Purification and electron cryomicroscopy of coronavirus particles.

Intact, enveloped coronavirus particles vary widely in size and contour, and are thus refractory to study by traditional structural means such as X-ray crystallography. Electron microscopy (EM) overcomes some problems associated with particle variability and has been an important tool for investigating coronavirus ultrastructure. However, EM sample preparation requires that the specimen be dried onto a carbon support film before imaging, collapsing internal particle structure in the case of coronaviruses. Moreover, conventional EM achieves image contrast by immersing the specimen briefly in heavy-metal-containing stain, which reveals some features while obscuring others. Electron cryomicroscopy (cryo-EM) instead employs a porous support film, to which the specimen is adsorbed and flash-frozen. Specimens preserved in vitreous ice over holes in the support film can then be imaged without additional staining. Cryo-EM, coupled with single-particle image analysis techniques, makes it possible to examine the size, structure and arrangement of coronavirus structural components in fully hydrated, native virions. Two virus purification procedures are described

Cervical Deformity Patients Have Baseline Swallowing Dysfunction but Surgery Does Not Increase Dysphagia at 3 Months: Results From a Prospective Cohort Study.

Study designProspective cohort study.ObjectivesMost studies of dysphagia in the cervical spine have focused on a degenerative patient population; the rate of dysphagia following surgery for cervical deformity (CD) is unknown. This study aims to investigate if surgery for cervical deformity results in postoperative dysphagia.MethodsPatients with CD undergoing surgery from 2013 to 2015 were prospectively enrolled to evaluate dysphagia. Demographic, operative, and radiographic variables were analyzed. The Quality of Life in Swallowing Disorders (SWAL-QoL) was used to measure dysphagia. Paired t test, independent t tests, and bivariate Pearson correlations were performed.ResultsA total of 88 CD patients, aged 61.52 ± 10.52 years, were enrolled. All patients (100%) had 3-month SWAL-QoL for analysis. The baseline preoperative SWAL-QoL was 78.35. This is roughly the same level of dysphagia as an anterior cervical discectomy patient that is 3 weeks removed from surgery. Increasing body mass index (BMI) was correlated with decreased SWAL-QoL score (r = -0.30, P = .001). Age, gender, smoking, and Charlson Comorbidity Index (CCI) showed no significant correlations with preoperative SWAL-QoL. Patients with prior cervical surgery had a lower preoperative SWAL-QoL (P = .04). While 11 patients had acute postoperative dysphagia, CD surgery did not result in lower SWAL-QoL at 3 months (77.26 vs 78.35, P = .53). Surgical variables, including estimated blood loss (EBL), anterior or posterior fusion levels, steroid use, preoperative traction, staged surgery, surgical approach, anterior corpectomy, posterior osteotomy, and UIV (upper instrumented vertebrae) location, showed no impact on postoperative SWAL-QoL. Correction of cervical kyphosis was not correlated to 3-month SWAL-QoL scores or the change in SWAL-QoL scores.ConclusionsWhile patients undergoing surgery for cervical deformity had swallowing dysfunction at baseline, we did not observe a significant decline in SWAL-QoL scores at 3 months. Patients with prior cervical surgery and higher BMI had a lower baseline SWAL-QoL. There were no surgical or radiographic variables correlated to a change in SWAL-QOL score

The posterior use of BMP-2 in cervical deformity surgery does not result in increased early complications: A prospective multicenter study

Study designProspective cohort study.ObjectivesTo describe the rate of short-term complications following the posterior use of recombinant human bone morphogenetic protein-2 (rhBMP-2) in cervical deformity (CD) surgery.MethodsCD patients from 2013 to 2015 were enrolled in a prospective, multicenter database. Patients were divided into those receiving rhBMP-2 (BMP) and no rhBMP-2 (NOBMP). The relationship between BMP use, demographic variables surgical variables, radiographic parameters and complications was evaluated.ResultsA total of 100 patients (47 BMP, 53 NOBMP) were included. Follow-up time averaged 7.6 months (range 3-12 months). An average of 13.6mg of BMP was used per person with 1.49 mg per level. Compared with the NOBMP group, patients in the BMP group were older (P = .03). BMP was more commonly used in patients that and had longer prior fusions (6.0 vs 2.5, P < .01). There were no differences between groups with regards to a history of surgery, Charlson Comorbidity Index, estimated blood loss, operation time, fusion levels, and surgical approach. The maintenance of radiographic parameters at 6-month follow-up was similar. There were no differences in terms of total complication incidence, total complications per person, major complications per person or any specific complication. Linear regression and Pearson correlation analysis did not reveal any strong r2 values (r2 = 0.09, 0.08, 0.06) between the use of BMP and complications (major or operative).ConclusionsBMP use was not directly associated with an increased incidence of early complications in this prospective cohort of operative adult CD patients. Its use was associated with increased number of levels instrumented and fused

Classifying Complications: Assessing Adult Spinal Deformity 2-Year Surgical Outcomes.

