI in the ipsilateral frontal, parietal, and occipital lobes. Results: The deep cerebral vein diameters, left and right DMV scores, and high-grade PVS differed between the healthy controls and WMH patients (P<0.05). Left DMV scores were independently associated with age {ß [95% confidence interval (CI)]: 0.050 (0.018, 0.082)}, high-grade PVS [ß (95% CI): 0.998 (0.262, 1.737)], and the diameters of the ipsilateral SVs [ß (95% CI): -1.114 (-1.754, -0.475)], SVs [ß (95% CI): -0.734 (-1.191, -0.277)], and LVVs [ß (95% CI): -0.921 (-1.567, -0.275)] [all false discovery rate (FDR)-corrected P<0.05]. Right DMV scores were independently associated with age [ß (95% CI): 0.071 (0.037, 0.105)], high-grade PVS [ß (95% CI): 0.873 (0.111, 1.635)], and the diameters of the ipsilateral SVs [ß (95% CI): -0.837 (-1.386, -0.289)], TSVs [ß (95% CI): -0.875 (-1.331, -0.419)], and LVVs [ß (95% CI): -1.813 (-2.484, -1.142)] (all FDR-corrected P<0.05). Conclusions: Decreased hypo-visibility of DMVs on SWI was associated with a higher age, the presence of high-grade PVS, and smaller diameters of the ipsilateral deep cerebral veins in individuals with WMH. Our findings provide novel insights into the probable mechanisms leading to high DMV scores.

Activation of ChAT+ neuron in dorsal motor vagus (DMV) increases blood glucose through the regulation of hepatic gene expression in mice.

The brain and peripheral organs communicate through hormones and neural connections. Proper communication is required to maintain normal whole-body energy homeostasis. In addition to endocrine system, from the perspective of neural connections for metabolic homeostasis, the role of the sympathetic nervous system has been extensively studied, but understanding of the parasympathetic nervous system is limited. The liver plays a central role in glucose and lipid metabolism. This study aimed to clarify the innervation of parasympathetic nervous system in the liver and its functional roles in metabolic homeostasis. The liver-specific parasympathetic nervous system innervation (PNS) was shown by tissue clearing, immunofluorescence and transgenic mice at the three-dimensional histological level. The parasympathetic efferent signals were manipulated using a chemogenetic technique and the activation of ChAT+ parasympathetic neurons in dorsal motor vagus (DMV) results in the increased blood glucose through the elevated hepatic gluconeogenic and lipogenic gene expression in the liver. Thus, our study showed the evidence of ChAT+ parasympathetic neurons in the liver and its role for hepatic parasympathetic nervous signaling in glucose homeostasis through the regulation of hepatic gene expression.

DMV biogenesis during ß-coronavirus infection requires autophagy proteins VMP1 and TMEM41B.

Upon entering host cells, ß-coronaviruses specifically induce generation of replication organelles (ROs) from the endoplasmic reticulum (ER) through their nonstructural protein 3 (nsp3) and nsp4 for viral genome transcription and replication. The most predominant ROs are double-membrane vesicles (DMVs). The ER-resident proteins VMP1 and TMEM41B, which form a complex to regulate autophagosome and lipid droplet (LD) formation, were recently shown to be essential for ß-coronavirus infection. Here we report that VMP1 and TMEM41B contribute to DMV generation but function at different steps. TMEM41B facilitates nsp3-nsp4 interaction and ER zippering, while VMP1 is required for subsequent closing of the paired ER into DMVs. Additionally, inhibition of phosphatidylserine (PS) formation by siPTDSS1 partially reverses the DMV and LD defects in VMP1 KO cells, suggesting that appropriate PS levels also contribute to DMV formation. This work provides clues to the mechanism of how host proteins collaborate with viral proteins for endomembrane reshaping to promote viral infection.

Comparative pathogenicity of infectious bronchitis virus Massachusetts and Delmarva (DMV/1639) genotypes in laying hens.

Infectious bronchitis (IB) is a highly contagious and acute viral disease of chicken caused by the infectious bronchitis virus (IBV) of the family Coronaviridae. Even with extensive vaccination against IB by the poultry industry, the occurrence of new IBV genotypes is a continuous challenge encountered by the global poultry industry. This experiment was designed to compare the pathogenicity of two IBV strains belonging to Massachusetts (Mass) and Delmarva DMV/1639 genotypes. Specific pathogen-free laying hens were challenged during the peak of production (30 weeks), keeping a mock-infected control group. During 21 days of observation following infection, a significant drop in egg production with miss-shaped and soft shells was observed in the DMV/1639 IBV-infected hens only. The DMV/1639 IBV infected group showed prolonged and higher cloacal viral shedding compared with the Mass IBV-infected group. At the end of the study (21 days post-infection), the viral genome loads in the respiratory, urogenital, and immune tissues were significantly higher in the DMV/1639 IBV-infected group compared with the Mass IBV-infected group. Macroscopic lesions such as distorted ova leading to egg peritonitis were observed only in the DMV/1639 IBV-infected group. Moreover, microscopic lesion scores were significantly higher in the lung, kidney, cecal tonsils, and oviduct of the DMV/1639 IBV-infected group compared with the Mass IBV-infected group. Finally, the apoptosis index in the kidney, ovary, magnum, isthmus, and shell gland was significantly higher in the DMV/1639 IBV-infected group compared with the control and Mass-infected groups. This study examined the pathogenicity of two IBV genotypes that are impacting the layer industry in North America.

