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Background Healthcare-associated (HCA) SARS-CoV-2 infection is a significant contributor to the spread of the 2020 pandemic. Timely review of HCA cases is essential to identify learning to inform infection prevention and control (IPC) policies and organisational response. Aim To identify key areas for improvement through rapid investigation of HCA SARS-CoV-2 cases and to implement change. Methods Cases were identified based on date of first positive SARS-CoV-2 PCR sample in relation to date of hospital admission. Cases were reviewed using a structured gap analysis tool to identify key learning points. These were discussed in weekly multidisciplinary meetings to gain consensus on learning outcomes, level of harm incurred by the patient and required actions. Learning was then promptly fed back to individual teams and the organisation. Findings Of the 489 SARS-CoV-2 cases admitted between 10th March and 23rd June 2020, 114 suspected HCA cases (23.3%) were reviewed; 58/489 (11.8%) were ultimately deemed to be HCA. Five themes were identified: individual patient vulnerability, communication, IPC implementation, policy issues and organisational response. Adaptations to policies based on these reviews were completed within the course of the initial phase of the pandemic. Conclusion This approach enabled timely learning and implementation of control measures and policy development.

BACKGROUND: There is currently no consensus on the diagnosis, definition, symptoms, or duration of COVID-19 illness. The diagnostic complexity of Long COVID is compounded in many patients who were or might have been infected with SARS-CoV-2 but not tested during the acute illness and/or are SARS-CoV-2 antibody negative. METHODS: Given the diagnostic conundrum of Long COVID, we set out to investigate SARS-CoV-2-specific T cell responses in patients with confirmed SARS-CoV-2 infection and/or Long COVID from a cohort of mostly non-hospitalised patients. FINDINGS: We discovered that IL-2 release (but not IFN-γ release) from T cells in response to SARS-CoV-2 peptides is both sensitive (75% +/-13%) and specific (88%+/-7%) for previous SARS-CoV-2 infection >6 months after a positive PCR test. We identified that 42-53% of patients with Long COVID, but without detectable SARS-CoV-2 antibodies, nonetheless have detectable SARS-CoV-2 specific T cell responses. INTERPRETATION: Our study reveals evidence (detectable T cell mediated IL-2 release) of previous SARS-CoV-2 infection in seronegative patients with Long COVID. FUNDING: This work was funded by the Addenbrooke's Charitable Trust (900276 to NS), NIHR award (G112259 to NS) and supported by the NIHR Cambridge Biomedical Research Centre. NJM is supported by the MRC (TSF MR/T032413/1) and NHSBT (WPA15-02). PJL is supported by the Wellcome Trust (PRF 210688/Z/18/Z, 084957/Z/08/Z), a Medical Research Council research grant MR/V011561/1 and the United Kingdom Research and a Innovation COVID Immunology Consortium grant (MR/V028448/1).

Summary There is urgent need for rapid SARS-CoV-2 testing in hospital to limit nosocomial spread. We report an evaluation of point of care (POC) nucleic acid amplification testing (NAAT) in 149 participants with parallel combined nasal/throat swabbing for POC versus standard lab RT-PCR testing. Median time to result is 2.6 (IQR 2.3 to 4.8) versus 26.4 hours (IQR 21.4 to 31.4, p, Graphical Abstract, Highlights • Point of care (POC) test highly sensitive and specific against RT-PCR • Time to result 2.6hrs for POC v 26.4hrs for standard lab RT-PCR. • SARS-CoV-2 POC test reduces median time-to-bed placement by approximately six hours • SARS-CoV-2 POC improves indices of hospital functioning and patient care, Collier et al report testing and implementation of an accurate SARS-CoV-2 point of care nucleic acid amplification test that leads to faster time to definitive bed placement from the ER, release of isolation rooms, avoidance of bed closures, and expedited discharges to care homes and access to hospital investigations/procedures.

