Med

Article DescriptionWe present a real-world deployment of a large language model-powered colonoscopy recall pipeline that structured over 100,000 patient records during an EHR transition. This end-to-end AI system demonstrated high fidelity, scalability, and a projected prevention of up to 6092 colorectal cancer cases and cost savings between 400 - 670 million dollars. The rise of structured data elements in Electronic Health Records (EHRs) is a key enabler of improving care quality. However, the transition towards routine use of these fields paradoxically heightens patient safety risks due to increased variability in documentation and the use of "placeholder" values pending manual review. For large clinical initiatives such as colon cancer screening and surveillance, misinterpretation of recorded clinical data can be particularly problematic, disrupting risk-adapted recall guidance and potentially exacerbating care gaps. This case study details the development and deployment of a Large Language Model (LLM)-driven workflow to extract and transfer unstructured colonoscopy recall recommendations as part of a larger EHR migration. Utilizing GPT-4 Turbo for the core inference step of a fully integrated pipeline--spanning custom SQL queries, Optical Character Recognition (OCR) of historical PDFs, LLM-based inference, and anomaly detection--we successfully structured and migrated population-wide colonoscopy recall data corresponding to over 100,00 patients and 10 years of clinical care. The pipeline demonstrated high accuracy (Macro F1=1.0 against clinician review), scalability, and cost efficiency. We estimate that use of this workflow--relative to the alternative of a default 10-year reminder from last colonoscopy--may prevent over 6,000 new colorectal cancer cases (a projected cost savings of $400-670 million). Key lessons from implementation include the importance of stakeholder alignment, the necessity of robust quality control at scale, and the technical challenges of expanding optimized LLM inference to a fully-fledged end-to-end clinical workflow.

Central nervous system (CNS) accessibility constitutes a major hurdle for drug development to treat neurological diseases. Existing drug delivery methods rely integrity of the blood-brain barrier (BBB) for CNS penetration. Here we showed that the microchannels between the skull marrow and the dura mater could be harnessed for drug delivery by intracalvariosseous (ICO) injection. Drugs administered via ICO injection were found to reach cranial bone marrow-dura-perivascular space, and the injection procedure did not cause osteomyelitis or BBB damage. To validate this approach, we examined the efficacy of two neuroprotective agents, NA-1 and Y-3, via ICO injection in rat model of stroke and found that ICO injection increased drug accumulation in the brain compared to intravenous injection, reduced infarct area and alleviated neurological deficits. We subsequently conducted a clinical trial to assess the safety of ICO in acute ischemic stroke patients (ClinicalTrials.gov identifier NCT05849805), showing that ICO injection was feasible and safe in humans and its therapeutic effects may be observed. Collectively, our study identifies that the microchannels between the skull bone marrow and the dura mater act as a new channel for CNS drug delivery to achieve high intracranial drug exposure in a short period of time. The safety of ICO injection makes it a promising route of drug administration for CNS diseases.

Fecal microbiota transplantation (FMT) is an effective therapy for recurrent Clostridioides difficile infection (rCDI) but has undefined composition and poor scalability. In vitro manufactured live biotherapeutic products (LBP) enable both scalability and defined strain composition but with higher manufacturing complexity, resulting in few LBP trials. We developed an accessible platform to produce human-grade LBPs. We provide regulatory documentation and manufacturing protocols to facilitate translating microbiome advances to human trials. With this platform, we conduct the first direct comparison of the same bacterial strains administered after in vitro manufacturing (LBP) compared to donor sourced (FMT) across two doses. In a phase 1b trial (n=18), an endoscopic dose of the 15-strain consortium MTC01 was safe with rCDI prevention eight weeks after dosing in seven out of nine LBP patients, similar to eight out of nine FMT patients. Notably, MTC01 strain engraftment was superior to FMT at higher doses.

