LINE1-Mediated Reverse Transcription and Genomic Integration of SARS-CoV-2 mRNA Detected in Virus-Infected but Not in Viral mRNA-Transfected Cells.

SARS-CoV-2 sequences can be reverse-transcribed and integrated into the genomes of virus-infected cells by a LINE1-mediated retrotransposition mechanism. Whole-genome sequencing (WGS) methods detected retrotransposed SARS-CoV-2 subgenomic sequences in virus-infected cells overexpressing LINE1, while an enrichment method (TagMap) identified retrotranspositions in cells that did not overexpress LINE1. LINE1 overexpression increased retrotranspositions about 1000-fold as compared to non-overexpressing cells. Nanopore WGS can directly recover retrotransposed viral and flanking host sequences, but its sensitivity depends on the depth of sequencing (a typical 20-fold sequencing depth would only examine 10 diploid cell equivalents). In contrast, TagMap enriches the host-virus junctions and can interrogate up to 20,000 cells and is able to detect rare viral retrotranspositions in LINE1 non-overexpressing cells. Although Nanopore WGS is 10-20-fold more sensitive per tested cell, TagMap can interrogate 1000-2000-fold more cells and, therefore, can identify infrequent retrotranspositions. When comparing SARS-CoV-2 infection and viral nucleocapsid mRNA transfection by TagMap, retrotransposed SARS-CoV-2 sequences were only detected in infected but not in transfected cells. Retrotransposition in virus-infected cells, in contrast to transfected cells, may be facilitated because virus infection, in contrast to viral RNA transfection, results in significantly higher viral RNA levels and stimulates LINE1 expression by causing cellular stress.

Procalcitonin to Guide Antibacterial Prescribing in Patients Hospitalised with COVID-19.

Antibacterial prescribing in patients presenting with COVID-19 remains discordant to rates of bacterial co-infection. Implementing diagnostic tests to exclude bacterial infection may aid reduction in antibacterial prescribing. (1) Method: A retrospective observational analysis was undertaken of all hospitalised patients with COVID-19 across a single-site NHS acute Trust (London, UK) from 1 December 2020 to 28 February 2021. Electronic patient records were used to identify patients, clinical data, and outcomes. Procalcitonin (PCT) serum assays, where available on admission, were analysed against electronic prescribing records for antibacterial prescribing to determine relationships with a negative PCT result (<25 mg/L) and antibacterial course length. (2) Results: Antibacterial agents were initiated on admission in 310/624 (49.7%) of patients presenting with COVID-19. A total of 33/74 (44.5%) patients with a negative PCT on admission had their treatment stopped within 24 h. A total of 6/49 (12.2%) patients were started on antibacterials, but a positive PCT saw their treatment stopped. Microbiologically confirmed bacterial infection was low (19/594; 3.2%) and no correlation was seen between PCT and culture positivity (p = 1). Lower mortality (15.6% vs. 31.4%; p = 0.049), length of hospital stay (7.9 days vs. 10.1 days; p = 0.044), and intensive care unit (ICU) admission (13.9% vs. 40.8%; p = 0.001) was noted among patients with low PCT. (3) Conclusions: This retrospective analysis of community acquired COVID-19 patients demonstrates the potential role of PCT in excluding bacterial co-infection. A negative PCT on admission correlates with shorter antimicrobial courses, early cessation of therapy, and predicts lower frequency of ICU admission. Low PCT may support decision making in cessation of antibacterials at the 48-72 h review.

A 24-Year-Old Man With Hemoptysis Found to Have a Chest Mass and Contralateral Axillary Lymphadenopathy.

CASE PRESENTATION: A 24-year-old man, never smoker, with no medical or surgical history, not currently on medications, presented to the ED with a second episode of gross hemoptysis, 4 months after an initial episode that had not previously been evaluated. He described the current episode of hemoptysis as "enough to fill the sink"; however, he did not further quantify. He has no history of recurrent epistaxis, hematemesis, or other evidence of clotting disorder. He denied any fevers, chills, night sweats, or recent travel. He denied any sick contacts and has no history of TB exposure or risk factors. The patient denied any shortness of breath, wheezing, or chest pain. He had no lower extremity pain or swelling. He routinely exercises and generally lives a healthy lifestyle. He is a health care worker who has not routinely worked with patients infected with SARS-CoV-2, although he received his second (of two) COVID-19 vaccines 4 days before presentation.

