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OBJECTIVE: Neonatal abstinence syndrome (NAS) is a set of drug withdrawal symptoms suffered by neonates exposed to drugs in utero. Several studies have widely described NAS incidence and treatment approach; however, little is known regarding the incidence and manifestations of this disease in Puerto Rico (PR). The principal aim of this study was to describe NAS incidence in the neonatal units of hospitals affiliated with the University of PR in terms of occurrence, clinical manifestations, and treatment approaches. METHODS: Our study evaluated the medical records of NAS babies diagnosed from 2018 through 2020 at 2 hospitals affiliated with the University of PR Medical Sciences Campus. Descriptive and inferential statistics were employed to analyze trends. RESULTS: We identified 12 neonates diagnosed with NAS, 5 with low birthweights (<2500 g); for a NAS incidence of 2 cases per 1000 admitted for the 3 years of recollected data. The urine toxicology results revealed that 9 had experienced intrauterine polydrug exposure. Phenobarbital loading dose were determined on the day of diagnosis (indicated by Finnegan score). The first manifestation of NAS symptoms varied: 8 neonates showed symptoms within 48 hours after birth, whereas 4 had withdrawal symptoms within 72-120 hours of their births. Differences between dosing practices and guidelines were observed, ranging from a 0.69% to a 25% difference during treatment initiation. CONCLUSION: Further research on the incidence of NAS in PR (national level) is needed for a deeper understanding that we hope will lead to the development of enhanced treatment protocols in PR.

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Supplementation with natural additives such as essential oils (EO) or probiotics has resulted in comparable growth performance to that of supplemental monensin in fattening lambs in hot environments. Supra-supplementation levels of vitamin D3 improved the carcass weight and dressing percentage of steers fattened under tropical conditions. We hypothesized that certain combinations of these natural additives could be complementary. For this reason, a feeding trial was carried out using 48 Pelibuey × Katahdin non-castrated male lambs (107 ± 14 d age; 17.9 ± 2.51 kg LW). Lambs were fed an 88:12 concentrate to forage ratio basal diet supplemented (dry matter basis, DMI) with: (1) no additive (CON); (2) 28 mg monensin/kg diet (MON); (3) 150 mg of essential oils containing a combination of thymol, eugenol, vanillin, guaiac, and limonene plus 0.12 mg vitamin D3 (EO + D3)/kg diet; and (4) 300 mg of essential oils containing a combination of carvacrol and cynamaldehyde plus 2 g probiotic (2.2 × 108 CFU of bacillus subtilis/kg diet, EO + BS). Lambs were grouped by initial weight and assigned within six weight groupings to 24 pens (2 lambs/pen, 6 replicas per treatment) in a randomized complete block design. The experiment lasted 121 days. Daily maximal THI exceeded the 80 "danger or "emergency" range for 119 days of the 121 days of the trial. Lambs supplemented with MON had similar DMI, growth performance, and dietary energetics to those of CON lambs. Lambs supplemented with EO + BS had a greater (9.2%, p ≤ 0.05) average daily gain (ADG) than the CON and MON groups due to enhanced (10.2%, p ≤ 0.05) dry matter intake. Thus, gain efficiency (GF) and estimated dietary energy were similar for CON, MON, and EO + BS. Lambs receiving EO + D3 had similar (0.254 vs. 0.262 kg/d) ADG but a lower DMI (8%, p < 0.05) compared with EO + BS lambs. Consequently, GF and estimated dietary net energy were greater (4.9 and 3.7%, respectively; p ≤ 0.05) for EO + D3 lambs. Even when ambient heat load was elevated, the efficiency of utilization of dietary energy (observed-to-expected dietary net energy) was close to 1.00 (0.992) expected for EO + D3 lambs. In contrast, efficiency of energy utilization was depressed by -4.4% for lambs on the other treatments. Compared with the other treatments, lambs receiving EO + D3 had greater longissimus muscle area (5.6%, p < 0.05) and lower kidney pelvic fat (21.8%, p ≤ 0.05). There were no treatment effects on shoulder tissue composition or whole cuts (expressed as % of cold carcass weight). Compared to CON, lambs that were fed with natural additives showed 3.5% lower (p ≤ 0.05) intestine mass. All supplemental additives decreased visceral fat mass, which was minimal with EO + D3 treatment. Combinations of essential oils with vitamins or probiotics were superior to antibiotic monensin in finishing diets for feedlot lambs. Combining EO with probiotics promoted DM intake and gain but not gain efficiency, while combining EO with vitamin D3 supra-supplementation increased dietary energy efficiency and improved some carcass characteristics in lambs fattening under high ambient heat loads.

