N-tert-Butoxycarbonyl-N-(2-(tritylthio)ethoxy)glycine as a Building Block for Peptide Ubiquitination.

N-Boc-N-(2-(tritylthio)ethoxy)glycine has been developed as a building block for peptide ubiquitination, which is fully compatible with solid-phase Fmoc chemistry and common peptide modifications including phosphorylation, methylation, acetylation, biotinylation, and fluorescence labeling. The optimal conditions for peptide cleavage and auxiliary removal were obtained. The utility of this building block in peptide ubiquitination was demonstrated by the synthesis of seven ubiquitinated histone and Tau peptides bearing various modifications. Cys residues were well tolerated and did not require orthogonal protection. The structural integrity and folding of the synthesized ubiquitinated peptides were confirmed by enzymatic deubiquitination of a fluorescently labeled ubiquitin conjugate. The synthetic strategy using this building block provides a practical approach for the preparation of ubiquitinated peptides with diverse modifications.

Applications of artificial intelligence and bioinformatics methodologies in the analysis of ocular biofluid markers: a scoping review.

PURPOSE: This scoping review summarizes the applications of artificial intelligence (AI) and bioinformatics methodologies in analysis of ocular biofluid markers. The secondary objective was to explore supervised and unsupervised AI techniques and their predictive accuracies. We also evaluate the integration of bioinformatics with AI tools. METHODS: This scoping review was conducted across five electronic databases including EMBASE, Medline, Cochrane Central Register of Controlled Trials, Cochrane Database of Systematic Reviews, and Web of Science from inception to July 14, 2021. Studies pertaining to biofluid marker analysis using AI or bioinformatics were included. RESULTS: A total of 10,262 articles were retrieved from all databases and 177 studies met the inclusion criteria. The most commonly studied ocular diseases were diabetic eye diseases, with 50 papers (28%), while glaucoma was explored in 25 studies (14%), age-related macular degeneration in 20 (11%), dry eye disease in 10 (6%), and uveitis in 9 (5%). Supervised learning was used in 91 papers (51%), unsupervised AI in 83 (46%), and bioinformatics in 85 (48%). Ninety-eight papers (55%) used more than one class of AI (e.g. > 1 of supervised, unsupervised, bioinformatics, or statistical techniques), while 79 (45%) used only one. Supervised learning techniques were often used to predict disease status or prognosis, and demonstrated strong accuracy. Unsupervised AI algorithms were used to bolster the accuracy of other algorithms, identify molecularly distinct subgroups, or cluster cases into distinct subgroups that are useful for prediction of the disease course. Finally, bioinformatic tools were used to translate complex biomarker profiles or findings into interpretable data. CONCLUSION: AI analysis of biofluid markers displayed diagnostic accuracy, provided insight into mechanisms of molecular etiologies, and had the ability to provide individualized targeted therapeutic treatment for patients. Given the progression of AI towards use in both research and the clinic, ophthalmologists should be broadly aware of the commonly used algorithms and their applications. Future research may be aimed at validating algorithms and integrating them in clinical practice.

Unlocking Continuous Improvement in Heart Failure Remote Monitoring: A Participatory Approach to Unveil Value Dimensions and Performance Indicators.

Introduction: Heart failure (HF) constitutes a public health concern affecting quality of life, survival, and costs. Remote patient monitoring (RPM) can enhance HF management, involving patients actively and improving follow-up. While current HF RPM assessments emphasise cost-effectiveness analysis, there is a need to consider wider RPM impacts and integrate stakeholders' perspectives into assessments for better comprehensiveness. Methods: We developed a four-stage participatory approach to select value dimensions and indicators for continuous HF RPM assessment: Stage 1 involved building a literature-informed initial list; Stage 2 utilized expert interviews for validation and list expansion; Stage 3 involved a web-Delphi process with Portuguese stakeholders and experts for agreement assessment; and Stage 4 included a conclusive expert interview. Results: A literature review identified fourteen studies on telehealth, RPM, and HF, informing an initial list of four value dimensions (Access, Clinical aspects, Acceptability, and Costs) and 22 indicators. Seven semistructured interviews validated and further adjusted the list to 38 indicators. Subsequently, the web-Delphi process engaged 29 stakeholders, giving their opinions regarding assessment aspects' relevance and proposing additional elements - 1 dimension and 12 indicators. Five value dimensions and 38 indicators (76.0%) reached group agreement for selection, while 12 did not reach an agreement. Upon expert appreciation, 5 dimensions, 43 indicators, and 6 case-mix parameters were considered relevant. Discussion: This comprehensive social approach captured diverse stakeholder perspectives, achieving agreement on pertinent HF RPM monitoring and evaluation indicators. Findings can inform visualization and management tool development, aiding day-to-day RPM evaluation and identification of improvement opportunities.