Surgical stabilization of multiple rib fractures in an Asian population: a systematic review and meta-analysis.

Background: There is no consensus on the effectiveness of surgical stabilization in multiple rib fractures in Asia, especially among patients with a non-flail rib fracture pattern. We aim to synthesize the evidence on the effectiveness of surgical stabilization of rib fractures (SSRF) in an Asian population with multiple non-flail rib fractures. Methods: The MEDLINE, Embase, Cochrane Central Register of Controlled Trials (CENTRAL), and Cochrane Database of Systematic Reviews were searched in this systematic literature review and meta-analysis to identify studies conducted in Asia that included patients with multiple non-flail rib fractures in at least one of their treatment groups. The intervention of interest was SSRF, and the comparator was a nonoperative treatment. The duration of mechanical ventilation (DMV) was the primary outcome. Posttreatment pain score, pneumonia, atelectasis, intensive care unit length of stay (ICU LOS), hospital length of stay (HLOS), need for tracheostomy, respiratory function, functional outcomes, quality of life (QoL), and mortality were identified as the secondary outcomes. A random effects model (REM) was used to pool data for outcomes reported in two or more studies. Results: A total of 12 studies (n=2,440 patients) were included. There was a significantly shorter DMV {mean difference (MD): -5.23 [95% confidence interval (CI): -9.64 to -0.81], P=0.02}, lower 4-week post-treatment pain score [standard mean difference (SMD): -2.24 (95% CI: -3.18 to -1.31), P<0.00001], lower risk for pneumonia [risk ratio (RR): 0.46 (95% CI: 0.23 to 0.95), P=0.04], lower risk for atelectasis [RR: 0.44, (95% CI: 0.29 to 0.65), P<0.0001], shorter ICU LOS [MD: -4.00 (95% CI: -6.33 to -1.66), P=0.0008], and shorter HLOS [MD: -6.54 (95% CI: -9.28 to -3.79), P<0.00001] in favor of SSRF. Effect estimates for the need for tracheostomy [RR: 0.67 (95% CI: 0.42 to 1.08), P=0.10] and mortality [RR: 0.94 (95% CI: 0.37 to 2.41), P=0.90] were nonsignificant. Conclusions: In the Asian population with mainly non-flail rib fracture patterns, SSRF was associated with shorter DMV, ICU LOS, and HLOS as well as lower risks for atelectasis and pneumonia, and pain scores after 4 weeks. The risk of mortality was comparable between treatment groups.

CNS sites controlling the gastric pyloric sphincter: Neuroanatomical and functional study in the rat.

The pyloric sphincter receives parasympathetic vagal innervation from the dorsal motor nucleus of the vagus (DMV). However, little is known about its higher-order neurons and the nuclei that engage the DMV neurons controlling the pylorus. The purpose of the present study was twofold. First, to identify neuroanatomical connections between higher-order neurons and the DMV. This was carried out by using the transneuronal pseudorabies virus PRV-152 injected into rat pylorus torus and examining the brains of these animals for PRV labeling. Second, to identify the specific sites within the DMV that functionally control the motility and tone of the pyloric sphincter. For these studies, experiments were performed to assess the effect of DMV stimulation on pylorus activity in urethane-anesthetized male rats. A strain gauge force transducer was sutured onto the pyloric tonus to monitor tone and motility. L-glutamate (500 pmol/30 nL) was microinjected unilaterally into the rostral and caudal areas of the DMV. Data from the first study indicated that neurons labeled with PRV occurred in the DMV, hindbrain raphe nuclei, midbrain Edinger-Westphal nucleus, ventral tegmental area, lateral habenula, and arcuate nucleus. Data from the second study indicated that microinjected L-glutamate into the rostral DMV results in contraction of the pylorus blocked by intravenously administered atropine and ipsilateral vagotomy. L-glutamate injected into the caudal DMV relaxed the pylorus. This response was abolished by ipsilateral vagotomy but not by intravenously administered atropine or L-NG-nitroarginine methyl ester (L-NAME). These findings identify the anatomical and functional brain neurocircuitry involved in controlling the pyloric sphincter. Our results also show that site-specific stimulation of the DMV can differentially influence the activity of the pyloric sphincter by separate vagal nerve pathways.

Endomembrane remodeling in SARS-CoV-2 infection.

During severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, the viral proteins intimately interact with host factors to remodel the endomembrane system at various steps of the viral lifecycle. The entry of SARS-CoV-2 can be mediated by endocytosis-mediated internalization. Virus-containing endosomes then fuse with lysosomes, in which the viral S protein is cleaved to trigger membrane fusion. Double-membrane vesicles generated from the ER serve as platforms for viral replication and transcription. Virions are assembled at the ER-Golgi intermediate compartment and released through the secretory pathway and/or lysosome-mediated exocytosis. In this review, we will focus on how SARS-CoV-2 viral proteins collaborate with host factors to remodel the endomembrane system for viral entry, replication, assembly and egress. We will also describe how viral proteins hijack the host cell surveillance system-the autophagic degradation pathway-to evade destruction and benefit virus production. Finally, potential antiviral therapies targeting the host cell endomembrane system will be discussed.