Viral load in upper respiratory tract of COVID-19 patients detected by digital PCR

Abstract Background: Upper respiratory tract specimens are widely applicable for the diagnosis of COVID-19. To date, no study has analyzed the actual viral loads in upper respiratory tract and its relationship with the severity of lung lesions, Ct value of RT-PCR and transmission capacity in COVID-19 patients.Methods: We retrospectively enrolled nine COVID-19 patients. Clinical data and close contacts of these patients were investigated. Respiratory samples were tested for SARS-CoV-2 with both normal RT-PCR and droplet digital PCR.Results: All the COVID-19 patients complicated with pneumonia. Viral loads in nasopharyngeal swabs were accurately quantified, and they had no direct correspondence with the severity of lung lesions. The Cycle Threshold (Ct) value of RT-PCR was approximately consistent with the absolute quantification of digital PCR. The spearman correlation coefficient between them was -0.952 with P value < 0.001. Close contacts of patients with very low viral load or no detected virus were not infected.Conclusions: Viral loads in nasopharyngeal swabs, could not predict the severity of lung lesions revealed by CT in COVID-19 patients. The infectious capacity of patients with low or absent viral load in upper respiratory tract was relatively weak, and wearing mask might be helpful for lower its spread.

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Induction of interferon response by high viral loads at early stage infection may protect against severe outcomes in COVID-19 patients

Scientific Reports ◽

10.1038/s41598-021-95197-y ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Eric C. Rouchka ◽

Julia H. Chariker ◽

Brian Alejandro ◽

Robert S. Adcock ◽

Richa Singhal ◽

...

Keyword(s):

Gene Expression ◽

Viral Load ◽

Respiratory Tract ◽

Disease Severity ◽

Critical Factor ◽

Interferon Response ◽

Upper Respiratory Tract ◽

Viral Loads ◽

Antiviral Responses ◽

Upper Respiratory

AbstractKey elements for viral pathogenesis include viral strains, viral load, co-infection, and host responses. Several studies analyzing these factors in the function of disease severity of have been published; however, no studies have shown how all of these factors interplay within a defined cohort. To address this important question, we sought to understand how these four key components interplay in a cohort of COVID-19 patients. We determined the viral loads and gene expression using high throughput sequencing and various virological methods. We found that viral loads in the upper respiratory tract in COVID-19 patients at an early phase of infection vary widely. While the majority of nasopharyngeal (NP) samples have a viral load lower than the limit of detection of infectious viruses, there are samples with an extraordinary amount of SARS-CoV-2 RNA and a high viral titer. No specific viral factors were identified that are associated with high viral loads. Host gene expression analysis showed that viral loads were strongly correlated with cellular antiviral responses. Interestingly, however, COVID-19 patients who experience mild symptoms have a higher viral load than those with severe complications, indicating that naso-pharyngeal viral load may not be a key factor of the clinical outcomes of COVID-19. The metagenomics analysis revealed that the microflora in the upper respiratory tract of COVID-19 patients with high viral loads were dominated by SARS-CoV-2, with a high degree of dysbiosis. Finally, we found a strong inverse correlation between upregulation of interferon responses and disease severity. Overall our study suggests that a high viral load in the upper respiratory tract may not be a critical factor for severe symptoms; rather, dampened antiviral responses may be a critical factor for a severe outcome from the infection.

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Paired nasopharyngeal and deep lung testing for SARS-CoV2 reveals a viral gradient in critically ill patients: a multi-centre study

10.1101/2020.07.19.20156869 ◽

2020 ◽

Author(s):

Islam Hamed ◽

Nesreen Shaban ◽

Marwan Nassar ◽

Sam Love ◽

Martin D Curran ◽

...

Keyword(s):