[Implications of cyberbullying in major depressive symptoms development in young people between 15 and 29 years of age in Chile]. / Implicancias del ciberacoso en el desarrollo de sintomatología depresiva mayor en jóvenes entre 15 y 29 años en Chile.

The use of digital means of communication and socialization among young people brings with it new stressors, risks, and forms of ag- gression, such as cyberbullying. AIM: To evaluate the relationship between cyberbullying and major depressive symptoms in young Chileans between 15 and 29 years. METHODS: Survey through the StatKnows platform. Sampling was probabilistic and stratified with allocation proportional to the size of biphasic selection. The sample and the data were accessed virtually, respecting the bioethical standards of research with humans. RESULTS: At all ages studied, the probability of experiencing depressive symptoms increases as levels of cyber aggression do, especially in the group between 19 and 24 years. CONCLUSIONS: The results reflect various factors of emerging adulthood that make this group more vulnerable, emphasizing the importance of studying the phenomenon at non-school ages since this form of aggression is not exclusive to adolescence, and neither are its effects on mental health.

Delayed sleep phase disorder during the COVID-19 pandemic and its health implications.

Circadian rhythm sleep disorders are alterations that are characterized by a shift in the sleep-wake cycle relative to day and night, such as the delayed sleep phase disorder (DSPD), which is a retard of at least 2 hours in the sleep start. Typically, the patient falls asleep after 2 a.m. and wakes up after 10 a.m. and with symptom of sleep onset insomnia. The prevalence of DSPD in young adults is 0.48%, increasing to 3.3% in adolescents. Interestingly, patients with COVID-19 infection report anxiety due to the intensive care unit lockdown and constant exposure to bright light. In addition, post-COVID patients have an increased risk of developing DSPD. For example, in adolescent post-COVID patients, the prevalence of DSPD increases to 63.3%. Patients with DSPD also have alterations in metabolic health, poor school performance, cognitive impairment, and a higher risk of developing other diseases. The objective of the present review is therefore to describe the characteristics of DSPD during the COVID-19 pandemic and to outline its possible implications for physical health (eg, metabolism) and mental health (eg, anxiety or depression).

Emerging applications of bioinformatics and artificial intelligence in the analysis of biofluid markers involved in retinal occlusive diseases: a systematic review.

PURPOSE: To review the literature on the application of bioinformatics and artificial intelligence (AI) for analysis of biofluid biomarkers in retinal vein occlusion (RVO) and their potential utility in clinical decision-making. METHODS: We systematically searched MEDLINE, Embase, Cochrane, and Web of Science databases for articles reporting on AI or bioinformatics in RVO involving biofluids from inception to August 2021. Simple AI was categorized as logistics regressions of any type. Risk of bias was assessed using the Joanna Briggs Institute Critical Appraisal Tools. RESULTS: Among 10,264 studies screened, 14 eligible articles, encompassing 578 RVO patients, met the inclusion criteria. The use and reporting of AI and bioinformatics was heterogenous. Four articles performed proteomic analyses, two of which integrated AI tools such as discriminant analysis, probabilistic clustering, and string pathway analysis. A metabolomic study used AI tools for clustering, classification, and predictive modeling such as orthogonal partial least squares discriminant analysis. However, most studies used simple AI (n = 9). Vitreous humor sample levels of interleukin-6 (IL-6), vascular endothelial growth factor (VEGF), and aqueous humor levels of intercellular adhesion molecule-1 and IL-8 were implicated in the pathogenesis of branch RVO with macular edema. IL-6 and VEGF may predict visual acuity after intravitreal injections or vitrectomy, respectively. Metabolomics and Kyoto Encyclopedia of Genes and Genomes enrichment analysis identified the metabolic signature of central RVO to be related to lower aqueous humor concentration of carbohydrates and amino acids. Risk of bias was low or moderate for included studies. CONCLUSION: Bioinformatics has applications for analysis of proteomics and metabolomics present in biofluids in RVO with AI for clinical decision-making and advancing the future of RVO precision medicine. However, multiple limitations such as simple AI use, small sample volume, inconsistent feasibility of office-based sampling, lack of longitudinal follow-up, lack of sampling before and after RVO, and lack of healthy controls must be addressed in future studies.

