Pandemic origins and a One Health approach to preparedness and prevention: Solutions based on SARS-CoV-2 and other RNA viruses.

COVID-19 is the latest zoonotic RNA virus epidemic of concern. Learning how it began and spread will help to determine how to reduce the risk of future events. We review major RNA virus outbreaks since 1967 to identify common features and opportunities to prevent emergence, including ancestral viral origins in birds, bats, and other mammals; animal reservoirs and intermediate hosts; and pathways for zoonotic spillover and community spread, leading to local, regional, or international outbreaks. The increasing scientific evidence concerning the origins of Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) is most consistent with a zoonotic origin and a spillover pathway from wildlife to people via wildlife farming and the wildlife trade. We apply what we know about these outbreaks to identify relevant, feasible, and implementable interventions. We identify three primary targets for pandemic prevention and preparedness: first, smart surveillance coupled with epidemiological risk assessment across wildlife-livestock-human (One Health) spillover interfaces; second, research to enhance pandemic preparedness and expedite development of vaccines and therapeutics; and third, strategies to reduce underlying drivers of spillover risk and spread and reduce the influence of misinformation. For all three, continued efforts to improve and integrate biosafety and biosecurity with the implementation of a One Health approach are essential. We discuss new models to address the challenges of creating an inclusive and effective governance structure, with the necessary stable funding for cross-disciplinary collaborative research. Finally, we offer recommendations for feasible actions to close the knowledge gaps across the One Health continuum and improve preparedness and response in the future.

Complete blood count parameters as biomarkers of retinopathy of prematurity: a Portuguese multicenter study.

PURPOSE: To evaluate complete blood count (CBC) parameters in the first week of life as predictive biomarkers for the development of retinopathy of prematurity (ROP). METHODS: Multicenter, prospective, observational study of a cohort of preterm infants born with gestational age (GA) < 32 weeks or birth weight < 1500 g in eight Portuguese neonatal intensive care units. All demographic, clinical, and laboratory data from the first week of life were collected. Univariate logistic regression was used to assess risk factors for ROP and then multivariate regression was performed. RESULTS: A total of 455 infants were included in the study. The median GA was 29.6 weeks, and the median birth weight was 1295 g. One hundred and seventy-two infants (37.8%) developed ROP. Median values of erythrocytes (p < 0.001), hemoglobin (p < 0.001), hematocrit (p < 0.001), mean corpuscular hemoglobin concentration (p < 0.001), lymphocytes (p = 0.035), and platelets (p = 0.003) of the group of infants diagnosed with ROP any stage were lower than those without ROP. Mean corpuscular volume (MCV) (p = 0.044), red blood cell distribution width (RDW) (p < 0.001), erythroblasts (p < 0.001), neutrophils (p = 0.030), neutrophils-lymphocytes ratio (p = 0.028), and basophils (p = 0.003) were higher in the ROP group. Higher values of MCV, erythroblasts, and basophils remained significantly associated with ROP after multivariate regression. CONCLUSION: In our cohort, the increase in erythroblasts, MCV, and basophils in the first week of life was significantly and independently associated with the development of ROP. These CBC parameters may be early predictive biomarkers for ROP.

Drosophila nasuta (Diptera, Drosophilidae) in Brazil: a decade of invasion and occupation of more than half of the country.

As a result of human activities and natural dispersal, exotic species can be brought to new areas, where they become established and spread, becoming invaders. These species are responsible for the loss of biodiversity and cause ecosystemic harm throughout the world. In this paper, we report the rapid, broad geographic expansion of the invasive fly Drosophila nasuta in Brazil. An 84% increase was found in its area of occupation in the country compared to previous studies. The present data reveal its arrival to the Pantanal wetlands in a location more than one thousand kilometers from the closest previous record in the Cerrado biome. We present the first record of D. nasuta in the Atlantic Forest in the states of Paraíba and Bahia. We report its introduction in the Amazon Forest in the state of Amazonas approximately 700 kilometers from previous records. The relative abundance of D. nasuta in this biome increased fivefold in comparison to a previous study. In the first decade of invasion in Brazil, D. nasuta has already colonized more than half of the country. The present data reveal its invasive potential and underscore the importance of following up the possible negative effects of this biological invasion.

