Insulin reference intervals in Brazilian adolescents by direct and indirect approaches: validation of a data mining method from laboratory data

Abstract Objective To determine reference intervals (RI) for fasting blood insulin (FBI) in Brazilian adolescents, 12 to 17 years old, by direct and indirect approaches, and to validate indirectly determined RI. Methods Two databases were used for RI determination. Database 1 (DB1), used to obtain RI through a posteriori direct method, consisted of prospectively selected healthy individuals. Database 2 (DB2) was retrospectively mined from an outpatient laboratory information system (LIS) used for the indirect method (Bhattacharya method). Results From DB1, 29345 individuals were enrolled (57.65 % female) and seven age ranges and sex partitions were statistically determined according to mean FBI values: females: 12 and 13 years-old, 14 years-old, 15 years-old, 16 and 17 years-old; and males: 12, 13 and 14 years-old, 15 years-old, 16 and 17 years-old. From DB2, 5465 adolescents (67.5 % female) were selected and grouped according to DB1 partitions. The mean FBI level was significantly higher in DB2, on all groups. The RI upper limit (URL) determined by Bhattacharya method was slightly lower than the 90 % CI URL directly obtained on DB1, except for group female 12 and 13 years old. High agreement rates for diagnosing elevated FBI in all groups on DB1 validated indirect RI presented. Conclusion The present study demonstrates that Bhattacharya indirect method to determine FBI RI in adolescents can overcome some of the difficulties and challenges of the direct approach.

Referencias argentinas de perímetro cefálico desde el nacimiento hasta los 19 años / Argentine reference charts for head circumference from birth to 19 years of age

Introducción. El perímetro cefálico (PC) es un indicador del crecimiento cerebral y es necesario contar con referencias de crecimiento que permitan determinar variaciones normales o patológicas. Objetivos. Presentar las primeras referencias argentinas de perímetro cefálico entre el nacimiento y los 19 años, y compararlas con las referencias de Nellhaus, utilizadas en nuestro país hasta la actualidad. Población y métodos. Para la construcción de estas referencias, se combinaron datos de la Encuesta Nacional de Nutrición y Salud 2018 y estudios transversales realizados entre 2004 y 2007 en las provincias de Buenos Aires y La Pampa, que incluyeron 8326 niños, niñas y adolescentes sanos. Las curvas de crecimiento fueron ajustadas con el método LMS. Para evaluar la magnitud de las diferencias entre estas referencias y las de Nellhaus, a diferentes edades, se graficaron los centilos 2, 50 y 98. Resultados. El PC mostró un incremento de tamaño variable con la edad, de mayor magnitud en los primeros años de vida, y un ligero incremento en la pubertad. Los valores del centilo 98 de las referencias argentinas fueron mayores que los de Nellhaus en todas las edades. Los valores del centilo 2 de la referencia nacional fueron menores que los de Nellhaus durante los primeros 2 años de vida, similares entre los 3 y 7 años, y mayores a partir de esta edad. Conclusiones. Las curvas argentinas describen adecuadamente el patrón de crecimiento del PC. Las diferencias halladas con la referencia de Nellhaus pueden atribuirse a cambios seculares.

Introduction. Head circumference (HC) is an indicator of brain growth; growth charts are necessary to determine normal or pathological variations. Objectives. To present the first Argentine HC reference charts between birth and 19 years of age and to compare them with the Nellhaus charts, which have been used in our country to date. Population and methods. These references were developed based on combined data from the National Survey on Nutrition and Health of 2018 and cross-sectional studies conducted between 2004 and 2007 in the provinces of Buenos Aires and La Pampa, which included 8326 healthy children and adolescents. Growth curves were adjusted using the LMS method. To assess the differences between these reference charts and the Nellhaus charts, at different ages, the 2nd, 50th, and 98th percentiles were plotted. Results. HC showed a variable increase in size with age, which was greater in the first years of life, and a slight increase at puberty. The values for the 98th percentile of the Argentine reference charts were higher than those of the Nellhaus charts at all ages. The values for the 2 nd percentile of the national reference were lower than those of the Nellhaus charts during the first 2 years of life, similar between 3 and 7 years of age, and higher after this age. Conclusions. The Argentine curves adequately describe the growth pattern of HC. The differences found with the Nellhaus charts may be attributed to secular changes.