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Introducción. La anestesia regional es una técnica importante, innovadora y popular para el manejo anestésico y analgésico. En el bloqueo supraclavicular del plexo braquial existe incidencia 50-60% de parálisis diafragmática. La presentación clínica es variable de acuerdo con factores externos e internos del paciente. Existen múltiples técnicas radiológicas en el diagnóstico, siendo de gran utilidad la ecografía por su fácil acceso. El objetivo es determinar la incidencia de parálisis diafragmática secundaria a bloqueo supraclavicular del plexo braquial guiado por ecografía en una institución de cuarto nivel. Metodología. Estudio analítico, longitudinal, prospectivo; se seleccionaron 110 pacientes. Criterios de inclusión: pacientes mayores de 18 años sometidos a cirugía de miembro superior con bloqueo supraclavicular de plexo braquial. Criterios exclusión: pacientes ASA (American Society of anesthesiologist) 4 y 5, gestantes, IMC >35 kg/m2, pacientes con antecedentes neuromusculares, enfermedad pulmonar restrictiva u obstructiva, parálisis nervio frénico o disfunción diafragmática. Se realizó un análisis mediante test estadísticos, describiendo los diferentes grados de parálisis diafragmática. Resultados. En los pacientes que cumplieron con los criterios de inclusión, la incidencia de parálisis diafragmática fue de 65% (37% parálisis total y 28% parcial), el 1.81% presentó síntomas respiratorios sin cambios hemodinámicos. Discusión. El bloqueo del plexo braquial por vía supraclavicular es una técnica relacionada con parálisis diafragmática, la ecografía ha permitido reducir la incidencia de esta complicación, es un método útil en el diagnóstico postoperatorio. Conclusiones. La parálisis diafragmática post bloqueo plexo braquial supraclavicular es una complicación observada principalmente en pacientes con previo compromiso pulmonar, por lo cual es de importancia la vigilancia estricta.

Introduction. Regional anesthesia is an important, innovative, and popular technique for anesthetic and painkiller management. In supraclavicular brachial plexus blockade, there is a 50-60% rate of diaphragmatic paralysis. Clinical presentation is variable according to the patient's internal and external factors. There are multiple radiological techniques in diagnosis, with ultrasounds being very useful due to their easy access. The objective is to determine the incidence of diaphragmatic paralysis secondary to supraclavicular brachial plexus blockade guided by ultrasound in a fourth level institution. Methodology. Prospective, longitudinal, analytical study. 110 patients were selected. Inclusion criteria: patients over 18 years of age subject to upper limb surgery with supraclavicular brachial plexus blockade. Exclusion criteria: ASA (American Society of Anesthesiologists) 4 and 5 patients, pregnant women BMI >35 kg/m2 patients with neuromuscular background, restrictive or obstructive pulmonary disease, phrenic nerve paralysis, or diaphragmatic dysfunction. An analysis was carried out via statistical tests, describing the different degrees of diaphragmatic paralysis. Results. In patients who met the inclusion criteria, the incidence of diaphragmatic paralysis was 65% (37% with total and 28% with partial paralysis), 1.81% showed respiratory symptoms without hemodynamic changes. Discussion. Supraclavicular brachial plexus blockade is a technique related to diaphragmatic paralysis. Ultrasound has allowed for the incidence of this complication to be reduced. It is a useful method in post-operative diagnosis. Conclusions. Diaphragmatic paralysis after supraclavicular brachial plexus blockade is a complication mainly observed in patients with previous pulmonary problems. Therefore, it must be strictly monitored.