Genetic Modulation of the Erythrocyte Phenotype Associated with Retinopathy of Prematurity-A Multicenter Portuguese Cohort Study.

The development of retinopathy of prematurity (ROP) may be influenced by anemia or a low fetal/adult hemoglobin ratio. We aimed to analyze the association between DNA methyltransferase 3 ß (DNMT3B) (rs2424913), methylenetetrahydrofolate reductase (MTHFR) (rs1801133), and lysine-specific histone demethylase 1A (KDM1A) (rs7548692) polymorphisms, erythrocyte parameters during the first week of life, and ROP. In total, 396 infants (gestational age < 32 weeks or birth weight < 1500 g) were evaluated clinically and hematologically. Genotyping was performed using a MicroChip DNA on a platform employing iPlex MassARRAY®. Multivariate regression was performed after determining risk factors for ROP using univariate regression. In the group of infants who developed ROP red blood cell distribution width (RDW), erythroblasts, and mean corpuscular volume (MCV) were higher, while mean hemoglobin and mean corpuscular hemoglobin concentration (MCHC) were lower; higher RDW was associated with KDM1A (AA), MTHFR (CC and CC + TT), KDM1A (AA) + MTHFR (CC), and KDM1A (AA) + DNMT3B (allele C); KDM1A (AA) + MTHFR (CC) were associated with higher RDW, erythroblasts, MCV, and mean corpuscular hemoglobin (MCH); higher MCV and MCH were also associated with KDM1A (AA) + MTHFR (CC) + DNMT3B (allele C). We concluded that the polymorphisms studied may influence susceptibility to ROP by modulating erythropoiesis and gene expression of the fetal/adult hemoglobin ratio.

Retinopathy of prematurity: contribution of inflammatory and genetic factors.

Retinopathy of prematurity (ROP) is a retinal vasoproliferative disorder that represents an important cause of childhood visual impairment and blindness. Although oxidative stress has long been implicated in ROP etiology, other prenatal and perinatal factors are also involved. This review focuses on current research involving inflammation and genetic factors in the pathogenesis of ROP. Increasing evidence suggests that perinatal inflammation or infection contributes to ROP pathogenesis. Cytokines and chemokines with a fundamental role in inflammatory responses and that significantly contributing to angiogenesis are analyzed. Microglia cells, the retinal-resident macrophages, are crucial for retinal homeostasis, however, under sustained pathological stimuli release exaggerated amounts of inflammatory mediators and can promote pathological neovascularization. Current modulation of angiogenic cytokines, such as treatment with antibodies to vascular endothelial growth factor (anti-VEGF), has shown efficacy in the treatment of ocular neovascularization; however, some patients are refractory to anti-VEGF agents, suggesting that other angiogenic or anti-angiogenic cytokines need to be identified. Much evidence suggests that genetic factors contribute to the phenotypic variability of ROP. Several studies have implicated the involvement of candidate genes from different signaling pathways in the development of ROP. However, a genetic component with a major impact on ROP has not yet been discovered. Most studies have limitations and did not replicate results. Future research involving bioinformatics, genomics, and proteomics may contribute to finding more genes associated with ROP and may allow discovering better solutions in the management and treatment of ROP.

A Delphi study on the clinical management of age-related macular degeneration.

PURPOSE: Age-related macular degeneration (AMD) is one of the main causes of blindness and visual impairment worldwide. As achieving a dry macula is one of the main objectives in AMD management, the purpose of this work was to reach a consensus on the relevance of retinal fluid in function, disease activity control and treatment patterns. METHODS: Forty-seven Portuguese ophthalmologists specialized in AMD participated in a DELPHI panel. Two rounds of presential meetings were conducted and a cut-off of 80% or more of votes was defined to consider answers consensual. RESULTS: Consensus was reached for 11 out of 18 questions. These questions focused on the impact of anatomical results on visual acuity, standards exams and parameters to assess disease activity, frequency and factors which influence disease activity assessment, criteria to use non-fixed treatment regimens, usefulness of individualized regimens and conditions for treatment interruption. No consensus was obtained for relevance of the different fluid types in AMD prognosis, frequency of fluid presence assessment, factors commonly associated with progression to geographic atrophy, ideal conditions for a fixed treatment regimen, date of first disease activity assessment and parameters to monitor disease activity. CONCLUSIONS: Consensus was achieved for over half of the questions assessed through this Delphi study. The questions for which no consensus was reached concerned either subjects that need further investigation or monitoring times which are influenced by resource availability. Raising awareness for these issues will allow the improvement of AMD management and treatment.

