Revisiting the origins of the Sobemovirus genus: A case for ancient origins of plant viruses.

The discrepancy between short- and long-term rate estimates, known as the time-dependent rate phenomenon (TDRP), poses a challenge to extrapolating evolutionary rates over time and reconstructing evolutionary history of viruses. The TDRP reveals a decline in evolutionary rate estimates with the measurement timescale, explained empirically by a power-law rate decay, notably observed in animal and human viruses. A mechanistic evolutionary model, the Prisoner of War (PoW) model, has been proposed to address TDRP in viruses. Although TDRP has been studied in animal viruses, its impact on plant virus evolutionary history remains largely unexplored. Here, we investigated the consequences of TDRP in plant viruses by applying the PoW model to reconstruct the evolutionary history of sobemoviruses, plant pathogens with significant importance due to their impact on agriculture and plant health. Our analysis showed that the Sobemovirus genus dates back over four million years, indicating an ancient origin. We found evidence that supports deep host jumps to Poaceae, Fabaceae, and Solanaceae occurring between tens to hundreds of thousand years ago, followed by specialization. Remarkably, the TDRP-corrected evolutionary history of sobemoviruses was extended far beyond previous estimates that had suggested their emergence nearly 9,000 years ago, a time coinciding with the Neolithic period in the Near East. By incorporating sequences collected through metagenomic analyses, the resulting phylogenetic tree showcases increased genetic diversity, reflecting a deep history of sobemovirus species. We identified major radiation events beginning between 4,600 to 2,000 years ago, which aligns with the Neolithic period in various regions, suggesting a period of rapid diversification from then to the present. Our findings make a case for the possibility of deep evolutionary origins of plant viruses.

Acute assessment of spinal cord injury in New South Wales: A retrospective study of current practice in two spinal cord injury referral centers.

INTRODUCTION: Interventions provided in the early phases after spinal cord injury (SCI) may improve neurological recovery and provide for best possible functional outcomes. Knowing this relies on early and clear documentation of the level and grade of the spinal cord injury. Guidelines advocate for early documentation of neurological status within 72 h of injury to allow early prognostication and to help guide initial management. It is unclear whether this is current practice in New South Wales (NSW). METHODS: Patients with acute SCI who were admitted to two SCI referral centers during 2018-2019 in NSW were included. Data relating to documentation of neurological status, timing of imaging, surgery and transfer to spinal cord injury center were collected and summarized using descriptive statistics. RESULTS: Only 18 percent of patients had an acceptable neurological examination according to the International Standards for Classification of Spinal Cord Injury (ISNCSCI) within 72 h of injury (either not done, or unable to determine the neurological level of injury). At the first neurological examination, the neurological level of injury and grade was unable to be determined in 26.8% of patients and 29.9% of patients respectively. At discharge from acute care and transfer to rehabilitation, the neurological level was undetermined in 28.9% of patients and grade undetermined in 26.8%. ISNCSCI examination was most commonly performed by spinal rehabilitation doctors after patients were discharged from the intensive care unit (ICU). CONCLUSIONS: Documentation of neurological level and grade of SCI within 72 h of injury is not being performed in the large majority of this cohort, which may impede evaluation of neurological improvement in response to acute treatment, and hinder prognostication.

COVID-19-associated pulmonary aspergillosis in mechanically ventilated patients: a prospective, multicentre UK study.

