Intranasal pediatric parainfluenza virus-vectored SARS-CoV-2 vaccine is protective in monkeys.

Pediatric SARS-CoV-2 vaccines are needed that elicit immunity directly in the airways as well as systemically. Building on pediatric parainfluenza virus vaccines in clinical development, we generated a live-attenuated parainfluenza-virus-vectored vaccine candidate expressing SARS-CoV-2 prefusion-stabilized spike (S) protein (B/HPIV3/S-6P) and evaluated its immunogenicity and protective efficacy in rhesus macaques. A single intranasal/intratracheal dose of B/HPIV3/S-6P induced strong S-specific airway mucosal immunoglobulin A (IgA) and IgG responses. High levels of S-specific antibodies were also induced in serum, which efficiently neutralized SARS-CoV-2 variants of concern of alpha, beta, and delta lineages, while their ability to neutralize Omicron sub-lineages was lower. Furthermore, B/HPIV3/S-6P induced robust systemic and pulmonary S-specific CD4+ and CD8+ T cell responses, including tissue-resident memory cells in the lungs. Following challenge, SARS-CoV-2 replication was undetectable in airways and lung tissues of immunized macaques. B/HPIV3/S-6P will be evaluated clinically as pediatric intranasal SARS-CoV-2/parainfluenza virus type 3 vaccine.

Recombinant OC43 SARS-CoV-2 spike replacement virus: An improved BSL-2 proxy virus for SARS-CoV-2 neutralization assays.

We generated a replication-competent OC43 human seasonal coronavirus (CoV) expressing the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike in place of the native spike (rOC43-CoV2 S). This virus is highly attenuated relative to OC43 and SARS-CoV-2 in cultured cells and animals and is classified as a biosafety level 2 (BSL-2) agent by the NIH biosafety committee. Neutralization of rOC43-CoV2 S and SARS-CoV-2 by S-specific monoclonal antibodies and human sera is highly correlated, unlike recombinant vesicular stomatitis virus-CoV2 S. Single-dose immunization with rOC43-CoV2 S generates high levels of neutralizing antibodies against SARS-CoV-2 and fully protects human ACE2 transgenic mice from SARS-CoV-2 lethal challenge, despite nondetectable replication in respiratory and nonrespiratory organs. rOC43-CoV2 S induces S-specific serum and airway mucosal immunoglobulin A and IgG responses in rhesus macaques. rOC43-CoV2 S has enormous value as a BSL-2 agent to measure S-specific antibodies in the context of a bona fide CoV and is a candidate live attenuated SARS-CoV-2 mucosal vaccine that preferentially replicates in the upper airway.

IL-10 suppresses T cell expansion while promoting tissue-resident memory cell formation during SARS-CoV-2 infection in rhesus macaques.

The regulation of inflammatory responses and pulmonary disease during SARS-CoV-2 infection is incompletely understood. Here we examine the roles of the prototypic pro- and anti-inflammatory cytokines IFNγ and IL-10 using the rhesus macaque model of mild COVID-19. We find that IFNγ drives the development of 18fluorodeoxyglucose (FDG)-avid lesions in the lungs as measured by PET/CT imaging but is not required for suppression of viral replication. In contrast, IL-10 limits the duration of acute pulmonary lesions, serum markers of inflammation and the magnitude of virus-specific T cell expansion but does not impair viral clearance. We also show that IL-10 induces the subsequent differentiation of virus-specific effector T cells into CD69+CD103+ tissue resident memory cells (Trm) in the airways and maintains Trm cells in nasal mucosal surfaces, highlighting an unexpected role for IL-10 in promoting airway memory T cells during SARS-CoV-2 infection of macaques.

Live-attenuated pediatric parainfluenza vaccine expressing 6P-stabilized SARS-CoV-2 spike protein is protective against SARS-CoV-2 variants in hamsters.

The pediatric live-attenuated bovine/human parainfluenza virus type 3 (B/HPIV3)-vectored vaccine expressing the prefusion-stabilized SARS-CoV-2 spike (S) protein (B/HPIV3/S-2P) was previously evaluated in vitro and in hamsters. To improve its immunogenicity, we generated B/HPIV3/S-6P, expressing S further stabilized with 6 proline mutations (S-6P). Intranasal immunization of hamsters with B/HPIV3/S-6P reproducibly elicited significantly higher serum anti-S IgA/IgG titers than B/HPIV3/S-2P; hamster sera efficiently neutralized variants of concern (VoCs), including Omicron variants. B/HPIV3/S-2P and B/HPIV3/S-6P immunization protected hamsters against weight loss and lung inflammation following SARS-CoV-2 challenge with the vaccine-matched strain WA1/2020 or VoCs B.1.1.7/Alpha or B.1.351/Beta and induced near-sterilizing immunity. Three weeks post-challenge, B/HPIV3/S-2P- and B/HPIV3/S-6P-immunized hamsters exhibited a robust anamnestic serum antibody response with increased neutralizing potency to VoCs, including Omicron sublineages. B/HPIV3/S-6P primed for stronger anamnestic antibody responses after challenge with WA1/2020 than B/HPIV3/S-2P. B/HPIV3/S-6P will be evaluated as an intranasal vaccine to protect infants against both HPIV3 and SARS-CoV-2.

