Outpatient COVID-19 convalescent plasma recipient antibody thresholds correlated to reduced hospitalizations within a randomized trial.

BACKGROUNDCOVID-19 convalescent plasma (CCP) virus-specific antibody levels that translate into recipient posttransfusion antibody levels sufficient to prevent disease progression are not defined.METHODSThis secondary analysis correlated donor and recipient antibody levels to hospitalization risk among unvaccinated, seronegative CCP recipients within the outpatient, double-blind, randomized clinical trial that compared CCP to control plasma. The majority of COVID-19 CCP arm hospitalizations (15/17, 88%) occurred in this unvaccinated, seronegative subgroup. A functional cutoff to delineate recipient high versus low posttransfusion antibody levels was established by 2 methods: (i) analyzing virus neutralization-equivalent anti-Spike receptor-binding domain immunoglobulin G (anti-S-RBD IgG) responses in donors or (ii) receiver operating characteristic (ROC) curve analysis.RESULTSSARS-CoV-2 anti-S-RBD IgG antibody was volume diluted 21.3-fold into posttransfusion seronegative recipients from matched donor units. Virus-specific antibody delivered was approximately 1.2 mg. The high-antibody recipients transfused early (symptom onset within 5 days) had no hospitalizations. A CCP-recipient analysis for antibody thresholds correlated to reduced hospitalizations found a statistical significant association between early transfusion and high antibodies versus all other CCP recipients (or control plasma), with antibody cutoffs established by both methods-donor-based virus neutralization cutoffs in posttransfusion recipients (0/85 [0%] versus 15/276 [5.6%]; P = 0.03) or ROC-based cutoff (0/94 [0%] versus 15/267 [5.4%]; P = 0.01).CONCLUSIONIn unvaccinated, seronegative CCP recipients, early transfusion of plasma units in the upper 30% of study donors' antibody levels reduced outpatient hospitalizations. High antibody level plasma units, given early, should be reserved for therapeutic use.TRIAL REGISTRATIONClinicalTrials.gov NCT04373460.FUNDINGDepartment of Defense (W911QY2090012); Defense Health Agency; Bloomberg Philanthropies; the State of Maryland; NIH (3R01AI152078-01S1, U24TR001609-S3, 1K23HL151826NIH); the Mental Wellness Foundation; the Moriah Fund; Octapharma; the Healthnetwork Foundation; the Shear Family Foundation; the NorthShore Research Institute; and the Rice Foundation.

Synthetic Glycans Reveal Determinants of Antibody Functional Efficacy against a Fungal Pathogen.

Antibodies play a vital role in the immune response to infectious diseases and can be administered passively to protect patients. In the case of Cryptococcus neoformans, a WHO critical priority fungal pathogen, infection results in antibodies targeting capsular glucuronoxylomannan (GXM). These antibodies yield protective, non-protective, and disease-enhancing outcomes when administered passively. However, it was unknown how these distinct antibodies recognized their antigens at the molecular level, leading to the hypothesis that they may target different GXM epitopes. To test this hypothesis, we constructed a microarray containing 26 glycans representative of those found in highly virulent cryptococcal strains and utilized it to study 16 well-characterized monoclonal antibodies. Notably, we found that protective and non-protective antibodies shared conserved reactivity to the M2 motif of GXM, irrespective of the strain used in infection or GXM-isolated to produce a conjugate vaccine. Here, only two antibodies, 12A1 and 18B7, exhibited diverse trivalent GXM motif reactivity. IgG antibodies associated with protective responses showed cross-reactivity to at least two GXM motifs. This molecular understanding of antibody binding epitopes was used to map the antigenic diversity of two Cryptococcus neoformans strains, which revealed the exceptional complexity of fungal capsular polysaccharides. A multi-GXM motif vaccine holds the potential to effectively address this antigenic diversity. Collectively, these findings underscore the context-dependent nature of antibody function and challenge the classification of anti-GXM epitopes as either "protective" or "non-protective".

Clinically relevant atovaquone-resistant human malaria parasites fail to transmit by mosquito.