Introdução. A anestesia regional é uma técnica importante, inovadora e popular para o manejo anestésico e analgésico. No bloqueio supraclavicular do plexo braquial há uma incidência de 50-60% de paralisia diafragmática. A apresentação clínica é variável de acordo com fatores externos e internos do paciente. Existem múltiplas técnicas radiológicas no diagnóstico, sendo a ultrassonografia muito útil devido ao seu fácil acesso. O objetivo é determinar a incidência de paralisia diafragmática secundária ao bloqueio supraclavicular do plexo braquial guiado por ultrassom em uma instituição de quarto nível. Metodologia. Estudo analítico, longitudinal, prospectivo; 110 pacientes foram selecionados. Critérios de inclusão: pacientes maiores de 18 anos submetidos à cirurgia de membro superior com bloqueio supraclavicular do plexo braquial. Critérios de exclusão: pacientes ASA (American Anesthesiology Society) 4 e 5, gestantes, IMC>35 kg/m2 pacientes com história neuromuscular, doença pulmonar restritiva ou obstrutiva, paralisia do nervo frênico ou disfunção diafragmática. Foi realizada uma análise por meio de testes estatísticos, descrevendo os diferentes graus de paralisia diafragmática. Resultados. Nos pacientes que atenderam aos critérios de inclusão, a incidência de paralisia diafragmática foi de 65% (37% paralisia total e 28% parcial), 1.81% apresentavam sintomas respiratórios sem alterações hemodinâmicas. Discussão. O bloqueio do plexo braquial supraclavicular é uma técnica relacionada à paralisia diafragmática, a ultrassonografia tem reduzido a incidência dessa complicação e é um método útil no diagnóstico pós-operatório. Conclusões. A paralisia diafragmática após bloqueio do plexo braquial supraclavicular é uma complicação observada principalmente em pacientes com envolvimento pulmonar prévio, pelo que é importante vigilância rigorosa.

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The danger of Sclerotium cepivorum lies in the strength of its survival structure: sclerotia. Sclerotia comprising hardened mycelium contains food reserves that allow it to remain dormant for long period, which makes the sclerotia-infested soil useless to grow any crop of the Allium species, including onion and garlic. This paper would be the first report on the application of two-photon fluorescence microscopy to the analysis of the structure of sclerotia from S. cepivorum. For this study and, in order to test the method, two different types of sclerotia were used: (1) sclerotia isolated from naturally infested soil and (2) sclerotia produced in vitro (from 20-day-old cultures). Both types of sclerotia were processed by cryopreservation and eight µm histological cuts were used to obtain an autofluorescence image. For both sclerotia, the fluorescence spectrum has three peak signals at their wall. Sclerotia from infested soil presented fluorescence peaks at 400-436, 436-475, and 515-575 nm, while signals from sclerotia produced in vitro presented fluorescence peaks at 400-442, 500-600, and 655-700 nm. Peaks at the violet electromagnetic region (400-436 and 400-442) are like that of the signals reported by the melanin. This study showed that two-photon microscopy is a novel and valuable tool for the study of sclerotia structure and their fluorescence signal, and the possibility of using it as a specific marker to direct detection in the field should be explored.

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Trehalose is an important disaccharide that can be found in bacteria, fungi, invertebrates and plants. In some Ascomycota fungal plant pathogens, the role of trehalose was recently studied and shown to be important for conferring protection against several environmental stresses and for virulence. In most of the fungi studied, two enzymes are involved in the synthesis of trehalose: trehalose-6-phosphate synthase (Tps1) and trehalose-6-phosphate phosphatase (Tps2). To study the role of trehalose in virulence and stress response in the Basidiomycota maize pathogen Ustilago maydis, Δtps2 deletion mutants were constructed. These mutants did not produce trehalose as confirmed by HPLC analysis, showing that the single gene disruption impaired its biosynthesis. The mutants displayed increased sensitivity to oxidative, heat, acid, ionic and osmotic stresses as compared to the wild-type strains. Virulence of Δtps2 mutants to maize plants was extremely reduced compared to wild-type strains, possibly due to reduced capability to deal with the hostile host environment. The phenotypic traits displayed by Δtps2 strains were fully restored to wild-type levels when complemented with the endogenous UmTPS2 gene, or a chimeric construct having the Saccharomyces cerevisiae TPS2 ORF. This report demonstrates the presence of a single biosynthetic pathway for trehalose, and its importance for virulence in this model Basidiomycota plant pathogen.