BACKGROUND: Invasive pulmonary aspergillosis is a complication of severe COVID-19, with regional variation in reported incidence and mortality. We describe the incidence, risk factors and mortality associated with COVID-19-associated pulmonary aspergillosis (CAPA) in a prospective, multicentre UK cohort. METHODS: From March 2020 to March 2021, 266 mechanically ventilated adults with COVID-19 were enrolled across 5 UK hospital intensive care units (ICUs). CAPA was defined using European Confederation for Medical Mycology and the International Society for Human and Animal Mycology criteria and fungal diagnostics performed on respiratory and serum samples. RESULTS: Twenty-nine of 266 patients (10.9%) had probable CAPA, 14 (5.2%) possible CAPA and none proven CAPA. Probable CAPA was diagnosed a median of 9 (IQR 7-16) days after ICU admission. Factors associated with probable CAPA after multivariable logistic regression were cumulative steroid dose given within 28 days prior to ICU admission (adjusted OR (aOR) 1.16; 95% CI 1.01 to 1.43 per 100 mg prednisolone-equivalent), receipt of an interleukin (IL)-6 inhibitor (aOR 2.79; 95% CI 1.22 to 6.48) and chronic obstructive pulmonary disease (COPD) (aOR 4.78; 95% CI 1.13 to 18.13). Mortality in patients with probable CAPA was 55%, vs 46% in those without. After adjustment for immortal time bias, CAPA was associated with an increased risk of 90-day mortality (HR 1.85; 95% CI 1.07 to 3.19); however, this association did not remain statistically significant after further adjustment for confounders (adjusted HR 1.57; 95% CI 0.88 to 2.80). There was no difference in mortality between patients with CAPA prescribed antifungals (9 of 17; 53%) and those who were not (7 of 12; 58%) (p=0.77). INTERPRETATION: In this first prospective UK study, probable CAPA was associated with corticosteroid use, receipt of IL-6 inhibitors and pre-existing COPD. CAPA did not impact mortality following adjustment for prognostic variables.

Not all cholecystitis is created equal: Disparities contributing to ED presentation and failure of the outpatient algorithm.

BACKGROUND: The majority of final surgical pathology (FSP) from both emergency department (ED) and outpatient clinic (OC) patients is chronic cholecystitis. We aimed to differentiate these presentations and identify disparities associated with ED utilization and OC failure. METHODS: Retrospective chart review of single institution ED and OC cholecystectomies for cholelithiasis. Clinical presentation, FSP, demographics, and zip code poverty (ZCP) levels were evaluated. Data analysis by summary statistics, bivariate comparisons, and logistic regression. RESULTS: Of 299 OC and 308 ED patients, OC was more likely to be Caucasian (78% vs 46%, p < 0.0001) and insured (89% vs. 32%, p < 0.0001). 71.8% of OC with ZCP <10% had insurance versus only 32.5% in ZCP >20%. Uninsured ED OR was 13.1 (95% CI 8.7-22.9). CONCLUSION: Uninsured ED patients are vulnerable to fail the outpatient algorithm, especially when misdiagnosed by US. Clinical diagnosis of cholecystitis in this population should warrant offering of urgent cholecystectomy.

Polyvalent immunization elicits a synergistic broadly neutralizing immune response to hypervariable region 1 variants of hepatitis C virus.

A hepatitis C virus (HCV) vaccine is urgently needed. Vaccine development has been hindered by HCV's genetic diversity, particularly within the immunodominant hypervariable region 1 (HVR1). Here, we developed a strategy to elicit broadly neutralizing antibodies to HVR1, which had previously been considered infeasible. We first applied a unique information theory-based measure of genetic distance to evaluate phenotypic relatedness between HVR1 variants. These distances were used to model the structure of HVR1's sequence space, which was found to have five major clusters. Variants from each cluster were used to immunize mice individually, and as a pentavalent mixture. Sera obtained following immunization neutralized every variant in a diverse HCVpp panel (n = 10), including those resistant to monovalent immunization, and at higher mean titers (1/ID50 = 435) than a glycoprotein E2 (1/ID50 = 205) vaccine. This synergistic immune response offers a unique approach to overcoming antigenic variability and may be applicable to other highly mutable viruses.

Anecdotes Drive Attitudes, Data Drives Decisions: Optimizing the Emergency Department Workup Prior to Surgical Consultations.

