Seroprevalence of West Nile Virus in Tampa Bay Florida Patients Admitted to Hospital during 2020-2021 for Respiratory Symptoms.

West Nile virus (WNV) is an arbovirus spread primarily by Culex mosquitoes, with humans being a dead-end host. WNV was introduced to Florida in 2001, with 467 confirmed cases since. It is estimated that 80 percent of cases are asymptomatic, with mild cases presenting as a non-specific flu-like illness. Currently, detection of WNV in humans occurs primarily in healthcare settings via RT-PCR or CSF IgM when patients present with severe manifestations of disease including fever, meningitis, encephalitis, or acute flaccid paralysis. Given the short window of detectable viremia and requirement for CSF sampling, most WNV infections never receive an official diagnosis. This study utilized enzyme-linked immunosorbent assay (ELISA) to detect WNV IgG antibodies in 250 patient serum and plasma samples collected at Tampa General Hospital during 2020 and 2021. Plaque reduction neutralization tests were used to confirm ELISA results. Out of the 250 patients included in this study, 18.8% of them were IgG positive, consistent with previous WNV exposure. There was no relationship between WNV exposure and age or sex.

Prothrombotic autoantibodies targeting platelet factor 4/polyanion are associated with pediatric cerebral malaria.

BACKGROUNDFeatures of consumptive coagulopathy and thromboinflammation are prominent in cerebral malaria (CM). We hypothesized that thrombogenic autoantibodies contribute to a procoagulant state in CM.METHODSPlasma from children with uncomplicated malaria (UM) (n = 124) and CM (n = 136) was analyzed by ELISA for a panel of 8 autoantibodies including anti-platelet factor 4/polyanion (anti-PF4/P), anti-phospholipid, anti-phosphatidylserine, anti-myeloperoxidase, anti-proteinase 3, anti-dsDNA, anti-ß-2-glycoprotein I, and anti-cardiolipin. Plasma samples from individuals with nonmalarial coma (NMC) (n = 49) and healthy controls (HCs) (n = 56) were assayed for comparison. Associations with clinical and immune biomarkers were determined using univariate and logistic regression analyses.RESULTSMedian anti-PF4/P and anti-PS IgG levels were elevated in individuals with malaria infection relative to levels in HCs (P < 0.001) and patients with NMC (PF4/P: P < 0.001). Anti-PF4/P IgG levels were elevated in children with CM (median = 0.27, IQR: 0.19-0.41) compared with those with UM (median = 0.19, IQR: 0.14-0.22, P < 0.0001). Anti-PS IgG levels did not differ between patients with UM and those with CM (P = 0.39). When patients with CM were stratified by malaria retinopathy (Ret) status, the levels of anti-PF4/P IgG correlated negatively with the peripheral platelet count in patients with Ret+ CM (Spearman's rho [Rs] = 0.201, P = 0.04) and associated positively with mortality (OR = 15.2, 95% CI: 1.02-275, P = 0.048). Plasma from patients with CM induced greater platelet activation in an ex vivo assay relative to plasma from patients with UM (P = 0.02), and the observed platelet activation was associated with anti-PF4/P IgG levels (Rs= 0.293, P = 0.035).CONCLUSIONSThrombosis mediated by elevated anti-PF4/P autoantibodies may be one mechanism contributing to the clinical complications of CM.

Autochthonous Plasmodium vivax Infections, Florida, USA, 2023.

During May-July 2023, a cluster of 7 patients at local hospitals in Florida, USA, received a diagnosis of Plasmodium vivax malaria. Whole-genome sequencing of the organism from 4 patients and phylogenetic analysis with worldwide representative P. vivax genomes indicated probable single parasite introduction from Central/South America.

Viral and Host Factors Are Associated With Mortality in Hospitalized Patients With COVID-19.