Establishing the health and wellbeing needs of mining host community in Brazil, Chile and Peru: a mixed-method approach to identify priority areas for action to help communities meet their SDG3 targets.

BACKGROUND: The global mining industry is an important partner in advancing the 2030 Agenda for Sustainable Development. In 2018, Anglo American plc published their Sustainable Mining Plan, containing a goal for improving health and wellbeing aligned with the Sustainable Development Goal 3 (SDG3) targets. Having formed an independent multidisciplinary research consortium, we designed and implemented a mixed-methods approach to attain a deeper understanding of SDG3 priorities within the local context of communities hosting Anglo American mining operations located in Latin America. METHODS: In 2019, within the host communities of three mining operations in Chile, three in Brazil, and one in Peru, we conducted a qualitative study which included stakeholder workshops and key informant interviews. We also quantitatively appraised existing health data. Findings emerging from the qualitative and quantitative assessments were compared to identify health and wellbeing priority areas for action relevant to each community. RESULTS: Across the three countries, 120 people took part in workshops and 35 in interviews. In these workshops and interviews, non-communicable diseases (SDG3.4), harmful alcohol consumption (SDG3.5), and pollution, particularly air pollution (SDG3.9), were consistently identified as areas for priority action. There were similarities in the reporting of individual, interpersonal, community, societal, and structural factors underlying these priority areas across the different communities. The availability of quantitative data was generally good at the state level, becoming increasing sparse as we focused on smaller geographies. The priorities identified in the quantitative assessments generally aligned with those highlighted in the qualitative data. CONCLUSIONS: We highlight the importance of engaging with local populations to understand and address health needs. To address the priorities identified, intervention packages tailored to the specific needs of host communities, that tackle associated upstream societal level factors, are required. To facilitate this, appropriate monitoring systems and epidemiological investigations should be implemented to better understand the local context and quantify health issues. In the host communities, it is essential for the mining sector to be a key health partner in promoting integrated programmes that contribute to achieving the priority objectives and targets aligned with the SDG3 agenda.

[Evidence map on post-acute COVID-19 sequelae and rehabilitation: Update as of July 2022Mapa de evidencia sobre las secuelas y la rehabilitación tras la COVID-19 aguda: versión actualizada en julio del 2022]. / Mapa de evidências sobre sequelas e reablitação da covid-19 pós-aguda: uma versão atualizada em julho de 2022.

Objective: To update the evidence map on the effects of interventions for post-acute COVID-19 rehabilitation. Method: The search scope was defined according to the population (patients with symptomatic COVID-19 and post-acute COVID sequelae), the context (interventions for rehabilitation), and the type of study (systematic reviews, rapid reviews, scoping reviews or overviews of reviews). Following a search in PubMed and the Virtual Health Library, two independent authors selected the articles for review. The map was updated on July 27, 2022, using the same procedures employed in the initial review. Results: The initial evidence map included 22 studies (four systematic reviews, four rapid reviews, four reviews of case reports, one scoping review, and nine systematic review protocols). In the present update, an additional 10 studies were included. The analysis revealed four groups of interventions (multimodal, therapeutic, complementary, and pharmacological) and seven groups of outcomes (pathological conditions, diseases/respiratory disorders, pain, physiological and metabolic markers, mental health/quality of life, sensory function, and mortality), totaling 166 associations between interventions and outcomes. The highest number of associations was observed for complementary therapies (n = 94). Among the outcomes, the highest number of associations was observed for physiological and metabolic markers, pathological conditions, and mental health/quality of life (44, 41, and 35 associations respectively). Conclusions: The map update involved the analysis of 69 associations, most notably exercise (isolated, multicomponent, or multimodal intervention), with 23 positive and four potentially positive effects) and pharmacologic and complementary therapies for sensorial functions (15 associations). The high number of systematic review protocols indicates that the literature is still incipient.