PET imaging of TSPO expression in immune cells can assess organ-level pathophysiology in high-consequence viral infections.

Ebola virus (EBOV) disease is characterized by lymphopenia, breach in vascular integrity, cytokine storm, and multiorgan failure. The pathophysiology of organ involvement, however, is incompletely understood. Using [18F]-DPA-714 positron emission tomography (PET) imaging targeting the translocator protein (TSPO), an immune cell marker, we sought to characterize the progression of EBOV-associated organ-level pathophysiology in the EBOV Rhesus macaque model. Dynamic [18F]-DPA-714 PET/computed tomography imaging was performed longitudinally at baseline and at multiple time points after EBOV inoculation, and distribution volumes (Vt) were calculated as a measure of peripheral TSPO binding. Using a mixed-effect linear regression model, spleen and lung Vt decreased, while the bone marrow Vt increased over time after infection. No clear trend was found for liver Vt. Multiple plasma cytokines correlated negatively with lung/spleen Vt and positively with bone marrow Vt. Multiplex immunofluorescence staining in spleen and lung sections confirmed organ-level lymphoid and monocytic loss/apoptosis, thus validating the imaging results. Our findings are consistent with EBOV-induced progressive monocytic and lymphocytic depletion in the spleen, rather than immune activation, as well as depletion of alveolar macrophages in the lungs, with inefficient reactive neutrophilic activation. Increased bone marrow Vt, on the other hand, suggests hematopoietic activation in response to systemic immune cell depletion and leukocytosis and could have prognostic relevance. In vivo PET imaging provided better understanding of organ-level pathophysiology during EBOV infection. A similar approach can be used to delineate the pathophysiology of other systemic infections and to evaluate the effectiveness of newly developed treatment and vaccine strategies.

Exploring the prospective acceptability of a healthy food incentive program from the perspective of people with type 2 diabetes and experiences of household food insecurity in Alberta, Canada.

OBJECTIVE: FoodRx is a 12-month healthy food prescription incentive program for people with type 2 diabetes (T2DM) and experiences of household food insecurity. In this study, we aimed to explore potential users' prospective acceptability (acceptability prior to program use) of the design and delivery of the FoodRx incentive and identify factors influencing prospective acceptability. DESIGN: We used a qualitative descriptive approach and purposive sampling to recruit individuals who were interested or uninterested in using the FoodRx incentive. Semi-structured interviews were guided by the theoretical framework of acceptability, and corresponding interview transcripts were analysed using differential qualitative analysis guided by the socioecological model. SETTING: Individuals living in Alberta, Canada. PARTICIPANTS: In total, fifteen adults with T2DM and experiences of household food insecurity. RESULTS: People who were interested in using the FoodRx incentive (n 10) perceived it to be more acceptable than those who were uninterested (n 5). We identified four themes that captured factors that influenced users' prospective acceptability: (i) participants' confidence, views and beliefs of FoodRx design and delivery and its future use (intrapersonal), (ii) the shopping routines and roles of individuals in participants' social networks (interpersonal), (iii) access to and experience with food retail outlets (community), and (iv) income and food access support to cope with the cost of living (policy). CONCLUSION: Future healthy food prescription programs should consider how factors at all levels of the socioecological model influence program acceptability and use these data to inform program design and delivery.

A single intranasal dose of a live-attenuated parainfluenza virus-vectored SARS-CoV-2 vaccine is protective in hamsters.

