Depletion of Bone Marrow Hematopoietic Cells in Ebolavirus-Infected Rhesus Macaques: A Possible Cause of Hematologic Abnormalities in Ebolavirus Disease.

The pathophysiology of long-recognized hematologic abnormalities in Ebolavirus (EBOV) disease (EVD) is unknown. From limited human sampling (of peripheral blood), it has been postulated that emergency hematopoiesis plays a role in severe EVD, but the systematic characterization of the bone marrow (BM) has not occurred in human disease or in nonhuman primate models. In a lethal rhesus macaque model of EVD, 18 sternal BM samples exposed to the Kikwit strain of EBOV were compared to those from uninfected controls (n = 3). Immunohistochemistry, RNAscope in situ hybridization, transmission electron microscopy, and confocal microscopy showed that EBOV infects BM monocytes/macrophages and megakaryocytes. EBOV exposure was associated with severe BM hypocellularity, including depletion of myeloid, erythroid, and megakaryocyte hematopoietic cells. These depletions were negatively correlated with cell proliferation (Ki67 expression) and were not associated with BM apoptosis during disease progression. In EBOV-infected rhesus macaques with terminal disease, BM showed marked hemophagocytosis, megakaryocyte emperipolesis, and the release of immature hematopoietic cells into the sinusoids. Collectively, these data demonstrate not only direct EBOV infection of BM monocytes/macrophages and megakaryocytes but also that disease progression is associated with hematopoietic failure, notably in peripheral cytopenia. These findings inform current pathophysiologic unknowns and suggest a crucial role for BM dysfunction and/or failure, including emergency hematopoiesis, as part of the natural history of severe human disease.

Ebola Virus Disease Features Hemophagocytic Lymphohistiocytosis/Macrophage Activation Syndrome in the Rhesus Macaque Model.

BACKGROUND: Ebola virus (EBOV) disease (EVD) is one of the most severe and fatal viral hemorrhagic fevers and appears to mimic many clinical and laboratory manifestations of hemophagocytic lymphohistiocytosis syndrome (HLS), also known as macrophage activation syndrome. However, a clear association is yet to be firmly established for effective host-targeted, immunomodulatory therapeutic approaches to improve outcomes in patients with severe EVD. METHODS: Twenty-four rhesus monkeys were exposed intramuscularly to the EBOV Kikwit isolate and euthanized at prescheduled time points or when they reached the end-stage disease criteria. Three additional monkeys were mock-exposed and used as uninfected controls. RESULTS: EBOV-exposed monkeys presented with clinicopathologic features of HLS, including fever, multiple organomegaly, pancytopenia, hemophagocytosis, hyperfibrinogenemia with disseminated intravascular coagulation, hypertriglyceridemia, hypercytokinemia, increased concentrations of soluble CD163 and CD25 in serum, and the loss of activated natural killer cells. CONCLUSIONS: Our data suggest that EVD in the rhesus macaque model mimics pathophysiologic features of HLS/macrophage activation syndrome. Hence, regulating inflammation and immune function might provide an effective treatment for controlling the pathogenesis of acute EVD.

Visual acuity and vision-related quality of life outcomes following cataract surgery in Ebola virus disease survivors.

