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Authors: Butler, DJ. et al

Link to paper: https://www.biorxiv.org/content/10.1101/2020.04.20.048066v4

Journal/ Pre-Print: bioRxiv

Tags: Bioinformatics, Diagnostics, Immunology/Immunity, Molecular biology, Statistics, Virology

Research Highlights

1. Development of a reproducible and fast colorimetric (LAMP, ~30 minutes) assay for SARS-CoV-2 detection

2. Discovery of a SARS-CoV-2 subclade that is enriched in New York City

Summary 

Butler et al developed a reproducible LAMP assay for a fast (~30 minute) discovery of SARS-CoV-2 (confirmed with qRT-PCR and RNA-seq). They also investigated the nasopharyngeal metatranscriptome and host transcriptional response of patients and found a correlation between viral load and expression of ACE2 and some interferon-response genes. Additionally, metatranscripome sequencing detected SARS-CoV-2 in some patients identified as negative by qRT-PCR. Reassembling 155 complete genomes they identified a dominant SARS-CoV-2 subclade in NYC, which originates from Western Europe. In a clinical analysis of 8856 patients they found that ACEI/ARB exposure increased COVID-19 risk, but showed equivocal effects for morbidity and mortality when accounting for hypertension.

Impact for SARS-CoV2/COVID19 research efforts

Understand the immune response to SARS-CoV2/COVID19

RNA-seq analysis of the host transcriptional responses with different viral load.

Develop diagnostic tools for SARS-CoV2/COVID19

Development of a LAMP assay for a fast (~30 min) detection of SARS-CoV-2.

Study Type

· In silico study / bioinformatics study: RNA-seq; phylogenetic

· In vitro study: LAMP and qRT-PCR of negative and positive patients.

Strengths and limitations of the paper

Novelty: Development of a one-tube, reproducible LAMP assay for a fast (~30 min) detection of SARS-CoV-2, without need for specialist equipment, and with a colorimetric read-out visible to the eye. Discovery of a highly enriched SARS-CoV-2 subclade

in NYC and presence of a three amino acid deletion in several subclades. ACE inhibitor (ACEI) or angiotensin receptor blocker (ARB) use in patients associated with testing positive for SARS-CoV-2, even after adjustment for age, sex, IL-6 levels, and other clinical covariates. Correlation between high viral load and high expression of ACE2 and some interferon response genes. Testing of surfaces (fomite transmission) in high-transit areas of the New York subway system found no detectable SARS-CoV-2 in 86 samples taken during 1 week in mid-March. Potentially highlights host downregulation of ALAS2 during infection, which has important roles in heme production during erythroblast development.

Standing in the field: The RNA-seq analysis of the host transcriptional response to SARS-CoV-2 gives a higher number of differentially expressed genes than a previous preprint found (Daniel Blanco-Melo et al, 2020). However, the current study uses a better approach and is therefore likely to be more meaningful (patient samples compared to cell culture, separation of the analysis based on viral load vs single viral load and time point). Other aspects of the study (development of the LAMP test, metatranscriptomics, phylogenetics, and results of the observational cohort) provide results in agreement with other studies.

Appropriate statistics: Yes. The number of samples and statistical tests used are described.

Viral model used: None

Translatability: Development of a reproducible LAMP test for a fast analysis (~ 30 minutes) of SARS-CoV-2 in humans. LAMP was performed on naso/oropharyngeal swab samples.

Analysis of an observational cohort (n=8,278) in NYC found that usage of ACE inhibitors confers an increased risk of intubation (HR=2.63 95%CI: 2.01-3.43, p=1.22E-12), mechanical respiration (HR=1.83 95%CI: 1.39-2.40, p=1.27E-05), and mortality (HR=1.68 95%CI: 1.22-2.31, p=1.42E-03). The ACE inhibitor, benazepril, was found to be associated with the highest risk of mortality (N=32, HR=2.37 95%CI: 1.05-5.35, p=3.70E-02).

Main limitations: The LAMP assay can only detect current or recent infection (ie primer-based, not antibody-based). Further work is needed to validate the assay with lower viral titres, and in samples easier to obtain than naso/oropharyngeal swabs (eg saliva, stool or buccal swab). More work is required to investigate the potential deleterious effects of ACE inhibitors on patients, and to understand the potential effects of SNPs on SARS-CoV-2 biology.