Single-dose vaccines with the ability to restrict SARS-CoV-2 replication in the respiratory tract are needed for all age groups, aiding efforts toward control of COVID-19. We developed a live intranasal vector vaccine for infants and children against COVID-19 based on replication-competent chimeric bovine/human parainfluenza virus type 3 (B/HPIV3) that express the native (S) or prefusion-stabilized (S-2P) SARS-CoV-2 S spike protein, the major protective and neutralization antigen of SARS-CoV-2. B/HPIV3/S and B/HPIV3/S-2P replicated as efficiently as B/HPIV3 in vitro and stably expressed SARS-CoV-2 S. Prefusion stabilization increased S expression by B/HPIV3 in vitro. In hamsters, a single intranasal dose of B/HPIV3/S-2P induced significantly higher titers compared to B/HPIV3/S of serum SARS-CoV-2-neutralizing antibodies (12-fold higher), serum IgA and IgG to SARS-CoV-2 S protein (5-fold and 13-fold), and IgG to the receptor binding domain (10-fold). Antibodies exhibited broad neutralizing activity against SARS-CoV-2 of lineages A, B.1.1.7, and B.1.351. Four weeks after immunization, hamsters were challenged intranasally with 104.5 50% tissue-culture infectious-dose (TCID50) of SARS-CoV-2. In B/HPIV3 empty vector-immunized hamsters, SARS-CoV-2 replicated to mean titers of 106.6 TCID50/g in lungs and 107 TCID50/g in nasal tissues and induced moderate weight loss. In B/HPIV3/S-immunized hamsters, SARS-CoV-2 challenge virus was reduced 20-fold in nasal tissues and undetectable in lungs. In B/HPIV3/S-2P-immunized hamsters, infectious challenge virus was undetectable in nasal tissues and lungs; B/HPIV3/S and B/HPIV3/S-2P completely protected against weight loss after SARS-CoV-2 challenge. B/HPIV3/S-2P is a promising vaccine candidate to protect infants and young children against HPIV3 and SARS-CoV-2.

Advances and Challenges in Molecular Imaging of Viral Infections.

Molecular imaging of viral infection, using a variety of advanced imaging techniques such as optical and nuclear imaging, can and has been used for direct visualization of the virus as well as assessment of virus-host interactions. Unlike imaging of other pathogens such as bacteria and fungi, challenging aspects of imaging viral infections include the small size of viruses, the complexity of viral infection animal models (eg, species dependence), and the high-level containment needs for many high-consequence pathogens, among others. In this review, using representative viral infections, we discuss how molecular imaging can reveal real-time infection dynamics, improve our understanding of disease pathogenesis, and guide optimization of treatment and prevention strategies. Key findings from human and animal studies are highlighted.

Endogenous and Therapeutic 25-Hydroxycholesterols May Worsen Early SARS-CoV-2 Pathogenesis in Mice.

Oxysterols (i.e., oxidized cholesterol species) have complex roles in biology. 25-Hydroxycholesterol (25HC), a product of the activity of cholesterol-25-hydroxylase (CH25H) on cholesterol, has recently been shown to be broadly antiviral, suggesting therapeutic potential against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). However, 25HC can also amplify inflammation and be converted by CYP7B1 (cytochrome P450 family 7 subfamily B member 1) to 7α,25-dihydroxycholesterol, a lipid with chemoattractant activity, via the G protein-coupled receptor EBI2 (Epstein-Barr virus-induced gene 2)/GPR183 (G protein-coupled receptor 183). Here, using in vitro studies and two different murine models of SARS-CoV-2 infection, we investigate the effects of these two oxysterols on SARS-CoV-2 pneumonia. We show that although 25HC and enantiomeric-25HC are antiviral in vitro against human endemic coronavirus-229E, they did not inhibit SARS-CoV-2; nor did supplemental 25HC reduce pulmonary SARS-CoV-2 titers in the K18-human ACE2 (angiotensin-converting enzyme 2) mouse model in vivo. Treatment with 25HC also did not alter immune cell influx into the airway, airspace cytokines, lung pathology, weight loss, symptoms, or survival but was associated with increased airspace albumin, an indicator of microvascular injury, and increased plasma proinflammatory cytokines. Conversely, mice treated with the EBI2/GPR183 inhibitor NIBR189 displayed a modest increase in lung viral load only at late time points but no change in weight loss. Consistent with these findings, although Ch25h and 25HC were upregulated in the lungs of SARS-CoV-2-infected wild-type mice, lung viral titers and weight loss in Ch25h-/- and Gpr183-/- mice infected with the ß variant were similar to those in control animals. Taken together, endogenous 25HCs do not significantly regulate early SARS-CoV-2 replication or pathogenesis, and supplemental 25HC may have proinjury rather than therapeutic effects in SARS-CoV-2 pneumonia.

Patents and regulatory exclusivities on FDA-approved insulin products: A longitudinal database study, 1986-2019.

