Doherty Institute - Research Publications
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ItemNo Preview AvailablePandemic printing: a novel 3D-printed swab for detecting SARS-CoV-2(WILEY, 2020-09)OBJECTIVES: To design and evaluate 3D-printed nasal swabs for collection of samples for SARS-CoV-2 testing. DESIGN: An iterative design process was employed. Laboratory evaluation included in vitro assessment of mock nasopharyngeal samples spiked with two different concentrations of gamma-irradiated SARS-CoV-2. A prospective clinical study compared SARS-CoV-2 and human cellular material recovery by 3D-printed swabs and standard nasopharyngeal swabs. SETTING, PARTICIPANTS: Royal Melbourne Hospital, May 2020. Participants in the clinical evaluation were 50 hospital staff members attending a COVID-19 screening clinic and two inpatients with laboratory-confirmed COVID-19. INTERVENTION: In the clinical evaluation, a flocked nasopharyngeal swab sample was collected with the Copan ESwab and a mid-nasal sample from the other nostril was collected with the 3D-printed swab. RESULTS: In the laboratory evaluation, qualitative agreement with regard to SARS-CoV-2 detection in mock samples collected with 3D-printed swabs and two standard swabs was complete. In the clinical evaluation, qualitative agreement with regard to RNase P detection (a surrogate measure of adequate collection of human cellular material) in samples collected from 50 hospital staff members with standard and 3D-printed swabs was complete. Qualitative agreement with regard to SARS-CoV-2 detection in three pairs of 3D-printed mid-nasal and standard swab samples from two inpatients with laboratory-confirmed SARS-CoV-2 was also complete. CONCLUSIONS: Using 3D-printed swabs to collect nasal samples for SARS-CoV-2 testing is feasible, acceptable to patients and health carers, and convenient.
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ItemValidation of a single-step, single-tube reverse transcription loop-mediated isothermal amplification assay for rapid detection of SARS-CoV-2 RNA(MICROBIOLOGY SOC, 2020)Introduction. The SARS-CoV-2 pandemic of 2020 has resulted in unparalleled requirements for RNA extraction kits and enzymes required for virus detection, leading to global shortages. This has necessitated the exploration of alternative diagnostic options to alleviate supply chain issues.Aim. To establish and validate a reverse transcription loop-mediated isothermal amplification (RT- LAMP) assay for the detection of SARS-CoV-2 from nasopharyngeal swabs.Methodology. We used a commercial RT-LAMP mastermix from OptiGene in combination with a primer set designed to detect the CDC N1 region of the SARS-CoV-2 nucleocapsid (N) gene. A single-tube, single-step fluorescence assay was implemented whereby 1 µl of universal transport medium (UTM) directly from a nasopharyngeal swab could be used as template, bypassing the requirement for RNA purification. Amplification and detection could be conducted in any thermocycler capable of holding 65 °C for 30 min and measure fluorescence in the FAM channel at 1 min intervals.Results. Assay evaluation by assessment of 157 clinical specimens previously screened by E-gene RT-qPCR revealed assay sensitivity and specificity of 87 and 100%, respectively. Results were fast, with an average time-to-positive (Tp) for 93 clinical samples of 14 min (sd±7 min). Using dilutions of SARS-CoV-2 virus spiked into UTM, we also evaluated assay performance against FDA guidelines for implementation of emergency-use diagnostics and established a limit-of-detection of 54 Tissue Culture Infectious Dose 50 per ml (TCID50 ml-1), with satisfactory assay sensitivity and specificity. A comparison of 20 clinical specimens between four laboratories showed excellent interlaboratory concordance; performing equally well on three different, commonly used thermocyclers, pointing to the robustness of the assay.Conclusion. With a simplified workflow, The N1 gene Single Tube Optigene LAMP assay (N1-STOP-LAMP) is a powerful, scalable option for specific and rapid detection of SARS-CoV-2 and an additional resource in the diagnostic armamentarium against COVID-19.
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ItemTracking the COVID-19 pandemic in Australia using genomics(NATURE PORTFOLIO, 2020-09-01)Genomic sequencing has significant potential to inform public health management for SARS-CoV-2. Here we report high-throughput genomics for SARS-CoV-2, sequencing 80% of cases in Victoria, Australia (population 6.24 million) between 6 January and 14 April 2020 (total 1,333 COVID-19 cases). We integrate epidemiological, genomic and phylodynamic data to identify clusters and impact of interventions. The global diversity of SARS-CoV-2 is represented, consistent with multiple importations. Seventy-six distinct genomic clusters were identified, including large clusters associated with social venues, healthcare and cruise ships. Sequencing sequential samples from 98 patients reveals minimal intra-patient SARS-CoV-2 genomic diversity. Phylodynamic modelling indicates a significant reduction in the effective viral reproductive number (Re) from 1.63 to 0.48 after implementing travel restrictions and physical distancing. Our data provide a concrete framework for the use of SARS-CoV-2 genomics in public health responses, including its use to rapidly identify SARS-CoV-2 transmission chains, increasingly important as social restrictions ease globally.