PURPOSE: Emergency department (ED) wait times and ED length of stay (LOS) have a significant impact on patient morbidity and mortality and patient satisfaction. Consultation-to-decision time can contribute to increased wait times and LOS in the ED. Up to 40% of patients presenting to the ED require consultation from subspecialty services. We hypothesize that in surgical patients, completion of workup prior to consultation will decrease consultation-to-decision time, ED wait times, and LOS in the ED. METHODS: A retrospective review was conducted at a single site including all overnight general surgery consultations from the ED over 2-months. Data collected included wait times, LOS, and workup completed prior to consultation. Summary statistics were calculated and bivariate tests were performed using t-tests for continuous variables. RESULTS: Time to final surgical plan and LOS in ED were evaluated for 137 patients comparing "complete" and "incomplete" workups at time of consultation. It was considered a "complete" workup if labs and imaging were resulted prior to time of consult. If any baseline tests were not ordered prior to time of consult, it was considered an "incomplete" workup. Analysis demonstrated an average time of 4.9 and 2.5 hours for consultation-to-decision time for "incomplete" and "complete" workups respectively (p < 0.0001). For LOS in ED, there was an average of 11.4 and 7.9 hours for "incomplete" and "complete" workups respectively (p < 0.0001). CONCLUSIONS: There is a significant difference in consultation-to-decision time and LOS in the ED when consultation is performed following a complete versus incomplete workup for surgical patients independent of inherent wait times for testing to result and the need for additional testing requested from consulting services. Developing strategies to optimize workups from the ED prior to surgical consultation, including the development of care pathways, could significantly decrease patient wait times and LOS in the ED.

Arbovirus circulation, epidemiology and spatiotemporal distribution in Uganda.

Background: Arboviruses are endemic in Uganda; however, little is known about their epidemiology, seasonality and spatiotemporal distribution. Our study sought to provide information on arbovirus outbreaks from acute clinical presentations. Methods: Immunoglobulin M (IgM) and confirmatory Plaque Reduction Neutralisation Test (PRNT) results for arbovirus diagnosis of samples collected from patients attending sentinel sites from 2016-19 were analysed retrospectively. Demographic data were analysed with SaTScan and SPSS software to determine the epidemiology and spatiotemporal distribution of arboviruses. Results: Arbovirus activity peaked consistently during March-May rainy seasons. Overall, arbovirus seroprevalence was 9.5%. Of 137 IgM positives, 52.6% were confirmed by PRNT, of which 73.6% cases were observed in central Uganda with Yellow Fever Virus had the highest prevalence (27.8%). The 5-14 age group were four times more likely to be infected with an arbovirus p=0.003, 4.1 (95% CI 1.3-12.3). Significant arboviral activity was observed among outdoor workers(p=0.05) . Spatiotemporal analysis indicated arboviral activity in 23 of the 85 districts analysed.. Interpretation: Our study shows that arbovirus activity peaks during the March-May rainy season and highlights the need for YFV mass vaccination to reduce the clinical burden of arboviruses transmitted within the region.

Scavenger receptor class B type I genetic variants associated with disease severity in chronic hepatitis C virus infection.

Analysis of host genetic polymorphisms is an increasingly important tool for understanding and predicting pathogenesis and treatment response of viral diseases. The gene locus of scavenger receptor class B type I (SR-BI), encoding a cell entry factor and receptor for hepatitis C virus (HCV), contains several genetic polymorphisms. We applied a probe extension assay to determine the frequency of six single nucleotide polymorphisms (SNPs) within the SR-BI gene locus in 374 individuals with history of HCV infection. In addition, SR-BI messenger RNA (mRNA) levels were analyzed in liver biopsy specimens of chronically infected HCV subjects. The rs5888 variant allele T was present at a higher frequency in subjects with advanced fibrosis (χ2 , p = 0.016) and after adjusting for age, duration of infection and alcohol intake as confounding factors. Haplotype analysis of SNP frequencies showed that a haplotype consisting of rs61932577 variant allele C and rs5888 variant allele T was associated with an increased risk of advanced liver fibrosis (defined by an Ishak score 4-6) (adjusted odds ratio 2.81; 95% confidence interval 1.06-7.46. p = 0.038). Carriers of the rs5888 variant allele T displayed reduced SR-BI mRNA expression in liver biopsy specimens. In conclusion the rs5888 polymorphism variant is associated with decreased SR-BI expression and an increased risk of development of advanced fibrosis in chronic HCV infection. These findings provide further evidence for a role of SR-BI in HCV pathogenesis and provides a genetic marker for prediction of those infected individuals at greater risk of developing severe disease.