BACKGROUND: Insulin is the primary treatment for type 1 and some type 2 diabetes but remains costly in the United States, even though it was discovered more than a century ago. High prices can lead to nonadherence and are often sustained by patents and regulatory exclusivities that limit competition on brand-name products. We sought to examine how manufacturers have used patents and regulatory exclusivities on insulin products approved from 1986 to 2019 to extend periods of market exclusivity. METHODS AND FINDINGS: We used the publicly available Food and Drug Administration (FDA) Approved Drug Products with Therapeutic Equivalence Evaluations (Orange Book) to identify all approved biosynthetic insulin products. Individual products approved under the same New Drug Application (NDA)-e.g., a vial and pen-were considered as separate products for the purposes of analysis. We recorded all patents and regulatory exclusivities listed in the Orange Book on each product and used Google Patents to extract the timing of patent application and whether patents were obtained on delivery devices or others aspects of the product. The primary outcome was the duration of expected protection, which was determined by subtracting the FDA approval date for each product from its last-to-expire patent or regulatory exclusivity (whichever occurred later). We performed a secondary analysis that considered overall protection on insulin lines-defined as groups of products approved under the same NDA with the same active ingredients manufactured by the same company. We also examined competition from follow-on insulin products-defined as products approved with the same active ingredients as originators but manufactured by different companies (approved via a specific drug approval pathway under section 505(b)(2) of the Food, Drug, and Cosmetic Act). During the study period, the FDA approved 56 individual products across 25 different insulin lines and 5 follow-ons across 3 different insulin lines. Thirty-three (59%) of the 56 products were drug-device combinations. Manufacturers of 9 products approved during the study period obtained patents filed after FDA approval that extended their duration of expected protection (by a median of 6 years). Approximately 63% of all patents on drug-device combinations approved during the study period were related to delivery devices. The median duration of expected protection on insulin products was 16.0 years, and the median protection on insulin lines was 17.6 years. An important limitation of our analysis is that manufacturers may continue to add patents on existing insulin products while competitors may challenge patents; therefore, periods of protection may change over time. CONCLUSIONS: Among several strategies that insulin manufacturers have employed to extend periods of market exclusivity on brand-name insulin products are filing patents after FDA approval and obtaining a large number of patents on delivery devices. Policy reforms are needed to promote timely competition in the pharmaceutical market and ensure that patients have access to low-cost drugs.

Four challenges to an effective national nature assessment.

Comprehensive biodiversity assessments play an essential role in strengthening global and national conservation strategies. The recently announced first U.S. National Nature Assessment (NNA) provides an unparalleled opportunity to comprehensively review status and trends of biodiversity at all levels. This broad context can help in the coordination of actions to conserve individual species and ecosystems. The scientific assessments that informed the Kunming-Montreal Global Biodiversity Framework adopted at the 2022 Convention on Biological Diversity (CBD) conference of parties provide models for synthesizing information on trends at multiple levels of biodiversity, including decline in abundance and distribution of species, loss of populations and genetic diversity, and degradation and loss of ecosystems and their services. The assessments then relate these trends to data on drivers of biodiversity loss and pathways to their mitigation. The U.S. NNA can augment such global analyses and avoid the pitfalls encountered by previous U.S. efforts by ensuring policy-relevant design, data accessibility, and inclusivity in process and product and by incorporating spatial data relevant to national and subnational audiences. Although the United States is not formally a CBD party, an effective NNA should take full advantage of the global context by including indicators adopted at the 2022 meeting and incorporating an independent review mechanism that supports periodic stocktaking and ratcheting up of ambition in response to identified shortfalls in stemming biodiversity loss. The challenges to design of an effective U.S. assessment are relevant globally as nations develop assessments and reporting to support the new global biodiversity framework's targets. By considering and incorporating the diverse ways in which society values and benefits from nature, such assessments can help bridge the gap between research and conservation practice and communicate the extent of the biodiversity crisis to the public, fostering broad-based support for transformative change in humanity's relationship to the natural world.

Cuatro obstáculos para una eficaz evaluación nacional de la naturaleza Resumen Las evaluaciones completas de la biodiversidad tienen un papel esencial en el fortalecimiento de las estrategias de conservación nacional y mundial. La recién anunciada Evaluación Nacional de la Naturaleza (ENN) de los EE. UU. proporciona una oportunidad sin precedentes para revisar de manera completa el estado y las tendencias de la biodiversidad en todos los niveles. Este contexto generalizado puede ayudar a la coordinación de acciones para la conservación de especies individuales y ecosistemas. Las evaluaciones científicas que guiaron el Marco Mundial de la Biodiversidad de Kumming-Montreal adoptado en la conferencia de las partes de la Convención sobre la Diversidad Biológica (CBD) de 2022 proporcionan modelos para sintetizar la información sobre las tendencias de la biodiversidad a varios niveles, incluyendo la declinación en abundancia y distribución de especies, pérdida de poblaciones y diversidad genética y la degradación y pérdida de los ecosistemas y sus servicios. Después de esto, las evaluaciones relacionan estas tendencias con la información sobre los causantes de la pérdida de la biodiversidad y las maneras de mitigarla. La ENN de los EE. UU. pueden aumentar estos análisis mundiales y evitar las dificultades enfrentadas por los esfuerzos previos al garantizar un diseño relevante para las políticas, la disponibilidad de datos y la inclusión en el proceso y el producto y también mediante la incorporación de datos espaciales relevantes para el público nacional y subnacional. Aunque los EE. UU. no son una parte formal de la CBD, una ENN efectiva debería aprovechar de lleno el contexto global al incluir los indicadores adoptados en la reunión de 2022 e incorporar un mecanismo independiente de revisión que respalde el balance periódico y el aumento de la ambición en respuesta a las deficiencias detectadas en la contención de la pérdida de biodiversidad. Los retos para diseñar una evaluación estadunidense son relevantes a nivel mundial ya que los países evalúan y reportan para mantener los objetivos mundiales de biodiversidad post-2020. Si consideramos las diferentes maneras en las que la sociedad valora y se beneficia de la naturaleza, dichas evaluaciones pueden ayudar a cerrar la brecha entre la investigación y la práctica de la conservación y a comunicarle al público el nivel de la crisis de la biodiversidad, lo que fomenta el apoyo generalizado para transformar la relación entre la humanidad y el mundo natural.