Long-acting injectable medications, such as atovaquone, offer the prospect of a "chemical vaccine" for malaria, combining drug efficacy with vaccine durability. However, selection and transmission of drug-resistant parasites is of concern. Laboratory studies have indicated that atovaquone resistance disadvantages parasites in mosquitoes, but lack of data on clinically relevant Plasmodium falciparum has hampered integration of these variable findings into drug development decisions. Here we generate atovaquone-resistant parasites that differ from wild type parent by only a Y268S mutation in cytochrome b, a modification associated with atovaquone treatment failure in humans. Relative to wild type, Y268S parasites evidence multiple defects, most marked in their development in mosquitoes, whether from Southeast Asia (Anopheles stephensi) or Africa (An. gambiae). Growth of asexual Y268S P. falciparum in human red cells is impaired, but parasite loss in the mosquito is progressive, from reduced gametocyte exflagellation, to smaller number and size of oocysts, and finally to absence of sporozoites. The Y268S mutant fails to transmit from mosquitoes to mice engrafted with human liver cells and erythrocytes. The severe-to-lethal fitness cost of clinically relevant atovaquone resistance to P. falciparum in the mosquito substantially lessens the likelihood of its transmission in the field.

The Influenza B Virus Victoria and Yamagata Lineages Display Distinct Cell Tropism and Infection-Induced Host Gene Expression in Human Nasal Epithelial Cell Cultures.

Understanding Influenza B virus infections is of critical importance in our efforts to control severe influenza and influenza-related diseases. Until 2020, two genetic lineages of influenza B virus-Yamagata and Victoria-circulated in the population. These lineages are antigenically distinct, but the differences in virus replication or the induction of host cell responses after infection have not been carefully studied. Recent IBV clinical isolates of both lineages were obtained from influenza surveillance efforts of the Johns Hopkins Center of Excellence in Influenza Research and Response and characterized in vitro. B/Victoria and B/Yamagata clinical isolates were recognized less efficiently by serum from influenza-vaccinated individuals in comparison to the vaccine strains. B/Victoria lineages formed smaller plaques on MDCK cells compared to B/Yamagata, but infectious virus production in primary human nasal epithelial cell (hNEC) cultures showed no differences. While ciliated epithelial cells were the dominant cell type infected by both lineages, B/Victoria lineages had a slight preference for MUC5AC-positive cells, and B/Yamagata lineages infected more basal cells. Finally, while both lineages induced a strong interferon response 48 h after infection of hNEC cultures, the B/Victoria lineages showed a much stronger induction of interferon-related signaling pathways compared to B/Yamagata. This demonstrates that the two influenza B virus lineages differ not only in their antigenic structure but also in their ability to induce host innate immune responses.

The Influenza B Virus Victoria and Yamagata Lineages Display Distinct Cell Tropism and Infection Induced Host Gene Expression in Human Nasal Epithelial Cell Cultures.

Understanding Influenza B virus infections is of critical importance in our efforts to control severe influenza and influenza-related disease. Until 2020, two genetic lineages of influenza B virus - Yamagata and Victoria - circulated in the population. These lineages are antigenically distinct but differences in virus replication or the induction of host cell responses after infection have not been carefully studied. Recent IBV clinical isolates of both lineages were obtained from influenza surveillance efforts of the Johns Hopkins Center of Excellence in Influenza Research and Response and characterized in vitro . B/Victoria and B/Yamagata clinical isolates were recognized less efficiently by serum from influenza-vaccinated individuals in comparison to the vaccine strains. B/Victoria lineages formed smaller plaques on MDCK cells compared to B/Yamagata, but infectious virus production in primary human nasal epithelial cell (hNEC) cultures showed no differences. While ciliated epithelial cells were the dominant cell type infected by both lineages, B/Victoria lineages had a slight preference for MUC5AC-positive cells, while B/Yamagata lineages infected more basal cells. Finally, while both lineages induced a strong interferon response 48 hours after infection of hNEC cultures, the B/Victoria lineages showed a much stronger induction of interferon related signaling pathways compared to B/Yamagata. This demonstrates that the two influenza B virus lineages differ not only in their antigenic structure but in their ability to induce host innate immune responses.

SARS-CoV-2 infection alters mitochondrial and cytoskeletal function in human respiratory epithelial cells mediated by expression of spike protein.

Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2, SCV2), which has resulted in higher morbidity and mortality rate than other respiratory viral infections, such as Influenza A virus (IAV) infection. Investigating the molecular mechanisms of SCV2-host infection vs IAV is vital in exploring antiviral drug targets against SCV2. We assessed differential gene expression in human nasal cells upon SCV2 or IAV infection using RNA sequencing. Compared to IAV, we observed alterations in both metabolic and cytoskeletal pathways suggestive of epithelial remodeling in the SCV2-infected cells, reminiscent of pathways activated as a response to chronic injury. We found that spike protein interaction with the epithelium was sufficient to instigate these epithelial responses using a SCV2 spike pseudovirus. Specifically, we found downregulation of the mitochondrial markers SIRT3 and TOMM22. Moreover, SCV2 spike infection increased extracellular acidification and decreased oxygen consumption rate in the epithelium. In addition, we observed cytoskeletal rearrangements with a reduction in the actin-severing protein cofilin-1 and an increase in polymerized actin, indicating epithelial cytoskeletal rearrangements. This study revealed distinct epithelial responses to SCV2 infection, with early mitochondrial dysfunction in the host cells and evidence of cytoskeletal remodeling that could contribute to the worsened outcome in COVID-19 patients compared to IAV patients. These changes in cell structure and energetics could contribute to cellular resilience early during infection, allowing for prolonged cell survival and potentially paving the way for more chronic symptoms. IMPORTANCE COVID-19 has caused a global pandemic affecting millions of people worldwide, resulting in a higher mortality rate and concerns of more persistent symptoms compared to influenza A. To study this, we compare lung epithelial responses to both viruses. Interestingly, we found that in response to SARS-CoV-2 infection, the cellular energetics changed and there were cell structural rearrangements. These changes in cell structure could lead to prolonged epithelial cell survival, even in the face of not working well, potentially contributing to the development of chronic symptoms. In summary, these findings represent strategies utilized by the cell to survive the infection but result in a fundamental shift in the epithelial phenotype, with potential long-term consequences, which could set the stage for the development of chronic lung disease or long COVID-19.

2019-2020 H1N1 clade A5a.1 viruses have better in vitro fitness compared with the co-circulating A5a.2 clade.

Surveillance for emerging human influenza virus clades is important for identifying changes in viral fitness and assessing antigenic similarity to vaccine strains. While fitness and antigenic structure are both important aspects of virus success, they are distinct characteristics and do not always change in a complementary manner. The 2019-2020 Northern Hemisphere influenza season saw the emergence of two H1N1 clades: A5a.1 and A5a.2. While several studies indicated that A5a.2 showed similar or even increased antigenic drift compared with A5a.1, the A5a.1 clade was still the predominant circulating clade that season. Clinical isolates of representative viruses from these clades were collected in Baltimore, Maryland during the 2019-2020 season and multiple assays were performed to compare both antigenic drift and viral fitness between clades. Neutralization assays performed on serum from healthcare workers pre- and post-vaccination during the 2019-2020 season show a comparable drop in neutralizing titers against both A5a.1 and A5a.2 viruses compared with the vaccine strain, indicating that A5a.1 did not have antigenic advantages over A5a.2 that would explain its predominance in this population. Plaque assays were performed to investigate fitness differences, and the A5a.2 virus produced significantly smaller plaques compared with viruses from A5a.1 or the parental A5a clade. To assess viral replication, low MOI growth curves were performed on both MDCK-SIAT and primary differentiated human nasal epithelial cell cultures. In both cell cultures, A5a.2 yielded significantly reduced viral titers at multiple timepoints post-infection compared with A5a.1 or A5a. Receptor binding was then investigated through glycan array experiments which showed a reduction in receptor binding diversity for A5a.2, with fewer glycans bound and a higher percentage of total binding attributable to the top three highest bound glycans. Together these data indicate that the A5a.2 clade had a reduction in viral fitness, including reductions in receptor binding, that may have contributed to the limited prevalence observed after emergence.

Outpatient COVID-19 convalescent plasma recipient antibody thresholds correlated to reduced hospitalizations within a randomized trial.