Hepatitis C subtyping assay failure in UK patients born in sub-Saharan Africa: Implications for global treatment and elimination.

BACKGROUND AND AIMS: The newly developed direct-acting antivirals have revolutionized the treatment of chronic hepatitis C virus (HCV), with cure rates as high as 98% in some cohorts. Although genome sequencing has demonstrated that some subtypes of HCV naturally harbor drug resistance associated substitutions (RAS), these are often overlooked as "rarities." Furthermore, commercial subtyping assays and associated epidemiological findings are skewed towards Western cohorts and whole-genome sequencing can be problematic to deploy without significant infrastructure and training support. We thus aimed to develop a simple, robust and accurate HCV subtyping pipeline, to optimize and streamline molecular detection and sequence-based typing of diverse RAS-containing subtypes. METHODS: HCV serum derived from 146 individuals, whose likely source of infection was from sub-Saharan Africa (SSA) was investigated with a novel panel of single round polymerase chain reaction (PCR) assays targeting NS5B and NS5A genomic regions. Virus subtype assignments were determined by pairwise-distance analysis and compared to both diagnostic laboratory assignments and free-to-use online typing tools. RESULTS: Partial NS5A and NS5B sequences were respectively obtained from 131 to 135 HCV-positive patients born in 19 different countries from SSA but attending clinics in the UK. We determined that routine clinical diagnostic methods incorrectly subtyped 59.0% of samples, with a further 6.8% incorrectly genotyped. Of five commonly used online tools, Geno2Pheno performed most effectively in determining a subtype in agreement with pairwise distance analysis. CONCLUSION: This study provides a simple low-cost pathway to accurately subtype in SSA, guide regional therapeutic choice and assist global surveillance and elimination initiatives.

Optimization of the pseudoparticle system for standardized assessments of neutralizing antibodies against hepatitis C virus.

A better understanding of the antibody response during natural infection and the effect on disease progression and reinfection is necessary for the development of a protective hepatitis C virus (HCV) vaccine. The HCV pseudoparticle (HCVpp) system enables the study of viral entry and inhibition by antibody neutralization. A robust and comparable neutralization assay is crucial for the development and evaluation of experimental vaccines.With the aim of optimizing the HCVpp-murine leukaemia virus (MLV) system, we tested the neutralization of HCVpp-harbouring E1E2 from 21 HCV isolates representing 6 different genotypes by several monoclonal antibodies (mAbs). HCVpps are generated by expressing functional envelope glycoproteins (E1E2) onto pseudoparticles derived from env-deleted MLV. Adjustments of E1E2, gag-pol and luciferase plasmid ratios resulted in increased yields for most HCVpps and recovery of one non-infectious HCVpp. We simplified and improved the protocol to achieve higher signal/noise ratios and minimized the amount of HCVpps and mAbs needed for the detection of neutralization. Using our optimized protocol, we demonstrated comparable results to previously reported data with both diluted and freeze-thawed HCVpps.In conclusion, we successfully established a simplified and reproducible HCVpp neutralization protocol for studying a wide range of HCV variants. This simplified protocol provides highly consistent results and could be easily adopted by others to evaluate precious biological material. This will contribute to a better understanding of the antibody response during natural infection and help evaluate experimental HCV vaccines.