STUDY DESIGN: Retrospective review of prospective database. OBJECTIVE: Complication rates for adult spinal deformity (ASD) surgery vary widely because there is no accepted system for categorization. Our objective was to identify the impact of complication occurrence, minor-major complication, and Clavien-Dindo complication classification (Cc) on clinical variables and patient-reported outcomes. METHODS: Complications in surgical ASD patients with complete baseline and 2-year data were considered intraoperatively, perioperatively (\u3c6 \u3eweeks), and postoperatively (\u3e6 weeks). Primary outcome measures were complication timing and severity according to 3 scales: complication presence (yes/no), minor-major, and Cc score. Secondary outcomes were surgical outcomes (estimated blood loss [EBL], length of stay [LOS], reoperation) and health-related quality of life (HRQL) scores. Univariate analyses determined complication presence, type, and Cc grade impact on operative variables and on HRQL scores. RESULTS: Of 167 patients, 30.5% (n = 51) had intraoperative, 48.5% (n = 81) had perioperative, and 58.7% (n = 98) had postoperative complications. Major intraoperative complications were associated with increased EBL ( CONCLUSION: The Cc Scale was most useful in predicting changes in patient outcomes; at 2 years, patients with raised perioperative Cc scores and postoperative complications saw reduced HRQL improvement. Intraoperative and perioperative complications were associated with worse short-term surgical and inpatient outcomes

Evidence for a Transport-Trap Mode of Drosophila melanogaster gurken mRNA Localization

The Drosophila melanogaster gurken gene encodes a TGF alpha-like signaling molecule that is secreted from the oocyte during two distinct stages of oogenesis to define the coordinate axes of the follicle cell epithelium that surrounds the oocyte and its 15 anterior nurse cells. Because the gurken receptor is expressed throughout the epithelium, axial patterning requires region-specific secretion of Gurken protein, which in turn requires subcellular localization of gurken transcripts. The first stage of Gurken signaling induces anteroposterior pattern in the epithelium and requires the transport of gurken transcripts from nurse cells into the oocyte. The second stage of Gurken signaling induces dorsovental polarity in the epithelium and requires localization of gurken transcripts to the oocyte's anterodorsal corner. Previous studies, relying predominantly on real-time imaging of injected transcripts, indicated that anterodorsal localization involves transport of gurken transcripts to the oocyte's anterior cortex followed by transport to the anterodorsal corner, and anchoring. Such studies further indicated that a single RNA sequence element, the GLS, mediates both transport steps by facilitating association of gurken transcripts with a cytoplasmic dynein motor complex. Finally, it was proposed that the GLS somehow steers the motor complex toward that subset of microtubules that are nucleated around the oocyte nucleus, permitting directed transport to the anterodorsal corner. Here, we re-investigate the role of the GLS using a transgenic fly assay system that includes use of the endogenous gurken promoter and biological rescue as well as RNA localization assays. In contrast to previous reports, our studies indicate that the GLS is sufficient for anterior localization only. Our data support a model in which anterodorsal localization is brought about by repeated rounds of anterior transport, accompanied by specific trapping at the anterodorsal cortex. Our data further indicate that trapping at the anterodorsal corner requires at least one as-yet-unidentified gurken RLE

Analysis of long-term observations of NOx and CO in megacities and application to constraining emissions inventories

Long-term atmospheric NOx/CO enhancement ratios in megacities provide evaluations of emission inventories. A fuel-based emission inventory approach that diverges from conventional bottom-up inventory methods explains 1970–2015 trends in NOx/CO enhancement ratios in Los Angeles. Combining this comparison with similar measurements in other U.S. cities demonstrates that motor vehicle emissions controls were largely responsible for U.S. urban NOx/CO trends in the past half century. Differing NOx/CO enhancement ratio trends in U.S. and European cities over the past 25 years highlights alternative strategies for mitigating transportation emissions, reflecting Europe's increased use of light-duty diesel vehicles and correspondingly slower decreases in NOx emissions compared to the U.S. A global inventory widely used by global chemistry models fails to capture these long-term trends and regional differences in U.S. and Europe megacity NOx/CO enhancement ratios, possibly contributing to these models' inability to accurately reproduce observed long-term changes in tropospheric ozone

Clinical Features and Pathobiology of Chordoma

Chordoma is the most common primary malignant bone tumor found in the spine and sacrum. While these tumors are relatively slow growing, they do have the potential to recur and metastasize. Chordoma was described histologically long before it was realized that they were probably derived from notochordal precursor cells. Early reports dating back to Virchow in 1857 describe a vacuolated cell type seen in these tumors. These cells were described as physaliferous, from Greek for “having bubbles.” It was thought at that early time that they were a cartilaginous tumor, which may have been a result of evaluation of a chondroid variant tumor. By 1923, Burrow and Stewart recognized that chordomas were a “lowly malignant tumor of slow growth, locally invasive and destructive, and only rarely giving rise to metastases.” By then, the location at either end of the spine correlated well with contemporary descriptions of the location of vestigial notochordal remnants by Muller. These observations led to the hypothesis that chordomas were not tumors of the intervertebral disc but rather malignant transformation of these notochordal remnants. In 1858, Muller coined the current name by proposing the following hypothesis: “A direct relation of these growths to the chorda dorsalis cannot be overlooked and I consider them to be excessively growing remnants of the chorda. Whosoever likes the name may designate these masses as chordoid tumors, or chordomas.” Since these early descriptions, this concept has been supported by significant indirect evidence, although there is a paucity of direct proof