Objectives: The objectives of this study were to assess relationships between vision-related quality of life (QoL) and visual acuity (VA) in Ebola virus disease (EVD) survivors after cataract surgery in the Ebola Viral Persistence in Ocular Tissues and Fluids (EVICT) Study. Materials and Methods: EVD survivors with undetectable Ebola virus (EBOV) ribonucleic acid in their aqueous humour were eligible to receive manual small-incision cataract surgery (MSICS). Among those that received surgery, assessments of VA and vision-related QoL were assessed pre-and post-cataract surgery. VA was converted from units on a tumbling 'E' chart to the logarithm of the minimal angle of resolution VA (logMAR VA). Vision-related QoL was assessed using the 25-item National Eye Institute Visual Function Questionnaire (NEI VFQ-25). Linear regression was used to evaluate the associations between VA and vision-related QoL. P = 0.05 was considered statistically significant for all analyses. Results: Thirty-four EVD survivors underwent cataract surgery in the EVICT study. Before MSICS, the mean logMAR VA was 2.24 (standard deviation [SD]: 0.98), and the mean NEI-VFQ-25 composite score was 54 (SD: 15); however, there was no significant association between the pre-surgery measurements (average difference in VA/10 unit increase in NEI-VFQ-25: -0.04, 95% confidence interval (CI): -0.33-0.26, P = 0.80). There was a significant improvement in logMAR VA after MSICS (mean: 1.6, P < 0.001), but there was no significant change in the NEI-VFQ-25 composite (-0.87, 95% (CI): -10.32-8.59, P = 0.85). None of the subscales showed significant improvements (P > 0.12 for all); however, the magnitude of the mean change for distance activities (6.65), near activities (6.76), general vision (-7.69), social functioning (-9.13) and colour vision (13.33) met the criteria for a clinically meaningful difference (4-6). In the subset with paired measurements (n = 16), there were no significant association changes in logMAR VA and NEI VFQ-25 composite scores (P > 0.12 for all). Conclusion: Following cataract surgery, VA in EVD survivors improved, but these improvements were not reflected in NEI VFQ-25 composite scores or specific subscales; however, the small sample size limits generalizability absent more research. Differences in sociocultural context and activities that affect the QoL in resource-limited areas may contribute to the limitations seen with NEI VFQ-25. In addition, better eye dominance could contribute to any lack of association as NEI VFQ-25 evaluates vision as a whole. Further, assessment of factors contributing to improved QoL may help to define the impact of vision health in varied environments.

A Novel Visually Guided Radiofrequency Balloon Ablation Catheter for Pulmonary Vein Isolation: One-Year Outcomes of the Multicenter AF-FICIENT I Trial.

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Implementation of the Ebola Virus Persistence in Ocular Tissues and Fluids (EVICT) study: Lessons learned for vision health systems strengthening in Sierra Leone.

BACKGROUND: Following the West African Ebola virus disease (EVD) outbreak of 2013-2016 and more recent EVD outbreaks in the Democratic Republic of Congo, thousands of EVD survivors are at-risk for sequelae including uveitis, which can lead to unremitting inflammation and vision loss from cataract. Because of the known risk of Ebola virus persistence in ocular fluid and the need to provide vision-restorative, safe cataract surgery, the Ebola Virus Persistence in Ocular Tissues and Fluids (EVICT) Study was implemented in Sierra Leone. During implementation of this multi-national study, challenges included regulatory approvals, mobilization, community engagement, infection prevention and control, and collaboration between multiple disciplines. In this report, we address the multifacted approach to address these challenges and the impact of implementation science research to address an urgent clinical subspecialty need in an outbreak setting. METHODOLOGY/PRINCIPAL FINDINGS: Given the patient care need to develop a protocol to evaluate ocular fluid for Ebola virus RNA persistence prior to cataract surgery, as well as protocols to provide reassurance to ophthalmologists caring for EVD survivors with cataracts, the EVICT study was designed and implemented through the work of the Ministry of Health, Sierra Leone National Eye Programme, and international partnerships. The EVICT study showed that all 50 patients who underwent ocular fluid sampling at 19 and 34 months, respectively, tested negative for Ebola virus RNA. Thirty-four patients underwent successful cataract surgery with visual acuity improvement. Here we describe the methodology for study implementation, challenges encountered, and key issues that impacted EVD vision care in the immediate aftermath of the EVD outbreak. Key aspects of the EVICT study included defining the pertinent questions and clinical need, partnership alignment with key stakeholders, community engagement with EVD survivor associations, in-country and international regulatory approvals, study site design for infection prevention and control, and thorough plans for EVD survivor follow-up care and monitoring. Challenges encountered included patient mobilization owing to transportation routes and distance of patients in rural districts. Strong in-country partnerships and multiple international organizations overcame these challenges so that lessons learned could be applied for future EVD outbreaks in West and Central Africa including EVD outbreaks that are ongoing in Guinea and Democratic Republic of Congo. CONCLUSIONS/SIGNIFICANCE: The EVICT Study showed that cataract surgery with a protocol-driven approach was safe and vision-restorative for EVD survivors, which provided guidance for EVD ophthalmic surgical care. Ophthalmologic care remains a key aspect of the public health response for EVD outbreaks but requires a meticulous, yet partnered approach with international and local in-country partners. Future efforts may build on this framework for clinical care and to improve our understanding of ophthalmic sequelae, develop treatment paradigms for EVD survivors, and strengthen vision health systems in resource-limited settings.