Workforce Experiences of a Rapidly Established SARS-CoV-2 Asymptomatic Testing Service in a Higher Education Setting: A Qualitative Study.

The aim of the study was to explore workforce experiences of the rapid implementation of a SARS-CoV-2 asymptomatic testing service (ATS) in a higher education setting during the COVID-19 pandemic. The setting was a multi-campus university in the UK, which hosted a testing service for employees and students over two years. Qualitative semi-structured videoconference interviews were conducted. We contacted 58 participants and 25 were interviewed (43% response rate). Data were analysed thematically. The analysis produced four overarching themes: (1) feelings relating to their involvement in the service, (2) perceptions of teamwork, (3) perceptions of ATS leadership, (4) valuing the opportunity for career development. Agile and inclusive leadership style created psychological safety and team cohesion, which facilitated participants in the implementation of a rapid mitigation service, at pace and scale. Specific features of the ATS (shared vision, collaboration, networking, skills acquisition) instilled self-confidence, value and belonging, meaningfully impacting on professional development and career opportunities. This is the first qualitative study to explore the experiences of university employees engaged in the rapid deployment of a service as part of a pandemic outbreak and mitigation strategy within a higher education setting. Despite pressures and challenges of the task, professional growth and advancement were universal. This has implications for workforce engagement and creating workplaces across the sector that are well-prepared to respond to future pandemics and other disruptive events.

Evaluation of the relative potential for contact and doffing transmission of SARS-CoV-2 by a range of personal protective equipment materials.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-the causative agent of coronavirus disease 2019 (COVID-19)-has caused a global public health emergency. Personal protective equipment (PPE) is the primary defence against viral exposure in healthcare and community settings. However, the surfaces of PPE materials may trap virus for contact transmission or through laden aerosols generated during removal of PPE, through cleaning or during movement. In this study, the relative efficacy of current PPE materials in terms of virion adsorption to materials and their antiviral potency, has been evaluated on a wide range of PPE for the first time, including four polymer glove types, two types of scrubs, apron material, a mask, visor and a selection of other commercial polymers and products. Although differences in virion adsorption to the test materials were observed, none of the existing polymer-based PPE resulted in more than tenfold reduction in the SARS-CoV-2 titre within either 10 min or 30 min contact period. The wettability and surface chemistry of the test materials were analysed to investigate any correlations with their surface physicochemical properties. While no correlation was found between wettability and viral retention under air flow challenge, one secondary ion of m/z 101.03 (+) and three secondary ions of m/z 31.98 (-), 196.93 (-) and 394.33 (+) in ToF-SIMS data of the test materials showed positive and negative correlations with the viral retention, respectively, which was identified by PLS regression model, suggesting that the surface chemistry plays a role in determining the extent of virion adsorption. Our findings outline the material aspects that influence the efficacy of current PPE against SARS-CoV-2 transmission and give suggestions on the development of novel simple polymer-based PPE for better infection protection.

A Qualitative Evaluation of the Barriers and Enablers for Implementation of an Asymptomatic SARS-CoV-2 Testing Service at the University of Nottingham: A Multi-Site Higher Education Setting in England.

