Dengue virus infection is associated with the upregulation of metabolic pathways within infected cells. This effect is common to infection by a broad array of viruses. These metabolic changes, including increased glucose metabolism, oxidative phosphorylation and autophagy, support the demands of viral genome replication and infectious particle formation. The mechanisms by which these changes occur are known to be, in part, directed by viral nonstructural proteins that contact and control cellular structures and metabolic enzymes. We investigated the roles of host proteins with overarching control of metabolic processes, the transcriptional regulators, cyclin-dependent kinase 8 (CDK8) and its paralog, CDK19, as mediators of virally induced metabolic changes. Here, we show that expression of CDK8, but not CDK19, is increased during dengue virus infection in Huh7 human hepatocellular carcinoma cells, although both are required for efficient viral replication. Chemical inhibition of CDK8 and CDK19 with Senexin A during infection blocks virus-induced expression of select metabolic and autophagic genes, hexokinase 2 (HK2) and microtubule-associated protein 1 light chain 3 (LC3), and reduces viral genome replication and infectious particle production. The results further define the dependence of virus replication on increased metabolic capacity in target cells and identify CDK8 and CDK19 as master regulators of key metabolic genes. The common inhibition of CDK8 and CDK19 offers a host-directed therapeutic intervention that is unlikely to be overcome by viral evolution.

Enterovirus D68 (EV-D68) caused large biennial cyclical outbreaks of respiratory disease and cases of acute flaccid myelitis from 2014 to 2018 in the USA. An anticipated outbreak did not occur in 2020, likely due to non-pharmaceutical interventions targeting the COVID-19 pandemic. A large respiratory disease outbreak occurred again in 2022, but uncertainty remained regarding if circulation of EV-D68 would return to the pre-pandemic patterns. We conducted prospective active surveillance of clinical respiratory specimens from Colorado children for EV-D68 in 2023 and 2024. A subset of residual specimens positive for rhinovirus/enterovirus (RV/EV) were tested for EV-D68 via a validated in-house EV-D68 reverse transcription–PCR assay. During epi weeks 18–44 in 2023, 525 residual specimens positive for RV/EV all tested negative for EV-D68. In 2024, during epi weeks 18–44, 10 (1.8%) of the 546 RV/EV-positive specimens were EV-D68-positive. The EV-D68-positive cases were predominantly young children (median age 4.8 years) receiving treatment with asthma medications. Following the 2022 EV-D68 outbreak, an anticipated outbreak did not occur in 2023. While EV-D68 was detected in 2024, the number of cases was not as significant as in prior outbreak years. Continued surveillance for EV-D68 will be important to understand the future dynamics of EV-D68 circulation and prepare for future outbreaks.

As part of research and wildlife disease surveillance efforts, we performed necropsy examinations of 125 free-ranging (n = 114) and captive (n = 11) prairie dogs in Colorado from 2009 to 2017. From these cases, we identified three cases of thymic lymphoma in free-ranging Gunnison’s prairie dogs (Cynomys gunnisoni), and we identified a novel retroviral sequence associated with these tumors. The viral sequence is 7700 nucleotides in length and exhibits a genetic organization that is consistent with the characteristics of a type D betaretrovirus. The proposed name of this virus is Gunnison’s prairie dog retrovirus (GPDRV). We screened all 125 prairie dogs for the presence of GPDRV using PCR with envelope-specific primers and DNA extracted from spleen samples. Samples were from Gunnison’s prairie dogs (n = 59), black-tailed prairie dogs (Cynomys ludovicianus) (n = 40), and white-tailed prairie dogs (Cynomys leucurus) (n = 26). We identified GPDRV in a total of 7/125 (5.6%) samples including all three of the prairie dogs with thymic lymphoma, as well as spleen from an additional four Gunnison’s prairie dogs with no tumors recognized at necropsy. None of the GPDRV-negative Gunnison’s prairie dogs had thymic lymphomas. We also identified a related, apparently endogenous retroviral sequence in all prairie dog samples. These results suggest that GPDRV infection may lead to development of thymic lymphoma in Gunnison’s prairie dogs.

