Quantifying the reduction of airborne infectious viral load using a ventilated patient hood.

BACKGROUND: Healthcare workers treating SARS-CoV-2 patients are at risk of infection by respiratory exposure to patient-emitted, virus-laden aerosols. Source control devices such as ventilated patient isolation hoods have been shown to limit the dissemination of non-infectious airborne particles in laboratory tests, but data on their performance in mitigating the airborne transmission risk of infectious viruses are lacking. AIM: We used an infectious airborne virus to quantify the ability of a ventilated hood to reduce infectious virus exposure in indoor environments. METHODS: We nebulized 109 plaque forming units (pfu) of bacteriophage PhiX174 virus into a ∼30-m3 room when the hood was active or inactive. The airborne concentration of infectious virus was measured by BioSpot-VIVAS and settle plates using plaque assay quantification on the bacterial host Escherichia coli C. The airborne particle number concentration (PNC) was also monitored continuously using an optical particle sizer. FINDINGS: The median airborne viral concentration in the room reached 1.41 × 105 pfu/m3 with the hood inactive. When active, the hood reduced infectious virus concentration in air samples by 374-fold. The deposition of infectious virus on the surface of settle plates was reduced by 87-fold. This was associated with a 109-fold reduction in total airborne particle number escape rate. CONCLUSION: A personal ventilation hood significantly reduced airborne particle escape, considerably lowering infectious virus contamination in an indoor environment. Our findings support the further development of source control devices to mitigate nosocomial infection risk among healthcare workers exposed to airborne viruses in clinical settings.

Core protocol for the adaptive Platform Trial In COVID-19 Vaccine priming and BOOsting (PICOBOO).

BACKGROUND: The need for coronavirus 2019 (COVID-19) vaccination in different age groups and populations is a subject of great uncertainty and an ongoing global debate. Critical knowledge gaps regarding COVID-19 vaccination include the duration of protection offered by different priming and booster vaccination regimens in different populations, including homologous or heterologous schedules; how vaccination impacts key elements of the immune system; how this is modified by prior or subsequent exposure to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and future variants; and how immune responses correlate with protection against infection and disease, including antibodies and effector and T cell central memory. METHODS: The Platform Trial In COVID-19 priming and BOOsting (PICOBOO) is a multi-site, multi-arm, Bayesian, adaptive, randomised controlled platform trial. PICOBOO will expeditiously generate and translate high-quality evidence of the immunogenicity, reactogenicity and cross-protection of different COVID-19 priming and booster vaccination strategies against SARS-CoV-2 and its variants/subvariants, specific to the Australian context. While the platform is designed to be vaccine agnostic, participants will be randomised to one of three vaccines at trial commencement, including Pfizer's Comirnaty, Moderna's Spikevax or Novavax's Nuvaxovid COVID-19 vaccine. The protocol structure specifying PICOBOO is modular and hierarchical. Here, we describe the Core Protocol, which outlines the trial processes applicable to all study participants included in the platform trial. DISCUSSION: PICOBOO is the first adaptive platform trial evaluating different COVID-19 priming and booster vaccination strategies in Australia, and one of the few established internationally, that is designed to generate high-quality evidence to inform immunisation practice and policy. The modular, hierarchical protocol structure is intended to standardise outcomes, endpoints, data collection and other study processes for nested substudies included in the trial platform and to minimise duplication. It is anticipated that this flexible trial structure will enable investigators to respond with agility to new research questions as they arise, such as the utility of new vaccines (such as bivalent, or SARS-CoV-2 variant-specific vaccines) as they become available for use. TRIAL REGISTRATION: Australian and New Zealand Clinical Trials Registry ACTRN12622000238774. Registered on 10 February 2022.