Viral Load ◽

Respiratory Tract ◽

Critically Ill ◽

Critically Ill Patients ◽

Lower Respiratory Tract ◽

Limit Of Detection ◽

Upper Respiratory Tract ◽

Rt Pcr ◽

Multi Centre Study ◽

Upper Respiratory

Introduction Samples for diagnostic tests for SARS-CoV-2 can be obtained from the upper (nasopharyngeal/oropharyngeal swabs) or lower respiratory tract (sputum or tracheal aspirate or broncho-alveolar lavage - BAL). Data from different testing sites indicates different rates of positivity. Reverse-transcriptase polymerase chain reaction (RT-PCR) allows for semi-quantitative estimates of viral load as time to crossing threshold (Ct) is inversely related to viral load. Objectives The objective of our study was to evaluate SARS-CoV2 RNA loads between paired nasopharyngeal (NP) and deep lung (endotracheal aspirate or BAL) samples from critically ill patients. Methods SARS-CoV-2 RT-PCR results were retrospectively reviewed for 51 critically ill patients from 5 intensive care units in 3 hospitals ; Addenbrookes Hospital Cambridge (3 units), Royal Papworth Cambridge (1 unit), and Royal Sunderland Hospital (1 unit). At the times when paired NP and deep lung samples were obtained, one patient had been on oxygen only, 6 patients on non-invasive ventilation, 18 patients on ECMO, and 26 patients mechanically ventilated. Results Results collected showed significant gradient between NP and deep lung viral loads. Median Ct value was 29 for NP samples and 24 for deep lung samples. Of 51 paired samples, 16 were negative (below limit of detection) on NP swabs but positive (above limit of detection) on deep lung sample, whilst 2 were negative on deep sample but positive on NP (both patients were on ECMO). Conclusions It has been suggested that whilst SARS-CoV1 tends to replicate in the lower respiratory tract, SARS-CoV2 replicates more vigorously in the upper respiratory tract. These data challenge that assumption. These data suggest that viral migration to, and proliferation in, the lower respiratory tract may be a key factor in the progression to critical illness and the development of severe acute respiratory syndrome (SARS). Factors which promote this migration should be examined for association with severe COVID-19. From a practical point of view, patients with suspected severe COVID-19 should have virological samples obtained from the lower respiratory tract where-ever possible, as upper respiratory samples have a significant negative rate.

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Investigation of Viral Load Cycle Threshold Values in Patients with SARS-CoV-2 Associated Pneumonia with Real-Time PCR Method

The Journal of Infection in Developing Countries ◽

10.3855/jidc.14281 ◽

2021 ◽

Vol 15 (10) ◽

pp. 1408-1414

Author(s):

Ayfer Bakir ◽

Tugrul Hosbul ◽

Ferhat Cuce ◽

Cumhur Artuk ◽

Gurhan Taskin ◽

...

Keyword(s):

Viral Load ◽

Respiratory Tract ◽

Load Cycle ◽

Cross Sectional Study ◽

Chest Ct ◽

Upper Respiratory Tract ◽

Cross Sectional ◽

Cycle Threshold ◽

Viral Loads ◽

Ct Value

Introduction: In this study, we aimed investigate the relationship of SARS-CoV-2 viral load cycle threshold (Ct) values with pneumonia. Methodology: A total of 158 patients in whom SARS-CoV-2 was confirmed in upper respiratory tract (URT) samples with molecular method and who had computed tomography (CT) of the chest, between April 2020 and June 2020 were included in this retrospective cross-sectional study. Results: Mean age of 158 PCR positive patients was 45.22 ± 17.89 and 60.8% of them were male. Pneumonia was detected in 40.5% of the patients on their chest CT. A weak but significant correlation was found between SARS-CoV-2 Ct value detected with PCR in analysis of oropharyngeal/ nasopharyngeal (OP/NP) samples and chest CT score (Pearson’s r: 0.197, p = 0.01). No correlation was found between the first detected viral load Ct value and age, gender and mortality. There was no significant correlation between chest CT score and mortality. While the areas remaining under ROC curve for Ct value in analysis of OP/NP samples in prediction of chest CT score ≥ 1, ≥ 5 and ≥ 10 were 0.564, 0.640 and 0.703 respectively. Conclusions: We found that the amount of SARS-CoV-2 viral load (inverse relationship with Ct) detected in OP/NP samples of patients with COVID-19 pneumonia did not reflect the increasing severity of pulmonary lesions on chest CT. Although primary target of SARS-CoV-2 is all epithelial cells of the respiratory tract we believe studies comparing viral loads in lower respiratory tract samples are needed to determine the severity of pulmonary disease.

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Longitudinal Analysis of Inflammatory Response to SARS-CoV-2 in the Upper Respiratory Tract Reveals an Association with Viral Load, Independent of Symptoms

Journal of Clinical Immunology ◽

10.1007/s10875-021-01134-z ◽

2021 ◽

Author(s):

Diem-Lan Vu ◽

Paola Martinez-Murillo ◽

Fiona Pigny ◽

Maria Vono ◽

Benjamin Meyer ◽

...