Objetivo: Actualizar el mapa de evidencia de los efectos de las intervenciones de rehabilitación tras la COVID-19 aguda. Métodos: El alcance de la búsqueda se definió en función de la población (pacientes que habían tenido COVID-19 sintomática y secuelas tras un cuadro agudo de la enfermedad), el contexto (intervenciones de recuperación de las secuelas) y el tipo de estudio (revisión sistemática, revisión sistemática rápida, revisión del alcance o revisión de revisiones). Después de realizar búsquedas en PubMed y en la Biblioteca Virtual de Salud, dos autores independientes seleccionaron los estudios de revisión. En la actualización del mapa realizada el 27 de julio del 2022 se siguieron los mismos procedimientos descritos anteriormente. Resultados: El mapa de evidencia inicial contenía 22 estudios (cuatro revisiones sistemáticas, cuatro revisiones rápidas, cuatro revisiones de estudios de casos, una revisión del alcance y nueve protocolos de revisión sistemática). En esta actualización se incluyeron otros 10 estudios. Se encontraron cuatro grupos de intervenciones (multimodales y terapéuticas, y tratamientos complementarios y farmacológicos) y siete grupos de resultados (afecciones patológicas, enfermedades y trastornos respiratorios, dolor, indicadores fisiológicos y metabólicos, salud mental/calidad de vida, funciones sensoriales y mortalidad), con un total de 166 asociaciones entre las intervenciones y los resultados. Los tratamientos complementarios presentaron más asociaciones con los resultados (n = 94). Entre los resultados, se destacaron los indicadores fisiológicos y metabólicos, las afecciones patológicas y la salud mental/calidad de vida (44, 41 y 35 asociaciones, respectivamente). Conclusiones: En la actualización del mapa, se analizaron 69 asociaciones entre las cuales se destacan el ejercicio (aislado o compuesto o una intervención multimodal, con 23 efectos positivos y cuatro potencialmente positivos) y las intervenciones farmacológicas y los tratamientos complementarios para las funciones sensoriales (15 asociaciones). El elevado número de protocolos indica que la bibliografía sigue siendo incipiente.

Parasomnias in patients with addictions-a systematic review.

Parasomnias are involuntary behaviors or subjective experiences during sleep. Our objective was to review existing information on the presence of parasomnias in patients with addictions or during treatment for addictions. Information about parasomnias related to rapid-eye-movement (REM) and non-REM sleep in patients with addictions, while using substances or in abstinence, was reviewed. A systematic search of published articles reporting parasomnias as a consequence of drug use or abuse was conducted in the PubMed and SciELO databases. The search for the studies was performed in three phases: (1) by title, (2) by abstract, and (3) by complete text. The search was performed independently by two researchers, who then compared their results from each screening phase. Seventeen articles were found. The consumption of alcohol was reported in association with arousal disorders, such as sexsomnia and sleep-related eating disorder; and REM sleep behavior disorder was reported during alcohol withdrawal. Cocaine abuse was associated with REM sleep behavior disorder with drug consumption dream content. Overall, we found that several types of parasomnias were very frequent in patients with addictions. To avoid accidents in bedroom, legal problems, and improve evolution and prognosis; must be mandatory to include security measures related to sleep period; avoid pharmacological therapy described as potential trigger factor; improve sleep hygiene; and give pharmacological and behavioral treatments for patients with these comorbid sleep disorders.

Effects of medicinal plants and natural compounds in models of prostate cancer related to sex steroids: A systematic review.

Prostate cancer remains a health problem for men. Targeting androgen (AR) and estrogen (ER) receptors improves the outcomes of the disease, and many medicinal plants exert their effects by modulating these pathways. Therefore, a systematic review was conducted to identify medicinal plants and their natural compounds that may modulate the AR and/or ER pathways in cell and animal models. A search was conducted across EMBASE, LILACS, PubMed, Scopus, and Web of Science, with grey literature from Google SCHOLAR and ProQuest. Two authors independently selected eligible studies based on their titles and abstracts, and a third author resolved conflicts. Then, data from the full text of eligible studies were extracted and synthesized. In total, 75 studies were included. Results showed the effects of several different medicinal plants and natural compounds in reduction of AR and/or ER transcription and translation and AR secondary effects: cell growth reduction, induction of apoptosis, and cell cycle arrest. In animal models, tumor size reduction, increase in apoptosis, and downregulation of AR expression in tumors were also observed. No single phytochemical group concentrating molecules with anti-AR and/or ER activity was identified. Nevertheless, several phytochemical compounds showed potential for future clinical studies in the management of the disease.