Differences in percentage predicted spirometric values obtained with thirteen reference equations applied to healthy children.

BACKGROUND: There is limited information on the discrepancy between the spirometric values calculated with diverse spirometric prediction equations in normal children. OBJECTIVE: To determine the differences in percentage predicted (PP) values calculated through thirteen prediction equations (PEs) in healthy Chilean children. METHODS: We compared the PP values for FVC, FEV1, FEF25-75% and FEV1/FVC obtained by applying thirteen PEs, using GLI-2012 as a gold standard reference equation and including race neutral GLI-2022 in the analysis; PP values are from a group of 208 healthy Chilean children aged 7 to 17 years. RESULTS: In boys, the range of differences for FVC between PEs compared to the reference group went from -5.47% to 20.82% and from -6.4% to 19.74% in girls. For FEV1, in girls, the range of differences went from -5.96% to 18.01% and from -5,04% to 20.67% in boys. Significant differences between GLI-2012 PPs and the other PEs were observed more frequently in girls than in boys (p < 0.001). CONCLUSIONS: There is a wide difference between the spirometric PPs values by different PEs, including race neutral GLI-2022. Our findings, especially given the potential interpretative and clinical implications, might suggest the need for conducting standardized, multicenter studies in Latin America to determine which PEs would better fit pediatric populations in this region.

Detección de macroenzimas después de la precipitación con polietilenglicol: búsqueda de la población de referencia en el caso particular de las aminotransferasas / Detection of macroenzymes for aminotransferases after polyethylene glycol precipitation: search for the reference population / Detecção de macroenzimas após a precipitação com polietilenoglicol: busca da população de referência no caso particular das aminotransferases

Resumen Las macroenzimas son complejos de elevado peso molecular que podrían incrementar la actividad enzimática sérica en ausencia de signos y síntomas. Se pueden detectar al ser precipitadas con polietilenglicol. El objetivo del trabajo fue determinar la actividad de las aminotransferasas por el método IFCC, calcular y comparar la media del porcentaje de actividad precipitable (x̄), su intervalo de confianza del 95% (IC95%) y el desvío estándar (DE). Se trabajó con individuos con (n=42) y sin hipertransaminasemia (n=22). Los resultados para uno y otro fueron: porcentaje de actividad precipitable con polietilenglicol (%PPA) (x̄, DE, IC95%) = (28%; 1,82; 27,45%-28,55% y 44%; 24,52; 32,84%-55,16%) y (15%; 13,03; 11,01%-18,99% y 25%; 9,1; 20,96%-29,04%) para ALT y AST, respectivamente (p=0,003 y p=0,001; p<0,05). En conclusión, la estimación de la media poblacional podría ser más precisa en individuos con hipertransaminasemia.

Abstract Macroenzymes are high-molecular-mass complexes that might increase the serum enzymatic activity in the absence of symptoms. An easy-touse method to detect them is the polyethylene glycol precipitation. The aim of this study was to determine aminotransferases activity using the IFCC method, and to calculate the mean percentage of precipitable activity (x̄), its 95% confidence interval (CI95%), and the standard deviation (SD). The study included individuals with (n=42) and without hypertransaminasemia (n=22). The results were: percentaje of precipitable activity (%PPA) (x̄, SD, CI95%) = (28%; 1.82; 27.45%-28.55% and 44%; 24.52; 32.84%-55.16%) and (15%; 13.03; 11.01%-18.99% and 25%; 9.1; 20.96%-29.04%) for ALT and AST, respectively (p=0.003 and p=0.001; p<0.05). In conclusion, the estimation of the population mean could be more precise in individuals with hypertransaminasemia.