BackgroundThe extracellular microenvironment modulates cancer behavior. Although radiographic contrast enhancement is an ominous finding in gliomas, it remains unclear if the associated blood-brain barrier disruption merely reflects or functionally supports tumor aggressiveness. MethodsWe utilized intra-operative microdialysis to sample the extracellular metabolome of radiographically diverse regions during fifteen neurosurgical resections. The global extracellular metabolome of recovered microdialysate was evaluated via ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) and assessed using enrichment and correlation analyses. ResultsAmong 162 named metabolites identified via ultra-performance liquid chromatography tandem mass spectrometry, guanidinoacetate (GAA), was 126.32x higher in enhancing tumor than in adjacent brain. 48 additional metabolites were 2.05-10.18x more abundant in enhancing tumor than brain. With exception of GAA, and 2-HG in IDH-mutant gliomas, differences between non-enhancing tumor and brain microdialysate were comparatively modest and less consistent. The enhancing but not the non-enhancing glioma metabolome was significantly enriched for plasma-associated metabolites largely comprising amino acids and carnitines. ConclusionsOur findings suggest that metabolite diffusion through a disrupted blood-brain barrier may largely define the enhancing extracellular glioma metabolome. Future studies are needed to determine how the altered extracellular metabolome impacts glioma behavior.

Despite the existence of an effective vaccine, the United States continues to experience outbreaks of hepatitis A, including in Massachusetts (MA) and New Hampshire (NH) in 2018 and again in MA in 2023. To clarify the relationship between these outbreaks and better understand their drivers, we generated hepatitis A virus whole genome sequences from reported cases and analyzed them using open-source genotyping tools developed and released as part of this study. We found that the 2018 and 2023 outbreaks were caused by distinct viral strains, despite affecting individuals with similar demographic characteristics and reported risk factors. Detailed analysis of genomic and epidemiologic data further resolved transmission patterns within and across outbreaks, showing that experiencing homelessness and prior use of drugs were associated with increased transmission while also revealing transmission between individuals with and without these risk factors, as well as spread across state borders. Together, these findings demonstrate the value of broadly accessible genomic tools for understanding hepatitis A outbreaks and illustrate how whole genome analysis can complement epidemiological investigation by resolving transmission patterns and outbreak drivers that can inform public health interventions.

Individuals with asymptomatic SARS-CoV-2 infection can unknowingly transmit the virus, yet identifying such infections in vaccinated populations remains challenging. We conducted a longitudinal study of 129 vaccine recipients immunized with various combinations of SARS-CoV-2 spike (S) protein vaccine platforms. Sera were collected before the first dose (v1), at 2 weeks (v7) and 6 months (v8) after the third dose. Taiwans first major COVID-19 outbreak occurred between v7 and v8. We measured anti-nucleocapsid (anti-N) and anti-S IgG antibody titers by ELISA and assessed virus-neutralizing activity using live virus and pseudovirus assays. By developing an iterative serial screening method, we identified asymptomatic infections among unconfirmed cases. Our v7-v8 paired cohort resolved into three distinct groups: confirmed cases (21%), asymptomatic infections (17%), and uninfected cases (62%). In normalized v8 sera, confirmed cases exhibited an anti-S+++/anti-N+++ phenotype, while uninfected cases showed an anti-S+/anti-N+ phenotype. Statistical analysis validated a distinct asymptomatic group characterized by intermediate anti-S but baseline anti-N antibody levels (anti-S++/anti-N+). This approach enables more accurate estimates of infection prevalence and vaccine efficacy.

As the COVID-19 pandemic continues, long-term immunity against SARS-CoV-2 will be globally important. Official weekly cases have not dropped below 2 million since September of 2020, and continued emergence of novel variants have created a moving target for our immune systems and public health alike. The temporal aspects of COVID-19 immunity, particularly from repeated vaccination and infection, are less well understood than short-term vaccine efficacy. In this study, we explore the impact of combined vaccination and infection, also known as hybrid immunity, and the timing thereof on the quality and quantity of antibodies produced by a cohort of 96 health care workers. We find robust neutralizing antibody responses among those with hybrid immunity against all variants, including Omicron BA.2, and we further found significantly improved neutralizing titers with longer vaccine-infection intervals up to 400 days. These results indicate that anti-SARS-CoV-2 antibody responses undergo continual maturation following primary exposure by either vaccination or infection for at least 400 days after last antigen exposure. We show that neutralizing antibody responses improved upon secondary boosting with greater impact seen after extended intervals. Our findings may also extend to booster vaccine doses, a critical consideration in future vaccine campaign strategies.

