Towards Equitable Access to Public Health Pathogen Genomics in the Western Pacific.

The COVID-19 pandemic has increased interest and understanding of the utility of pathogen genomics across the Western Pacific region. Access to genomic data enhances surveillance and response to COVID-19, and will also support surveillance of other infectious diseases and antimicrobial resistant pathogens. Models of access can be determined based on intended purpose, use and sustainability. Achieving equitable access to genomics across the Western Pacific will contribute to the development of a regional public health genomics network to respond to major disease threats in the future.

Navigating Pregnancy and the Healthcare System during COVID-19: A Qualitative Study with Perinatal Women of Color.

OBJECTIVE: To address health disparities in the perinatal period (i.e., during pregnancy and through one year after birth) by exploring the intersectional experiences of perinatal Black, Indigenous, and other People of Color (BIPOC) women during the COVID-19 pandemic. In this study, participants were asked if and how COVID-19 had impacted their experiences of receiving healthcare, whether they had faced any challenges during this time, how they had navigated these challenges, and what recommendations they had for improving perinatal healthcare. METHODS: Between November 2021 and March 2022 our team conducted eight virtual focus groups comprising perinatal BIPOC women. A semi-structured interview protocol was used, and interviews were voice recorded and transcribed verbatim. The data were analyzed using reflexive thematic analysis. RESULTS: Three major themes common in BIPOC perinatal healthcare experiences during COVID-19 were generated through engaging in reflexive thematic analysis: (1) an overwhelming lack of support from providers, (2) experiences of blame and shame, and (3) difficulties navigating institutional policies that were unclear or ever-changing during the COVID-19 pandemic. Recommendations from participants included greater empathic communication from providers in the face of uncertainty during COVID-19, greater access to information and guidance for caring for themselves and their babies, and an overall request for greater compassion while navigating an exciting and busy time. RELEVANCE: These findings have implications for trauma-informed and inclusive perinatal care that can reduce the impacts of systemic inequalities for perinatal BIPOC women. This study offers a discussion of implications for future training for maternal health providers and implications for community-based programs.

Feasibility of a refurbished shipping container as a transportable laboratory for rapid SARS-CoV-2 diagnostics.

Background: Australia's response to the coronavirus disease 2019 (COVID-19) pandemic relies on widespread availability of rapid, accurate testing and reporting of results to facilitate contact tracing. The extensive geographical area of Australia presents a logistical challenge, with many of the population located distant from a laboratory capable of robust severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) detection. A strategy to address this is the deployment of a mobile facility utilizing novel diagnostic platforms. This study aimed to evaluate the feasibility of a fully contained transportable SARS-CoV-2 testing laboratory using a range of rapid point-of-care tests. Method: A 20 ft (6.1 m) shipping container was refurbished (GeneWorks, Adelaide, South Australia) with climate controls, laboratory benches, hand-wash station and a class II biosafety cabinet. Portable marquees situated adjacent to the container served as stations for registration, sample acquisition and personal protective equipment for staff. Specimens were collected and tested on-site utilizing either the Abbott ID NOW or Abbott Panbio rapid tests. SARS-CoV-2 positive results from the rapid platforms or any participants reporting symptoms consistent with COVID-19 were tested on-site by GeneXpert Xpress RT-PCR. All samples were tested in parallel with a standard-of-care RT-PCR test (Panther Fusion SARS-CoV-2 assay) performed at the public health reference laboratory. In-laboratory environmental conditions and data management-related factors were also recorded. Results: Over a 3 week period, 415 participants were recruited for point-of-care SARS-CoV-2 testing. From time of enrolment, the median result turnaround time was 26 min for the Abbott ID NOW, 32 min for the Abbott Panbio and 75 min for the Xpert Xpress. The environmental conditions of the refurbished shipping container were found to be suitable for all platforms tested, although humidity may have produced condensation within the container. Available software enabled turnaround times to be recorded, although technical malfunction resulted in incomplete data capture. Conclusion: Transportable container laboratories can enable rapid COVID-19 results at the point of care and may be useful during outbreak settings, particularly in environments that are physically distant from centralized laboratories. They may also be appropriate in resource-limited settings. The results of this pilot study confirm feasibility, although larger trials to validate individual rapid point-of-care testing platforms in this environment are required.

Lab-in-a-van: Rapid SARS-CoV-2 testing response with a mobile laboratory.