The developmental and genetic basis of 'clubfoot' in the peroneal muscular atrophy mutant mouse.

Genetic factors underlying the human limb abnormality congenital talipes equinovarus ('clubfoot') remain incompletely understood. The spontaneous autosomal recessive mouse 'peroneal muscular atrophy' mutant (PMA) is a faithful morphological model of human clubfoot. In PMA mice, the dorsal (peroneal) branches of the sciatic nerves are absent. In this study, the primary developmental defect was identified as a reduced growth of sciatic nerve lateral motor column (LMC) neurons leading to failure to project to dorsal (peroneal) lower limb muscle blocks. The pma mutation was mapped and a candidate gene encoding LIM-domain kinase 1 (Limk1) identified, which is upregulated in mutant lateral LMC motor neurons. Genetic and molecular analyses showed that the mutation acts in the EphA4-Limk1-Cfl1/cofilin-actin pathway to modulate growth cone extension/collapse. In the chicken, both experimental upregulation of Limk1 by electroporation and pharmacological inhibition of actin turnover led to defects in hindlimb spinal motor neuron growth and pathfinding, and mimicked the clubfoot phenotype. The data support a neuromuscular aetiology for clubfoot and provide a mechanistic framework to understand clubfoot in humans.

First detection of highly pathogenic avian influenza virus in Norway.

BACKGROUND: Several outbreaks of highly pathogenic avian influenza (HPAI) caused by influenza A virus of subtype H5N8 have been reported in wild birds and poultry in Europe during autumn 2020. Norway is one of the few countries in Europe that had not previously detected HPAI virus, despite widespread active monitoring of both domestic and wild birds since 2005. RESULTS: We report detection of HPAI virus subtype H5N8 in a wild pink-footed goose (Anser brachyrhynchus), and several other geese, ducks and a gull, from south-western Norway in November and December 2020. Despite previous reports of low pathogenic avian influenza (LPAI), this constitutes the first detections of HPAI in Norway. CONCLUSIONS: The mode of introduction is unclear, but a northward migration of infected geese or gulls from Denmark or the Netherlands during the autumn of 2020 is currently our main hypothesis for the introduction of HPAI to Norway. The presence of HPAI in wild birds constitutes a new, and ongoing, threat to the Norwegian poultry industry, and compliance with the improved biosecurity measures on poultry farms should therefore be ensured. [MK1]Finally, although HPAI of subtype H5N8 has been reported to have very low zoonotic potential, this is a reminder that HPAI with greater zoonotic potential in wild birds may pose a threat in the future. [MK1]Updated with a sentence emphasizing the risk HPAI pose to poultry farms, both in the Abstract and in the Conclusion-section in main text, as suggested by Reviewer 1 (#7).

Retroauricular endoscope-assisted versus conventional submandibular gland excision for benign and malignant tumors.