Patents and Regulatory Exclusivities on GLP-1 Receptor Agonists.

Importance: Glucagon-like peptide 1 (GLP-1) receptor agonists were first approved for the treatment of type 2 diabetes in 2005. Demand for these drugs has increased rapidly in recent years, as indications have expanded, but they remain expensive. Objective: To analyze how manufacturers of brand-name GLP-1 receptor agonists have used the patent and regulatory systems to extend periods of market exclusivity. Evidence Review: The annual US Food and Drug Administration's (FDA) Approved Drug Products With Therapeutic Equivalence Evaluations was used to identify GLP-1 receptor agonists approved from 2005 to 2021 and to record patents and nonpatent statutory exclusivities listed for each product. Google Patents was used to extract additional data on patents, including whether each was obtained on the delivery device or another aspect of the product. The primary outcome was the duration of expected protection from generic competition, defined as the time elapsed from FDA approval until expiration of the last-to-expire patent or regulatory exclusivity. Findings: On the 10 GLP-1 receptor agonists included in the cohort, drug manufacturers listed with the FDA a median of 19.5 patents (IQR, 9.0-25.8) per product, including a median of 17 patents (IQR, 8.3-22.8) filed before FDA approval and 1.5 (IQR, 0-2.8) filed after FDA approval. Fifty-four percent of all patents listed on GLP-1 receptor agonists were on the delivery devices rather than active ingredients. Manufacturers augmented patent protection with a median of 2 regulatory exclusivities (IQR, 0-3) obtained at approval and 1 (IQR, 0.3-4.3) added after approval. The median total duration of expected protection after FDA approval, when accounting for both preapproval and postapproval patents and regulatory exclusivities, was 18.3 years (IQR, 16.0-19.4). No generic firm has successfully challenged patents on GLP-1 receptor agonists to gain FDA approval. Conclusions and Relevance: Patent and regulatory reform is needed to ensure timely generic entry of GLP-1 receptor agonists to the market.

Premarket Development Times for Innovative Vaccines--To What Extent Are the Coronavirus Disease 2019 (COVID-19) Vaccines Outliers?

One reason expressed in surveys of people reporting coronavirus disease 2019 (COVID-19) vaccine hesitancy is how rapidly these vaccines have reached the market. To estimate the length of time the COVID-19 vaccine spent in research and development as compared to other novel vaccines, we apply previously established methods for estimating medical product development times, using the key associated patent filings cited by the manufacturer as the marker of when commercial development activity began. Applying these methods to a cohort of recently approved innovative vaccines and comparing them to the first-approved COVID-19 vaccine (BioNTech/Pfizer), we found key patent filings for the technology in this COVID-19 vaccine occurred 10.0 years prior to regulatory authorization. By this metric, the development timelines for innovative vaccines have been shortening since the 1980s, and the COVID-19 vaccine comfortably fits within this pattern. Vaccine development timelines have now even drawn to parity with many of the most commonly used drugs.

T-Cell Immunoglobulin and Mucin Domain-Containing Protein-4 Is Critical for Kupffer Cell Homeostatic Function in the Activation and Resolution of Liver Ischemia Reperfusion Injury.

BACKGROUND AND AIMS: Liver ischemia reperfusion injury (IRI) remains an unresolved clinical problem. This study dissected roles of liver-resident macrophage Kupffer cells (KCs), with a functional focus on efferocytosis receptor T-cell immunoglobulin and mucin domain-containing protein-4 (TIM-4), in both the activation and resolution of IRI in a murine liver partial warm ischemia model. APPROACH AND RESULTS: Fluorescence-activated cell sorting results showed that TIM-4 was expressed exclusively by KCs, but not infiltrating macrophages (iMФs), in IR livers. Anti-TIM-4 antibody depleted TIM-4+ macrophages in vivo, resulting in either alleviation or deterioration of liver IRI, which was determined by the repopulation kinetics of the KC niche with CD11b+ macrophages. To determine the KC-specific function of TIM-4, we reconstituted clodronate-liposome-treated mice with exogenous wild-type or TIM-4-deficient KCs at either 0 hour or 24 hours postreperfusion. TIM-4 deficiency in KCs resulted in not only increases in the severity of liver IRI (at 6 hours postreperfusion), but also impairment of the inflammation resolution (at 7 days postreperfusion). In vitro analysis revealed that TIM-4 promoted KC efferocytosis to regulate their Toll-like receptor response by up-regulating IL-10 and down-regulating TNF-α productions. CONCLUSIONS: TIM-4 is critical for KC homeostatic function in both the activation and resolution of liver IRI by efferocytosis.