Background HIV-1 does not encode a helicase and hijacks those of the cell for efficient replication. We and others previously showed that the DEAD box helicase, DDX5, is an essential HIV dependency factor. DDX5 was recently shown to be associated with the 7SK snRNP. Cellular positive transcription elongation factor b (P-TEFb) is bound in an inactive form with HEXIM1/2 on 7SK snRNP. The Tat/P-TEFb complex is essential for efficient processivity of Pol II in HIV-1 transcription elongation and Tat competes with HEXIM1/2 for P-TEFb. We investigated the precise role of DDX5 in HIV replication using siRNA mediated knockdown and rescue with DDX5 mutants which prevent protein–protein interactions and RNA and ATP binding. Results We demonstrate a critical role for DDX5 in the Tat/HEXIM1 interaction. DDX5 acts to potentiate Tat activity and can bind both Tat and HEXIM1 suggesting it may facilitate the dissociation of HEXIM1/2 from the 7SK-snRNP complex, enhancing Tat/P-TEFb availability. We show knockdown of DDX5 in a T cell line significantly reduces HIV-1 infectivity and viral protein production. This activity is unique to DDX5 and cannot be substituted by its close paralog DDX17. Overexpression of DDX5 stimulates the Tat/LTR promoter but suppresses other cellular and viral promoters. Individual mutations of conserved ATP binding, RNA binding, helicase related or protein binding motifs within DDX5 show that the N terminal RNA binding motifs, the Walker B and the glycine doublet motifs are essential for this function. The Walker A and RNA binding motifs situated on the transactivation domain are however dispensable. Conclusion DDX5 is an essential cellular factor for efficient HIV transcription elongation. It interacts with Tat and may potentiate the availability of P-TEFb through sequestering HEXIM1.

HIV splicing involves five splice donor and eight splice acceptor sequences which, together with cryptic splice sites, generate over 100 mRNA species. Ninety percent of both partially spliced and fully spliced transcripts utilize the intrinsically weak A4/A5 3' splice site cluster. We show that DDX17, but not its close paralog DDX5, specifically controls the usage of this splice acceptor group. In its absence, production of the viral envelope protein and other regulatory and accessory proteins is grossly reduced, while Vif, which uses the A1 splice acceptor, is unaffected. This is associated with a profound decrease in viral export from the cell. Loss of Vpu expression causing upregulation of cellular Tetherin compounds the phenotype. DDX17 utilizes distinct RNA binding motifs for its role in efficient HIV replication, and we identify RNA binding motifs essential for its role, while the Walker A, Walker B (DEAD), Q motif and the glycine doublet motif are all dispensable. We show that DDX17 interacts with SRSF1/SF2 and the heterodimeric auxiliary factor U2AF65/35, which are essential splicing factors in the generation of Rev and Env/Vpu transcripts.

HIV splicing involves five splice donor and eight splice acceptor sequences which, together with cryptic splice sites, generate over 100 mRNA species. Ninety percent of both partially spliced and fully spliced transcripts utilize the intrinsically weak A4/A5 3′ splice site cluster. We show that DDX17, but not its close paralog DDX5, specifically controls the usage of this splice acceptor group. In its absence, production of the viral envelope protein and other regulatory and accessory proteins is grossly reduced, while Vif, which uses the A1 splice acceptor, is unaffected. This is associated with a profound decrease in viral export from the cell. Loss of Vpu expression causing upregulation of cellular Tetherin compounds the phenotype. DDX17 utilizes distinct RNA binding motifs for its role in efficient HIV replication, and we identify RNA binding motifs essential for its role, while the Walker A, Walker B (DEAD), Q motif and the glycine doublet motif are all dispensable. We show that DDX17 interacts with SRSF1/SF2 and the heterodimeric auxiliary factor U2AF65/35, which are essential splicing factors in the generation of Rev and Env/Vpu transcripts., Graphical Abstract Unlabelled Image, Highlights • Regulation of HIV splicing is significantly dependent on host cellular factors. • DDX17 interacts with splicing factors and is essential for efficient HIV-1 splicing. • DDX17 utilizes distinct RNA/ATPase motifs and acts independently of DDX5. • DDX17 C-terminal domain is critical for the role of DDX17 in HIV-1 replication.