Keyword(s):

Viral Load ◽

Respiratory Tract ◽

Upper Respiratory Tract ◽

University Hospitals ◽

Viral Loads ◽

Gm Csf ◽

Cytokine Levels ◽

Control Virus ◽

Upper Respiratory ◽

Local Recruitment

Abstract Background SARS-CoV-2 infection leads to high viral loads in the upper respiratory tract that may be determinant in virus dissemination. The extent of intranasal antiviral response in relation to symptoms is unknown. Understanding how local innate responses control virus is key in the development of therapeutic approaches. Methods SARS-CoV-2-infected patients were enrolled in an observational study conducted at the Geneva University Hospitals, Switzerland, investigating virological and immunological characteristics. Nasal wash and serum specimens from a subset of patients were collected to measure viral load, IgA specific for the S1 domain of the spike protein, and a cytokine panel at different time points after infection; cytokine levels were analyzed in relation to symptoms. Results Samples from 13 SARS-CoV-2-infected patients and six controls were analyzed. We found an increase in CXCL10 and IL-6, whose levels remained elevated for up to 3 weeks after symptom onset. SARS-CoV-2 infection also induced CCL2 and GM-CSF, suggesting local recruitment and activation of myeloid cells. Local cytokine levels correlated with viral load but not with serum cytokine levels, nor with specific symptoms, including anosmia. Some patients had S1-specific IgA in the nasal cavity while almost none had IgG. Conclusion The nasal epithelium is an active site of cytokine response against SARS-CoV-2 that can last more than 2 weeks; in this mild COVID-19 cohort, anosmia was not associated with increases in any locally produced cytokines.

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Duration of infectiousness and correlation with RT-PCR cycle threshold values in cases of COVID-19, England, January to May 2020

Eurosurveillance ◽

10.2807/1560-7917.es.2020.25.32.2001483 ◽

2020 ◽

Vol 25 (32) ◽

Cited By ~ 43

Author(s):

Anika Singanayagam ◽

Monika Patel ◽

Andre Charlett ◽

Jamie Lopez Bernal ◽

Vanessa Saliba ◽

...

Keyword(s):

Viral Load ◽

Severe Acute Respiratory Syndrome ◽

Respiratory Tract ◽

Symptom Onset ◽

Infectious Virus ◽

Upper Respiratory Tract ◽

Threshold Values ◽

Rt Pcr ◽

Cycle Threshold ◽

Upper Respiratory

Severe acute respiratory syndrome coronavirus 2 viral load in the upper respiratory tract peaks around symptom onset and infectious virus persists for 10 days in mild-to-moderate coronavirus disease (n = 324 samples analysed). RT-PCR cycle threshold (Ct) values correlate strongly with cultivable virus. Probability of culturing virus declines to 8% in samples with Ct > 35 and to 6% 10 days after onset; it is similar in asymptomatic and symptomatic persons. Asymptomatic persons represent a source of transmissible virus.

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Association between upper respiratory tract viral load, comorbidities, disease severity and outcome of patients with SARS-CoV-2 infection

The Journal of Infectious Diseases ◽

10.1093/infdis/jiaa804 ◽

2021 ◽

Author(s):

Helena C Maltezou ◽

Vasilios Raftopoulos ◽

Rengina Vorou ◽

Kalliopi Papadima ◽

Kassiani Mellou ◽

...

Keyword(s):

Viral Load ◽

Respiratory Tract ◽

Disease Severity ◽

High Viral Load ◽

P Value ◽

Upper Respiratory Tract ◽

Limited Information ◽

Symptomatic Disease ◽

Viral Loads ◽

Upper Respiratory

Abstract Background There is limited information on the association between upper respiratory tract (URT) viral loads, host factors, and disease severity in SARS-CoV-2 infected patients. Methods We studied 1,122 patients (mean age: 46 years) diagnosed by PCR. URT viral load, measured by PCR cycle threshold, was categorized as high, moderate or low. Results There were 336 (29.9%) patients with comorbidities; 309 patients (27.5%) had high, 316 (28.2%) moderate, and 497 (44.3%) low viral load. In univariate analyses, compared to patients with moderate or low viral load, patients with high viral load were older, had more often comorbidities, developed symptomatic disease, were intubated and died; in addition, patients with high viral load had longer stay in intensive care unit and longer intubation compared to patients with low viral load (p-values <0.05 for all). Patients with chronic cardiovascular disease, hypertension, chronic pulmonary disease, immunosuppression, obesity and chronic neurological disease had more often high viral load (p-value<0.05 for all). Multivariate analysis found that a high viral load was associated with COVID-19. The level of viral load was not associated with any other outcome. Conclusions URT viral load could be used to identify patients at higher risk for morbidity or severe outcome.

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Longitudinal Analysis of Inflammatory Response to SARS-CoV-2 in the Upper Respiratory Tract Reveals an Association With Viral Load, Independent of Symptoms

10.21203/rs.3.rs-203414/v1 ◽

2021 ◽

Author(s):

Arnaud Didierlaurent ◽

Vu Diem-Lan ◽

Paola Martinez Murillo ◽

Fiona Pigny ◽

Maria Vono ◽

...