BACKGROUND: Persistent mortality in adults hospitalized due to acute COVID-19 justifies pursuit of disease mechanisms and potential therapies. The aim was to evaluate which virus and host response factors were associated with mortality risk among participants in Therapeutics for Inpatients with COVID-19 (TICO/ACTIV-3) trials. METHODS: A secondary analysis of 2625 adults hospitalized for acute SARS-CoV-2 infection randomized to 1 of 5 antiviral products or matched placebo in 114 centers on 4 continents. Uniform, site-level collection of participant baseline clinical variables was performed. Research laboratories assayed baseline upper respiratory swabs for SARS-CoV-2 viral RNA and plasma for anti-SARS-CoV-2 antibodies, SARS-CoV-2 nucleocapsid antigen (viral Ag), and interleukin-6 (IL-6). Associations between factors and time to mortality by 90 days were assessed using univariate and multivariable Cox proportional hazards models. RESULTS: Viral Ag ≥4500 ng/L (vs <200 ng/L; adjusted hazard ratio [aHR], 2.07; 1.29-3.34), viral RNA (<35 000 copies/mL [aHR, 2.42; 1.09-5.34], ≥35 000 copies/mL [aHR, 2.84; 1.29-6.28], vs below detection), respiratory support (<4 L O2 [aHR, 1.84; 1.06-3.22]; ≥4 L O2 [aHR, 4.41; 2.63-7.39], or noninvasive ventilation/high-flow nasal cannula [aHR, 11.30; 6.46-19.75] vs no oxygen), renal impairment (aHR, 1.77; 1.29-2.42), and IL-6 >5.8 ng/L (aHR, 2.54 [1.74-3.70] vs ≤5.8 ng/L) were significantly associated with mortality risk in final adjusted analyses. Viral Ag, viral RNA, and IL-6 were not measured in real-time. CONCLUSIONS: Baseline virus-specific, clinical, and biological variables are strongly associated with mortality risk within 90 days, revealing potential pathogen and host-response therapeutic targets for acute COVID-19 disease.

Diabetic Retinopathy during pregnancy in Hispanic women with latent Toxoplasma gondii infection.

BACKGROUND: Retinal photography was performed in pregnancy and postpartum in pregnant Hispanic women with latent Toxoplasma gondii (TG) infection in order to screen for characteristic retinal lesions or the particular scars found in people with active T. gondii infection. A comparison group of TG negative women was included in the study but they did not have retinal photography. OBJECTIVE: The goal of the parent study was to assess for adverse pregnancy events and evidence for parasite reactivation in TG positive (TG + ) women, through examination of the eyes for characteristic lesions. Retinal photography, usually at prenatal visits 2 (17 +/- 3.35 weeks) and 3 (26.3+/-1.75) weeks, was done on TG + women. Fifty-six of these women also (43 %) had retinal photography at the postpartum visit. Health and demographic data were obtained at the first prenatal visit for all women. STUDY DESIGN: From the 690 recruited at the first prenatal visit, 128 TG- women and 158 TG + women were enrolled in a prospective study through pregnancy and the postpartum. All TG- women (n = 532) provided data at the first prenatal visit and throughout their pregnancy and birth through the EHR. This allowed comparison of health and outcome data for the TG + compared to a larger number of TG- Hispanic pregnant women. RESULTS: While there was no evidence of ocular toxoplasmosis during pregnancy, there was a surprisingly large number (42 %) of TG + women with diabetic retinopathy (DR). We also observed that TG + women had a 20 % incidence of gestational diabetes mellitus (GDM) compared to 11.3 % in the TG- women (p = 0.01). At postpartum (mean 5.6 weeks), 23 of 30 women with pregnancy DR showed no DR in the postpartum. CONCLUSIONS: No characteristic T. gondii lesions were discovered. Retinal photography serendipitously revealed DR in these T. gondii positive women. It was also found that latent TG infection was associated with increased incidence of GDM. Hispanic pregnant women's increased risk for latent TG infection, GDM and DR are underappreciated. Retinal photography may need to be considered an innovative approach to screening.

Draft genome sequence of a highly proteolytic Staphylococcus aureus USA300 isolate from human urine.

The secreted proteases of Staphylococcus aureus have been shown to be critical during infection. Here, we present the draft genome sequence of S. aureus TGH337, a hyper-proteolytic USA300 strain isolated from human urine.

Emergent Risk Group-4 (RG-4) Filoviruses: A paradox in progress.