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The arils of Bixa orellana L. seeds contain carotenoid storage cells (CSCs). The main compounds in these cells include bixin and norbixin, which are important pigments in the food and pharmaceutical industries. Although many studies have been conducted on these chemical constituents, the cellular events that occur during the development of the carotenoid-accumulating cells in the arils and their relationship with the final carotenoid accumulation in the vacuoles remain unknown. In this study, the development of the CSCs in B. orellana arils was analyzed by light and transmission electron microscopy. Carotenoids formed in specialized cells, whose number and size increased during aril development. At various stages of development, the cytoplasm of the CSCs contained chromoplasts that held an extensive network of tubules and plastoglobules. Next to the chromoplasts, lipid droplets may fuse one another to form osmiophilic bodies. In addition, vesicles were observed next to the tonoplast. At the final stages of development, both the osmiophilic bodies and vesicles, which became quadrangular or rectangular, were stored in the vacuoles of the CSCs. This study reported for the first time the occurrence of different storage unit types within the vacuole of carotenoid storage cells.

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Objective. To determine the effect of pH, and exposure time over the inactivation of aflatoxin B1 (AFB1) during the tortilla making process as well as the degradative molecules generated. Materials and methods. Inactivation of AFB1 in maize-dough with alkaline pH and in alkaline methanolic solutions was determined by HPLC. Kinetics of time exposure of AFB1 in methanolic solution and the degradative products were analyzed by direct injection electrospray mass spectometry (DIESI-MS). Results. The alkaline pH of the maize-dough after nixtamalización between 10.2, and 30-40 minutes of resting at room temperature allows the 100% reduction of AFB1. DIESI-MS analysis of the extracts indicated the presence of two degradation molecules from AFB1. Conclusion. The alkaline pH of maize-dough and resting time are the principal factors involved in diminishing AFB1 levels in tortillas. A procedure to the tortilla making process is proposed, which allows the reduction of remnant AFB1, avoiding the accumulative effect over consumers.

Objetivo. Determinar el efecto del pH alcalino de la masa de maíz y el tiempo de exposición sobre la aflatoxina B1 (AFB1) durante la producción de tortillas e identificar los posibles productos de degradación mediante DIESI-MS. Material y métodos. La inactivación de la AFB1 a pH alcalino y diferentes tiempos de exposición en masa nixtamalizada y en soluciones metanólicas fueron determinadas por HPLC. La cinética de degradación de AFB1, y los productos de degradación en soluciones metanólicas se determinaron por DIESI-MS. Resultados. El pH alcalino de la masa y 30 a 40 minutos de reposo redujeron en 100% la AFB1 adicionada. Se identificaron dos moléculas de degradación. Conclusión. Los principales factores involucrados en la disminución de la AFB1 durante la producción de tortillas son la hidrólisis alcalina y el tiempo de reposo. Se propone un procedimiento para la producción de tortilla que reducirá la AFB1 residual evitando el efecto acumulativo en los consumidores.

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OBJECTIVE: To determine the effect of pH, and exposure time over the inactivation of aflatoxin B1 (AFB1) during the tortilla making process as well as the degradative molecules generated. MATERIALS AND METHODS: Inactivation of AFB1 in maize-dough with alkaline pH and in alkaline methanolic solutions was determined by HPLC. Kinetics of time exposure of AFB1 in methanolic solution and the degradative products were analyzed by direct injection electrospray mass spectometry (DIESI-MS). RESULTS: The alkaline pH of the maize-dough after nixtamalización between 10.2, and 30-40 minutes of resting at room temperature allows the 100% reduction of AFB1. DIESI-MS analysis of the extracts indicated the presence of two degradation molecules from AFB1. CONCLUSION: The alkaline pH of maize-dough and resting time are the principal factors involved in diminishing AFB1 levels in tortillas. A procedure to the tortilla making process is proposed, which allows the reduction of remnant AFB1, avoiding the accumulative effect over consumers.