Messenger RNA (mRNA) based vaccines (Pfizer/BioNTech and Moderna) are highly effective at providing immunity against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). However, there is uncertainty about the duration of immunity, evolution of IgG antibody levels and IgG avidity (an index of antibody-antigen binding strength), and differences in the immune responses between vaccines. Here we performed a prospective pilot study of 71 previously COVID-19 free subjects upon receiving both doses of either the Pfizer (n = 54) or Moderna (n = 17) mRNA vaccine. Anti-spike protein receptor binding domain (RBD) IgG antibodies were measured longitudinally using a qualitative finger stick MidaSpot rapid test at the point-of-care for initial screening and a quantitative dry blood spot-based pGOLD laboratory test over [~] four months post-vaccination. The average anti-RBD IgG antibody levels peaked at [~] two weeks after the second dose of the vaccine and declined thereafter, while antibody avidity increased, suggesting antibody maturation. Moderna vaccine recipients compared to Pfizer vaccine recipients exhibited higher side effect severity, higher peak anti-RBD IgG antibody levels, and higher avidity up to the 90 days period. Differences in antibody levels diminished at [~] 120 days post-vaccination, in line with the similar efficacy observed in the two vaccines. The MidaSpot rapid test detected 100% anti-SARS-CoV-2 RBD positivity for fully vaccinated subjects in both Pfizer and Moderna cohorts post full vaccination but turned negative greater than 90 days post-vaccination for 5.4% of subjects in the Pfizer cohort, whose quantitative anti-IgG were near the minimum levels of the group. Immune responses were found to vary greatly among vaccinees. Personalized longitudinal monitoring of antibodies could be necessary to assess the immunity duration of vaccinated individuals.

Current guidelines recommend that individuals who have had COVID-19 should receive the identical vaccine regimen as those who have not had the infection. This includes two doses of the mRNA platform vaccines (BNT162b2/Pfizer; mRNA-1273/Moderna) that are approved for use in the United States. In this brief report, we show that after a single dose of the Pfizer SARS-CoV-2 vaccine, individuals that had prior SARS-CoV-2 infection had significantly higher antibody levels than individuals that had no history of infection. This provides the rationale for changing vaccination policy to deliver only a single dose to individuals with recent SARS-CoV-2 infection that may free up additional doses for individuals that have no preexisting immunity to the virus. Future study of other immune parameters such as T cell response and durability of immune response should be rapidly undertaken in individuals that had COVID-19 prior to vaccination.

Hurricane Harvey caused record-breaking, catastrophic flooding across the city of Houston. After floodwaters receded, several health concerns arose, including the potential adverse impact of exposure to mold in flooded homes. We rapidly launched the Houston Hurricane Harvey Health Study to evaluate if microbiome sampling in the wake of a disaster could inform flood-associated environmental exposures and adverse health outcomes. We enrolled a total of 347 subjects at 1-month and 12-months post-Harvey, collecting human (stool, nasal, saliva) and environmental (house swab) samples to profile the bacterial and fungal microbiota. Here we show reported exposure to mold was associated with increased risk of allergic symptoms for up to one year post-disaster, and that butyrate-producing bacteria in the gut were linked to protection from allergic symptoms in mold-exposed individuals. Together, these data provide new insights into how microbiome:environment interactions may influence health in the setting of a flood-related disaster.