BACKGROUND: The COVID-19 convalescent plasma (CCP) viral specific antibody levels that translate into recipient post-transfusion antibody levels sufficient to prevent disease progression is not defined. METHODS: This secondary analysis correlated donor and recipient antibody levels to hospitalization risk among unvaccinated, seronegative CCP recipients within the outpatient, double blind, randomized clinical trial that compared CCP to control plasma. The majority of COVID-19 CCP arm hospitalizations (15/17, 88%) occurred in this unvaccinated, seronegative subgroup. A functional cutoff to delineate recipient high versus low post-transfusion antibody levels was established by two methods: 1) analyzing virus neutralization-equivalent anti-S-RBD IgG responses in donors or 2) receiver operating characteristic (ROC) analysis. RESULTS: SARS-CoV-2 anti-S-RBD IgG antibody was diluted by a factor of 21.3 into post-transfusion seronegative recipients from matched donor units. Viral specific antibody delivered approximated 1.2 mg. The high antibody recipients transfused early (symptom onset within 5 days) had no hospitalizations. A CCP recipient analysis for antibody thresholds correlated to reduced hospitalizations found a significant association with Fisher's exact test between early and high antibodies versus all other CCP recipients (or control plasma) with antibody cutoffs established by both methods-donor virus neutralization-based cutoff: (0/85; 0% versus 15/276; 5.6%) p=0.03 or ROC based cutoff: (0/94; 0% versus 15/267; 5.4%) p=0.01. CONCLUSION: In unvaccinated, seronegative CCP recipients, early transfusion of plasma units corresponding to the upper 30% of all study donors reduced outpatient hospitalizations. These high antibody level plasma units, given early, should be reserved for therapeutic use.Trial registration: NCT04373460. FUNDING: Defense Health Agency and others.

2019-20 H1N1 clade A5a.1 viruses have better in vitro replication compared with the co-circulating A5a.2 clade.

Surveillance for emerging human influenza virus clades is important for identifying changes in viral fitness and assessing antigenic similarity to vaccine strains. While fitness and antigenic structure are both important aspects of virus success, they are distinct characteristics and do not always change in a complementary manner. The 2019-20 Northern Hemisphere influenza season saw the emergence of two H1N1 clades: A5a.1 and A5a.2. While several studies indicated that A5a.2 showed similar or even increased antigenic drift compared with A5a.1, the A5a.1 clade was still the predominant circulating clade that season. Clinical isolates of representative viruses from these clades were collected in Baltimore, Maryland during the 2019-20 season and multiple assays were performed to compare both antigenic drift and viral fitness between clades. Neutralization assays performed on serum from healthcare workers pre- and post-vaccination during the 2019-20 season show a comparable drop in neutralizing titers against both A5a.1 and A5a.2 viruses compared with the vaccine strain, indicating that A5a.1 did not have antigenic advantages over A5a.2 that would explain its predominance in this population. Plaque assays were performed to investigate fitness differences, and the A5a.2 virus produced significantly smaller plaques compared with viruses from A5a.1 or the parental A5a clade. To assess viral replication, low MOI growth curves were performed on both MDCK-SIAT and primary differentiated human nasal epithelial cell cultures. In both cell cultures, A5a.2 yielded significantly reduced viral titers at multiple timepoints post-infection compared with A5a.1 or A5a. Receptor binding was then investigated through glycan array experiments which showed a reduction in receptor binding diversity for A5a.2, with fewer glycans bound and a higher percentage of total binding attributable to the top three highest bound glycans. Together these data indicate that the A5a.2 clade had a reduction in viral fitness, including reductions in receptor binding, that may have contributed to the limited prevalence observed after emergence.

Transmissibility of clinically relevant atovaquone-resistant Plasmodium falciparum by anopheline mosquitoes.