Asymptomatic testing for SARS-CoV-2 RNA has been used to prevent and manage COVID-19 outbreaks in university settings, but few studies have explored their implementation. The aim of the study was to evaluate how an accredited asymptomatic SARS-CoV-2 testing service (ATS) was implemented at the University of Nottingham, a multi-campus university in England, to identify barriers and enablers of implementation and to draw out lessons for implementing pandemic response initiatives in higher education settings. A qualitative interview study was conducted with 25 ATS personnel between May and July 2022. Interviews were conducted online, audio-recorded, and transcribed. Participants were asked about their experience of the ATS, barriers and enablers of implementation. Transcripts were thematically analysed. There were four overarching themes: (1) social responsibility and innovation, (2) when, how and why people accessed testing, (3) impact of the ATS on the spread of COVID-19, and (4) lessons learned for the future. In establishing the service, the institution was seen to be valuing its community and socially responsible. The service was viewed to be broadly successful as a COVID-19 mitigation approach. Challenges to service implementation were the rapidly changing pandemic situation and government advice, delays in service accreditation and rollout to staff, ambivalence towards testing and isolating in the target population, and an inability to provide follow-up support for positive cases within the service. Facilitators included service visibility, reduction in organisational bureaucracy and red tape, inclusive leadership, collaborative working with regular feedback on service status, flexibility in service delivery approaches and simplicity of saliva testing. The ATS instilled a perception of early 'return to normality' and impacted positively on staff feelings of safety and wellbeing, with wider benefits for healthcare services and local communities. In conclusion, we identified common themes that have facilitated or hindered the implementation of a SARS-CoV-2 testing service at a university in England. Lessons learned from ATS implementation will inform future pandemic response interventions in higher education settings.

Exploring the Psychological Impacts of COVID-19 Social Restrictions on International University Students: A Qualitative Study.

The global COVID-19 pandemic has impacted on the mental well-being of university students, but little attention has been given to international students, who may have a unique experience and perspective. The aim of this study was to explore the views of international students and university staff towards COVID-19 restrictions, self-isolation, their well-being, and support needs, through eight online focus groups with international students (n = 29) and semi-structured interviews with university staff (n = 17) at a higher education institution in England. Data were analysed using an inductive thematic approach, revealing three key themes and six subthemes: (1) practical, academic, and psychological challenges faced during self-isolation and the COVID-19 pandemic; (2) coping strategies to self-isolation and life during the pandemic; and (3) views on further support needed for international students. International students faced practical, academic, and psychological challenges during the COVID-19 pandemic, particularly relating to the rapid transition to online learning and the impact of social restrictions on integration with peers and well-being. Online social connections with peers, family, or new acquaintances reduced feelings of isolation and encouraged involvement in university life. Despite raising mental health concerns, most international students did not access mental health support services. Staff related this to perceived stigma around mental health in certain cultural groups. In conclusion, international students experienced specific practical and emotional challenges during the pandemic, and are at risk of mental ill-health, but may not actively seek out support from university services. Proactive and personalised approaches to student support will be important for positive student experiences and the retention of students who are studying abroad in the UK higher education system.

In vitro evolution predicts emerging SARS-CoV-2 mutations with high affinity for ACE2 and cross-species binding.

Emerging SARS-CoV-2 variants are creating major challenges in the ongoing COVID-19 pandemic. Being able to predict mutations that could arise in SARS-CoV-2 leading to increased transmissibility or immune evasion would be extremely valuable in development of broad-acting therapeutics and vaccines, and prioritising viral monitoring and containment. Here we use in vitro evolution to seek mutations in SARS-CoV-2 receptor binding domain (RBD) that would substantially increase binding to ACE2. We find a double mutation, S477N and Q498H, that increases affinity of RBD for ACE2 by 6.5-fold. This affinity gain is largely driven by the Q498H mutation. We determine the structure of the mutant-RBD:ACE2 complex by cryo-electron microscopy to reveal the mechanism for increased affinity. Addition of Q498H to SARS-CoV-2 RBD variants is found to boost binding affinity of the variants for human ACE2 and confer a new ability to bind rat ACE2 with high affinity. Surprisingly however, in the presence of the common N501Y mutation, Q498H inhibits binding, due to a clash between H498 and Y501 side chains. To achieve an intermolecular bonding network, affinity gain and cross-species binding similar to Q498H alone, RBD variants with the N501Y mutation must acquire instead the related Q498R mutation. Thus, SARS-CoV-2 RBD can access large affinity gains and cross-species binding via two alternative mutational routes involving Q498, with route selection determined by whether a variant already has the N501Y mutation. These mutations are now appearing in emerging SARS-CoV-2 variants where they have the potential to influence human-to-human and cross-species transmission.