ObjectiveLittle is known about the interplay between gut microbiome and SARS-CoV-2 vaccine immunogenicity. In this prospective observational study, we investigated associations between the gut microbiome, habitual dietary fibre intake, and mRNA vaccine-elicited immune responses, including anti-Spike IgG, avidity, and ACE-2 competition (surrogate neutralization). Design16S rRNA sequencing and short-chain fatty acid analyses were undertaken using stool samples collected from 48 healthy individuals at baseline and twelve-weeks after 1st BNT162b2 SARS-CoV-2 vaccine dose. Associations between gut microbiome data and SARS-CoV-2 spike and RBD IgG levels, competitive binding antibodies, and anti-SARS-CoV-2 spike total relative fractional avidity assays were evaluated. A validated dietary fibre intake food frequency questionnaire was also used to correlate habitual dietary fibre intakes with vaccine responses. ResultsOur data revealed several baseline bacterial taxa, including Prevotella, Haemophilus and Veillonella (p<0.01), associated with BNT162b2 vaccine responses. Several Bacteroides spp. (p<0.01) as well as Bifidobacterium animalis, (p=0.003), amongst others, were positively associated with antibody avidity. Conversely, concentrations of isovaleric and isobutyric acid were higher in individuals with the lowest SARS-CoV-2 vaccine responses (p<0.01). Classifying participants based on habitual dietary fibre intake identified distinct avidity responses. ConclusionWe showed associations between baseline gut microbiota composition and immunogenicity of BNT162b2 vaccine responses, particularly avidity maturation. We also demonstrate that branched-chain fatty acids and habitual dietary fibre intakes are associated with BNT162b2 vaccine immunogenicity. Together these findings indicate a link between gut microbiome, diet and antibody immunity to SARS-CoV-2 spike protein, suggesting interventions which modulate the gut microbiome could enhance COVID-19 vaccine responses. SIGNIFICANCE OF THIS STUDYO_ST_ABSWhat is already known on this subject?C_ST_ABSO_LIStrength and persistence of the SARS-CoV-2 BNT162b2 vaccine is variable between individuals. C_LIO_LITo date, only one study has demonstrated that baseline gut microbiota can predict SARS-CoV-2 vaccine response. C_LI What are the new findings?O_LIFor the first time we showed that the higher concentrations of branched-chain fatty acids, isovaleric and isobutyric acids, are negatively associated with SARS-CoV-2 BNT162b2 vaccine responses. C_LIO_LIWe revealed that habitual dietary fibre intake led to variability in the strength of antibody binding after the BNT162b2 vaccine. Specifically, high dietary fibre consumers displayed a significant increase in antibody avidity between in their 1st and 2nd dose. C_LI How this study might affect research, practice, or policyO_LIOur data suggests that therapeutic interventions which target the gut microbiome, including dietary modification, as well as pre-, pro-, and post-biotics, could enhance BNT162b2 vaccine immunogenicity, thus helping in the fight against COVID-19. C_LI

The microvasculature and hemodynamics of human brain tumors and other lesions have remained largely unexplored in vivo due to the limited resolution of conventional imaging techniques, and may present new opportunities in biomarker identification and neurosurgeries. Ultrasound Localization Microscopy (ULM), which tracks freely circulating intravascular microbubbles used as contrast agents, overcomes these limitations and has been used to visualize vascular structures and flow. In this study, we performed intraoperative ULM during brain surgeries involving resection of brain tumors (N = 3 meningiomas, N = 3 brain metastasis, N = 3 high grade gliomas) and an arteriovenous malformation (AVM) (N = 1). ULM provided microvascular images of the human brain, resolving vessels down to 35 m, revealing clear differences in structure and dynamics between tumor and surrounding healthy tissue. By following individual microbubbles, vessel connectivity was probed and used to identify feeding, draining, and non-tumoral vessels. In one case, a 3D ULM map of an AVM was generated with 226 m resolution, allowing us to resolve its complex internal structure, including feeding and draining vessels. Intraoperative ULM enables visualization of human brain vasculature and hemodynamics at unprecedented resolutions, and may directly aid tumor and AVM resections by providing flow paths and speeds through compact niduses. One Sentence SummaryUltrasound localization microscopy during human brain surgery revealed vascular structure and dynamics of brain lesions at sub-millimeter resolution.