Filoviruses, categorized as World Health Organization (WHO) Risk Group 4 (RG-4) pathogens, represent significant global health risks due to their extraordinary virulence. The Filoviridae family encompasses Ebola strains such as Sudan, Zaire, Bundibugyo, Tai Forest (formerly known as Ivory Coast), Reston, and Bombali, in addition to the closely related Marburg and Ravn virus strains. Filoviruses originated from a common ancestor about 10,000 years ago and displayed remarkable consistency in genetic heterogeneity until the 20th century. However, they overcame a genetic bottleneck by mid-century. Paradoxically, this resulted in the emergence of boosted virulent strains from the 1970's onward. Filovirus research is included in the NIAID Biodefense Program and utilizes the highest level specialized protective laboratories, Biosafety Laboratory (BSL)-4. The spread of Filoviruses as well as other RG-4 pathogens within Africa poses a significant health threat increasingly both in Africa and out of Africa.

MalariaSED: a deep learning framework to decipher the regulatory contributions of noncoding variants in malaria parasites.

Malaria remains one of the deadliest infectious diseases. Transcriptional regulation effects of noncoding variants in this unusual genome of malaria parasites remain elusive. We developed a sequence-based, ab initio deep learning framework, MalariaSED, for predicting chromatin profiles in malaria parasites. The MalariaSED performance was validated by published ChIP-qPCR and TF motifs results. Applying MalariaSED to ~ 1.3 million variants shows that geographically differentiated noncoding variants are associated with parasite invasion and drug resistance. Further analysis reveals chromatin accessibility changes at Plasmodium falciparum rings are partly associated with artemisinin resistance. MalariaSED illuminates the potential functional roles of noncoding variants in malaria parasites.

Tryptophan metabolism and immune alterations in pregnant Hispanic women with chronic Toxoplasma gondii infection.

PROBLEM: Pregnancy markedly modifies women's metabolism and immune functions. We hypothesized that pregnancy might alter the immune and metabolic responses to chronic Toxoplasma gondii infection in pregnancy. METHOD OF STUDY: A population of 690 pregnant Hispanic women were screened for antibodies to T. gondii and 158 women were positive (23% positivity) with 83% showing high avidity indices. These seropositive women were followed through their pregnancies with four data collection time points and a postpartum collection at two clinics in Tampa, Florida. A T. gondii seronegative group (N = 128) was randomly selected to serve as a control group and measured along pregnancy in the same way. Serum levels of tryptophan, kynurenine, and their ratio, phenylalanine, tyrosine and their ratio, neopterin, and nitrite were measured through pregnancy and the postpartum. A plasma cytokine panel (IFN-γ, TNFα, IL-2, IL-10, IL-12, IL-6, IL-17) was analyzed in parallel. RESULTS: The major findings suggest that indoleamine 2,3-dioxygenase (IDO-1) was less activated in T. gondii seropositive pregnant Hispanic women with chronic infection. Evidence for IDO-1 suppression was that tryptophan catabolism was less pronounced and there were lower levels of multiple inflammatory cytokines including IFN-γ, which is the major inducer of IDO-1, and higher nitrite concentration, a surrogate marker for nitric oxide, an inhibitor of IDO. CONCLUSIONS: Latent T. gondii infection was associated with higher plasma tryptophan levels, and lower inflammatory cytokines across pregnancy, suggesting suppression of the IDO-1 enzyme, and possible T cell exhaustion during pregnancy.

Evaluation of severe acute respiratory syndrome coronavirus 2 monoclonal antibodies in high-risk solid organ transplant recipients across three major coronavirus disease 2019 variant waves.