Simultaneous determination of HCV genotype and NS5B resistance associated substitutions using dried serum spots from São Paulo state, Brazil.

Hepatitis C virus (HCV) is responsible for more than 180 million infections worldwide, and about 80 % of infections are reported in Low and Middle-income countries (LMICs). Therapy is based on the administration of interferon (INF), ribavirin (RBV) or more recently Direct-Acting Antivirals (DAAs). However, amino acid substitutions associated with resistance (RAS) have been extensively described and can contribute to treatment failure, and diagnosis of RAS requires considerable infrastructure, not always locally available. Dried serum spots (DSS) sampling is an alternative specimen collection method, which embeds drops of serum onto filter paper to be transported by posting to a centralized laboratory. Here, we assessed feasibility of genotypic analysis of HCV from DSS in a cohort of 80 patients from São Paulo state Brazil. HCV RNA was detected on DSS specimens in 83 % of samples of HCV infected patients. HCV genotypes 1a, 1b, 2a, 2c and 3a were determined using the sequence of the palm domain of NS5B region, and RAS C316N/Y, Q309R and V321I were identified in HCV 1b samples. Concerning therapy outcome, 75 % of the patients who used INF +RBV as a previous protocol of treatment did not respond to DAAs, and 25 % were end-of-treatment responders. It suggests that therapy with INF plus RBV may contribute for non-response to a second therapeutic protocol with DAAs. One patient that presented RAS (V321I) was classified as non-responder, and combination of RAS C316N and Q309R does not necessarily imply in resistance to treatment in this cohort of patients. Data presented herein highlights the relevance of studying circulating variants for a better understanding of HCV variability and resistance to the therapy. Furthermore, the feasibility of carrying out genotyping and RAS phenotyping analysis by using DSS card for the potential of informing future treatment interventions could be relevant to overcome the limitations of processing samples in several location worldwide, especially in LMICs.

Human parainfluenza 2 & 4: Clinical and genetic epidemiology in the UK, 2013-2017, reveals distinct disease features and co-circulating genomic subtypes.

BACKGROUND: Human Parainfluenza viruses (HPIV) comprise of four members of the genetically distinct genera of Respirovirus (HPIV1&3) and Orthorubulavirus (HPIV2&4), causing significant upper and lower respiratory tract infections worldwide, particularly in children. However, despite frequent molecular diagnosis, they are frequently considered collectively or with HPIV4 overlooked entirely. We therefore investigated clinical and viral epidemiological distinctions of the relatively less prevalent Orthorubulaviruses HPIV2&4 at a regional UK hospital across four autumn/winter epidemic seasons. METHODS: A retrospective audit of clinical features of all HPIV2 or HPIV4 RT-PCR-positive patients, diagnosed between 1st September 2013 and 12th April 2017 was undertaken, alongside sequencing of viral genome fragments in a representative subset of samples. RESULTS: Infection was observed across all age groups, but predominantly in children under nine and adults over 40, with almost twice as many HPIV4 as HPIV2 cases. Fever, abnormal haematology, elevated C-reactive protein and hospital admission were more frequently seen in HPIV2 than HPIV4 infection. Each of the four seasonal peaks of either HPIV2, HPIV4 or both, closely matched that of RSV, occurring in November and December and preceding that of Influenza A. A subset of viruses were partially sequenced, indicating co-circulation of multiple subtypes of both HPIV2&4, but with little variation between each epidemic season or from limited global reference sequences. CONCLUSIONS: Despite being closest known genetic relatives, our data indicates a potential difference in associated disease between HPIV2 and HPIV4, with more hospitalisation seen in HPIV2 mono-infected individuals, but a greater overall number of HPIV4 cases.