BACKGROUND: High testing rates and rapid contact tracing have been key interventions to control COVID-19 in Victoria, Australia. A mobile laboratory (LabVan), for rapid SARS-CoV-2 diagnostics, was deployed at sites deemed critical by the Victorian State Department of Health as part of the response. We describe the process of design, implementation, and performance benchmarked against a central reference laboratory. METHODS: A BSL2 compliant laboratory, complete with a class II biological safety cabinet, was built within a Mercedes-Benz Sprinter Panel Van. Swabs were collected by on-site collection teams, registered using mobile internet-enabled tablets and tested using the Xpert® Xpress SARS-CoV-2 assay. Results were reported remotely via HL7 messaging to Public Health Units. Patients with negative results were automatically notified by mobile telephone text messaging (SMS). FINDINGS: A pilot trial of the LabVan identified a median turnaround time (TAT) from collection to reporting of 1:19 h:mm (IQR 0:18, Range 1:03-18:32) compared to 9:40 h:mm (IQR 8:46, Range 6:51-19:30) for standard processing within the central laboratory. During deployment in nine rural and urban COVID-19 outbreaks the median TAT was 2:18 h:mm (IQR 1:18, Range 0:50-16:52) compared to 19:08 h:mm (IQR 5:49, Range 1:36-58:52) for samples submitted to the central laboratory. No quality control issues were identified in the LabVan. INTERPRETATION: The LabVan is an ISO15189 compliant testing facility fully operationalized for mobile point-of-care testing that significantly reduces TAT for result reporting, facilitating rapid public health actions. FUNDING: This work was supported by the Department of Health, Victoria State Government, Australia.

Genomics-informed responses in the elimination of COVID-19 in Victoria, Australia: an observational, genomic epidemiological study.

BACKGROUND: A cornerstone of Australia's ability to control COVID-19 has been effective border control with an extensive supervised quarantine programme. However, a rapid recrudescence of COVID-19 was observed in the state of Victoria in June, 2020. We aim to describe the genomic findings that located the source of this second wave and show the role of genomic epidemiology in the successful elimination of COVID-19 for a second time in Australia. METHODS: In this observational, genomic epidemiological study, we did genomic sequencing of all laboratory-confirmed cases of COVID-19 diagnosed in Victoria, Australia between Jan 25, 2020, and Jan 31, 2021. We did phylogenetic analyses, genomic cluster discovery, and integrated results with epidemiological data (detailed information on demographics, risk factors, and exposure) collected via interview by the Victorian Government Department of Health. Genomic transmission networks were used to group multiple genomic clusters when epidemiological and genomic data suggested they arose from a single importation event and diversified within Victoria. To identify transmission of emergent lineages between Victoria and other states or territories in Australia, all publicly available SARS-CoV-2 sequences uploaded before Feb 11, 2021, were obtained from the national sequence sharing programme AusTrakka, and epidemiological data were obtained from the submitting laboratories. We did phylodynamic analyses to estimate the growth rate, doubling time, and number of days from the first local infection to the collection of the first sequenced genome for the dominant local cluster, and compared our growth estimates to previously published estimates from a similar growth phase of lineage B.1.1.7 (also known as the Alpha variant) in the UK. FINDINGS: Between Jan 25, 2020, and Jan 31, 2021, there were 20 451 laboratory-confirmed cases of COVID-19 in Victoria, Australia, of which 15 431 were submitted for sequencing, and 11 711 met all quality control metrics and were included in our analysis. We identified 595 genomic clusters, with a median of five cases per cluster (IQR 2-11). Overall, samples from 11 503 (98·2%) of 11 711 cases clustered with another sample in Victoria, either within a genomic cluster or transmission network. Genomic analysis revealed that 10 426 cases, including 10 416 (98·4%) of 10 584 locally acquired cases, diagnosed during the second wave (between June and October, 2020) were derived from a single incursion from hotel quarantine, with the outbreak lineage (transmission network G, lineage D.2) rapidly detected in other Australian states and territories. Phylodynamic analyses indicated that the epidemic growth rate of the outbreak lineage in Victoria during the initial growth phase (samples collected between June 4 and July 9, 2020; 47·4 putative transmission events, per branch, per year [1/years; 95% credible interval 26·0-85·0]), was similar to that of other reported variants, such as B.1.1.7 in the UK (mean approximately 71·5 1/years). Strict interventions were implemented, and the outbreak lineage has not been detected in Australia since Oct 29, 2020. Subsequent cases represented independent international or interstate introductions, with limited local spread. INTERPRETATION: Our study highlights how rapid escalation of clonal outbreaks can occur from a single incursion. However, strict quarantine measures and decisive public health responses to emergent cases are effective, even with high epidemic growth rates. Real-time genomic surveillance can alter the way in which public health agencies view and respond to COVID-19 outbreaks. FUNDING: The Victorian Government, the National Health and Medical Research Council Australia, and the Medical Research Future Fund.