BACKGROUND: Coronavirus disease 2019 (COVID-19) continues to impact immunocompromised populations including solid organ transplant recipients (SOTRs). Monoclonal antibodies (mAbs) have shown effectiveness in reducing COVID-19-related hospitalizations and emergency department (ED) visits in SOTRs at different time frames in the COVID-19 pandemic; however, less data exist on the impact of mAbs for SOTRs across variant waves and with the advent of available COVID-19 vaccines. METHODS: This retrospective study included SOTR outpatients who tested positive for SARS-CoV-2 and received mAbs from December 2020 to February 2022 (n = 233); using in-house sequencing of clinical samples, we monitored the emergence of Alpha, Delta, and Omicron variants. The primary outcome was a composite of 29-day COVID-19-related hospitalizations and ED visits. Prespecified secondary outcomes included individual components of the primary endpoint; for patients requiring hospitalization post-mAb administration, we describe their inpatient treatment. RESULTS: A low percentage of SOTRs treated with mAb required hospitalization or an ED visit (14.6% overall); this did not differ across COVID-19 variants (p = .152). Hospitalization and ED visits did not significantly differ between abdominal and cardiothoracic SOTRs. For hospitalized patients, the majority received treatment with corticosteroids and few required intensive care unit (ICU) care. CONCLUSION: Among SOTR outpatients with mild or moderate COVID-19 symptoms, early administration of mAb minimizes the need for hospital care. For patients requiring hospitalization, corticosteroids were common but patients experienced low rates of oxygen supplementation and ICU care. Use of mAbs in SOTRs should be considered early in the disease when therapy is available.

Histone variant H2B.Z acetylation is necessary for maintenance of Toxoplasma gondii biological fitness.

Through regulation of DNA packaging, histone proteins are fundamental to a wide array of biological processes. A variety of post-translational modifications (PTMs), including acetylation, constitute a proposed histone code that is interpreted by "reader" proteins to modulate chromatin structure. Canonical histones can be replaced with variant versions that add an additional layer of regulatory complexity. The protozoan parasite Toxoplasma gondii is unique among eukaryotes in possessing a novel variant of H2B designated H2B.Z. The combination of PTMs and the use of histone variants are important for gene regulation in T. gondii, offering new targets for drug development. In this work, T. gondii parasites were generated in which the 5 N-terminal acetylatable lysines in H2B.Z were mutated to either alanine (c-Myc-A) or arginine (c-Myc-R). The c-Myc-A mutant displayed no phenotype over than a mild defect in its ability to kill mice. The c-Myc-R mutant presented an impaired ability to grow and an increase in differentiation to latent bradyzoites. The c-Myc-R mutant was also more sensitive to DNA damage, displayed no virulence in mice, and provided protective immunity against future infection. While nucleosome composition was unaltered, key genes were abnormally expressed during in vitro bradyzoite differentiation. Our results show that regulation of the N-terminal positive charge patch of H2B.Z is important for these processes. We also show that acetylated N-terminal H2B.Z interacts with some unique proteins compared to its unacetylated counterpart; the acetylated peptide pulled down proteins associated with chromosome maintenance/segregation and cell cycle, suggesting a link between H2B.Z acetylation status and mitosis.

Clinical Spectrum, Radiological Findings, and Outcomes of Severe Toxoplasmosis in Immunocompetent Hosts: A Systematic Review.