Bacteraemia variation during the COVID-19 pandemic; a multi-centre UK secondary care ecological analysis.

BACKGROUND: We investigated for change in blood stream infections (BSI) with Enterobacterales, coagulase negative staphylococci (CoNS), Streptococcus pneumoniae, and Staphylococcus aureus during the first UK wave of SARS-CoV-2 across five London hospitals. METHODS: A retrospective multicentre ecological analysis was undertaken evaluating all blood cultures taken from adults from 01 April 2017 to 30 April 2020 across five acute hospitals in London. Linear trend analysis and ARIMA models allowing for seasonality were used to look for significant variation. RESULTS: One hundred nineteen thousand five hundred eighty-four blood cultures were included. At the height of the UK SARS-CoV-2 first wave in April 2020, Enterobacterales bacteraemias were at an historic low across two London trusts (63/3814, 1.65%), whilst all CoNS BSI were at an historic high (173/3814, 4.25%). This differed significantly for both Enterobacterales (p = 0.013), CoNS central line associated BSIs (CLABSI) (p < 0.01) and CoNS non-CLABSI (p < 0.01), when compared with prior periods, even allowing for seasonal variation. S. pneumoniae (p = 0.631) and S. aureus (p = 0.617) BSI did not vary significant throughout the study period. CONCLUSIONS: Significantly fewer than expected Enterobacterales BSI occurred during the UK peak of the COVID-19 pandemic; identifying potential causes, including potential unintended consequences of national self-isolation public health messaging, is essential. High rates of CoNS BSI, with evidence of increased CLABSI, but also likely contamination associated with increased use of personal protective equipment, may result in inappropriate antimicrobial use and indicates a clear area for intervention during further waves.

Reverse-transcribed SARS-CoV-2 RNA can integrate into the genome of cultured human cells and can be expressed in patient-derived tissues.

Prolonged detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA and recurrence of PCR-positive tests have been widely reported in patients after recovery from COVID-19, but some of these patients do not appear to shed infectious virus. We investigated the possibility that SARS-CoV-2 RNAs can be reverse-transcribed and integrated into the DNA of human cells in culture and that transcription of the integrated sequences might account for some of the positive PCR tests seen in patients. In support of this hypothesis, we found that DNA copies of SARS-CoV-2 sequences can be integrated into the genome of infected human cells. We found target site duplications flanking the viral sequences and consensus LINE1 endonuclease recognition sequences at the integration sites, consistent with a LINE1 retrotransposon-mediated, target-primed reverse transcription and retroposition mechanism. We also found, in some patient-derived tissues, evidence suggesting that a large fraction of the viral sequences is transcribed from integrated DNA copies of viral sequences, generating viral-host chimeric transcripts. The integration and transcription of viral sequences may thus contribute to the detection of viral RNA by PCR in patients after infection and clinical recovery. Because we have detected only subgenomic sequences derived mainly from the 3' end of the viral genome integrated into the DNA of the host cell, infectious virus cannot be produced from the integrated subgenomic SARS-CoV-2 sequences.

Assessing the Impact of COVID-19 on Antimicrobial Stewardship Activities/Programs in the United Kingdom.