Efficacy and Safety Outcomes of Cataract Surgery in Survivors of Ebola Virus Disease: 12-Month Results From the PREVAIL VII Study.

Purpose: In survivors of Ebola virus disease (EVD), intraocular viral persistence raises questions about the timing and safety of cataract surgery. To the best of our knowledge, this is the first controlled study evaluating Ebola virus persistence and cataract surgery safety and outcomes in EVD survivors. Methods: Seropositive EVD survivors and seronegative controls with vision worse than 20/40 from cataract and without active intraocular inflammation were enrolled. Aqueous humor from survivors was tested with reverse transcription-polymerase chain reaction for Ebola viral RNA. Participants underwent manual small-incision cataract surgery and 1 year of follow-up examinations. Results: Twenty-two eyes of 22 survivors and 12 eyes of eight controls underwent cataract surgery. All of the aqueous samples tested negative for Ebola viral RNA. Median visual acuity improved from 20/200 at baseline to 20/25 at 1 year in survivors and from count fingers to 20/50 in controls (overall, P < 0.001; between groups, P = 0.07). After a 1-month course of topical corticosteroids, 55% of survivors and 67% of controls demonstrated at least 1+ anterior chamber cell. Twelve months after surgery, optical coherence tomography revealed a median increase in macular central subfield thickness of 42 µm compared with baseline (overall, P = 0.029; between groups, P = 0.995). Conclusions: EVD survivors and controls demonstrated significant visual improvement from cataract surgery. The persistence of intraocular inflammation highlights the importance of follow-up. The absence of detectable intraocular Ebola viral RNA provides guidance regarding the safety of eye surgery in Ebola survivors. Translational Relevance: These findings demonstrate the safety and efficacy of cataract surgery in Ebola survivors and will inform ocular surgery guidelines in this population.

Approach to Cataract Surgery in an Ebola Virus Disease Survivor with Prior Ocular Viral Persistence.

A 46-year-old patient with previously documented Ebola virus persistence in his ocular fluid, associated with severe panuveitis, developed a visually significant cataract. A multidisciplinary approach was taken to prevent and control infection. Ebola virus persistence was assessed before and during the operation to provide safe, vision-restorative phacoemulsification surgery.

A Randomized, Controlled Trial of Ebola Virus Disease Therapeutics.