BACKGROUND: Surgical scars are a significant cosmetic problem, especially when in exposed areas such as the anterior neck. To avoid or reduce visible scarring, diverse innovative surgical approaches to the neck have been introduced. The purpose of this study was to evaluate the feasibility and safety of the endoscopic resection using the retroauricular approach for submandibular gland excision. METHODS: The present study enrolled 48 patients who underwent conventional transcervical submandibular gland excision and 23 patients who underwent endoscope-assisted retroauricular approach submandibular gland excision, from February 2014 through February 2018 at the Department of Head and Neck Surgery and Otorhinolaryngology of the AC Camargo Cancer Center, in Sao Paulo, Brazil. The surgical outcomes were retrospectively reviewed. RESULTS: In the conventional group, 26 (54%) patients were male. The mean age was 49.3 years (range 22-81). Twenty-two patients (46%) had sialoadenitis; twenty-one (44%) had benign and five (10%) had malignant tumors. The mean total surgical time was 86.4 min (range 40-180), and the mean total length of hospital stay was 1.3 days. Twenty-seven (56%) patients suffered from local postoperative complications in the neck. In the retroauricular group, 14 (61%) patients were male. The mean age was 44.1 years (range 24-71 years). Seven patients (31%) had sialoadenitis, twelve (53%) had benign tumors and four (16%) had malignant tumors. The mean total surgical time was 86.4 min (range 75-300 min), and the mean total length of hospital stay was 1.2 days. Twelve (53%) patients suffered from local postoperative complications in the neck. No surgical site infections or systemic complications were described. CONCLUSIONS: The retroauricular endoscopic-assisted submandibular gland resection is feasible, with excellent cosmetic results and no significant complication rate increase, and can be a safe potential surgical alternative for patients who are motivated to avoid a visible neck scar.

INVERTED INTERNAL LIMITING MEMBRANE FLAP TECHNIQUES AND OUTER RETINAL LAYER STRUCTURES.

PURPOSE: To examine the influence of the inverted flap (IF) internal limiting membrane (ILM) technique in macular hole (MH) closure on outer retinal layers after MH surgery. METHODS: Retrospective study. Postoperative position of ILM, recovery rate of external limiting membrane and ellipsoid zone, and best-corrected visual acuity were evaluated. The Inserted group, where the IF is placed inside the hole, was compared with the Cover group, where the IF completely covers the hole. RESULTS: Sixty-two eyes of 58 patients who underwent vitrectomy and ILM peeling with the IF technique for large MHs (>400 µm) with successful MH closure and a follow-up of 12 months were evaluated. In the 24 eyes of the Inserted group, there was no regeneration of external limiting membrane or ellipsoid zone after 12 months. In the 38 eyes of Cover group, external limiting membrane recovered in 55.3% of patients 1 month after surgery, and in 86.1% after 12 months. The elipsoid zone layer was present in 58% of the patients. CONCLUSION: Poorer anatomical and visual results were associated with the IF technique where ILM insertion occurs compared with ILM placed over the hole. These findings suggest that insertion of the ILM in the hole might prevent outer retinal layers realignment and visual recovery in MH surgery.

The Outcomes of Switching from Short- to Long-Term Intravitreal Corticosteroid Implant Therapy in Patients with Diabetic Macular Edema.

BACKGROUND: First-line treatment for diabetic macular edema (DME) is usually with antivascular endothelial growth factor agents, followed by intravitreal corticosteroids as a second-line treatment option. Long-term corticosteroids may offer quality of life and effectiveness benefits over short-term implants. OBJECTIVES: To evaluate outcomes of patients with persistent or recurrent DME who switched from a short-term (dexamethasone) to a long-term (fluocinolone acetonide, FAc) corticosteroid intravitreal implant in a real-world setting. METHODS: This is a retrospective study in 9 Portuguese centers. An FAc intravitreal implant was administered according to product labeling. Effectiveness outcomes were mean change in visual acuity (VA; ETDRS letters), central retinal thickness (CRT; µm), and macular volume (MV; mm3). The safety outcome was mean change in intraocular pressure (IOP; mm Hg). All were analyzed at months 1 and 3, and then quarterly until month 24 after implantation. RESULTS: Forty-four eyes from 36 patients were analyzed. Mean duration of DME was 3.3 ± 1.9 years, and mean follow-up was 8 months. From baseline following FAc implantation, VA increased significantly at months 1 and 6 (mean +6.82 and +13.02 letters, respectively; p = 0.005), and last observation carried forward (LOCF; mean +8.3 letters; p = 0.002). CRT improved significantly at months 1 and 6 (mean -71.81 and -170.77 µm, respectively; p = 0.001), and LOCF (mean -121.46 µm; p = 0.001). MV was consistently, but not significantly, decreased from baseline to LOCF (mean -0.69 mm3; p = 0.062). The mean change in IOP was -0.25 and +0.88 mm Hg at months 1 and 6, respectively (p = 0.268), and +1.86 mm Hg at LOCF (p = 0.036). Increases were controlled with topical medication in most cases. CONCLUSIONS: The FAc intravitreal implant is effective in patients previously treated with short-term corticosteroid implants. Thus, after a suboptimal response to antiangiogenics or a short-term corticosteroid, a single FAc implant may be considered an effective and tolerable treatment that can improve long-term outcomes for patients with sight-threatening DME.