Rising numbers of malaria cases and deaths underscore the need for new interventions. Long-acting injectable medications, such as those now in use for HIV prophylaxis, offer the prospect of a malaria "chemical vaccine", combining the efficacy of a drug (like atovaquone) with the durability of a biological vaccine. Of concern, however, is the possible selection and transmission of drug-resistant parasites. We addressed this question by generating clinically relevant, highly atovaquone-resistant, Plasmodium falciparum mutants competent to infect mosquitoes. Isogenic paired strains, that differ only by a single Y268S mutation in cytochrome b, were evaluated in parallel in southeast Asian (Anopheles stephensi) or African (Anopheles gambiae) mosquitoes, and thence in humanized mice. Fitness costs of the mutation were evident along the lifecycle, in asexual parasite growth in vitro and in a progressive loss of parasites in the mosquito. In numerous independent experiments, microscopic exam of salivary glands from hundreds of mosquitoes failed to detect even one Y268S sporozoite, a defect not rescued by coinfection with wild type parasites. Furthermore, despite uniformly successful transmission of wild type parasites from An. stephensi to FRG NOD huHep mice bearing human hepatocytes and erythrocytes, multiple attempts with Y268S-fed mosquitoes failed: there was no evidence of parasites in mouse tissues by microscopy, in vitro culture, or PCR. These studies confirm a severe-to-lethal fitness cost of clinically relevant atovaquone-resistant P. falciparum in the mosquito, and they significantly lessen the likelihood of their transmission in the field.

Transfusing Convalescent Plasma as Post-Exposure Prophylaxis Against Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Infection: A Double-Blinded, Phase 2 Randomized, Controlled Trial.

BACKGROUND: The efficacy of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) convalescent plasma (CCP) for preventing infection in exposed, uninfected individuals is unknown. CCP might prevent infection when administered before symptoms or laboratory evidence of infection. METHODS: This double-blinded, phase 2 randomized, controlled trial (RCT) compared the efficacy and safety of prophylactic high titer (≥1:320 by Euroimmun ELISA) CCP with standard plasma. Asymptomatic participants aged ≥18 years with close contact exposure to a person with confirmed coronavirus disease 2019 (COVID-19) in the previous 120 hours and negative SARS-CoV-2 test within 24 hours before transfusion were eligible. The primary outcome was new SARS-CoV-2 infection. RESULTS: In total, 180 participants were enrolled; 87 were assigned to CCP and 93 to control plasma, and 170 transfused at 19 sites across the United States from June 2020 to March 2021. Two were excluded for screening SARS-CoV-2 reverse transcription polymerase chain reaction (RT-PCR) positivity. Of the remaining 168 participants, 12/81 (14.8%) CCP and 13/87 (14.9%) control recipients developed SARS-CoV-2 infection; 6 (7.4%) CCP and 7 (8%) control recipients developed COVID-19 (infection with symptoms). There were no COVID-19-related hospitalizations in CCP and 2 in control recipients. Efficacy by restricted mean infection free time (RMIFT) by 28 days for all SARS-CoV-2 infections (25.3 vs 25.2 days; P = .49) and COVID-19 (26.3 vs 25.9 days; P = .35) was similar for both groups. CONCLUSIONS: Administration of high-titer CCP as post-exposure prophylaxis, although appearing safe, did not prevent SARS-CoV-2 infection. CLINICAL TRIALS REGISTRATION: NCT04323800.

Cryptococcus neoformans releases proteins during intracellular residence that affect the outcome of the fungal-macrophage interaction.

Cryptococcus neoformans is a facultative intracellular pathogen that can replicate and disseminate in mammalian macrophages. In this study, we analyzed fungal proteins identified in murine macrophage-like cells after infection with C. neoformans. To accomplish this, we developed a protocol to identify proteins released from cryptococcal cells inside macrophage-like cells; we identified 127 proteins of fungal origin in infected macrophage-like cells. Among the proteins identified was urease, a known virulence factor, and others such as transaldolase and phospholipase D, which have catalytic activities that could contribute to virulence. This method provides a straightforward methodology to study host-pathogen interactions. We chose to study further Yeast Oligomycin Resistance (Yor1), a relatively uncharacterized protein belonging to the large family of ATP binding cassette transporter (ABC transporters). These transporters belong to a large and ancient protein family found in all extant phyla. While ABC transporters have an enormous diversity of functions across varied species, in pathogenic fungi they are better studied as drug efflux pumps. Analysis of C. neoformans yor1Δ strains revealed defects in nonlytic exocytosis, capsule size, and dimensions of extracellular vesicles, when compared to wild-type strains. We detected no difference in growth rates and cell body size. Our results indicate that C. neoformans releases a large suite of proteins during macrophage infection, some of which can modulate fungal virulence and are likely to affect the fungal-macrophage interaction.