Resumo As macroenzimas são complexos de alto peso molecular que poderiam aumentar a atividade enzimática sérica na ausência de sinais e sintomas. Podem ser detectadas ao precipitar com polietilenoglicol. O objetivo do trabalho foi determinar a atividade das aminotransferases pelo método IFCC, calcular e comparar a média da porcentagem de atividade precipitável (x̄), seu intervalo de confiança de 95% (IC95%) e o desvio padrão (DP). O trabalho foi realizado com indivíduos com (n=42) e sem hipertransaminasemia (n=22). Os resultados para os dois foram: porcentagem de atividade precipitável com polietilenoglicol (%PPA) (x̄, DP, IC95%)=(28%; 1,82; 27,45%- 28,55% e 44%; 24,52; 32,84%- 55,16%) e (15%; 13,03; 11,01%-18,99% e 25%; 9,1; 20,96%-29,04%) para ALT e AST, respectivamente (p=0,003 e p=0,001; p<0,05). Concluindo, a estimativa da média populacional poderia ser mais precisa em indivíduos com hipertransaminasemia.

Análisis de los niveles de ácidos orgánicos en orina de la primera infancia y recién nacidos: un estudio de cromatografía gaseosa acoplada a espectrometría de masas con evaluación de rangos de referencia / Analysis of urinary organic acid levels in early childhood and newborns: a gas chromatography-mass spectrometry study with evaluation of reference ranges / Análise dos níveis de ácidos orgânicos na urina da primeira infância e dos recém-nascidos: um estudo por cromatografia gasosa acoplada a espectrometria de massa com avaliação de intervalos de referência

Resumen El análisis de los ácidos orgánicos urinarios juega un papel crucial en el diagnóstico y seguimiento de pacientes con trastornos metabólicos congénitos. La falta de datos completos, las variaciones en los hábitos alimentarios entre países y el aumento del consumo de alimentos procesados subrayan la necesidad de realizar investigaciones actualizadas. Con el fin de establecer los intervalos de confianza y medianas de ácidos orgánicos urinarios se evaluaron mediante cromatografía gaseosa acoplada a espectrometría de masas, 125 muestras de orina de pacientes sanos, con edades comprendidas entre 2 días y 13 años. Los resultados fueron analizados teniendo en cuenta el grupo etario, y evidenciaron que las concentraciones de la mayoría de los ácidos orgánicos urinarios varían de acuerdo a la edad, lo que enfatiza la importancia de los valores de referencia emparejados con la edad para interpretar los datos de los pacientes. Existen pocos informes en esta área; sin embargo, la comparación de estos resultados con los valores de referencia informados por otros trabajos muestra una concordancia razonable y las pocas disparidades podrían atribuirse a factores genéticos o influencias dietéticas. Se presentan resultados e interpretaciones de niños previamente diagnosticados con trastornos metabólicos y otras afecciones, lo que confirma la confiabilidad de nuestros datos y métodos analíticos. Este estudio proporciona datos de referencia esenciales para los profesionales clínicos, destaca la importancia de los valores de referencia específicos de la edad para diagnosticar y tratar con precisión a pacientes con trastornos metabólicos y sirve como recurso fundamental para futuras investigaciones en este campo.

Abstract Analysis of urinary organic acids plays a crucial role in the diagnosis and monitoring of patients with congenital metabolic disorders. The lack of complete data, variations in dietary habits between countries, and increasing consumption of processed foods highlight the need for up-to-date research. In order to establish the confidence intervals and medians of urinary organic acids, 125 urine samples from healthy patients, aged between 2 days and 13 years, were evaluated by gas chromatography coupled to mass spectrometry. The results were analysed taking into account the age group, showing that the concentrations of most urinary organic acids vary according to age, which emphasises the importance of paired age reference values to interpret patient data. There are few reports in this area; however, comparing our results with those reference values reported by other papers demonstrates reasonable agreement, and minor disparities could be attributed to genetic factors or dietary influences. Results and interpretations from children previously diagnosed with metabolic disorders and other conditions are presented, confirming the reliability of our data and analytical methods. This study provides essential reference data for clinicians, highlighting the importance of age-specific reference values to accurately diagnose and treat patients with metabolic disorders and serves as a critical resource for future research in this field.