Access to Primary Care for Persons Recently Released From Prison.

We aimed to determine if a history of recent imprisonment affects access to primary care. Using patient roles, we telephoned to request an initial appointment with all family physicians (n = 339) who were accepting new patients in British Columbia, Canada. We sequentially assigned patient scenarios: male or female recently released from prison; male or female control. Controls were 1.98 (95% CI, 1.59-2.46) times as likely to be offered an appointment compared with persons recently released from prison, with an absolute risk difference of 41.8% (95% CI, 31.0-52.5). Our study suggests discrimination is a barrier to primary care for people released from prison, even with universal health insurance. We need to improve access to primary care during the high-risk period following prison release.

Molecular identification and microscopic characterization of poxvirus in a Guiana dolphin and a common bottlenose dolphin, Brazil.

The poxviruses identified in cetaceans are associated with characteristic tattoo or ring skin lesions. However, little is known regarding the prevalence and progression of these lesions and the molecular characterization of cetacean poxviruses in the Southern Hemisphere. This manuscript describes the progression of poxvirus-like skin lesions in 5 free-ranging Guiana dolphins Sotalia guianensis. Additionally, 151 skin samples from 113 free-ranging cetaceans from Brazil, including 4 animals with tattoo skin lesions, were selected for poxvirus testing. Poxviral DNA polymerase gene PCR amplification was used to detect the virus in ß-actin-positive samples (145/151). DNA topoisomerase I gene PCR was then used in Cetaceanpoxvirus (CePV)-positive cases (n = 2), which were further evaluated by histopathology and electron microscopy. Based on photo-identification, adult Guiana dolphins presented regressing or healed poxvirus-like lesions (2/2), while juveniles presented persistent (2/3) or healed and progressive lesions (1/3). CePV DNA was amplified in a common bottlenose dolphin Tursiops truncatus and in a Guiana dolphin. Intracytoplasmic inclusion bodies and viral particles consistent with poxvirus were identified by histology and electron microscopy, respectively. CePV-specific amino acid motifs were identified through phylogenetic analysis. Our findings corroborate previous studies that suggest the placement of poxviruses from cetaceans within the novel CePV genus. This is the first molecular identification of poxvirus in South American odontocetes.

Intra- and Inter-Rater Agreement of Anterior Lamina Cribrosa Depth Measurements Using Enhanced-Depth Imaging Optical Coherence Tomography.

PURPOSE: To determine intra- and inter-rater agreement of anterior lamina cribrosa depth (ALCD) manual measurements using enhanced-depth imaging optical coherence tomography (EDI-OCT). METHODS: Observational study. EDI-OCT of the optic nerve head was performed in healthy subjects. ALCD was defined as the perpendicular distance between the line connecting both edges of Bruch's membrane opening and the anterior border of the lamina cribrosa (LC), at the maximum depth point. Two experienced operators performed manual ALCD measurements of the LC independently, and one of the observers performed the same measurements twice. Intra- and inter-rater agreement was evaluated using intraclass correlation coefficients (ICC) and concordance correlation coefficients (CCC) for the right eye vertical (REV) and horizontal (REH) scans and left eye vertical (LEV) and horizontal (LEH) scans. RESULTS: 120 eyes of 61 subjects, with a mean age of 62.1 ± 15.0 years. The mean REV, REH, LEV and LEH ALCD were 456.2 ± 84.3, 444.5 ± 92.2, 436.7 ± 81.6, 427.6 ± 82.7 µm, respectively. ALCD intra- and inter-rater ICC and intra- and inter-rater CCC varied between 0.85-0.95, 0.84-0.93, 0.85-0.95 and 0.84-0.93, respectively. CONCLUSIONS: ALCD manual intra- and inter-rater measurements with EDI-OCT showed high agreement. EDI-OCT is a reliable tool for ALCD measurement, which can provide potentially useful information for integrated glaucoma management.