The temporal dynamics of SARS-CoV-2 infectivity in immunocompromised children (IC) are unknown but may have important infection control implications. We evaluated SARS-CoV-2 viral persistence and assessed factors associated with viral persistence and cycle threshold (CT) values as a surrogate of viral load for IC. We conducted a retrospective cohort study of SARS-CoV-2-positive IC at a large quaternary pediatric hospital from March 2020-2021. Immunocompromised status was defined as primary or secondary/acquired immunodeficiencies due to comorbidities or immunosuppressive treatment. The primary outcome was time to first-of-two consecutively negative SARS-CoV-2 PCR tests ≥ 24 hours apart. Polymerase chain reaction (PCR) testing of sequential patient samples was conducted using the Centers for Disease Control 2019-nCoV Real-Time RT-PCR Diagnostic Panel (CDC assay). Chi-square, Fisher exact, and Wilcoxon tests were used to compare demographic and clinical characteristics. Kaplan-Meier curve median event times and log-rank tests were used to compare outcomes. Subjects without 2 consecutive negative tests censored at the last test. Analyses were conducted using SAS v 9.4. Ninety-one children met inclusion criteria, and 67 children had more than 1 test (Figure 1). Median age was 15.5 years (IQR 8-18 yrs), 64% were male, 58% of children were white, and 43% were Latinx. Most (67%) were tested in outpatient settings, and 58% of children were asymptomatic. The median time to two negative tests was 42 days (IQR 25.0,55.0), with no difference in duration of positivity with specific diagnoses, degree of lymphopenia, or symptomatic vs asymptomatic illness. Five of 7 (71%) children with samples available for repeat testing had initial C_T values < 30, indicating a moderate to high viral load, and of these, 4 (57%) had repeat testing 21 to 30 days later with C_T values < 30 (Figure 2), suggesting persistence of moderate to high viral loads. Figure 1. Plot of immunocompromised children in cohort with positive SARS CoV2 PCR and subsequent testing (n = 67). Timelines of immunocompromised children in cohort with positive SARS CoV2 PCR and subsequent testing, grouped by immunocompromising condition. Each line represents an individual patient. Positive results are shown in light grey, negative results are shown in black. Figure 2. Plot of CT values from SARS-CoV-2 PCR testing over time among children with sequential samples available for retesting (n = 7) Plot of CT values (y axis) from SARS-CoV-2 PCR testing on the CDC assay over time (x axis) in days from initial positive test. Repeated testing which yielded a negative result on the CDC assay or intermittent negative results on clinical testing represented as CT value of 40. Each line represents a unique patient. The median duration of viral persistence among IC with SARS-CoV-2 infection was 6 weeks, with no significant difference in immunocompromised diagnoses or clinical presentation, with over half of children with testing on the same platform having moderate to high viral loads after 3 weeks, suggesting potential transmission risk. Samuel R. Dominguez, MD, PhD, BioFire Diagnostics (Consultant, Research Grant or Support)DiaSorin Molecular (Consultant)Pfizer (Grant/Research Support) Samuel R. Dominguez, MD, PhD, BioFire (Individual(s) Involved: Self): Consultant, Research Grant or Support; DiaSorin Molecular (Individual(s) Involved: Self): Consultant; Pfizer (Individual(s) Involved: Self): Grant/Research Support Suchitra Rao, MBBS, MSCS, BioFire (Research Grant or Support)

Background: Enterovirus D68 (EV-D68) causes cyclical outbreaks worldwide. Though EV-D68 can rarely cause acute flaccid myelitis, it commonly causes respiratory disease. In particular, EV-D68 causes outbreaks of severe asthma-like respiratory disease, but few systems have the capability to detect EV-D68 cases. Methods: Active, prospective EV-D68 surveillance at Children’s Hospital Colorado has been performed since 2014, where a random subset of residual clinical respiratory specimens, primarily from inpatient settings, and positive for rhinovirus/enterovirus are tested for EV-D68 via reverse transcription-polymerase chain reaction. Data via chart review from this surveillance were analyzed from 2014 to 2025 to characterize the clinical characteristics of EV-D68 respiratory disease in children (<18 years). Results: From 2014 to 2025, EV-D68 was detected in 308 patients, of which 271/308 (88%%) were children with respiratory disease. Median age of children with respiratory disease was 6 [interquartile range (IQR): 3.1–10.4] years; 162/271 (59.8%) were males. A majority of children admitted required intensive care unit admission (206/258, 79.8%). Though only 174/271 (64.2%) children had an atopic history, a significant majority required bronchodilator therapy (249/271, 91.9%) and systemic steroids (229/271, 84.5%). Median intensive care unit and hospital length of stay were 2 (IQR: 1–3) and 3 (IQR: 2–5) days, respectively. Conclusions: Children with EV-D68 respiratory disease commonly presented with severe asthma-like respiratory disease, even in those without atopy. Despite the severity of disease, children quickly improved over short hospitalizations when managed with bronchodilators, steroids, and respiratory support. With improved diagnostics to detect and recognize the true burden of EV-D68, additional studies evaluating short- and long-term management of EV-D68 respiratory disease are needed.