Downregulation of transcriptional activity, increased inflammation, and damage in the placenta following in utero Zika virus infection is associated with adverse pregnancy outcomes.

Zika virus (ZIKV) infection during pregnancy causes serious adverse outcomes to the developing fetus, including fetal loss and birth defects known as congenital Zika syndrome (CZS). The mechanism by which ZIKV infection causes these adverse outcomes and specifically, the interplay between the maternal immune response and ZIKV replication has yet to be fully elucidated. Using an immunocompetent mouse model of transplacental ZIKV transmission and adverse pregnancy outcomes, we have previously shown that Asian lineage ZIKV disrupts placental morphology and induces elevated secretion of IL-1ß. In the current manuscript, we characterized placental damage and inflammation during in utero African lineage ZIKV infection. Within 48 hours after ZIKV infection at embryonic day 10, viral RNA was detected in placentas and fetuses from ZIKA infected dams, which corresponded with placental damage and reduced fetal viability as compared with mock infected dams. Dams infected with ZIKV had reduced proportions of trophoblasts and endothelial cells and disrupted placental morphology compared to mock infected dams. While placental IL-1ß was increased in the placenta, but not the spleen, within 3 hours post infection, this was not caused by activation of the NLRP3 inflammasome. Using bulk mRNAseq from placentas of ZIKV and mock infected dams, ZIKV infection caused profound downregulation of the transcriptional activity of genes that may underly tissue morphology, neurological development, metabolism, cell signaling and inflammation, illustrating that in utero ZIKV infections causes disruption of pathways associated with CZS in our model.

Adaptive immune responses in vaccinated patients with symptomatic SARS-CoV-2 Alpha infection.

Benchmarks for protective immunity from infection or severe disease after SARS-CoV-2 vaccination are still being defined. Here, we characterized virus neutralizing and ELISA antibody levels, cellular immune responses, and viral variants in 4 separate groups: healthy controls (HCs) weeks (early) or months (late) following vaccination in comparison with symptomatic patients with SARS-CoV-2 after partial or full mRNA vaccination. During the period of the study, most symptomatic breakthrough infections were caused by the SARS-CoV-2 Alpha variant. Neutralizing antibody levels in the HCs were sustained over time against the vaccine parent virus but decreased against the Alpha variant, whereas IgG titers and T cell responses against the parent virus and Alpha variant declined over time. Both partially and fully vaccinated patients with symptomatic infections had lower virus neutralizing antibody levels against the parent virus than the HCs, similar IgG antibody titers, and similar virus-specific T cell responses measured by IFN-γ. Compared with HCs, neutralization activity against the Alpha variant was lower in the partially vaccinated infected patients and tended to be lower in the fully vaccinated infected patients. In this cohort of breakthrough infections, parent virus neutralization was the superior predictor of breakthrough infections with the Alpha variant of SARS-CoV-2.

The effect of mindfulness-based stress reduction on the urinary microbiome in interstitial cystitis.

INTRODUCTION AND HYPOTHESIS: The objective was to investigate the impact of mindfulness-based stress reduction therapy on the urinary microbiome of patients with interstitial cystitis/bladder pain syndrome. METHODS: In this Institutional Review Board-approved prospective cohort study, patients with interstitial cystitis/bladder pain syndrome were recruited to attend an 8-week mindfulness-based stress reduction course involving yoga and meditation. Eligible participants were English-speaking women aged 18 or older with interstitial cystitis/bladder pain syndrome. All participants had a negative urinalysis within 2 months of enrollment and were currently undergoing first- or second-line treatment at the time of recruitment. The mindfulness-based stress reduction course met weekly for 1 h. A straight-catheter urine sample was obtained prior to and following the mindfulness-based stress reduction series. DNA from urine samples underwent bacterial 16S ribosomal gene sequencing at Johns Hopkins University Laboratories followed by taxonomic abundance and diversity analysis by Resphera Biosciences Laboratory. Participants completed validated symptom questionnaires pre- and post-intervention. RESULTS: A total of 12 participants completed the 8-week course and were included in the analysis. The average age was 59 and the majority identified as white. Patient symptoms, measured by the Urogenital Distress Inventory Short Form and Interstitial Cystitis Symptom and Pain Indices, improved significantly (all p < 0.05). Overall composition of the urinary microbiome changed significantly (p < 0.01) and demonstrated an increase in diversity following the intervention. CONCLUSIONS: Mindfulness-based stress reduction therapy improves patient symptoms and was associated with significant changes in the urinary microbiome in patients with interstitial cystitis/bladder pain syndrome.