Chemical analysis of irradiated root dentin and its interaction with resin cements.

OBJECTIVES: To investigate the chemical changes in root dentin submitted to ionizing radiation and how it affects the interaction with resin cements. MATERIALS AND METHODS: Forty human premolars were randomly divided into two groups (n = 20): non-irradiated and irradiated. They were randomly subdivided according to the type of resin cement (n = 10): conventional (RelyX ARC, 3 M ESPE) or self-adhesive (RelyX U200, 3 M ESPE). After cementation of the fiberglass posts, the roots were sectioned to be analyzed by Fourier transform infrared spectroscopy (FTIR), Raman spectroscopy, and confocal laser scanning microscopy (CLSM). The data obtained from FTIR and Raman were analyzed using two-way ANOVA followed Tukey's test (α = 0.05). For CLSM, a descriptive analysis was performed. RESULTS: In the FTIR, there was a significant difference between the non-irradiated and irradiated groups for phosphate (p = 0.011), carbonate (p < 0.001), amide III (p = 0.038), and carbonate/mineral ratio (p < 0.001). Regarding the root third, there was a difference for amide I (p = 0.002), mineral/matrix ratio (p = 0.001), and amide I/CH2 (p = 0.026) between the cervical and the others. Raman spectroscopy revealed no difference between groups for 961/1458 cm-1 in the diffusion zone. CLSM showed a different interaction pattern for the two cements with the irradiated dentin from the cervical third. CONCLUSIONS: Ionizing radiation altered the chemical composition of root dentin, especially in the cervical third. The resin cements showed less interaction with the irradiated root dentin. CLINICAL RELEVANCE: As radiotherapy alters the chemical composition of root dentin, the interaction of resin cement with dentin can compromise the success of rehabilitation with fiberglass posts.

Development of Neurogenic Detrusor Overactivity after Thoracic Spinal Cord Injury Is Accompanied by Time-Dependent Changes in Lumbosacral Expression of Axonal Growth Regulators.

Thoracic spinal cord injury (SCI) results in urinary dysfunction, which majorly affects the quality of life of SCI patients. Abnormal sprouting of lumbosacral bladder afferents plays a crucial role in this condition. Underlying mechanisms may include changes in expression of regulators of axonal growth, including chondroitin sulphate proteoglycans (CSPGs), myelin-associated inhibitors (MAIs) and repulsive guidance molecules, known to be upregulated at the injury site post SCI. Here, we confirmed lumbosacral upregulation of the growth-associated protein GAP43 in SCI animals with bladder dysfunction, indicating the occurrence of axonal sprouting. Neurocan and Phosphacan (CSPGs), as well as Nogo-A (MAI), at the same spinal segments were upregulated 7 days post injury (dpi) but returned to baseline values 28 dpi. In turn, qPCR analysis of the mRNA levels for receptors of those repulsive molecules in dorsal root ganglia (DRG) neurons showed a time-dependent decrease in receptor expression. In vitro assays with DRG neurons from SCI rats demonstrated that exposure to high levels of NGF downregulated the expression of some, but not all, receptors for those regulators of axonal growth. The present results, therefore, show significant molecular changes at the lumbosacral cord and DRGs after thoracic lesion, likely critically involved in neuroplastic events leading to urinary impairment.

Global trends in research output by zoos and aquariums.