BACKGROUND: Accumulating evidence suggests that toxoplasmosis in immunocompetent hosts can be severe and life-threatening. METHODS: We performed a systematic review of severe toxoplasmosis cases in immunocompetent patients to gain insight into the epidemiology, clinical characteristics, radiological findings, and outcomes of these cases. We classified severe toxoplasmosis as cases with the symptomatic involvement of target organs (the lungs, central nervous system (CNS), and heart), disseminated disease, prolonged disease (>3 months), or a fatal outcome. Our primary analysis focused on cases published from 1985-2022 to avoid confounding with cases in AIDS patients. RESULTS: We identified 82 pertinent articles (1985-2022) with a total of 117 eligible cases; the top five countries for these cases were French Guiana (20%), France (15%), Colombia (9%), India (9%), and Brazil (7%). Overall, 44% (51/117) of cases had pulmonary involvement, 39% (46/117) CNS, 31% (36/117) cardiac, 24% (28/117) disseminated disease, 2% (2/117) had prolonged disease, and 8% (9/117) of patients died. More than one organ was involved in 26% (31/117) of cases. Eighty-four percent (98/117) of cases occurred in the context of a recent acute primary Toxoplasma infection; for the remaining, the exact timing of infection was unclear. Genotyping data were very sparse. Among those reporting genotyping data, 96% (22/23) were caused by atypical non-type II strains; one case was caused by a type-II strain. Only half of the cases reported risk factors. The most common risk factors were eating raw/undercooked meat or eating game meat (47% (28/60)), drinking untreated water (37% (22/60)), or living in a toxoplasmosis high-prevalence area (38% (23/60)). For the 51 pulmonary cases, the main clinical presentation was pneumonia or pleural effusions in 94% (48/51) and respiratory failure in 47% (24/51). For the 46 CNS cases, the main clinical presentation was encephalitis in 54% (25/46), meningitis in 13% (6/46), focal neurologic findings in 24% (11/46), cranial nerve palsies in 17% (8/46), Guillain-Barre syndrome or Miller Fisher syndrome in 7% (3/46), and Brown-Sequard syndrome in 2% (1/46) of cases; more than one clinical manifestation could also be present. Among the 41 CNS cases reporting the CNS imaging findings, 68% (28/41) had focal supratentorial lesions and 7% (3/41) had focal infratentorial lesions. Brain abscess-like/mass-like lesions were seen in 51% (21/41) of cases. For the 36 cardiac cases, the main clinical presentation was myocarditis in 75% (27/36), pericarditis in 50% (18/36), heart failure and/or cardiogenic shock in 19% (7/36), and cardiac arrhythmias in 22% (8/36); more than one manifestation could also be present. Illness was critical in 49% (44/90) of cases intensive care unit care was needed in 54% (29/54) of cases among those reporting this information, and 9 patients died. CONCLUSION: The diagnosis of severe toxoplasmosis in immunocompetent hosts can be challenging. Toxoplasmosis should be considered in the differential diagnosis of immunocompetent patients presenting with severe illness of unclear etiology with pulmonary, cardiac, CNS, or multiorgan involvement/failure, or prolonged febrile illness, even in the absence of common exposure risk factors or common manifestations of toxoplasmosis (e.g., fever, mononucleosis-like illness, lymphadenopathy, and chorioretinitis). Fatal outcomes can also rarely occur in immunocompetent patients. Prompt initiation of anti-Toxoplasma treatment can be lifesaving.

Histone variant H2B.Z acetylation is necessary for maintenance of Toxoplasma gondii biological fitness.

Through regulation of DNA packaging, histone proteins are fundamental to a wide array of biological processes. A variety of post-translational modifications (PTMs), including acetylation, constitute a proposed histone code that is interpreted by "reader" proteins to modulate chromatin structure. Canonical histones can be replaced with variant versions that add an additional layer of regulatory complexity. The protozoan parasite Toxoplasma gondii is unique among eukaryotes in possessing a novel variant of H2B designated H2B.Z. The combination of PTMs and the use of histone variants is important for gene regulation in T. gondii, offering new targets for drug development. In this work, T. gondii parasites were generated in which the 5 N-terminal acetylatable lysines in H2B.Z were mutated to either alanine (c-Myc-A) or arginine (c-Myc-R). c-Myc-A mutant only displayed a mild effect in its ability to kill mice. c-Myc-R mutant presented an impaired ability to grow and an increase in differentiation to latent bradyzoites. This mutant line was also more sensitive to DNA damage, displayed no virulence in mice, and provided protective immunity against future infection. While nucleosome composition was unaltered, key genes were abnormally expressed during in vitro bradyzoite differentiation. Our results show that the N-terminal positive charge patch of H2B.Z is important for these procceses. Pull down assays with acetylated N-terminal H2B.Z peptide and unacetylated one retrieved common and differential interactors. Acetylated peptide pulled down proteins associated with chromosome maintenance/segregation and cell cycle, opening the question of a possible link between H2B.Z acetylation status and mitosis.

Adverse pregnancy outcomes in Toxoplasma gondii seropositive Hispanic women.