Progression and Resolution of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Infection in Golden Syrian Hamsters.

To catalyze severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) research, including development of novel interventive and preventive strategies, the progression of disease was characterized in a robust coronavirus disease 2019 (COVID-19) animal model. In this model, male and female golden Syrian hamsters were inoculated intranasally with SARS-CoV-2 USA-WA1/2020. Groups of inoculated and mock-inoculated uninfected control animals were euthanized at 2, 4, 7, 14, and 28 days after inoculation to track multiple clinical, pathology, virology, and immunology outcomes. SARS-CoV-2-inoculated animals consistently lost body weight during the first week of infection, had higher lung weights at terminal time points, and developed lung consolidation per histopathology and quantitative image analysis measurements. High levels of infectious virus and viral RNA were reliably present in the respiratory tract at days 2 and 4 after inoculation, corresponding with widespread necrosis and inflammation. At day 7, when the presence of infectious virus was rare, interstitial and alveolar macrophage infiltrates and marked reparative epithelial responses (type II hyperplasia) dominated in the lung. These lesions resolved over time, with only residual epithelial repair evident by day 28 after inoculation. The use of quantitative approaches to measure cellular and morphologic alterations in the lung provides valuable outcome measures for developing therapeutic and preventive interventions for COVID-19 using the hamster COVID-19 model.

Randomized controlled trial transfusing convalescent plasma as post-exposure prophylaxis against SARS-CoV-2 infection.

BACKGROUND: The efficacy of SARS-CoV-2 convalescent plasma (CCP) for preventing infection in exposed, uninfected individuals is unknown. We hypothesized that CCP might prevent infection when administered before symptoms or laboratory evidence of infection. METHODS: This double-blinded, phase 2 randomized, controlled trial (RCT) compared the efficacy and safety of prophylactic high titer (≥1:320) CCP with standard plasma. Asymptomatic participants aged ≥18 years with close contact exposure to a person with confirmed COVID-19 in the previous 120 hours and negative SARS-CoV-2 test within 24 hours before transfusion were eligible. The primary outcome was development of SARS-CoV-2 infection. RESULTS: 180 participants were enrolled; 87 were assigned to CCP and 93 to control plasma, and 170 transfused at 19 sites across the United States from June 2020 to March 2021. Two were excluded for SARS-CoV-2 RT-PCR positivity at screening. Of the remaining 168 participants, 12/81 (14.8%) CCP and 13/87 (14.9%) control recipients developed SARS-CoV-2 infection; 6 (7.4%) CCP and 7 (8%) control recipients developed COVID-19 (infection with symptoms). There were no COVID-19-related hospitalizations in CCP and 2 in control recipients. There were 28 adverse events in CCP and 58 in control recipients. Efficacy by restricted mean infection free time (RMIFT) by 28 days for all SARS-CoV-2 infections (25.3 vs. 25.2 days; p=0.49) and COVID-19 (26.3 vs. 25.9 days; p=0.35) were similar for both groups. CONCLUSION: In this trial, which enrolled persons with recent exposure to a person with confirmed COVID-19, high titer CCP as post-exposure prophylaxis appeared safe, but did not prevent SARS-CoV-2 infection. TRIAL REGISTRATION: Clinicaltrial.gov number NCT04323800 .

Transcriptomics Underlying Pulmonary Ozone Pathogenesis Regulated by Inflammatory Mediators in Mice.