Resumo A análise dos ácidos orgânicos urinários desempenha um papel crucial no diagnóstico e monitoramento de pacientes com distúrbios metabólicos congênitos. A falta de dados completos, as variações nos hábitos alimentares entre países e o aumento do consumo de alimentos processados destacam a necessidade de realizar pesquisas atualizadas. Para estabelecer os intervalos de confiança e medianas de ácidos orgânicos urinários, 125 amostras de urina de pacientes saudáveis, com idades entre 2 dias e 13 anos, foram avaliadas por cromatografia gasosa acoplada à espectrometria de massa. Os resultados foram analisados levando em consideração o grupo etário, evidenciando que as concentrações da maioria dos ácidos orgânicos urinários variam de acordo com a idade, o que destaca a importância dos valores de referência pareados com a idade para interpretar os dados dos pacientes. Há poucos relatos nesta área; no entanto, a comparação destes resultados com os valores de referência informados por outros trabalhos demonstra uma concordância razoável, e as poucas disparidades poderiam ser atribuídas a fatores genéticos ou influências dietéticas. São apresentados resultados e interpretações de crianças previamente diagnosticadas com distúrbios metabólicos e outras condições, o que confirma a confiabilidade de nossos dados e métodos analíticos. Este estudo fornece dados de referência essenciais para profissionais clínicos, enfatizando a importância dos valores de referência específicos da idade para diagnosticar e tratar com precisão pacientes com distúrbios metabólicos, e serve como recurso fundamental para futuras pesquisas nesse campo.

Development of a new reference material for accurate measurements of lithium in Li-clays.

Currently, there are no national and international certified reference materials (CRM) in lithium clays that can make reliable and traceable lithium measurements for the International System of Units (SI). Hence, it is necessary to have references to meet the needs in terms of mining and activities that involve the use of lithium to favor the economy derived from its multiple uses and associated benefits in the exploration, exploitation, and handling of lithium ore. In this study, a candidate for reference material (RM) of Li in clays was developed and certified based on the provisions of ISO 17034:2016 and ISO Guide 35:2017. Different mass sizes of the RM (0.05, 0.1, and 0.25 g) were used to evaluate homogeneity. An isochronous study (short-term stability) was carried out in the assessment of stability, influenced by the effects of transport at different temperatures (20, 40, and 50 °C) for a determined time of 6 weeks, in addition to a classic (long-term) study for 19 weeks. The sample was treated using microwave-assisted acid digestion and Li measurements were performed using the analytical technique of Flame Atomic Absorption Spectrometry (FAAS). The CRM is homogeneous for the sample mass sizes of 0.05 and 0.1 g, and the mass fraction of w(Li) was stable in the RM for temperatures of 20, 40, and 50 °C. The determined period of validity was 3 years.

A New Design for Reference Values Assignment in Proficiency Testing for Fat and Crude Protein in Raw Milk for a Limited Number of Participants.

Proficiency testing (PT) allows food laboratories to endorse their competency to provide food safety guarantees to producers and consumers. One of the recommended methods for assigning reference values in PT with a small number of participants consists in considering the results that a laboratory obtains by means of a calibration test based on certified reference material (CRM). The present study delves into the results from eight PT rounds on the determination of fat and crude protein from raw milk, with modifications in the number of samples and the analysis sessions from that required by the ISO 13528:2022. The uncertainty criterion of the assigned value established by the ISO 13528:2022 standard was met by 93% of the participating laboratories, which allowed most participants to be evaluated through z-score. The assigned values were generally compatible with the results obtained by the participants. Thus, it can be concluded that the design for the assignment of the reference value is appropriate for PT with a limited number of participants. It is recommended for future PT to limit the uncertainty of the CRM according to their availability and to update the standard deviation of the proficiency assessment for the Mid-Infrared Spectroscopy method (MIR).