Accessible Platform for High-Throughput COVID-19 Molecular Diagnostics and Genome Sequencing Using a Repurposed 3D Printer for RNA Extraction.

The COVID-19 pandemic has exposed the dependence of diagnostic laboratories on a handful of large corporations with market monopolies on the worldwide supply of reagents, consumables, and hardware for molecular diagnostics. Global shortages of key consumables for RT-qPCR detection of SARS-CoV-2 RNA have impaired the ability to run essential, routine diagnostic services. Here, we describe a workflow for rapid detection of SARS-CoV-2 RNA in upper respiratory samples including nasal swabs and saliva, utilizing low-cost equipment and readily accessible reagents. Using repurposed Creality3D Ender-3 three-dimensional (3D) printers, we built a semiautomated paramagnetic bead RNA extraction platform. The hardware for the system was built for $300 USD, and the material cost per reaction was $1 USD. Named the Ender VX500, instrument performance when paired with RT-qPCR for SARS-CoV-2 detection in nasal and saliva specimens was two virus copies per microliter. There was a high-performance agreement (assessed using 458 COVID-19 nasal swab specimens) with the Aptima SARS-CoV-2 assay run on the Hologic Panther, a commercial automated RNA extraction and detection platform. Inter- and intrainstrument precision was excellent (coefficients of variation (CoV) of 1.10 and 0.66-1.32%, respectively) across four instruments. The platform is scalable with throughput ranging from 23 specimens on a single instrument run by one user in 50 min to 364 specimens on four instruments run by four users in 190 min. Step-by-step instructions and protocols for building and running the Ender VX500 have been made available without restriction.

Use of emerging testing technologies and approaches for SARS-CoV-2: review of literature and global experience in an Australian context.

Emerging testing technologies for detection of SARS-CoV-2 include those that are rapid and can be used at point-of-care (POC), and those facilitating high throughput laboratory-based testing. Tests designed to be performed at POC (such as antigen tests and molecular assays) have the potential to expedite isolation of infectious patients and their contacts, but most are less sensitive than standard-of-care reverse transcription polymerase chain reaction (RT-PCR). Data on clinical performance of the majority of emerging assays are limited with most evaluations performed on contrived or stored laboratory samples. Further evaluations of these assays are required, particularly when performed at POC on symptomatic and asymptomatic patients and at various time-points after symptom onset. A few studies have so far shown several of these assays have high specificity. However, large prospective evaluations are needed to confirm specificity, particularly before the assays are implemented in low prevalence settings or asymptomatic populations. High throughput laboratory-based testing includes the use of new sample types (e.g., saliva to increase acceptability) or innovative uses of existing technology (e.g., sample pooling). Information detailing population-wide testing strategies for SARS-COV-2 is largely missing from peer-reviewed literature. Logistics and supply chains are key considerations in any plan to 'scale up' testing in the Australian context. The strategic use of novel assays will help strike the balance between achieving adequate test numbers without overwhelming laboratory capacity. To protect testing of high-risk populations, the aims of testing with respect to the phase of the pandemic must be considered.

An implementation science approach to evaluating pathogen whole genome sequencing in public health.

BACKGROUND: Pathogen whole genome sequencing (WGS) is being incorporated into public health surveillance and disease control systems worldwide and has the potential to make significant contributions to infectious disease surveillance, outbreak investigation and infection prevention and control. However, to date, there are limited data regarding (i) the optimal models for integration of genomic data into epidemiological investigations and (ii) how to quantify and evaluate public health impacts resulting from genomic epidemiological investigations. METHODS: We developed the Pathogen Genomics in Public HeAlth Surveillance Evaluation (PG-PHASE) Framework to guide examination of the use of WGS in public health surveillance and disease control. We illustrate the use of this framework with three pathogens as case studies: Listeria monocytogenes, Mycobacterium tuberculosis and SARS-CoV-2. RESULTS: The framework utilises an adaptable whole-of-system approach towards understanding how interconnected elements in the public health application of pathogen genomics contribute to public health processes and outcomes. The three phases of the PG-PHASE Framework are designed to support understanding of WGS laboratory processes, analysis, reporting and data sharing, and how genomic data are utilised in public health practice across all stages, from the decision to send an isolate or sample for sequencing to the use of sequence data in public health surveillance, investigation and decision-making. Importantly, the phases can be used separately or in conjunction, depending on the need of the evaluator. Subsequent to conducting evaluation underpinned by the framework, avenues may be developed for strategic investment or interventions to improve utilisation of whole genome sequencing. CONCLUSIONS: Comprehensive evaluation is critical to support health departments, public health laboratories and other stakeholders to successfully incorporate microbial genomics into public health practice. The PG-PHASE Framework aims to assist public health laboratories, health departments and authorities who are either considering transitioning to whole genome sequencing or intending to assess the integration of WGS in public health practice, including the capacity to detect and respond to outbreaks and associated costs, challenges and facilitators in the utilisation of microbial genomics and public health impacts.