BACKGROUND: Although several experimental therapeutics for Ebola virus disease (EVD) have been developed, the safety and efficacy of the most promising therapies need to be assessed in the context of a randomized, controlled trial. METHODS: We conducted a trial of four investigational therapies for EVD in the Democratic Republic of Congo, where an outbreak began in August 2018. Patients of any age who had a positive result for Ebola virus RNA on reverse-transcriptase-polymerase-chain-reaction assay were enrolled. All patients received standard care and were randomly assigned in a 1:1:1:1 ratio to intravenous administration of the triple monoclonal antibody ZMapp (the control group), the antiviral agent remdesivir, the single monoclonal antibody MAb114, or the triple monoclonal antibody REGN-EB3. The REGN-EB3 group was added in a later version of the protocol, so data from these patients were compared with those of patients in the ZMapp group who were enrolled at or after the time the REGN-EB3 group was added (the ZMapp subgroup). The primary end point was death at 28 days. RESULTS: A total of 681 patients were enrolled from November 20, 2018, to August 9, 2019, at which time the data and safety monitoring board recommended that patients be assigned only to the MAb114 and REGN-EB3 groups for the remainder of the trial; the recommendation was based on the results of an interim analysis that showed superiority of these groups to ZMapp and remdesivir with respect to mortality. At 28 days, death had occurred in 61 of 174 patients (35.1%) in the MAb114 group, as compared with 84 of 169 (49.7%) in the ZMapp group (P = 0.007), and in 52 of 155 (33.5%) in the REGN-EB3 group, as compared with 79 of 154 (51.3%) in the ZMapp subgroup (P = 0.002). A shorter duration of symptoms before admission and lower baseline values for viral load and for serum creatinine and aminotransferase levels each correlated with improved survival. Four serious adverse events were judged to be potentially related to the trial drugs. CONCLUSIONS: Both MAb114 and REGN-EB3 were superior to ZMapp in reducing mortality from EVD. Scientifically and ethically sound clinical research can be conducted during disease outbreaks and can help inform the outbreak response. (Funded by the National Institute of Allergy and Infectious Diseases and others; PALM ClinicalTrials.gov number, NCT03719586.).

Longitudinal Analysis of the Human B Cell Response to Ebola Virus Infection.

Ebola virus (EBOV) remains a public health threat. We performed a longitudinal study of B cell responses to EBOV in four survivors of the 2014 West African outbreak. Infection induced lasting EBOV-specific immunoglobulin G (IgG) antibodies, but their subclass composition changed over time, with IgG1 persisting, IgG3 rapidly declining, and IgG4 appearing late. Striking changes occurred in the immunoglobulin repertoire, with massive recruitment of naive B cells that subsequently underwent hypermutation. We characterized a large panel of EBOV glycoprotein-specific monoclonal antibodies (mAbs). Only a small subset of mAbs that bound glycoprotein by ELISA recognized cell-surface glycoprotein. However, this subset contained all neutralizing mAbs. Several mAbs protected against EBOV disease in animals, including one mAb that targeted an epitope under evolutionary selection during the 2014 outbreak. Convergent antibody evolution was seen across multiple donors, particularly among VH3-13 neutralizing antibodies specific for the GP1 core. Our study provides a benchmark for assessing EBOV vaccine-induced immunity.

Ebola Virus Persistence in Ocular Tissues and Fluids (EVICT) Study: Reverse Transcription-Polymerase Chain Reaction and Cataract Surgery Outcomes of Ebola Survivors in Sierra Leone.

BACKGROUND: Ebola virus disease (EVD) survivors are at risk for uveitis during convalescence. Vision loss has been observed following uveitis due to cataracts. Since Ebola virus (EBOV) may persist in the ocular fluid of EVD survivors for an unknown duration, there are questions about the safety and feasibility of vision restorative cataract surgery in EVD survivors. METHODS: We conducted a cross-sectional study of EVD survivors anticipating cataract surgery and patients with active uveitis to evaluate EBOV RNA persistence in ocular fluid, as well as vision outcomes post cataract surgery. Patients with aqueous humor that tested negative for EBOV RNA were eligible to proceed with manual small incision cataract surgery (MSICS). FINDINGS: We screened 137 EVD survivors from June 2016 - August 2017 for enrolment. We enrolled 50 EVD survivors; 46 with visually significant cataract, 1 with a subluxated lens, 2 with active uveitis and 1 with a blind painful eye due to uveitis. The median age was 24.0years (IQR 17-35) and 35 patients (70%) were female. The median logMAR visual acuity (VA) was 3.0 (Snellen VA Hand motions; Interquartile Range, IQR: 1.2-3.0, Snellen VA 20/320 - Hand motions). All patients tested negative for EBOV RNA by RT-PCR in aqueous humor/vitreous fluid and conjunctiva at a median of 19months (IQR 18-20) from EVD diagnosis in Phase 1 of ocular fluid sampling and 34months (IQR 32-36) from EVD diagnosis in Phase 2 of ocular fluid sampling. Thirty-four patients underwent MSICS, with a preoperative median VA improvement from hand motions to 20/30 at three-month postoperative follow-up (P<0.001). INTERPRETATION: EBOV persistence by RT-PCR was not identified in ocular fluid or conjunctivae of fifty EVD survivors with ocular disease. Cataract surgery can be performed safely with vision restorative outcomes in patients who test negative for EBOV RNA in ocular fluid specimens. These findings impact the thousands of West African EVD survivors at-risk for ocular complications who may also require eye surgery during EVD convalescence.