ImportanceThe antigen-based rapid diagnostic test (Ag-RDT), using saliva specimens, is fast, non-invasive and suitable for SARS-CoV-2 self-testing, unlike nasopharyngeal swab (NPS) testing. ObjectiveTo assess the diagnostic sensitivity of a novel Beanguard gargle (BG)-based virus detection method for early diagnosis of COVID-19. DesignThis clinical trial was conducted at Gunsan Medical Center, Namwon Medical Center, and Jeonbuk National University Hospital, between May 7 and July 7, 2021. SettingPaired NPS and BG-based saliva specimens collected from COVID-19 patients and healthy individuals were analyzed using NPS-RT-PCR, BG-RT-PCR, and BG-Ag-RDTs. ParticipantsThe study comprised 102 COVID-19-positive patients hospitalized after governmental screening process and 100 healthy individuals. Forty-five COVID-19 patients were sampled within 6 days of illness and 57 within 7-15 days; 27 were categorized as asymptomatic and 75, as symptomatic. Eight and 2 patients carried the SARS-CoV-2 Alpha and Delta variants, respectively. InterventionThe diagnostic performances of BG-Ag-RDT, BG-RT-PCR, and NPS-RT-PCR for detecting SARS-CoV-2 were compared. Main outcomesThe sensitivities of BG-Ag-RDT and BG-RT-PCR towards salivary viral detection were highly concordant, with no discrimination between symptomatic, asymptomatic, or SARS-CoV-2 variant cases. ResultsAmong total participants (mean age, 43.7 years), 51% were women. BG-Ag-RDTs showed high sensitivity (97.8%, [95% CI, 88.4% to 99.6%]) and specificity (100%, [95% CI, 96.3% to 100%) in 45 patients within 6 days of illness and could detect all cases of SARS-CoV-2 Alpha and Delta variants. In 11 asymptomatic early-stage cases, both BG-Ag-RDTs and BG-RT-PCR showed excellent sensitivity and specificity of 100% (95% CI, 74.1% to 100% and 95% CI, 20.7% to 100%, respectively). The interaction between SARS-CoV-2 spike proteins and truncated canavalin, an active ingredient from bean extract (BE) and the ultrastructural features of SARS-CoV-2 particles coated with BE were observed. The detachment of the SARS-CoV-2 receptor-binding domain from hACE2 increased as the BE concentration increased, allowing the release of the virus from hACE2 for early diagnosis. Conclusions and RelevanceUsing BG-based saliva remarkably enhances the Ag-RDT diagnostic performance as an alternative to NPS and enables rapid and accurate COVID-19 self-testing and mass screening, supporting efficient COVID-19 management. Trial RegistrationKCT0006438 Key PointsO_ST_ABSQuestionC_ST_ABSHow can we collect SARS-CoV-2 from oral cavity to improve the sensitivity of antigen-based rapid diagnostic test (Ag-RDT)? FindingsIn this clinical study involving 102 hospitalized COVID-19 patients, the Ag-RDT test using Beanguard gargle-based saliva specimens showed significantly enhanced sensitivity and specificity towards detection of SARS-CoV-2 along with Alpha and Delta variants in all patients tested within 6 days of illness. MeaningOur self-testing method represents an attractive alternative to nasopharyngeal swab RT-PCR for the early diagnosis of symptomatic and asymptomatic COVID-19 cases.