AIMS: Chronic Toxoplasma gondii infection is not thought to affect pregnancy or birth outcomes, but there are few prospective studies. The study aims were T. gondii immunoglobulin G measurement and relationship of chronic T. gondii infection with gestational age at birth and adverse pregnancy outcomes in 690 Hispanic women in Tampa, Florida. METHODS: Hispanic women, born either in the United States or in Latin America or the Caribbean had a venous blood sample drawn to measure T. gondii IgG and T. gondii serotype at the first prenatal visit, along with collection of demographic and health-related measures. Seropositive and seronegative women were followed throughout their pregnancy. Gestational age, infant weights, and adverse pregnancy outcomes (miscarriages, preterm births) were compared in the two groups. RESULTS: There were 740 women of self-reported Hispanic ethnicity screened and enrolled in Tampa, Florida, with 690 having birth data extracted from the electronic health record (538 T. gondii negative and 152 T. gondii seropositive). T. gondii seropositivity was 22.4% and the majority (83%) had high avidity titers, indicating chronic infection. Compared to T. gondii seronegative Hispanic women, seroseropositive women had more smaller for gestational age infants and higher prevalences of miscarriages and preterm birth. CONCLUSION: This is one of the largest longitudinal cohort studies of women with chronic T. gondii infection followed through pregnancy. There was a higher percentages of adverse pregnancy outcomes in this group compared to T. gondii seronegative controls. The mechanism for this is unknown and warrants reexamination of the dogma that chronic T. gondii infection in pregnant women has no significant clinical consequences.

Risk of hemolysis in Plasmodium vivax malaria patients receiving standard primaquine treatment in a population with high prevalence of G6PD deficiency.

BACKGROUND: Primaquine is essential for the radical cure of Plasmodium vivax malaria, but it poses a potential danger of severe hemolysis in G6PD-deficient (G6PDd) patients. This study aimed to determine whether primaquine is safe in a population with high G6PD prevalence but lacking G6PD diagnosis capacity. METHODS: In Myanmar, 152 vivax patients were gender- and age-matched at 1:3 for G6PDd versus G6PD-normal (G6PDn). Their risk of acute hemolysis was followed for 28 days after treatment with the standard chloroquine and 14-day primaquine (0.25 mg/kg/day) regimen. RESULTS: Patients anemic and non-anemic at enrollment showed a rising and declining trend in the mean hemoglobin level, respectively. In males, the G6PDd group showed substantially larger magnitudes of hemoglobin reduction and lower hemoglobin nadir levels than the G6PDn group, but this trend was not evident in females. Almost 1/3 of the patients experienced clinically concerning declines in hemoglobin, with five requiring blood transfusion. CONCLUSIONS: The standard 14-day primaquine regimen carries a significant risk of acute hemolytic anemia (AHA) in vivax patients without G6PD testing in a population with a high prevalence of G6PD deficiency and anemia. G6PD testing would avoid most of the clinically significant Hb reductions and AHA in male patients.

Architecture, Chromatin and Gene Organization of Toxoplasma gondii Subtelomeres.

Subtelomeres (ST) are chromosome regions that separate telomeres from euchromatin and play relevant roles in various biological processes of the cell. While their functions are conserved, ST structure and genetic compositions are unique to each species. This study aims to identify and characterize the subtelomeric regions of the 13 Toxoplasma gondii chromosomes of the Me49 strain. Here, STs were defined at chromosome ends based on poor gene density. The length of STs ranges from 8.1 to 232.4 kbp, with a gene density of 0.049 genes/kbp, lower than the Me49 genome (0.15 kbp). Chromatin organization showed that H3K9me3, H2A.X, and H3.3 are highly enriched near telomeres and the 5' end of silenced genes, decaying in intensity towards euchromatin. H3K4me3 and H2A.Z/H2B.Z are shown to be enriched in the 5' end of the ST genes. Satellite DNA was detected in almost all STs, mainly the sat350 family and a novel satellite named sat240. Beyond the STs, only short dispersed fragments of sat240 and sat350 were found. Within STs, there were 12 functional annotated genes, 59 with unknown functions (Hypothetical proteins), 15 from multigene FamB, and 13 from multigene family FamC. Some genes presented low interstrain synteny associated with the presence of satellite DNA. Orthologues of FamB and FamC were also detected in Neospora caninum and Hammondia hammondi. A re-analysis of previous transcriptomic data indicated that ST gene expression is strongly linked to the adaptation to different situations such as extracellular passage (evolve and resequencing study) and changes in metabolism (lack of acetyl-CoA cofactor). In conclusion, the ST region of the T. gondii chromosomes was defined, the STs genes were determined, and it was possible to associate them with high interstrain plasticity and a role in the adaptability of T. gondii to environmental changes.