High-Resolution Infrared Thermography of Esophageal Temperature During Radiofrequency Ablation of Atrial Fibrillation.

BACKGROUND: Catheter ablation for atrial fibrillation has potential to cause esophageal thermal injury. Esophageal temperature monitoring during ablation is commonly used; however, it has not eliminated thermal injuries, possibly because conventional sensors have poor spatial sampling and response characteristics. To enhance understanding of temperature dynamics that may underlie esophageal injury, we tested a high-resolution, intrabody, infrared thermography catheter to continuously image esophageal temperatures during ablation. METHODS AND RESULTS: Atrial fibrillation ablation patients were instrumented with a flexible, 9F infrared temperature catheter inserted nasally (n=8) or orally (n=8) into the esophagus adjacent to the left atrium. Ablation was performed while the infrared catheter continuously recorded surface temperatures from 7680 points per second circumferentially over a 6-cm length of esophagus. Physicians were blinded to temperature data. Endoscopy was performed within 24 hours to document esophageal injury. Thermal imaging showed that most patients (10/16) experienced ≥1 events where peak esophageal temperature was >40°C. Three patients experienced temperatures >50°C; and 1 experienced >60°C. Analysis of temperature data for each subject's maximum thermal event revealed high gradients (2.3±1.4°C/mm) and rates of change (1.5±1.3°C/s) with an average length of esophageal involvement of 11.0±5.4 mm. Endoscopy identified 3 distinct thermal lesions, all in patients with temperatures >50°C; all resolved within 2 weeks. CONCLUSIONS: Infrared thermography provided dynamic, high-resolution mapping of esophageal temperatures during cardiac ablation. Esophageal thermal injury occurred with temperatures >50°C and was associated with large spatiotemporal gradients. Additional studies are warranted to determine the relationships between thermal parameters and esophageal injury.

Surgical radiofrequency ablation for atrial fibrillation: the Christchurch, New Zealand experience.

AIMS: To report the long-term results following surgical radiofrequency ablation (RFA) for atrial fibrillation as an adjunct to other cardiac surgery at Christchurch Hospital. METHODS: A retrospective observational audit review of outcomes. The sample population included all patients identified as having undergone surgical RFA for atrial fibrillation at Christchurch Hospital, between the first procedure performed on 2 July 2001 and 28 January 2009. RESULTS: A total of 44 patients underwent surgical RFA between 2 July 2001 and 28 January 2009. Postoperatively there were three deaths prior to discharge (7%). Pacemakers were required in four patients (9%), and two patients subsequently underwent catheter ablation for atrial arrhythmias. In the immediate postoperative period only three patients remained in atrial fibrillation. At last follow-up up to 102 months from surgery (45 plus or minus 29 months), 27 patients had developed persistent atrial fibrillation and four persistent atrial flutter. Persisting long-term benefit was seen in seven patients (18%, 7/38); five patients were in stable sinus rhythm, one had paroxysmal atrial fibrillation and one paroxysmal atrial flutter. CONCLUSIONS: Whilst the procedure was effectively acutely, the recurrence of atrial fibrillation was high and development of new atrial flutter common over long-term follow-up.