Establishment of a Reference Material in Quality Control for Use in Infectivity and Identity Assays of Recombinant COVID-19 Vaccine, in Accordance with International Standards Organization Guidance.

The COVID-19 pandemic, caused by the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), began in 2019. One of the strategies for pandemic control was mass vaccination. In Brazil, the recombinant COVID-19 vaccine (RCV) was produced on a large scale and offered at no charge to the population. The specifications for quality control analyses of RCV included identity and infectivity determination. To validate the results, a reference material (RM) must be analyzed in parallel with the sample vaccine. This research aimed to establish the RM for use in the identity and infectivity assay for RCV. The candidate RM was analyzed using homogeneity and stability studies. The RM was considered homogeneous for identity (cycle threshold (Ct) ≤ 25.19) and infectivity (average x- was 9.25 log10 infectious units/mL). The RM was considered adequately stable for identity during the total period in all studies, being stable at -70, 5, and 22.5 °C for 380, 313, and 14 days, respectively (Ct ≤ 21.81). For infectivity, the RM was stable at -70, 5, and 22.5 °C for 380, 97, and three days, respectively. Since the property identity and infectivity values of the RM were established, the new RM could be used in quality control analysis.

Reconstructing 90 years of anthropogenic activities in a subtropical reservoir: a chemometric and paleolimnological perspective.

Environmental reference conditions (RC) and historical trends are crucial for assessing the degree of freshwater impact and formulating restoration measures. This is particularly relevant for freshwater systems used as potable water sources. Using sediments from the Carlos Maggiolo reservoir (Minas, Uruguay), located in a watershed with a mining history, this study aimed to (1) establish metal (As, Cr, Cu, Ni, Pb, and Zn) RC using a predicted interval (PI) chemometric technique; (2) evaluate metal enrichment and toxicity over time and space; and (3) assess environmental changes examining geochemical proxies. Surface sediments from 29 stations were used to establish RC and a core from the dam area was sampled to infer past conditions. The sediments were submitted to partial digestion and analyzed by inductively coupled plasma optical emission spectrometry- ICP-OES. Enrichment factor (EF) calculated using both PI and bottom core values did not show significant differences over time. Over space and time, most metals primarily originated from natural sources (EF < 2). The PI in mg/kg was Cr: 23.74-37.32; Cu: 25.75-48.99; Ni: 16.29-25.55; Pb: 7.63-13.75; and Zn: 94.34-174.80. A stratigraphically constrained cluster analysis corroborated by a permutational multivariate analysis of variance categorized the reservoir into two zones: Zone I, before reservoir operation to ~ 1996, and Zone II from ~ 1997 to 2017. The average concentrations of the main metals of toxicological interest in zones I and II, respectively, were as follows: Cr 37.60 ± 1.59, 34.54 ± 1.49 mg/kg; Cu 49.76 ± 2.84, 44.55 ± 2.70 mg/kg; Ni 24.11 ± 0.67, 22.53 ± 1.22 mg/kg; Pb 12.40 ± 0.63, 13.52 ± 0.82 mg/kg; Zn 99.25 ± 3.12, 93.86 ± 4.42 mg/kg; Mn 1160.56 ± 68.88, 1441.61 ± 83.55 mg/kg; and P 1243.21 ± 271.56, 1128.42 ± 183.10 mg/kg. According to a principal component analysis, the period preceding reservoir operation until ~ 1985 was linked to mining activities and application of Cu2SO4, and the period from 1985s to 2000 was influenced by C and P concentrations due to increasing agricultural and afforestation activities in the watershed that consume P fertilizers. The most recent period, from 2000 to ~ 2017, was characterized by an increase in sedimentation rates mainly associated with erosion, particularly in the agricultural areas. This subsequent soil loss in the watershed could compromise the reservoir's useful life. This study contributes to a better understanding of metal geochemistry in subtropical reservoirs and aids in formulating effective recovery and restoration measures.

Selection of references for quantitative real-time PCR analysis of microRNAs in Nile tilapia (Oreochromis niloticus) under osmotic stress.