The Association of Baseline Plasma SARS-CoV-2 Nucleocapsid Antigen Level and Outcomes in Patients Hospitalized With COVID-19.

BACKGROUND: Levels of plasma SARS-CoV-2 nucleocapsid (N) antigen may be an important biomarker in patients with COVID-19 and enhance our understanding of the pathogenesis of COVID-19. OBJECTIVE: To evaluate whether levels of plasma antigen can predict short-term clinical outcomes and identify clinical and viral factors associated with plasma antigen levels in hospitalized patients with SARS-CoV-2. DESIGN: Cross-sectional study of baseline plasma antigen level from 2540 participants enrolled in the TICO (Therapeutics for Inpatients With COVID-19) platform trial from August 2020 to November 2021, with additional data on day 5 outcome and time to discharge. SETTING: 114 centers in 10 countries. PARTICIPANTS: Adults hospitalized for acute SARS-CoV-2 infection with 12 days or less of symptoms. MEASUREMENTS: Baseline plasma viral N antigen level was measured at a central laboratory. Delta variant status was determined from baseline nasal swabs using reverse transcriptase polymerase chain reaction. Associations between baseline patient characteristics and viral factors and baseline plasma antigen levels were assessed using both unadjusted and multivariable modeling. Association between elevated baseline antigen level of 1000 ng/L or greater and outcomes, including worsening of ordinal pulmonary scale at day 5 and time to hospital discharge, were evaluated using logistic regression and Fine-Gray regression models, respectively. RESULTS: Plasma antigen was below the level of quantification in 5% of participants at enrollment, and 1000 ng/L or greater in 57%. Baseline pulmonary severity of illness was strongly associated with plasma antigen level, with mean plasma antigen level 3.10-fold higher among those requiring noninvasive ventilation or high-flow nasal cannula compared with room air (95% CI, 2.22 to 4.34). Plasma antigen level was higher in those who lacked antispike antibodies (6.42 fold; CI, 5.37 to 7.66) and in those with the Delta variant (1.73 fold; CI, 1.41 to 2.13). Additional factors associated with higher baseline antigen level included male sex, shorter time since hospital admission, decreased days of remdesivir, and renal impairment. In contrast, race, ethnicity, body mass index, and immunocompromising conditions were not associated with plasma antigen levels. Plasma antigen level of 1000 ng/L or greater was associated with a markedly higher odds of worsened pulmonary status at day 5 (odds ratio, 5.06 [CI, 3.41 to 7.50]) and longer time to hospital discharge (median, 7 vs. 4 days; subhazard ratio, 0.51 [CI, 0.45 to 0.57]), with subhazard ratios similar across all levels of baseline pulmonary severity. LIMITATIONS: Plasma samples were drawn at enrollment, not hospital presentation. No point-of-care test to measure plasma antigen is currently available. CONCLUSION: Elevated plasma antigen is highly associated with both severity of pulmonary illness and clinically important patient outcomes. Multiple clinical and viral factors are associated with plasma antigen level at presentation. These data support a potential role of ongoing viral replication in the pathogenesis of SARS-CoV-2 in hospitalized patients. PRIMARY FUNDING SOURCE: U.S. government Operation Warp Speed and National Institute of Allergy and Infectious Diseases.

Real-world effectiveness of early remdesivir and sotrovimab in the highest-risk COVID-19 outpatients during the Omicron surge.