Laparoscopic and robotic pyeloplasty as minimally invasive alternatives to the open approach for the treatment of uretero-pelvic junction obstruction in infants: a multi-institutional comparison of outcomes and learning curves.

BACKGROUND: Since the development of minimally invasive surgery (MIS), laparoscopic and robotic approaches have been widely adopted. However, little has been published detailing the learning curve of MIS, especially in infants. OBJECTIVE: To quantify the learning curve of laparoscopic (LP) and robot-assisted laparoscopic pyeloplasty (RAL-P) for treatment of uretero-pelvic junction obstruction (UPJO) in infants evidenced by number of cases, operative time, success and complications. PATIENTS AND METHODS: Between 2009 and 2017, we retrospectively reviewed pyeloplasty cases for treatment of UPJO in infants at three academic institutions. The primary outcome was success. Secondary outcomes were UPJO recurrence, complications, and operative time as a surrogate of skill acquisition. Continuous variables were analyzed by t test, Welch-test, and one-way ANOVA. Non-continuous variables were analyzed by Chi-squared test or Fisher's exact test. Learning curves (LC) were studied by r-to-z transformation and CUSUM. RESULTS: Thirty-nine OP, 26 LP, and 39 RAL-P had mean operative times (OT) of 106, 121, and 151 min, respectively. LCs showed plateau in OT after 18 and 13 cases for LP and RAL-P, respectively. RAL-P showed a second phase of further improvements after 37 cases. At 16 months follow-up, there were similar rates of success and complications between the three groups. CONCLUSIONS: Despite different duration of learning phases, proficiency was achieved in both LP and RAL-P as evidenced by stabilization of operative time and similar success rates and complications to OP. Before and after achievement of proficiency, LP and RAL-P can be safely learned and implemented for treatment of UPJO in infants.

COVID-19 and the prevalence of drug shortages in Canada: a cross-sectional time-series analysis from April 2017 to April 2022.

BACKGROUND: In March 2020, the Government of Canada introduced measures to reduce intensifying shortages of prescription drugs during the beginning of the COVID-19 pandemic. We sought to assess the extent to which a decline in drug shortages was observed in the months after this policy change. METHODS: Our data source was the Drug Shortages Canada Database, which reports shortages by drug product, including shortage start and duration. Using a cross-sectional design, we tracked shortage rates of drug products using a 30-day moving average from Apr. 15, 2017, to Apr. 1, 2022. We used autoregressive integrated moving average modelling with a ramp function to determine the significance of trend changes after policy implementation. RESULTS: We found that of the 13 329 drug products at risk for shortage, 44.7% (n = 5953) had at least 1 shortage event in the past 5 years. Average daily shortage prevalence rates rose from 901 in April 2017 to a peak of 2345 by April 2020. Significant declines (p = 0.02) ensued shortly thereafter, dropping to a rate of 1611 shortages by the end of the first year after policy implementation. However, we did not observe a significant reduction in shortage rates in the second year (p = 0.2), with rates plateauing below 1500 and then rising back above 1600 by the end of March 2022. INTERPRETATION: Drug shortages are common in Canada, including during the initial months of the COVID-19 pandemic. We observed substantial improvements after the implementation of the new measures, but gains appear to have plateaued. Continued vigilance is needed to sustain improvements.

How percentage-protected targets can support positive biodiversity outcomes.

Global targets for the percentage area of land protected, such as 30% by 2030, have gained increasing prominence, but both their scientific basis and likely effectiveness have been questioned. As with emissions-reduction targets based on desired climate outcomes, percentage-protected targets combine values and science by estimating the area over which conservation actions are required to help achieve desired biodiversity outcomes. Protected areas are essential for achieving many biodiversity targets, in part because many species are highly sensitive to human-associated disturbance. However, because the contribution of protected areas to biodiversity outcomes is contingent on their location, management, governance, threats, and what occurs across the broader landscape matrix, global percentage-protected targets are unavoidably empirical generalizations of ecological patterns and processes across diverse geographies. Percentage-protected targets are insufficient in isolation but can complement other actions and contribute to biodiversity outcomes within a framework that balances accuracy and pragmatism in a global context characterized by imperfect biodiversity data. Ideally, percentage-protected targets serve as anchors that strengthen comprehensive national biodiversity strategies by communicating the level of ambition necessary to reverse current trends of biodiversity loss. If such targets are to fulfill this role within the complex societal process by which both values and science impel conservation actions, conservation scientists must clearly communicate the nature of the evidence base supporting percentage-protected targets and how protected areas can function within a broader landscape managed for sustainable coexistence between people and nature. A new paradigm for protected and conserved areas recognizes that national coordination, incentives, and monitoring should support rather than undermine diverse locally led conservation initiatives. However, the definition of a conserved area must retain a strong focus on biodiversity to remain consistent with the evidence base from which percentage-protected targets were originally derived.