Ozone (O3) is the predominant oxidant air pollutant associated with airway inflammation, lung dysfunction, and the worsening of preexisting respiratory diseases. We previously demonstrated the injurious roles of pulmonary immune receptors, tumor necrosis factor receptor (TNFR), and toll-like receptor 4, as well as a transcription factor NF-κB, in response to O3 in mice. In the current study, we profiled time-dependent and TNFR- and NF-κB-regulated lung transcriptome changes by subacute O3 to illuminate the underlying molecular events and downstream targets. Mice lacking Tnfr1/Tnfr2 (Tnfr-/-) or Nfkb1 (Nfkb1-/-) were exposed to air or O3. Lung RNAs were prepared for cDNA microarray analyses, and downstream and upstream mechanisms were predicted by pathway analyses of the enriched genes. O3 significantly altered the genes involved in inflammation and redox (24 h), cholesterol biosynthesis and vaso-occlusion (48 h), and cell cycle and DNA repair (48-72 h). Transforming growth factor-ß1 was a predicted upstream regulator. Lack of Tnfr suppressed the immune cell proliferation and lipid-related processes and heightened epithelial cell integrity, and Nfkb1 deficiency markedly suppressed lung cell cycle progress during O3 exposure. Common differentially regulated genes by TNFR and NF-κB1 (e.g., Casp8, Il6, and Edn1) were predicted to protect the lungs from cell death, connective tissue injury, and inflammation. Il6-deficient mice were susceptible to O3-induced protein hyperpermeability, indicating its defensive role, while Tnf-deficient mice were resistant to overall lung injury caused by O3. The results elucidated transcriptome dynamics and provided new insights into the molecular mechanisms regulated by TNFR and NF-κB1 in pulmonary subacute O3 pathogenesis.

Sex Differences in Lung Imaging and SARS-CoV-2 Antibody Responses in a COVID-19 Golden Syrian Hamster Model.

In the coronavirus disease 2019 (COVID-19) pandemic caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), more severe outcomes are reported in males than in females, including hospitalizations and deaths. Animal models can provide an opportunity to mechanistically interrogate causes of sex differences in the pathogenesis of SARS-CoV-2. Adult male and female golden Syrian hamsters (8 to 10 weeks of age) were inoculated intranasally with 105 50% tissue culture infective dose (TCID50) of SARS-CoV-2/USA-WA1/2020 and euthanized at several time points during the acute (i.e., virus actively replicating) and recovery (i.e., after the infectious virus has been cleared) phases of infection. There was no mortality, but infected male hamsters experienced greater morbidity, losing a greater percentage of body mass, developed more extensive pneumonia as noted on chest computed tomography, and recovered more slowly than females. Treatment of male hamsters with estradiol did not alter pulmonary damage. Virus titers in respiratory tissues, including nasal turbinates, trachea, and lungs, and pulmonary cytokine concentrations, including interferon-ß (IFN-ß) and tumor necrosis factor-α (TNF-α), were comparable between the sexes. However, during the recovery phase of infection, females mounted 2-fold greater IgM, IgG, and IgA responses against the receptor-binding domain of the spike protein (S-RBD) in both plasma and respiratory tissues. Female hamsters also had significantly greater IgG antibodies against whole-inactivated SARS-CoV-2 and mutant S-RBDs as well as virus-neutralizing antibodies in plasma. The development of an animal model to study COVID-19 sex differences will allow for a greater mechanistic understanding of the SARS-CoV-2-associated sex differences seen in the human population. IMPORTANCE Men experience more severe outcomes from coronavirus disease 2019 (COVID-19) than women. Golden Syrian hamsters were used to explore sex differences in the pathogenesis of a human isolate of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). After inoculation, male hamsters experienced greater sickness, developed more severe lung pathology, and recovered more slowly than females. Sex differences in disease could not be reversed by estradiol treatment in males and were not explained by either virus replication kinetics or the concentrations of inflammatory cytokines in the lungs. During the recovery period, antiviral antibody responses in the respiratory tract and plasma, including to newly emerging SARS-CoV-2 variants, were greater in female than in male hamsters. Greater lung pathology during the acute phase combined with lower antiviral antibody responses during the recovery phase of infection in males than in females illustrate the utility of golden Syrian hamsters as a model to explore sex differences in the pathogenesis of SARS-CoV-2 and vaccine-induced immunity and protection.