Zoos and aquaria, often regarded as preservation-cum-entertainment enterprises, are also actors in the effort to curb the biodiversity crisis: raising awareness, supporting conservation, and conducting research. We assessed trends in zoo and aquaria research topics and study organisms over time worldwide. For the zoos and aquaria registered in the Species360's Zoological Information Management System and the World Association of Zoos and Aquariums, we compiled metadata on their research published in the peer-reviewed literature indexed in Scopus and carried out a keyword frequency analysis. The production of scientific papers by zoos increased at a much faster rate than the average accrual of scientific papers in the literature. Evolution of research themes ran parallel to that of biological sciences (e.g., development of molecular genetics or increased awareness about conservation). The focus of 48.5% of zoo-led research was on vertebrates, of which mammal research was 33.7%. Whether zoos are effectively contributing to conservation may still be debatable, but our results highlight their institutional efforts to increase knowledge about the species in their care.

Tendencias Mundiales en la Producción Investigativa de los Zoológicos y Acuarios Resumen Los zoológicos y los acuarios, con frecuencia considerados empresas de conservación y entretenimiento, también son actores en el esfuerzo por reducir la crisis de la biodiversidad al crear conciencia, apoyar a la conservación y llevar a cabo investigaciones. Evaluamos las tendencias en los temas de investigación y los organismos de estudios en los zoológicos y los acuarios en todo el mundo a través del tiempo. Compilamos los metadatos de las investigaciones realizadas en los zoológicos y acuarios registrados en el Sistema de Manejo de Información Zoológica de Species360 y en la Asociación Mundial de Zoológicos y Acuarios que han sido publicadas en la literatura revisada por pares indexada en Scopus y realizamos un análisis de frecuencias de palabras clave. La producción de artículos científicos por zoológicos se incrementó a una tasa mucho más rápida que la acumulación promedio de artículos científicos en la literatura. La evolución de los temas de investigación fue paralela a la de las ciencias biológicas (p. ej.: desarrollo de la genética molecular o incremento en la conciencia por la conservación). El enfoque del 48.5% de las investigaciones conducidas por los zoológicos estuvo sobre los vertebrados. De este 48.5%, el 33.7% fueron investigaciones sobre mamíferos. Todavía puede debatirse si los zoológicos están contribuyendo efectivamente a la conservación, pero nuestros resultados resaltan sus esfuerzos institucionales por incrementar el conocimiento sobre las especies bajo su cuidado.

Correlates of Bullying Victimization and Sexual Harassment: Implications for Life Satisfaction in Late Adolescents.

Bullying and sexual harassment are considered widespread public health concerns because they may have negative effects on physical and mental health. However, more studies are necessary that relate these forms of victimization and their overlap with subjective well-being. This study explores the prevalence and association between different forms of bullying victimization, sexual harassment, and life satisfaction using a sample of 47,114 students aged 16-18 years and from 646 Peruvian educational institutions. Face-to-face bullying was the most reported type of victimization, followed by cyberbullying. There was a large degree of overlap between these two forms of bullying as well as between traditional bullying and sexual harassment. This overlap causes a decrease in life satisfaction in late adolescents. Thus, the need of preventing the negative dynamics of violence in order to prevent the overlapping of different violence forms in adolescence is discussed. Finally, implications for school nurses are outlined.

Health-Related Quality of Life in Neurological Disorders Most Commonly Associated With Zika-Virus Infection: A Systematic Review.

OBJECTIVES: In this systematic review, we synthesize the current evidence on health-related quality of life (HRQoL) for the two of the most relevant outcomes of Zika virus infection in humans, microcephaly and Guillain-Barré Syndrome (GBS). METHODS: We searched the following databases: MEDLINE, Embase, CINAHL, LILACS, WHO's ICTRP clinical trials registries database and PROSPERO. Search terms included quality of life, microcephaly, and Guillain-Barré Syndrome. We included primary studies where HRQoL was quantitatively assessed for microcephaly and GBS using validated instruments. We used the Joanna Briggs Institute Critical Appraisal Tools to assess the risk of bias of individual studies. RESULTS: From a total of 1,657 abstracts screened and 66 full texts reviewed, 21 studies met the eligibility criteria; one study for microcephaly and 20 for GBS. Adjusted disutilities for microcephaly compared to a normative childhood utility ranged from -0.745 to -0.820. For GBS, time traded-off the expected lifetime ranged from 16 days to 3 years. HRQoL follows the clinical course of GBS, with lower scores in the first months, recovery within the first year post onset, and stabilization after one year. CONCLUSIONS: Included studies reported a wide range of HRQoL for GBS, due in part to a high level of heterogeneity in methods, inclusion criteria, follow-up and reporting of results. Opportunities exist for primary studies assessing the longitudinal HRQoL over the entire course of the diseases to inform clinical practice, economic evaluations and health policy.