RESUMEN: Las metas globales del porcentaje de área de suelo protegido, como el de 30% para el 2030, han obtenido una prominencia incrementada, a pesar de que se les cuestionen sus bases científicas y la probabilidad de su efectividad. Así como las metas de reducción de emisiones, las metas de porcentaje de protección combinan valores y ciencia mediante la estimación del área que requiere acciones de conservación para ayudar a lograr los resultados deseados de biodiversidad. Las áreas protegidas son esenciales para alcanzar muchas metas de biodiversidad, en parte porque muchas especies son altamente sensibles a las perturbaciones asociadas con el humano. Sin embargo, debido a que la contribución de las áreas protegidas a los resultados de biodiversidad depende de su ubicación, gestión, manejo, amenazas y lo que ocurra a lo largo de la amplia matriz del paisaje, las metas de porcentaje de protección son generalizaciones empíricas inevitables de los patrones y procesos ecológicos en la geografía diversa. Las metas de porcentaje de protección son insuficientes por sí solas, pero pueden complementar a otras acciones y contribuir a los resultados de biodiversidad dentro de un marco de trabajo que balancee la exactitud y el pragmatismo dentro de un contexto global caracterizado por datos imperfectos de la biodiversidad. Idealmente, las metas de porcentaje de protección fungen como pilares que fortalecen las estrategias nacionales integrales de biodiversidad mediante la comunicación del nivel de ambición necesaria para revertir las tendencias actuales de pérdida de la biodiversidad. Si se espera que dichas metas realicen este papel dentro del complejo proceso social en el que tanto los valores como la ciencia impulsan las acciones de conservación, los científicos de la conservación deben comunicar claramente la naturaleza de la base de evidencias que respalda las metas de porcentaje de protección y cómo las áreas protegidas pueden funcionar dentro de un paisaje más amplio gestionado por la coexistencia sustentable entre la naturaleza y las personas. Un nuevo paradigma para las áreas protegidas y conservadas reconoce que la coordinación nacional, los incentivos y el monitoreo deberían respaldar, y no debilitar, a las diferentes iniciativas de conservación llevadas por la población local. Sin embargo, la definición de un área conservada debe mantener un enfoque sólido sobre la biodiversidad para seguir siendo coherente con la base de evidencias de la cual derivaron originalmente las metas de porcentaje de protección.

Filoviruses Infect Rhesus Macaque Synoviocytes in Vivo and Primary Human Synoviocytes in Vitro.

The most commonly reported symptom of post-Ebola virus disease syndrome in survivors is arthralgia, yet involvement of the joints in acute or convalescent Ebola virus infection is not well characterized in human patients or animal models. Through immunohistochemistry, we found that the lining synovial intima of the stifle (knee) is a target for acute infection by Ebola virus/Kikwit, Ebola virus/Makona-C05, and Marburg virus/Angola in the rhesus macaque model. Furthermore, histologic analysis, immunohistochemistry, RNAscope in situ hybridization, and transmission electron microscopy showed that synoviocytes of the stifle, shoulder, and hip are a target for mouse-adapted Ebola virus/Yambuku-Mayinga infection during acute disease in rhesus macaques. A time course of infection study with Ebola virus/Kikwit found that the large joint synovium became immunopositive beginning on postinfection day 6. In total, the synovium of 28 of 30 rhesus macaques with terminal filovirus disease had evidence of infection (64 of 96 joints examined). On the basis of immunofluorescence, infected cell types included CD68+ type A (macrophage-like) synoviocytes and CD44+ type B (fibroblast-like) synoviocytes. Cultured primary human fibroblast-like synoviocytes were permissive to infection with Ebola and Marburg viruses in vitro. Because synovial joints include immune privileged sites, these findings are significant for future investigations of filovirus pathogenesis and persistence as well as arthralgias in acute and convalescent filovirus disease.

Reimagining Pharmaceutical Market Exclusivities: Should the Duration of Guaranteed Monopoly Periods Be Value Based?

