The total number and mass of SARS-CoV-2 virions

Sender, Ron; Bar-On, Yinon M.; Flamholz, Avi I.; Gleizer, Shmuel; Bernsthein, Biana; Phillips, Rob

doi:10.1101/2020.11.16.20232009

Cited by 61 publications

(73 citation statements)

References 79 publications

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“…The cytopathic effect was observed from approx. 10 4 RNA copies/mL, which, with the infectious particles:RNA ratio in the tissues ranging typically between 1:1000 and 1:10,000 [ 10 , 11 ] corresponds to 1–10 infectious particles/mL.…”

Section: Methodsmentioning

confidence: 99%

Five Antigen Tests for SARS-CoV-2: Virus Viability Matters

Homza

Zelená

Janošek

et al. 2021

Viruses

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Antigen testing for SARS-CoV-2 (AGT) is generally considered inferior to RT-PCR testing in terms of sensitivity. However, little is known about the infectiousness of RT-PCR positive patients who pass undetected by AGT. In a screening setting for mildly symptomatic or asymptomatic patients with high COVID-19 prevalence (30–40%), 1141 patients were tested using one of five AGTs and RT-PCR. Where the results differed, virus viability in the samples was tested on cell culture (CV-1 cells). The test battery included AGTs by JOYSBIO, Assure Tech, SD Biosensor, VivaChek Biotech and NDFOS. Sensitivities of the ATGs compared to RT-PCR ranged from 42% to 76%. The best test yielded a 76% sensitivity, 97% specificity, 92% positive, and 89% negative predictive values, respectively. However, in the best performing ATG tests, almost 90% of samples with “false negative” AGT results contained no viable virus. Corrected on the virus viability, sensitivities grew to 81%–97% and, with one exception, the tests yielded high specificities >96%. Performance characteristics of the best test after adjustment were 96% sensitivity, 97% specificity, 92% positive, and 99% negative predictive values (high prevalence population). We, therefore, believe that virus viability should be considered when assessing the AGT performance. Also, our results indicate that a well-performing antigen test could in a high-prevalence setting serve as an excellent tool for identifying patients shedding viable virus. We also propose that the high proportion of RT-PCR-positive samples containing no viable virus in the group of “false negatives” of the antigen test should be further investigated with the aim of possibly preventing needless isolation of such patients.

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Section: Methodsmentioning

confidence: 99%

Five Antigen Tests for SARS-CoV-2: Virus Viability Matters

Homza

Zelená

Janošek

et al. 2021

Viruses

View full text Add to dashboard Cite

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“…The method sensitivity was verified through a serial dilution of the virus stock suspension (3 × 10 11 RNA copies/ml), both without and after freezing at −80 °C. The cytopathic effect was observed from ∼10 4 RNA copies/ml, which corresponds to 1–10 infectious particles/ml [ 14 , 15 ]. To verify that samples after freezing were not compromised, we performed (besides the above-described verification of the method sensitivity) also an analysis before and after freezing on 10 real-world samples with cycle thresholds 25–30 (5 samples) and 30–40 (5 samples).…”

Section: Methodsmentioning

confidence: 99%

Covid-19 antigen testing: better than we know? A test accuracy study

Homza

Zelená

Janošek

et al. 2021

Infectious Diseases

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Background Antigen testing for SARS-CoV-2 is considered to be less sensitive than the standard reference method – real-time PCR (RT-PCR). It has been suggested that many patients with positive RT-PCR ‘missed’ by antigen testing might be non-infectious. Methods In a real-world high-throughput setting for asymptomatic or mildly symptomatic patients, 494 patients were tested using RT-PCR as well as a single lateral flow antigen test (Ecotest, AssureTech, China). Where the results differed, virus viability was evaluated by cell culture. The test parameters were calculated with RT-PCR and RT-PCR adjusted on viability as reference standards. Results The overall sensitivity of the used antigen test related to the RT-PCR only was 76.2%, specificity was 97.3%. However, 36 out of 39 patients ‘missed’ by the antigen test contained no viable virus. After adjusting on that, the sensitivity grew to 97.7% and, more importantly for disease control purposes, the negative predictive value reached 99.2%. Conclusions We propose that viability testing should be always performed when evaluating a new antigen test. A well-chosen and validated antigen test provides excellent results in identifying patients who are shedding viable virus (although some caveats still remain) in the real-world high-throughput setting of asymptomatic or mildly symptomatic individuals.

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“…The 46-kDa SARS-CoV-2 Nucleocapsid (N) protein is 89.1% identical to SARS-CoV N. Genomic analysis of human coronaviruses indicated that N might be the major factor conferring the enhanced pathogenicity to SARS-CoV-2 [4]. N represents the most abundant viral protein in the infected cell [12][13][14], with each assembled virion containing approximately one thousand molecules of N [15]. Given that each infected cell can contain up to 10 5 virions (infectious, defective and incomplete overall) [14], the number of N molecules in an infected cell can reach 10 8 , accounting for ~1% of a total number of cellular proteins (~10 10 [16]).…”

Section: Introductionmentioning

confidence: 99%

The mechanism of SARS-CoV-2 nucleocapsid protein recognition by the human 14-3-3 proteins

Tugaeva

Hawkins

Smith

et al. 2020

Preprint

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The coronavirus nucleocapsid protein (N) controls viral genome packaging and contains numerous phosphorylation sites located within unstructured regions. Binding of phosphorylated SARS-CoV N to the host 14-3-3 protein in the cytoplasm was reported to regulate nucleocytoplasmic N shuttling. All seven isoforms of the human 14-3-3 are abundantly present in tissues vulnerable to SARS-CoV-2, where N can constitute up to ~1% of expressed proteins during infection. Although the association between 14-3-3 and SARS-CoV-2 N proteins can represent one of the key host-pathogen interactions, its molecular mechanism and the specific critical phosphosites are unknown. Here, we show that phosphorylated SARS-CoV-2 N protein (pN) dimers, reconstituted via bacterial co-expression with protein kinase A, directly associate, in a phosphorylation-dependent manner, with the dimeric 14-3-3 protein, but not with its monomeric mutant. We demonstrate that pN is recognized by all seven human 14-3-3 isoforms with various efficiencies and deduce the apparent KD to selected isoforms, showing that these are in a low micromolar range. Serial truncations pinpointed a critical phosphorylation site to Ser197, which is conserved among related zoonotic coronaviruses and located within the functionally important, SR-rich region of N. The relatively tight 14-3-3/pN association can regulate nucleocytoplasmic shuttling and other functions of N via occlusion of the SR-rich region, while hijacking cellular pathways by 14-3-3 sequestration. As such, the assembly may represent a valuable target for therapeutic intervention.HighlightsSARS-CoV-2 nucleocapsid protein (N) binds to all seven human 14-3-3 isoforms. This association with 14-3-3 strictly depends on phosphorylation of N. The two proteins interact in 2:2 stoichiometry and with the Kd in a μM range. Affinity of interaction depends on the specific 14-3-3 isoform. Conserved Ser197-phosphopeptide of N is critical for the interaction.

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The total number and mass of SARS-CoV-2 virions

Cited by 61 publications

References 79 publications

Five Antigen Tests for SARS-CoV-2: Virus Viability Matters

Five Antigen Tests for SARS-CoV-2: Virus Viability Matters

Covid-19 antigen testing: better than we know? A test accuracy study

The mechanism of SARS-CoV-2 nucleocapsid protein recognition by the human 14-3-3 proteins

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