RNA-binding specificity landscapes of designer pentatricopeptide repeat proteins elucidate principles of PPR-RNA interactions.

Pentatricopeptide repeat (PPR) proteins are helical-repeat proteins that offer a promising scaffold for the engineering of proteins to bind specified RNAs. PPR tracts bind RNA in a modular 1-repeat, 1-nucleotide fashion. An amino acid code specifying the bound nucleotide has been elucidated. However, this code does not fully explain the sequence specificity of native PPR proteins. Furthermore, it does not address nuances such as the contribution toward binding affinity of various repeat-nucleotide pairs or the impact of mismatches between a repeat and aligning nucleotide. We used an in vitro bind-n-seq approach to describe the population of sequences bound by four artificial PPR proteins built from consensus scaffolds. The specificity of these proteins can be accounted for by canonical code-based nucleotide recognition. The results show, however, that interactions near the 3'-end of binding sites make less contribution to binding affinity than do those near the 5'-end, that proteins with 11 and 14 repeats exhibit similar affinity for their intended targets but 14-repeats are more permissive for mismatches, and that purine-binding repeats are less tolerant of transversion mismatches than are pyrimidine-binding motifs. These findings have implications for mechanisms that establish PPR-RNA interactions and for optimizing PPR design to minimize off-target interactions.

Unexpected functional versatility of the pentatricopeptide repeat proteins PGR3, PPR5 and PPR10.

Pentatricopeptide repeat (PPR) proteins are a large family of helical repeat proteins that bind RNA in mitochondria and chloroplasts. Sites of PPR action have been inferred primarily from genetic data, which have led to the view that most PPR proteins act at a very small number of sites in vivo. Here, we report new functions for three chloroplast PPR proteins that had already been studied in depth. Maize PPR5, previously shown to promote trnG splicing, is also required for rpl16 splicing. Maize PPR10, previously shown to bind the atpI-atpH and psaJ-rpl33 intercistronic regions, also stabilizes a 3'-end downstream from psaI. Arabidopsis PGR3, shown previously to bind upstream of petL, also binds the rpl14-rps8 intercistronic region where it stabilizes a 3'-end and stimulates rps8 translation. These functions of PGR3 are conserved in maize. The discovery of new functions for three proteins that were already among the best characterized members of the PPR family implies that functional repertoires of PPR proteins are more complex than have been appreciated. The diversity of sequences bound by PPR10 and PGR3 in vivo highlights challenges of predicting binding sites of native PPR proteins based on the amino acid code for nucleotide recognition by PPR motifs.

Ceftolozane/tazobactam and ceftazidime/avibactam antimicrobial activity against clinically relevant gram-negative bacilli isolated in Mexico.

INTRODUCTION: There is limited information on the effectiveness of ceftolozane/tazobactam and ceftazidime/avibactam combinations on clinically relevant strains isolated in Mexico. OBJECTIVE: To determine the antimicrobial profile of both antibiotic combinations in our community. METHOD: The present research study was prospective, descriptive and cross-sectional. Clinically relevant strains isolated from pure-strain cultures were included during the period from August 2018 to January 2019 in Mexicali, Baja California, Mexico. RESULTS: 74 enterobacteriaceae and 19 Pseudomonas aeruginosa strains were analyzed; the percentage of sensitivity of ceftazidime/avibactam was 100 % for enterobacteriaceae and 72.7 % for Pseudomonas aeruginosa; the percentage of sensitivity of ceftolozane/tazobactam for enterobacteriaceae was 90.5 % and 72.7 % for Pseudomonas aeruginosa. CONCLUSIONS: The ceftolozane/tazobactam and ceftazidime/avibactam combinations offer good antimicrobial sensitivity in vitro, both for ESBL-producing enterobacteriaceae and Pseudomonas aeruginosa. More data are required to assess clinical response in patients receiving these antibiotic combinations. INTRODUCCIÓN: Existe poca información acerca de la efectividad de las combinaciones ceftolozano/tazobactam y ceftazidima/avibactam en cepas clínicamente relevantes aisladas en México. OBJETIVO: Determinar el perfil antimicrobiano de ambos antibióticos en nuestra comunidad. MÉTODO: El presente estudio de investigación fue prospectivo, descriptivo y transversal. Se incluyeron cepas clínicamente relevantes aisladas a partir de cultivos de cepa pura durante el periodo de agosto de 2018 a enero de 2019 en Mexicali, Baja California, México. RESULTADOS: Se analizaron 74 cepas de enterobacterias y 19 cepas de Pseudomonas aeruginosa; el porcentaje de sensibilidad de ceftazidima/avibactam fue de 100 % contra enterobacterias y de 72.7 % contra Pseudomonas aeruginosa; el porcentaje de sensibilidad de ceftolozano/tazobactam fue de 90.5 % para enterobacterias y de 72.7 % para Pseudomonas aeruginosa. CONCLUSIONES: Las combinaciones ceftolozano/tazobactam y ceftazidima/avibactam ofrecen buena sensibilidad antimicrobiana in vitro, tanto contra enterobacterias productoras de betalactamasas de espectro extendido como contra Pseudomonas aeruginosa. Se requieren más datos para valorar la respuesta clínica en pacientes que reciben esas combinaciones de antibióticos.