OMERO: flexible, model-driven data management for experimental biology.

Data-intensive research depends on tools that manage multidimensional, heterogeneous datasets. We built OME Remote Objects (OMERO), a software platform that enables access to and use of a wide range of biological data. OMERO uses a server-based middleware application to provide a unified interface for images, matrices and tables. OMERO's design and flexibility have enabled its use for light-microscopy, high-content-screening, electron-microscopy and even non-image-genotype data. OMERO is open-source software, available at http://openmicroscopy.org/.

Cataract surgery and intraocular pressure.

Cataract surgery is one of the most performed surgeries in the developed world. In addition to its significant impact on visual acuity, phacoemulsification has been hailed as a potential intraocular pressure (IOP)-lowering procedure. While current evidence suggests an overall significant and sustained decrease in IOP to exist after cataract surgery, the specific ocular characteristics that could help predict which patients are likely to benefit from this IOP-lowering effect remain unclear. This definition is important in glaucoma patients if this surgery is to be used in the treatment for this disease. Our review aims to summarize the literature on the subject, depicting possible mechanisms behind this IOP decrease, which type of patients are more likely to benefit from this surgery for IOP-lowering purposes and ultimately help optimizing disease management for the increasing number of patients with concomitant glaucoma and cataract.

Review on Dynamic Contour Tonometry and Ocular Pulse Amplitude.

Intraocular pressure (IOP) measurement is the cornerstone of the management of glaucoma patients. The gold standard for assessing IOP is Goldmann applanation tonometry (GAT). Recently, the dynamic contour tonometer (DCT) has become available. While both devices provide reliable IOP measurements, the results are not interchangeable. DCT has the advantage of measuring an additional parameter: ocular pulse amplitude (OPA). OPA is defined as the difference between systolic and diastolic IOP and represents the pulsatile wave front produced by the varying amount of blood in the eye during the cardiac cycle. It has been shown to vary with ocular structural parameters, such as axial length, corneal thickness, and ocular rigidity, as well as with systemic variables like heart rate, blood pressure, and left ventricular ejection fraction. Although the existence of some of these associations is still controversial, the clinical relevance of OPA has been consistently suggested, especially in glaucoma. Further research on this intriguing parameter could not only provide insight into glaucoma pathophysiology but also help integrate this variable into clinical practice.

A comprehensive study of small non-frameshift insertions/deletions in proteins and prediction of their phenotypic effects by a machine learning method (KD4i).

BACKGROUND: Small insertion and deletion polymorphisms (Indels) are the second most common mutations in the human genome, after Single Nucleotide Polymorphisms (SNPs). Recent studies have shown that they have significant influence on genetic variation by altering human traits and can cause multiple human diseases. In particular, many Indels that occur in protein coding regions are known to impact the structure or function of the protein. A major challenge is to predict the effects of these Indels and to distinguish between deleterious and neutral variants. When an Indel occurs within a coding region, it can be either frameshifting (FS) or non-frameshifting (NFS). FS-Indels either modify the complete C-terminal region of the protein or result in premature termination of translation. NFS-Indels insert/delete multiples of three nucleotides leading to the insertion/deletion of one or more amino acids. RESULTS: In order to study the relationships between NFS-Indels and Mendelian diseases, we characterized NFS-Indels according to numerous structural, functional and evolutionary parameters. We then used these parameters to identify specific characteristics of disease-causing and neutral NFS-Indels. Finally, we developed a new machine learning approach, KD4i, that can be used to predict the phenotypic effects of NFS-Indels. CONCLUSIONS: We demonstrate in a large-scale evaluation that the accuracy of KD4i is comparable to existing state-of-the-art methods. However, a major advantage of our approach is that we also provide the reasons for the predictions, in the form of a set of rules. The rules are interpretable by non-expert humans and they thus represent new knowledge about the relationships between the genotype and phenotypes of NFS-Indels and the causative molecular perturbations that result in the disease.