BACKGROUND: Remdesivir and sotrovimab both have clinical trial data in the outpatient setting demonstrating reduction in the risk of hospitalizations and emergency department (ED) visits related to COVID-19. OBJECTIVES: To evaluate the effectiveness of remdesivir in comparison with sotrovimab and matched high-risk control patients in preventing COVID-19-related hospitalizations and ED visits during the Omicron B.1.1.529 surge. PATIENTS AND METHODS: This retrospective cohort study included outpatients positive for SARS-CoV-2, with non-severe symptoms for ≤7 days and deemed high-risk for severe COVID-19 by an internal scoring matrix. Patients who received remdesivir or sotrovimab from 27/12/2021 to 04/02/2022 were included (n = 82 and n = 88, respectively). These were compared with a control cohort of high-risk COVID-19 outpatients who did not receive therapy (n = 90). The primary outcome was a composite of 29 day COVID-19-related hospitalizations and/or ED visits. Pre-specified secondary outcomes included components of the primary endpoint, 29 day all-cause mortality and serious adverse drug events. RESULTS: Patients treated with remdesivir were significantly less likely to be hospitalized or visit the ED within 29 days from symptom onset (11% versus 23.3%; OR = 0.41, 95% CI = 0.17-0.95). Patients receiving sotrovimab were also less likely to be hospitalized or visit the ED (8% versus 23.3%; OR = 0.28, 95% CI = 0.11-0.71). There was no difference in the incidence of hospitalizations/ED visits between sotrovimab and remdesivir. CONCLUSIONS: Our highest-risk outpatients with Omicron-related COVID-19 who received early sotrovimab or remdesivir had significantly lower likelihoods of a hospitalization and/or ED visit.

Restriction Checkpoint Controls Bradyzoite Development in Toxoplasma gondii.

Human toxoplasmosis is a life-threatening disease caused by the apicomplexan parasite Toxoplasma gondii. Rapid replication of the tachyzoite is associated with symptomatic disease, while suppressed division of the bradyzoite is responsible for chronic disease. Here, we identified the T. gondii cell cycle mechanism, the G1 restriction checkpoint (R-point), that operates the switch between parasite growth and differentiation. Apicomplexans lack conventional R-point regulators, suggesting adaptation of alternative factors. We showed that Cdk-related G1 kinase TgCrk2 forms alternative complexes with atypical cyclins (TgCycP1, TgCycP2, and TgCyc5) in the rapidly dividing developmentally incompetent RH and slower dividing developmentally competent ME49 tachyzoites and bradyzoites. Examination of cyclins verified the correlation of cyclin expression with growth dependence and development capacity of RH and ME49 strains. We demonstrated that rapidly dividing RH tachyzoites were dependent on TgCycP1 expression, which interfered with bradyzoite differentiation. Using the conditional knockdown model, we established that TgCycP2 regulated G1 duration in the developmentally competent ME49 tachyzoites but not in the developmentally incompetent RH tachyzoites. We tested the functions of TgCycP2 and TgCyc5 in alkaline induced and spontaneous bradyzoite differentiation (rat embryonic brain cells) models. Based on functional and global gene expression analyses, we determined that TgCycP2 also regulated bradyzoite replication, while signal-induced TgCyc5 was critical for efficient tissue cyst maturation. In conclusion, we identified the central machinery of the T. gondii restriction checkpoint comprised of TgCrk2 kinase and three atypical T. gondii cyclins and demonstrated the independent roles of TgCycP1, TgCycP2, and TgCyc5 in parasite growth and development. IMPORTANCE Toxoplasma gondii is a virulent and abundant human pathogen that puts millions of silently infected people at risk of reactivation of the chronic disease. Encysted bradyzoites formed during the chronic stage are resistant to current therapies. Therefore, insights into the mechanism of tissue cyst formation and reactivation are major areas of investigation. The fact that rapidly dividing parasites differentiate poorly strongly suggests that there is a threshold of replication rate that must be crossed to be considered for differentiation. We discovered a cell cycle mechanism that controls the T. gondii growth-rest switch involved in the conversion of dividing tachyzoites into largely quiescent bradyzoites. This switch operates the T. gondii restriction checkpoint using a set of atypical and parasite-specific regulators. Importantly, the novel T. gondii R-point network was not present in the parasite's human and animal hosts, offering a wealth of new and parasite-specific drug targets to explore in the future.