Since first identified in late 2019, the acute respiratory syndrome coronavirus (SARS-CoV2) and the resulting coronavirus disease (COVID-19) pandemic has overwhelmed healthcare systems worldwide, often diverting key resources in a bid to meet unprecedented challenges. To measure its impact on national antimicrobial stewardship (AMS) activities, a questionnaire was designed and disseminated to antimicrobialstewardship leads in the United Kingdom (UK). Most respondents reported a reduction in AMS activity with 64% (61/95) reporting that COVID-19 had a negative impact on routine AMS activities. Activities reported to have been negatively affected by the pandemic include audit, quality improvement initiatives, education, AMS meetings, and multidisciplinary working including ward rounds. However, positive outcomes were also identified, with technology being increasingly used as a tool to facilitate stewardship e.g., virtual meetings and ward rounds and increased the acceptance of using procalcitonin tests to distinguish between viral and bacterial infections. The COVID-19 pandemic has had a significant impact on the AMS activities undertaken across the UK. The long-term impact of the reduced AMS activities on incidence of AMR are not yet known. The legacy of innovation, use of technology, and increased collaboration from the pandemic could strengthen AMS in the post-pandemic era and presents opportunities for further development of AMS.

Presentation, Treatment Response and Short-Term Outcomes in Paediatric Multisystem Inflammatory Syndrome Temporally Associated with SARS-CoV-2 (PIMS-TS).

The novel Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is the pathogen responsible for Coronavirus Disease 2019 (COVID-19). Whilst most children and young people develop mild symptoms, recent reports suggest a novel paediatric inflammatory multisystem syndrome temporally associated with SARS-CoV-2 (PIMS-TS). Case definition and classification are preliminary, treatment is empiric and disease-associated outcomes are unclear. Here, we report 29 patients with PIMS-TS who were diagnosed, admitted and treated in the English North West between March and June 2020. Consistent with patterns observed internationally, cases peaked approximately 4 weeks after the initial surge of COVID-19-like symptoms in the UK population. Clinical symptoms included fever (100%), skin rashes (72%), cardiovascular involvement (86%), conjunctivitis (62%) and respiratory involvement (21%). Some patients had clinical features partially resembling Kawasaki disease (KD), toxic shock syndrome and cytokine storm syndrome. Male gender (69%), black, Asian and other minority ethnicities (BAME, 59%) were over-represented. Immune modulating treatment was used in all, including intravenous immunoglobulin (IVIG), corticosteroids and cytokine blockers. Notably, 32% of patients treated with IVIG alone went into remission. The rest required additional treatment, usually corticosteroids, with the exception of two patients who were treated with TNF inhibition and IL-1 blockade, respectively. Another patient received IL-1 inhibition as primary therapy, with associated rapid and sustained remission. Randomized and prospective studies are needed to investigate efficacy and safety of treatment, especially as resources of IVIG may be depleted secondary to high demand during future waves of COVID-19.

Investigating the association between ethnicity and health outcomes in SARS-CoV-2 in a London secondary care population.

BACKGROUND: Black, Asian and minority ethnic (BAME) populations are emerging as a vulnerable group in the severe acute respiratory syndrome coronavirus disease (SARS-CoV-2) pandemic. We investigated the relationship between ethnicity and health outcomes in SARS-CoV-2. METHODS AND FINDINGS: We conducted a retrospective, observational analysis of SARS-CoV-2 patients across two London teaching hospitals during March 1 -April 30, 2020. Routinely collected clinical data were extracted and analysed for 645 patients who met the study inclusion criteria. Within this hospitalised cohort, the BAME population were younger relative to the white population (61.70 years, 95% CI 59.70-63.73 versus 69.3 years, 95% CI 67.17-71.43, p<0.001). When adjusted for age, sex and comorbidity, ethnicity was not a predictor for ICU admission. The mean age at death was lower in the BAME population compared to the white population (71.44 years, 95% CI 69.90-72.90 versus, 77.40 years, 95% CI 76.1-78.70 respectively, p<0.001). When adjusted for age, sex and comorbidities, Asian patients had higher odds of death (OR 1.99: 95% CI 1.22-3.25, p<0.006). CONCLUSIONS: BAME patients were more likely to be admitted younger, and to die at a younger age with SARS-CoV-2. Within the BAME cohort, Asian patients were more likely to die but despite this, there was no difference in rates of admission to ICU. The reasons for these disparities are not fully understood and need to be addressed. Investigating ethnicity as a clinical risk factor remains a high public health priority. Studies that consider ethnicity as part of the wider socio-cultural determinant of health are urgently needed.