Combining viral genomics and clinical data to assess risk factors for severe COVID-19 (mortality, ICU admission, or intubation) amongst hospital patients in a large acute UK NHS hospital Trust.

Throughout the COVID-19 pandemic, valuable datasets have been collected on the effects of the virus SARS-CoV-2. In this study, we combined whole genome sequencing data with clinical data (including clinical outcomes, demographics, comorbidity, treatment information) for 929 patient cases seen at a large UK hospital Trust between March 2020 and May 2021. We identified associations between acute physiological status and three measures of disease severity; admission to the intensive care unit (ICU), requirement for intubation, and mortality. Whilst the maximum National Early Warning Score (NEWS2) was moderately associated with severe COVID-19 (A = 0.48), the admission NEWS2 was only weakly associated (A = 0.17), suggesting it is ineffective as an early predictor of severity. Patient outcome was weakly associated with myriad factors linked to acute physiological status and human genetics, including age, sex and pre-existing conditions. Overall, we found no significant links between viral genomics and severe outcomes, but saw evidence that variant subtype may impact relative risk for certain sub-populations. Specific mutations of SARS-CoV-2 appear to have little impact on overall severity risk in these data, suggesting that emerging SARS-CoV-2 variants do not result in more severe patient outcomes. However, our results show that determining a causal relationship between mutations and severe COVID-19 in the viral genome is challenging. Whilst improved understanding of the evolution of SARS-CoV-2 has been achieved through genomics, few studies on how these evolutionary changes impact on clinical outcomes have been seen due to complexities associated with data linkage. By combining viral genomics with patient records in a large acute UK hospital, this study represents a significant resource for understanding risk factors associated with COVID-19 severity. However, further understanding will likely arise from studies of the role of host genetics on disease progression.

Nanopore Sequencing for Mixed Samples.

Long read Nanopore sequencing can be utilised to determine the quality and accuracy of genetically engineered changes in animals, which often produce heterogenous samples. The protocol presented in this chapter can be used for a range of both low and high throughput sequencing applications. DNA must be repaired, barcoded and ligated to sequencing adapters prior to sequencing. Quality of sequencing data produced is dependent on stringent adherence to the protocol. However, nanopore sequencing is a fast moving field, therefore it is worth considering using the most up to date chemistry available.

Multiple pathways of SARS-CoV-2 nosocomial transmission uncovered by integrated genomic and epidemiological analyses during the second wave of the COVID-19 pandemic in the UK.

Introduction: Throughout the global COVID-19 pandemic, nosocomial transmission has represented a major concern for healthcare settings and has accounted for many infections diagnosed within hospitals. As restrictions ease and novel variants continue to spread, it is important to uncover the specific pathways by which nosocomial outbreaks occur to understand the most suitable transmission control strategies for the future. Methods: In this investigation, SARS-CoV-2 genome sequences obtained from 694 healthcare workers and 1,181 patients were analyzed at a large acute NHS hospital in the UK between September 2020 and May 2021. These viral genomic data were combined with epidemiological data to uncover transmission routes within the hospital. We also investigated the effects of the introduction of the highly transmissible variant of concern (VOC), Alpha, over this period, as well as the effects of the national vaccination program on SARS-CoV-2 infection in the hospital. Results: Our results show that infections of all variants within the hospital increased as community prevalence of Alpha increased, resulting in several outbreaks and super-spreader events. Nosocomial infections were enriched amongst older and more vulnerable patients more likely to be in hospital for longer periods but had no impact on disease severity. Infections appeared to be transmitted most regularly from patient to patient and from patients to HCWs. In contrast, infections from HCWs to patients appeared rare, highlighting the benefits of PPE in infection control. The introduction of the vaccine at this time also reduced infections amongst HCWs by over four-times. Discussion: These analyses have highlighted the importance of control measures such as regular testing, rapid lateral flow testing alongside polymerase chain reaction (PCR) testing, isolation of positive patients in the emergency department (where possible), and physical distancing of patient beds on hospital wards to minimize nosocomial transmission of infectious diseases such as COVID-19.