Transcriptome analysis of acerola fruit ripening: insights into ascorbate, ethylene, respiration, and softening metabolisms.

KEY MESSAGE: The first transcriptome coupled to metabolite analyses reveals major trends during acerola fruit ripening and shed lights on ascorbate, ethylene signalling, cellular respiration, sugar accumulation, and softening key regulatory genes. Acerola is a fast growing and ripening fruit that exhibits high amounts of ascorbate. During ripening, the fruit experience high respiratory rates leading to ascorbate depletion and a quickly fragile and perishable state. Despite its growing economic importance, understanding of its developmental metabolism remains obscure due to the absence of genomic and transcriptomic data. We performed an acerola transcriptome sequencing that generated over 600 million reads, 40,830 contigs, and provided the annotation of 25,298 unique transcripts. Overall, this study revealed the main metabolic changes that occur in the acerola ripening. This transcriptional profile linked to metabolite measurements, allowed us to focus on ascorbate, ethylene, respiration, sugar, and firmness, the major metabolism indicators for acerola quality. Our results suggest a cooperative role of several genes involved in AsA biosynthesis (PMM, GMP1 and 3, GME1 and 2, GGP1 and 2), translocation (NAT3, 4, 6 and 6-like) and recycling (MDHAR2 and DHAR1) pathways for AsA accumulation in unripe fruits. Moreover, the association of metabolites with transcript profiles provided a comprehensive understanding of ethylene signalling, respiration, sugar accumulation and softening of acerola, shedding light on promising key regulatory genes. Overall, this study provides a foundation for further examination of the functional significance of these genes to improve fruit quality traits.

RNA-binding specificity landscape of the pentatricopeptide repeat protein PPR10.

Pentatricopeptide repeat (PPR) proteins comprise a large family of helical repeat proteins that influence gene expression in mitochondria and chloroplasts. PPR tracts can bind RNA via a modular one repeat-one nucleotide mechanism in which the nucleotide is specified by the identities of several amino acids in each repeat. This mode of recognition, the so-called PPR code, offers opportunities for the prediction of native PPR binding sites and the design of proteins to bind specified RNAs. However, a deep understanding of the parameters that dictate the affinity and specificity of PPR-RNA interactions is necessary to realize these goals. We report a comprehensive analysis of the sequence specificity of PPR10, a protein that binds similar RNA sequences of ∼18 nucleotides (nt) near the chloroplast atpH and psaJ genes in maize. We assessed the contribution of each nucleotide in the atpH binding site to PPR10 affinity in vitro by analyzing the effects of single-nucleotide changes at each position. In a complementary approach, the RNAs bound by PPR10 from partially randomized RNA pools were analyzed by deep sequencing. The results revealed three patches in which nucleotide identity has a major impact on binding affinity. These include 5 nt for which protein contacts were not observed in a PPR10-RNA crystal structure and 4 nt that are not explained by current views of the PPR code. These findings highlight aspects of PPR-RNA interactions that pose challenges for binding site prediction and design.