Post-COVID-19 conditions, also known as "long COVID", has significantly impacted the lives of many individuals, but the risk factors for this condition are poorly understood. In this study, we performed a retrospective EHR analysis of 89,843 individuals at a multi-state health system in the United States with PCR-confirmed COVID-19, including 1,086 patients diagnosed with long COVID and 1,086 matched controls not diagnosed with long COVID. For these two cohorts, we evaluated a wide range of clinical covariates, including laboratory tests, medication orders, phenotypes recorded in the clinical notes, and outcomes. We found that chronic pulmonary disease (CPD) was significantly more common as a pre-existing condition for the long COVID cohort than the control cohort (odds ratio: 1.9, 95% CI: [1.5, 2.6]). Additionally, long-COVID patients were more likely to have a history of migraine (odds ratio: 2.2, 95% CI: [1.6, 3.1]) and fibromyalgia (odds ratio: 2.3, 95% CI: [1.3, 3.8]). During the acute infection phase, the following lab measurements were abnormal in the long COVID cohort: high triglycerides (meanlongCOVID: 278.5 mg/dL vs. meancontrol: 141.4 mg/dL), low HDL cholesterol levels (meanlongCOVID: 38.4 mg/dL vs. meancontrol: 52.5 mg/dL), and high neutrophil-lymphocyte ratio (meanlongCOVID: 10.7 vs. meancontrol: 7.2). The hospitalization rate during the acute infection phase was also higher in the long COVID cohort compared to the control cohort (ratelongCOVID: 5% vs. ratecontrol: 1%). Overall, this study suggests that the severity of acute infection and a history of CPD, migraine, CFS, or fibromyalgia may be risk factors for long COVID symptoms. Our findings motivate clinical studies to evaluate whether suppressing acute disease severity proactively, especially in patients at high risk, can reduce incidence of long COVID.

Recent estimates consider that Long COVID affects 400 million people worldwide, of which <10% recover after two years. Predictive biomarkers and effective therapies are urgently needed for the clinical management of Long COVID. Neurological manifestations (poor memory, brain fog) are frequent, while neurodamage and neuroinflammatory markers UCHL1, GFAP and NFL were found to be increased in some but not all Long COVID cohorts. However, a direct link between two major Long COVID disease mechanisms, viral persistence and coagulation defects, and CNS damage is unclear. Therefore, we investigated the effect of combined anticoagulant (Asaflow + Clopidogrel) vs. antiviral (Paxlovid) treatment upon clinical outcome and neurodamage markers in Long COVID patients. Using multivariate logistic regression, we found that second-line antiviral treatment is the single best predictor of clinical recovery in a real-world Long COVID cohort (n=106) with long-term (220 person-years) follow-up. Our results support a putative role for viral persistence in Long COVID pathogenesis but also indicate a large therapeutic window for Paxlovid and other antivirals. A rapid decline in neurodamage markers (GFAP/NFL) upon antiviral treatment suggests CNS damage in Long COVID might be therapeutically targeted and should be considered in ongoing and future clinical trials.

We longitudinally profiled immune responses in 102 adults who received BNT162b2 (Pfizer-BioNTech) or ChAdOx1-S (Oxford-AstraZeneca) as their primary vaccinations. Bloods were collected pre-vaccination, 1-7 days after the 1st, 2nd and 3rd doses (BNT162b2 or mRNA-1273) to assess innate and early adaptive responses, and [~]28 days after the 2nd and 3rd doses to assess immunogenicity. Using a multi-omics approach including RNAseq, cytokine multiplex assay, proteomics, lipidomics, and flow cytometry we identified key differences in the immune responses induced by the ChAdOx1-S and BNT162b2 vaccines that were correlated with subsequent antigen-specific antibody and T cell responses or vaccine reactogenicity. We observed that vaccination with ChAdOx1-S but not BNT162b2 induced a memory-like response after the first dose, which was correlated with the expression of several proteins involved in complement and coagulation. The COVID-19 Vaccine Immune Responses Study (COVIRS) thus represents a major resource to understand the immunogenicity and reactogenicity of these COVID-19 vaccines.

The COVID-19 pandemic has resulted in extensive surveillance of the genomic diversity of SARS-CoV-2. Sequencing data generated as part of these efforts can also capture the diversity of the SARS-CoV-2 virus populations replicating within infected individuals. To assess this within-host diversity of SARS-CoV-2 we quantified low frequency (minor) variants from deep sequence data of thousands of clinical samples collected by a large urban hospital system over the course of a year. Using a robust analytical pipeline to control for technical artefacts, we observe that at comparable viral loads, specimens from patients hospitalized due to COVID-19 had a greater number of minor variants than samples from outpatients. Since individuals with highly diverse viral populations could be disproportionate drivers of new viral lineages in the patient population, these results suggest that transmission control should pay special attention to patients with severe or protracted disease to prevent the spread of novel variants.