OBJECTIVES: To describe the main features of a pharmaceutical market in which the duration of guaranteed monopoly periods would correspond to a new pharmaceutical product's value. METHODS: After reviewing patent and regulatory exclusivity-based mechanisms for protecting prescription drug markets from competition to incentivize drug innovation in developed countries, we model market protection mechanisms within the current framework to give the longest-lasting market protections to drug developers that bring the most affordable products to market with highest public health and clinical value. RESULTS: An approach tying pharmaceutical market exclusivity to value would have 3 main features. First, it would be based on regulatory exclusivity (ie, the drug regulator refrains from authorizing generic entry for a certain amount of time), rather than patents. Second, the duration of exclusivity period would be pegged to the magnitude of a product's anticipated health impact and its proposed price by using modified methods from the field of health technology assessment. Third, the duration of the value-based exclusivity period would be reassessed routinely 3 years after the product's launch to account for its real-world effectiveness. CONCLUSIONS: Linking a drug's proposed price to the duration of its regulatory-based exclusivities would both incentivize the development of high impact, low-cost products and motivate drug developers to introduce these products at lower prices.

Rewilding in the face of climate change.

Expansion of the global protected-area network has been proposed as a strategy to address threats from accelerating climate change and species extinction. A key step in increasing the effectiveness of such expansion is understanding how novel threats to biodiversity from climate change alter concepts such as rewilding, which have underpinned many proposals for large interconnected reserves. We reviewed potential challenges that climate change poses to rewilding and found that the conservation value of large protected areas persists under climate change. Nevertheless, more attention should be given to protection of microrefugia, macrorefugia, complete environmental gradients, and areas that connect current and future suitable climates and to maintaining ecosystem processes and stabilizing feedbacks via conservation strategies that are resilient to uncertainty regarding climate trends. Because a major element of the threat from climate change stems from its novel geographic patterns, we examined, as an example, the implications for climate-adaptation planning of latitudinal, longitudinal (continental to maritime), and elevational gradients in climate-change exposure across the Yellowstone-to-Yukon region, the locus of an iconic conservation proposal initially designed to conserve wide-ranging carnivore species. In addition to a continued emphasis on conserving intact landscapes, restoration of degraded low-elevation areas within the region is needed to capture sites important for landscape-level climate resilience. Extreme climate exposure projected for boreal North America suggests the need for ambitious goals for expansion of the protected-area network there to include refugia created by topography and ecological features, such as peatlands, whose conservation can also reduce emissions from carbon stored in soil. Qualitative understanding of underlying reserve design rules and the geography of climate-change exposure can strengthen the outcomes of inclusive regional planning processes that identify specific sites for protection.

Retorno a la Vida Silvestre de frente al Cambio Climático Resumen La expansión de la red mundial de áreas protegidas ha sido propuesta como una estrategia para tratar con las amenazas del creciente cambio climático y la extinción de especies. Un paso importante para el incremento de la efectividad de dicha expansión es el entendimiento de cómo las amenazas novedosas para la biodiversidad que provienen del cambio climático alteran conceptos como el retorno a la vida silvestre, el cual ha apuntalado muchas propuestas de grandes reservas interconectadas. Revisamos los obstáculos potenciales que representan el cambio climático para el retorno a la vida silvestre y encontramos que el valor de conservación de las áreas protegidas grandes persiste bajo el cambio climático. Sin embargo, se le debería brindar mayor atención a la protección de los microrefugios, del gradiente ambiental completo y de las áreas que conectan climas adecuados actuales y futuros. También se le debe brindar atención al mantenimiento de los procesos ambientales y a la estabilización de la retroalimentación por medio las estrategias de conservación que son resilientes a la incertidumbre relacionada con las tendencias climáticas. Ya que un elemento principal de la amenaza que representa el cambio climático surge de sus patrones geográficos novedosos examinamos, como ejemplo, las implicaciones de los gradientes latitudinales, longitudinales (continental a marítima) y altitudinales para la planeación de la adaptación climática dentro de la exposición al cambio climático en toda la región de Yellowstone a Yukón, el sitio de una propuesta icónica de conservación diseñada inicialmente para conservar especies carnívoras de amplia distribución. Además de un énfasis continuo sobre la conservación intacta del paisaje, se requiere la restauración de las áreas degradadas de baja elevación dentro de la región para capturar los sitios importantes para la resiliencia climática a nivel de paisaje. La exposición climática extrema proyectada para la parte boreal de América del Norte sugiere que se necesitan metas ambiciosas para la expansión de la red de áreas protegidas que se encuentran allí para incluir también a los refugios creados por la topografía y las características ecológicas, como las turberas, cuya conservación también puede reducir las emisiones de carbono almacenado en el suelo. El entendimiento cualitativo de las reglas subyacentes de diseño de reservas y la geografía de la exposición al cambio climático puede fortalecer los resultados de los procesos de planeación regional incluyente para identificar los sitios específicos que requieren protección.