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1.
J Clin Microbiol ; 59(6)2021 05 19.
Article in English | MEDLINE | ID: covidwho-1158099

ABSTRACT

LamPORE is a novel diagnostic platform for the detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA combining loop-mediated isothermal amplification with nanopore sequencing, which could potentially be used to analyze thousands of samples per day on a single instrument. We evaluated the performance of LamPORE against reverse transcriptase PCR (RT-PCR) using RNA extracted from spiked respiratory samples and stored nose and throat swabs collected at two UK hospitals. The limit of detection of LamPORE was 10 genome copies/µl of extracted RNA, which is above the limit achievable by RT-PCR, but was not associated with a significant reduction of sensitivity in clinical samples. Positive clinical specimens came mostly from patients with acute symptomatic infection, and among them, LamPORE had a diagnostic sensitivity of 99.1% (226/228; 95% confidence interval [CI], 96.9% to 99.9%). Among negative clinical specimens, including 153 with other respiratory pathogens detected, LamPORE had a diagnostic specificity of 99.6% (278/279; 98.0% to 100.0%). Overall, 1.4% (7/514; 0.5% to 2.9%) of samples produced an indeterminate result on first testing, and repeat LamPORE testing on the same RNA extract had a reproducibility of 96.8% (478/494; 94.8% to 98.1%). LamPORE has a similar performance as RT-PCR for the diagnosis of SARS-CoV-2 infection in symptomatic patients and offers a promising approach to high-throughput testing.


Subject(s)
COVID-19 , Nanopore Sequencing , Humans , Molecular Diagnostic Techniques , Nucleic Acid Amplification Techniques , RNA, Viral/genetics , Reproducibility of Results , SARS-CoV-2 , Sensitivity and Specificity
2.
Science ; 372(6539)2021 04 16.
Article in English | MEDLINE | ID: covidwho-1125076

ABSTRACT

Extensive global sampling and sequencing of the pandemic virus severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have enabled researchers to monitor its spread and to identify concerning new variants. Two important determinants of variant spread are how frequently they arise within individuals and how likely they are to be transmitted. To characterize within-host diversity and transmission, we deep-sequenced 1313 clinical samples from the United Kingdom. SARS-CoV-2 infections are characterized by low levels of within-host diversity when viral loads are high and by a narrow bottleneck at transmission. Most variants are either lost or occasionally fixed at the point of transmission, with minimal persistence of shared diversity, patterns that are readily observable on the phylogenetic tree. Our results suggest that transmission-enhancing and/or immune-escape SARS-CoV-2 variants are likely to arise infrequently but could spread rapidly if successfully transmitted.


Subject(s)
COVID-19/transmission , COVID-19/virology , Genetic Variation , SARS-CoV-2/genetics , COVID-19/immunology , Coinfection/virology , Coronavirus Infections/virology , Coronavirus OC43, Human , Family Characteristics , Genome, Viral , Humans , Immune Evasion , Mutation , Phylogeny , RNA, Viral/genetics , RNA-Seq , SARS-CoV-2/pathogenicity , SARS-CoV-2/physiology , Selection, Genetic , Spike Glycoprotein, Coronavirus/genetics , United Kingdom , Viral Load
3.
BMC Infect Dis ; 21(1): 187, 2021 Feb 18.
Article in English | MEDLINE | ID: covidwho-1090687

ABSTRACT

BACKGROUND: Thresholds for SARS-CoV-2 antibody assays have typically been determined using samples from symptomatic, often hospitalised, patients. In this setting the sensitivity and specificity of the best performing assays can both exceed 98%. However, antibody assay performance following mild infection is less clear. METHODS: We assessed quantitative IgG responses in a cohort of healthcare workers in Oxford, UK, with a high pre-test probability of Covid-19, in particular the 991/11,475(8.6%) who reported loss of smell/taste. We use anosmia/ageusia and other risk factors as probes for Covid-19 infection potentially undiagnosed by immunoassays by investigating their relationship with antibody readings either side of assay thresholds. RESULTS: The proportion of healthcare workers reporting anosmia/ageusia increased at antibody readings below diagnostic thresholds using an in-house ELISA (n = 9324) and the Abbott Architect chemiluminescent microparticle immunoassay (CMIA; n = 11,324): 426/906 (47%) reported anosmia/ageusia with a positive ELISA, 59/449 (13.1%) with high-negative and 326/7969 (4.1%) with low-negative readings. Similarly, by CMIA, 518/1093 (47.4%) with a positive result reported anosmia/ageusia, 106/686 (15.5%) with a high-negative and 358/9563 (3.7%) with a low-negative result. Adjusting for the proportion of staff reporting anosmia/ageusia suggests the sensitivity of both assays in mild infection is lower than previously reported: Oxford ELISA 89.8% (95%CI 86.6-92.8%) and Abbott CMIA 79.3% (75.9-82.7%). CONCLUSION: Following mild SARS-CoV-2 infection 10-30% of individuals may have negative immunoassay results. While lowered diagnostic thresholds may result in unacceptable specificity, our findings have implications for epidemiological analyses and result interpretation in individuals with a high pre-test probability. Samples from mild PCR-confirmed infections should be included in SARS-CoV-2 immunoassay evaluations.


Subject(s)
Antibodies, Viral/analysis , COVID-19 Serological Testing/standards , COVID-19/diagnosis , Immunoglobulin G/analysis , Adult , Ageusia/virology , Anosmia/virology , Asymptomatic Infections , Enzyme-Linked Immunosorbent Assay/standards , Female , Health Personnel , Humans , Immunoassay/standards , Male , Middle Aged , Sensitivity and Specificity , Undiagnosed Diseases , United Kingdom
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