Vaccine Research and Development to Advance Pandemic and Seasonal Influenza Preparedness and Response: Lessons from COVID-19

The global response to COVID-19 has demonstrated the importance of vigilance and preparedness for infectious diseases, particularly influenza. There is a need for more effective influenza vaccines and modern manufacturing technologies that are adaptable and scalable to meet demand during a pandemic. The rapid development of COVID-19 vaccines has demonstrated what is possible with extensive data sharing, researchers who have the necessary resources and novel technologies to conduct and apply their research, rolling review by regulators, and public-private partnerships. As demonstrated throughout the response to COVID-19, the process of research and development of novel vaccines can be significantly optimized when stakeholders are provided with the resources and technologies needed to support their response. Vaccine Research and Development to Advance Pandemic and Seasonal Influenza Preparedness and Response focuses on how to leverage the knowledge gained from the COVID-19 pandemic to optimize vaccine research and development (R&D) to support the prevention and control of seasonal and pandemic influenza. The committee's findings address four dimensions of vaccine R&D: (1) basic and translational science, (2) clinical science, (3) manufacturing science, and (4) regulatory science. © 2022 by the National Academy of Sciences. All rights reserved.

Fibrin clot characteristics and anticoagulant response in a SARS-CoV-2 infected endothelial cell model

Background : SARS-CoV-2 infection is associated with risk of thrombosis however not all SARS-CoV-2 positive patients have equal risk of developing thrombi. Evidence of COVID-19 associated coagulation has not been demonstrated in children, whilst adults with a known vasculopathy, have been identified as high-risk. Currently the mechanism through which SARS-Cov-2 infection promotes thrombosis is unknown. Aims : We hypothesise that plasma factors interact with SARS-CoV-2 infected endothelium and play a significant role in this process. Methods : We developed a novel tool for modelling coagulation in vitro . Human Umbilical Vein Endothelial Cells (HUVECs) were infected with SARS-CoV-2;Uninfected cells and cells infected with Influenza A/Singapore/6/86 were used as controls. Wells were incubated with recalcified plasma from healthy children, healthy adults or vasculopathic adults (previous stroke) for 2 h. This experiment was repeated in the presence of anticoagulants: Bivalirudin (0.005 μg/μL), Defibrotide (0.02 μg/μL) and therapeutic and prophylactic doses of Low Molecular Weight Heparin (1 U/mL, 0.5 U/mL) and Unfractionated Heparin (0.7 U/mL, 0.4 U/mL). Fibrin clots were studied using scanning electron microscopy and fibrin characteristics were quantified manually using ImageJ software by two blinded reviewers. Results : Fibrin strands from child plasma had the largest thickness (0.32 μm), followed by adult plasma (0.30 μm) and finally vasculopath plasma (0.27 μm). Defibrotide and Bivalirudin had no effect, whilst Low Molecular Weight Heparin and Unfractionated Heparin completely eliminated clot formation at therapeutic and prophylactic doses. Conclusions : We demonstrate that SARS-CoV-2 infection of endothelial cells promotes thrombosis differently in plasma samples from children, adults and vasculopaths. Thinner fibrin strands in vasculopathy suggests formation of clots that are more resistant to lysis. Our results support that children are resistant to COVID-19 associated coagulation, and that adults with vasculopathies are an at-risk population. This novel cell-culture model for visualising coagulation is a valuable step towards incorporating the endothelium into haemostatic research.

Maturation-enhanced proximal tubules enable functionality, toxicity screening, and infectious disease modeling in kidney organoids

Background: The highly specialised proximal tubule (PT) nephron segment is responsible for most kidney functions in mammals and is acutely vulnerable to disease, making it a key objective for toxicity screening and disease research. While induced pluripotent stem cell (iPSC)-derived kidney organoids represent a promising approach, the PT remains immature with limited evidence of functional transporters. Here we report the development of PT-enhanced organoids with nephron functionality, enabling improved modelling of PT-relevant conditions including drug-induced toxicity and infection such as SARS-CoV-2. Methods: Standard and fluorescent reporter iPSC lines were subjected to prolonged monolayer differentiations (modified from: Howden et al. EMBO Rep 2019;Vanslambrouck et al. JASN 2019) and precisely-timed morphogens for targets such as WNT, BMP and NOTCH pathways prior to organoid generation (Takasato et al,. Nat Protoc 2016). Maturation was analysed via immunofluorescence, live confocal imaging of fluorescent reporters, transcriptional profiling, transporter function assays, and SARSCoV-2 infectivity. Results: Prolonged nephron progenitor differentiation with simultaneous prevention of spontaneous nephrogenesis resulted in PT-enhanced kidney organoids with elongated and aligned nephrons. Striking proximo-distal nephron orientation resulted from localised WNT antagonism. Improved upregulation of PT-specific markers compared to standard organoids was strengthened by evidence of transporter functionality, including uptake of albumin, organic cations, and cisplatin (eliciting appropriate KIM1 upregulation). This approach also improved expression of SARS-CoV-2 entry factors, confirmed by susceptibility to infection and viral replication. Conclusions: We describe enhanced kidney organoids with improved PT maturity arising from alterations to early mesodermal patterning and delayed nephron initiation. The enhanced conditions also provided more stringent control over nephron spatial arrangement. PT-enhanced organoids provide an ideal model to better understand human PT maturation, inherited and acquired PT disease, and drug toxicity implications.