SARS-CoV-2 lineage B.1.1.7 is associated with greater disease severity among hospitalised women but not men: multicentre cohort study.

BACKGROUND: SARS-CoV-2 lineage B.1.1.7 has been associated with an increased rate of transmission and disease severity among subjects testing positive in the community. Its impact on hospitalised patients is less well documented. METHODS: We collected viral sequences and clinical data of patients admitted with SARS-CoV-2 and hospital-onset COVID-19 infections (HOCIs), sampled 16 November 2020 to 10 January 2021, from eight hospitals participating in the COG-UK-HOCI study. Associations between the variant and the outcomes of all-cause mortality and intensive therapy unit (ITU) admission were evaluated using mixed effects Cox models adjusted by age, sex, comorbidities, care home residence, pregnancy and ethnicity. FINDINGS: Sequences were obtained from 2341 inpatients (HOCI cases=786) and analysis of clinical outcomes was carried out in 2147 inpatients with all data available. The HR for mortality of B.1.1.7 compared with other lineages was 1.01 (95% CI 0.79 to 1.28, p=0.94) and for ITU admission was 1.01 (95% CI 0.75 to 1.37, p=0.96). Analysis of sex-specific effects of B.1.1.7 identified increased risk of mortality (HR 1.30, 95% CI 0.95 to 1.78, p=0.096) and ITU admission (HR 1.82, 95% CI 1.15 to 2.90, p=0.011) in females infected with the variant but not males (mortality HR 0.82, 95% CI 0.61 to 1.10, p=0.177; ITU HR 0.74, 95% CI 0.52 to 1.04, p=0.086). INTERPRETATION: In common with smaller studies of patients hospitalised with SARS-CoV-2, we did not find an overall increase in mortality or ITU admission associated with B.1.1.7 compared with other lineages. However, women with B.1.1.7 may be at an increased risk of admission to intensive care and at modestly increased risk of mortality.

Estimation of ELISA results using a parallel curve analysis.

We introduce a new method for the analysis of enzyme-linked immunosorbent assay (ELISA) data. The new method can use data near the asymptotes and does not give undue weight to responses on the flatter parts of the dose-response curve. We apply it to simulated data and to two real-world assays and show it is more accurate and more precise than the traditional interpolation method. In particular, the new method works much better for very low-concentration samples for which the traditional method is often unable to give a result.

What makes a good new therapeutic L-asparaginase?

L-asparaginase is a critical part of the treatment of acute lymphoblastic leukaemia in children and adolescents, and has contributed to the improvement in patient outcomes over the last 40 years. The main products used in clinical treatment are L-asparaginase enzymes derived from Escherichia coli and Erwinia chrysanthemi. However, a very active area of research is the identification and characterisation of potential new L-asparaginase therapeutics, from existing or novel prokaryotic and eukaryotic sources, including mutations to improve function. In this review, we discuss the critical factors necessary to adequately characterise novel L-asparaginase therapeutic products, including enzyme kinetic parameters, glutaminase activity, and toxicity. One critical consideration is to ensure that the substrate affinity of novel enzymes, as measured by the Michaelis constant KM, is sufficiently low to enable efficient reaction rates in human clinical use. The activity of L-asparaginases towards glutamine as a substrate is discussed and reviewed in detail, as there is much debate in the scientific literature about the importance of this feature for therapeutic enzymes. The recent research in the area is reviewed, including identification of new sources of the enzyme, modulating glutaminase activity, and improving the thermal stability and immunogenic response. New research in the area may benefit from these considerations, to enable the next generation of therapeutic product design. Critical to future work in this area is a complete characterisation of novel enzymes with respect to performance for both L-asparagine and L-glutamine as substrates.