Keyword(s):

Viral Load ◽

Respiratory Tract ◽

Upper Respiratory Tract ◽

University Hospitals ◽

Viral Loads ◽

Gm Csf ◽

Cytokine Levels ◽

Control Virus ◽

Upper Respiratory ◽

Local Recruitment

Abstract BackgroundSARS-CoV-2 infection leads to high viral loads in the upper respiratory tract that may be determinant in virus dissemination. The extent of intranasal antiviral response in relation to symptoms is unknown. Understanding how local innate responses control virus is key in the development of therapeutic approaches.MethodsSARS-CoV-2-infected patients were enrolled in an observational study conducted at the Geneva University Hospitals, Switzerland, investigating virological and immunological characteristics. Nasal-wash and serum specimens from a subset of patients were collected to measure viral load and a cytokine panel at different time points after infection; cytokine levels were analyzed in relation to symptoms.ResultsSamples from 13 SARS-CoV-2-infected patients and six controls were analyzed. We found an increase in CXCL10 and IL-6, whose levels remained elevated for up to 3 weeks after symptom onset. SARS-CoV-2 infection also induced CCL2 and GM-CSF, suggesting local recruitment and activation of myeloid cells. Local cytokine levels correlated with viral load but not with serum cytokine levels, nor with specific symptoms, including anosmia.ConclusionsThe nasal epithelium is an active site of cytokine response against SARS-CoV-2 that can last more than 2 weeks; in this cohort, anosmia was not associated with increases in any locally produced cytokines.

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Kinetics of SARS-CoV-2 infection in the human upper and lower respiratory tracts and their relationship with infectiousness

10.1101/2020.09.25.20201772 ◽

2020 ◽

Cited By ~ 4

Author(s):

Ruian Ke ◽

Carolin Zitzmann ◽

Ruy M. Ribeiro ◽

Alan S. Perelson

Keyword(s):

Viral Load ◽

Respiratory Tract ◽

Dynamic Models ◽

Target Cells ◽

Upper Respiratory Tract ◽

Viral Kinetics ◽

Viral Loads ◽

Viral Load Data ◽

Upper Respiratory ◽

Kinetics Of

SARS-CoV-2 is a human pathogen that causes infection in both the upper respiratory tract (URT) and the lower respiratory tract (LRT). The viral kinetics of SARS-CoV-2 infection and how they relate to infectiousness and disease progression are not well understood. Here, we develop data-driven viral dynamic models of SARS-CoV-2 infection in both the URT and LRT. We fit the models to viral load data from patients with likely infection dates known, we estimated that infected individuals with a longer incubation period had lower rates of viral growth, took longer to reach peak viremia in the URT, and had higher chances of presymptomatic transmission. We then developed a model linking viral load to infectiousness. We found that to explain the substantial fraction of transmissions occurring presymptomatically, the infectiousness of a person should depend on a saturating function of the viral load, making the logarithm of the URT viral load a better surrogate of infectiousness than the viral load itself. Comparing the roles of target-cell limitation, the innate immune response, proliferation of target cells and spatial infection in the LRT, we found that spatial dissemination in the lungs is likely to be an important process in sustaining the prolonged high viral loads. Overall, our models provide a quantitative framework for predicting how SARS-CoV-2 within-host dynamics determine infectiousness and represent a step towards quantifying how viral load dynamics and the immune responses determine disease severity.

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Asymptomatic COVID-19 Adult Outpatients identified as Significant Viable SARS-CoV-2 Shedders

10.21203/rs.3.rs-403751/v1 ◽

2021 ◽

Author(s):

Marie Glenet ◽

Anne-Laure Lebreil ◽

Laetitia Heng ◽

Yohan N’Guyen ◽

Ittah Meyer ◽

...

Keyword(s):

Respiratory Tract ◽

Control Measures ◽

Upper Respiratory Tract ◽

Rna Detection ◽

Viral Loads ◽

Infection Control Measures ◽

Infected Adult ◽

Asymptomatic Adult ◽

Upper Respiratory ◽

Kinetics Of

Abstract Differential kinetics of RNA loads and infectious viral levels in the upper respiratory tract between asymptomatic and symptomatic SARS-CoV-2 infected adult outpatients remain unclear limiting recommendations that may guide clinical management, infection control measures and occupational health decisions. In the present investigation, 496 (2.5%) of 17,911 French adult outpatients were positive for an upper respiratory tract SARS-CoV-2 RNA detection by a quantitative RT-PCR assay, of which 180 (36.3%) were COVID-19 asymptomatic. Of these adult asymptomatic viral shedders, 84.4% had mean to high RNA viral loads (Ct values<30) which median value was significantly higher than that observed in symptomatic subjects (P=0.029), and 50.6% were positive by cell culture assays of their upper respiratory tract specimens. Our findings indicate that COVID-19 asymptomatic adult outpatients are significant viable SARS-CoV-2 shedders in their upper respiratory tract playing a major potential role as SARS-CoV-2 transmitters in various epidemiological transmission chains, promoting COVID-19 resurgence in populations.

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