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Polyamines are essential metabolites present in all living organisms, and this subject has attracted the attention of researchers worldwide interested in defining their mode of action in the variable cell functions in which they are involved, from growth to development and differentiation. Although the mechanism of polyamine synthesis is almost universal, different biological groups show interesting differences in this aspect that require to be further analyzed. For these studies, fungi represent interesting models because of their characteristics and facility of analysis. During the last decades fungi have contributed to the understanding of polyamine metabolism. The use of specific inhibitors and the isolation of mutants have allowed the manipulation of the pathway providing information on its regulation. During host-fungus interaction polyamine metabolism suffers striking changes in response to infection, which requires examination. Additionally the role of polyamine transporter is getting importance because of its role in polyamine regulation. In this paper we analyze the metabolism of polyamines in fungi, and the difference of this process with other biological groups. Of particular importance is the difference of polyamine biosynthesis between fungi and plants, which makes this process an attractive target for the control of phytopathogenic fungi.

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By means of an in silico analysis, we demonstrated that a previously described chimeric gene (Spe-Sdh) encoding spermidine synthase, a key enzyme involved in the synthesis of polyamines, and saccharopine dehydrogenase, an enzyme involved in lysine synthesis in fungi, were present exclusively in members of all Basidiomycota subphyla, but not in any other group of living organisms. We used this feature to design degenerated primers to amplify a specific fragment of the Spe-Sdh gene by PCR, as a tool to unequivocally identify Basidiomycota isolates. The specificity of this procedure was tested using different fungal species. As expected, positive results were obtained only with Basidiomycota species, whereas no amplification was achieved with species belonging to other fungal phyla.

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In previous communications the essential role of spermidine in Ustilago maydis was demonstrated by means of the disruption of the genes encoding ornithine decarboxylase (ODC) and spermidine synthase (SPE). However, the assignation of specific roles to each polyamine in different cellular functions was not possible because the spermidine added to satisfy the auxotrophic requirement of odc/spe double mutants is partly back converted into putrescine. In this study, we have approached this problem through the disruption of the gene-encoding polyamine oxidase (PAO), required for the conversion of spermidine into putrescine, and the construction of odc/pao double mutants that were unable to synthesize putrescine by either ornithine decarboxylation or retroconversion from spermidine. Phenotypic analysis of the mutants provided evidence that putrescine is only an intermediary in spermidine biosynthesis, and has no direct role in cell growth, dimorphic transition, or any other vital function of U. maydis. Nevertheless, our results show that putrescine may play a role in the protection of U. maydis against salt and osmotic stress, and possibly virulence. Evidence was also obtained that the retroconversion of spermidine into putrescine is not essential for U. maydis growth but may be important for its survival under natural conditions.

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To analyze the role of spermidine in cell growth and differentiation of Ustilago maydis, the gene encoding spermidine synthase (Spe) was isolated using PCR. We found that the enzyme is encoded by a chimeric bifunctional gene (Spe-Sdh) that also encodes saccharopine dehydrogenase (Sdh), an enzyme involved in lysine biosynthesis. The gene contains a 5' region encoding Spe, followed, without a termination signal or a second initiation codon, by a 3' region encoding Sdh, and directs the synthesis of a single transcript that hybridizes with 3' or 5' regions' probes of the gene. The gene could not be disrupted in a wild-type strain, but only in a mutant defective in the gene encoding ornithine decarboxylase (Odc). Single spe-sdh mutants were isolated after sexual recombination in planta with a compatible wild-type strain. Mutants were auxotrophic for lysine and spermidine, but not for putrescine, and contained putrescine and spermidine, but not spermine. Putrescine in double mutants is probably synthesized from spermidine by the concerted action of polyamine acetyl transferase and polyamine oxidase. spe-sdh mutants were sensitive to stress, unable to carry out the yeast-to-mycelium dimorphic transition, and showed attenuated virulence to maize. These phenotypic alterations were reverted by complementation with the wild-type gene.

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Background. Sevki's histochemical technique allows specific staining of catechomamine-containing cells, yet discrimiantion between adrenaline-(ADR-) cells and noradrenaline-(NOR-) cells is unrealiable, being based on hue differences. Methods. In this work, histochemical differentiation of ADR- and NOR-cells in rat adrenal medulla was carried out by introducing two modifications to Sevki's technique: 1) employment of aged Giemsa solution, and 2) addition of an alkaline differentiating step. Results. With these changes, ADR-cell stained brown, whereas NOR-cells were deep-green, resulting in a clear-cut differentiation. Conclusions. The modified technique permits to differentiate ADR- from NOR-cells in the adrenal medulla using only a bright field microscope without any sophisticated equipment. The present procedure is inexpensive and easy to carry out

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