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Human coronavirus NL63 (HCoV-NL63) is an enveloped pathogen of the family Coronaviridae that spreads worldwide and causes up to 10% of all annual respiratory diseases. HCoV-NL63 is typically associated with mild upper respiratory symptoms in children, elderly and immunocompromised individuals. It has also been shown to cause severe lower respiratory illness. NL63 shares ACE2 as a receptor for viral entry with SARS-CoV-1 and SARS-CoV-2. Here, we present the in situ structure of HCoV-NL63 spike (S) trimer at 3.4-Å resolution by single-particle cryo-EM imaging of vitrified virions without chemical fixative. It is structurally homologous to that obtained previously from the biochemically purified ectodomain of HCoV-NL63 S trimer, which displays a three-fold symmetric trimer in a single conformation. In addition to previously proposed and observed glycosylation sites, our map shows density at other sites, as well as different glycan structures. The domain arrangement within a protomer is strikingly different from that of the SARS-CoV-2 S and may explain their different requirements for activating binding to the receptor. This structure provides the basis for future studies of spike proteins with receptors, antibodies or drugs, in the native state of the coronavirus particles.
An in-house self-held respiration monitoring device (SHRMD) was developed for providing deep inspiration breath hold (DIBH) radiotherapy. The use of SHRMD is evaluated in terms of reproducibility, stability and heart dose reduction.
Methods and materials:
Sixteen patients receiving radiotherapy of left breast cancer were planned for treatment with both a free breathing (FB) scan and a DIBH scan. Both FB and DIBH plans were generated for comparison of the heart, left anterior descending (LAD) artery and lung dose. All patients received their treatments with DIBH using SHRMD. Megavoltage cine images were acquired during treatments for evaluating the reproducibility and stability of treatment position using SHRMD.
Compared with FB plans, the maximum dose to the heart by DIBH technique with SHRMD was reduced by 29·9 ± 15·6%; and the maximum dose of the LAD artery was reduced by 41·6 ± 18·3%. The inter-fractional overall mean error was 0·01 cm and the intra-fractional overall mean error was 0·04 cm.
This study demonstrated the potential benefits of using the SHRMD for DIBH to reduce the heart and LAD dose. The patients were able to perform stable and reproducible DIBHs.
Although acknowledged to be variable and subjective, manual annotation of cryo-electron tomography data is commonly used to answer structural questions and to create a “ground truth” for evaluation of automated segmentation algorithms. Validation of such annotation is lacking, but is critical for understanding the reproducibility of manual annotations. Here, we used voxel-based similarity scores for a variety of specimens, ranging in complexity and segmented by several annotators, to quantify the variation among their annotations. In addition, we have identified procedures for merging annotations to reduce variability, thereby increasing the reliability of manual annotation. Based on our analyses, we find that it is necessary to combine multiple manual annotations to increase the confidence level for answering structural questions. We also make recommendations to guide algorithm development for automated annotation of features of interest.