Human challenge models (CHIMs) are instrumental in advancing influenza research but have traditionally relied on intranasal inoculation, which does not mimic the natural aerosol transmission of the virus. We conducted a dose-escalation influenza CHIM study to evaluate the safety and feasibility of two modern aerosol delivery systems: a flow-focusing monodisperse aerosol generator (FMAG) and a medical nebulizer. Fourteen healthy adults aged 18-49 years were exposed to influenza A/Perth/16/2009 (H3N2) in a controlled inpatient setting. Infection rates were 75% (3/4) with FMAG and 50% (2/4) with the nebulizer at the higher dose. Infections were self-limited, with sinus congestion, rhinorrhea, and cough being the most common symptoms. No serious adverse events occurred. Viral shedding was reproducible across respiratory sites, and seroconversion occurred in 33% of infected participants. Symptom timing and viral kinetics were comparable to those observed in prior intranasal CHIMs. Participants receiving nebulizer-delivered virus showed earlier viral detection in the oral cavity, suggesting broader airway deposition. These findings demonstrate that aerosol influenza challenge is both safe and effective and can simulate natural infection more accurately than intranasal delivery. This reintroduction of aerosolized influenza challenge provides a robust platform for studying transmission dynamics, tissue-specific immune responses, and for evaluating next-generation vaccines and therapeutics under conditions that better approximate real-world exposure. One Sentence SummaryWe demonstrated controlled human infection with dose-escalated H3N2 influenza virus via two safe and effective aerosolization routes that provide more accurate simulation of real-world exposure than intranasal delivery for studying influenza transmission, tissue-specific immune responses, and vaccines/therapeutics.

Since the first report of SARS-CoV-2 in December 2019, genetic variants have continued to emerge, complicating strategies for mitigating the disease burden of COVID-19. Positive SARS-CoV-2 nasopharyngeal swabs (n=8,735) were collected from Missouri, USA, from March-October 2020, and viral genomes (n=178) were sequenced. Hospitalization status and length of stay were extracted from medical charts of 1,335 patients and integrated with emerging genetic variants and viral shedding analyses for assessment of clinical impacts. Multiple introductions of SARS-CoV-2 into Missouri, primarily from Australia, Europe, and domestic states, were observed. Four local lineages rapidly emerged and spread across urban and rural regions in Missouri. While the majority of Missouri viruses harbored Spike-D614G mutations, a large number of unreported mutations were identified among Missouri viruses, including seven in the RNA-dependent RNA polymerase complex and Spike protein that were positively selected. A 15.6-fold increase in viral RNA levels in swab samples occurred from March to May and remained elevated. Accounting for other comorbidities, individuals test-positive for COVID-19 with high viral loads were less likely to be hospitalized (odds ratio=0.39, 95% confidence interval [CI]=0.20, 0.77) and had shorter hospital stays (hazard ratio=0.34, p=0.003) than those with low viral loads. Overall, the first eight months of the pandemic in Missouri saw multiple locally acquired mutants emerge and dominate in urban and rural locations. Although we were unable to find associations between specific variants and greater disease severity, Missouri COVID-positive individuals that presented with increased viral shedding had less severe disease by several measures.

Meaningful metrics of antiviral activity are essential for determining the efficacy of therapeutics in human clinical trials. Molnupiravir (MOV) is a broadly acting antiviral nucleoside analog prodrug that acts as a competitive alternative substrate for the SARS-CoV-2 RNA-dependent RNA polymerase (RdRp). We developed an assay, Culture-PCR, to better understand the impact of MOV therapy on infectious SARS-CoV-2. Culture-PCR revealed MOV eliminated infectious virus within 48 hours in the nasopharyngeal compartment, the upper airway location with the greatest levels of infectious virus. MOV therapy was associated with increases in mutations across the viral genome but select regions were completely unaffected, thus identifying regions where mutation likely abrogates infectivity. MOV therapy did not alter the magnitude or neutralization capacity of the humoral immune response, a documented correlate of protection. Thus, we provide holistic insights into the function of MOV in adults with COVID-19.