Longitudinal evaluation of laboratory-based serological assays for SARS-CoV-2 antibody detection.

Serological assays for SARS-CoV-2 infection are now widely available for use in diagnostic laboratories. Limited data are available on the performance characteristics in different settings, and at time periods remote from the initial infection. Validation of the Abbott (Architect SARS-CoV-2 IgG), DiaSorin (Liaison SARS-CoV-2 S1/S2 IgG) and Roche (Cobas Elecsys Anti-SARS-CoV-2) assays was undertaken utilising 217 serum samples from 131 participants up to 7 months following COVID-19 infection. The Abbott and DiaSorin assays were implemented into routine laboratory workflow, with outcomes reported for 2764 clinical specimens. Sensitivity and specificity were concordant with the range reported by the manufacturers for all assays. Sensitivity across the convalescent period was highest for the Roche at 95.2-100% (95% CI 81.0-100%), then the DiaSorin at 88.1-100% (95% CI 76.0-100%), followed by the Abbott 68.2-100% (95% CI 53.4-100%). Sensitivity of the Abbott assay fell from approximately 5 months; on this assay paired serum samples for 45 participants showed a significant drop in the signal-to-cut-off ratio and 10 sero-reversion events. When used in clinical practice, all samples testing positive by both DiaSorin and Abbott assays were confirmed as true positive results. In this low prevalence setting, despite high laboratory specificity, the positive predictive value of a single positive assay was low. Comprehensive validation of serological assays is necessary to determine the optimal assay for each diagnostic setting. In this low prevalence setting we found implementation of two assays with different antibody targets maximised sensitivity and specificity, with confirmatory testing necessary for any sample which was positive in only one assay.

The threat of zoonotic coronaviruses

Since 2002, three zoonotic coronaviruses (CoV), SARS-CoV, MERS-CoV and SARS-CoV-2 have emerged in humans, establishing that emergence of coronaviruses from animal reservoirs represents a significant pandemic threat. SARS-CoV and MERS-CoV led to smaller epidemics with very high case fatality rates while SARS-CoV-2 resulted in a global pandemic. These zoonotic coronaviruses have their likely origins in bat species and they transmit to humans through intermediate hosts. Coronaviruses can occasionally jump between host species due to their high rate of recombination. Pandemic preparedness requires surveillance in animals and occupationally exposed humans and prevention and treatment strategies that have broad activity against coronaviruses.

Symptoms and laboratory manifestations of mild COVID-19 in a repatriated cruise ship cohort.

Much of our current understanding about novel coronavirus disease 2019 (COVID-19) comes from hospitalised patients. However, the spectrum of mild and subclinical disease has implications for population-level screening and control. Forty-nine participants were recruited from a group of 99 adults repatriated from a cruise ship with a high incidence of COVID-19. Respiratory and rectal swabs were tested by polymerase chain reaction (PCR) for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Sera were tested for anti-SARS-CoV-2 antibodies by enzyme-linked immunosorbent assay (ELISA) and microneutralisation assay. Symptoms, viral shedding and antibody response were examined. Forty-five participants (92%) were considered cases based on either positive PCR or positive ELISA for immunoglobulin G. Forty-two percent of cases were asymptomatic. Only 15% of symptomatic cases reported fever. Serial respiratory and rectal swabs were positive for 10% and 5% of participants respectively about 3 weeks after median symptom onset. Cycle threshold values were high (range 31-45). Attempts to isolate live virus were unsuccessful. The presence of symptoms was not associated with demographics, comorbidities or antibody response. In closed settings, incidence of COVID-19 could be almost double that suggested by symptom-based screening. Serology may be useful in diagnosis of mild disease and in aiding public health investigations.