Multi-site assessment of rapid, point-of-care antigen testing for the diagnosis of SARS-CoV-2 infection in a low-prevalence setting: A validation and implementation study.

BACKGROUND: In Australia, COVID-19 diagnosis relies on RT-PCR testing which is relatively costly and time-consuming. To date, few studies have assessed the performance and implementation of rapid antigen-based SARS-CoV-2 testing in a setting with a low prevalence of COVID-19 infections, such as Australia. METHODS: This study recruited participants presenting for COVID-19 testing at three Melbourne metropolitan hospitals during a period of low COVID-19 prevalence. The Abbott PanBioTM COVID-19 Ag point-of-care test was performed alongside RT-PCR. In addition, participants with COVID-19 notified to the Victorian Government were invited to provide additional swabs to aid validation. Implementation challenges were also documented. FINDINGS: The specificity of the Abbott PanBioTM COVID-19 Ag test was 99.96% (95% CI 99.73 - 100%). Sensitivity amongst participants with RT-PCR-confirmed infection was dependent upon the duration of symptoms reported, ranging from 77.3% (duration 1 to 33 days) to 100% in those within seven days of symptom onset. A range of implementation challenges were identified which may inform future COVID-19 testing strategies in a low prevalence setting. INTERPRETATION: Given the high specificity, antigen-based tests may be most useful in rapidly triaging public health and hospital resources while expediting confirmatory RT-PCR testing. Considering the limitations in test sensitivity and the potential for rapid transmission in susceptible populations, particularly in hospital settings, careful consideration is required for implementation of antigen testing in a low prevalence setting. FUNDING: This work was funded by the Victorian Department of Health and Human Services. The funder was not involved in data analysis or manuscript preparation.

Tracking the COVID-19 pandemic in Australia using genomics.

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.

Pandemic printing: a novel 3D-printed swab for detecting SARS-CoV-2.

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.

Validation of a single-step, single-tube reverse transcription loop-mediated isothermal amplification assay for rapid detection of SARS-CoV-2 RNA.

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.

Evaluation of Serological Tests for SARS-CoV-2: Implications for Serology Testing in a Low-Prevalence Setting.

BACKGROUND: Robust serological assays are essential for long-term control of the COVID-19 pandemic. Many recently released point-of-care (PoCT) serological assays have been distributed with little premarket validation. METHODS: Performance characteristics for 5 PoCT lateral flow devices approved for use in Australia were compared to a commercial enzyme immunoassay (ELISA) and a recently described novel surrogate virus neutralization test (sVNT). RESULTS: Sensitivities for PoCT ranged from 51.8% (95% confidence interval [CI], 43.1%-60.4%) to 67.9% (95% CI, 59.4%-75.6%), and specificities from 95.6% (95% CI, 89.2%-98.8%) to 100.0% (95% CI, 96.1%-100.0%). ELISA sensitivity for IgA or IgG detection was 67.9% (95% CI, 59.4%-75.6%), increasing to 93.8% (95% CI, 85.0%-98.3%) for samples >14 days post symptom onset. sVNT sensitivity was 60.9% (95% CI, 53.2%-68.4%), rising to 91.2% (95% CI, 81.8%-96.7%) for samples >14 days post symptom onset, with specificity 94.4% (95% CI, 89.2%-97.5%). CONCLUSIONS: Performance characteristics for COVID-19 serological assays were generally lower than those reported by manufacturers. Timing of specimen collection relative to onset of illness or infection is crucial in reporting of performance characteristics for COVID-19 serological assays. The optimal algorithm for implementing serological testing for COVID-19 remains to be determined, particularly in low-prevalence settings.