MicroRNAs play crucial regulatory roles in various aspects of development and physiology, including environmental adaptation and stress responses in teleosts. RT-qPCR is the most commonly used method for studying microRNA expression, with the accuracy and reliability of results depending on the use of an appropriate reference gene for normalization. This study aimed to evaluate seven miRNAs (U6, Let-7a, miR-23a, miR-25-3, miR-103, miR-99-5, and miR-455) expression stability in different tissues of Nile tilapia subjected to osmotic stress. Fish were divided into two groups: a control and an experimental group, raised in 0 and 12 ppt salinity water respectively. After 21 days, brain, gills, liver, and posterior intestine were collected for analysis. Different mathematical algorithms (geNorm, NormFinder, BestKeeper, and the comparative ΔCt method) were employed to identify the most suitable reference miRNAs. The results indicate that the miR-455/miR-23a combination is a robust reference for normalizing miRNA expression levels in studies of osmotic stress responses in Nile tilapia. The stability of miRNA expression can vary depending on specific stress conditions and biological processes, underscoring the necessity of selecting appropriate normalizing miRNAs for each experimental context. This study identifies reliable reference genes for future RT-qPCR analyses of miRNA expression, thereby enhancing our understanding of molecular responses in fish to environmental challenges. These insights are fundamental to the development of new technologies for the improved management and sustainability of aquaculture practices.

Unveiling the ethical void: Bias in reference citations and its academic ramifications.

Citation bias receives scant attention in discussions of ethics. However, inaccurate citation may lead to significant distortions in scientific understanding. Although ethnical and gender citation disparities have been proposed as critical aspects, there are other contributors to citation distortions, like region-based citation bias, that, although less recognized within the scientific community, are equally important. While the foundations of scientific citation include acknowledging pioneers, giving credit to related work, and providing background reading, other more subjective or even questionable criteria are often used when constructing a reference lists. Here, we discuss the potential causes and ethical concerns of citation bias, emphasizing the role of international- or region-based citation bias as one of the most harmful aspects of this ethical breach. We argue that the international scientific community should be aware of this problem and recognize its consequences, which include hindering the accurate dissemination of science, marginalizing underrepresented voices in academia, and impeding scientific progress. We advocate that scientists should compile their reference lists with the same seriousness and integrity they apply to all other aspects of their research.

Online Handwriting Recognition Method with a Non-Inertial Reference Frame Based on the Measurement of Linear Accelerations and Differential Geometry: An Alternative to Quaternions.

This work describes a mathematical model for handwriting devices without a specific reference surface (SRS). The research was carried out on two hypotheses: the first considers possible circular segments that could be made during execution for the reconstruction of the trace, and the second is the combination of lines and circles. The proposed system has no flat reference surface, since the sensor is inside the pencil that describes the trace, not on the surface as in tablets or cell phones. An inertial sensor was used for the measurements, in this case, a commercial Micro-Electro Mechanical sensor of linear acceleration. The tracking device is an IMU sensor and a processing card that allows inertial measurements of the pen during on-the-fly tracing. It is essential to highlight that the system has a non-inertial reference frame. Comparing the two proposed models shows that it is possible to construct shapes from curved lines and that the patterns obtained are similar to what is recognized; this method provides an alternative to quaternion calculus for poorly specified orientation problems.

Comparison of parasitological methods for the identification of soil-transmitted helminths, including Strongyloides stercoralis, in a regional reference laboratory in northwestern Argentina: An observational study.

Soil-transmitted helminths (STH) are a significant public health problem in impoverished communities of tropical and subtropical areas. Improved diagnostic methods are crucial for Neglected Tropical Diseases programs, particularly for S. stercoralis, as traditional methods are inadequate. Thus, it is important to identify the most accurate and efficient methods for the diagnosis of STH. We performed a retrospective study analyzing laboratory data at the Instituto de Investigaciones de Enfermedades Tropicales from 2010 to 2019. The study included data from outpatients referred for stool analysis and public health interventions from urban and rural communities in northern Salta province, Argentina. Samples were included in this analysis if processed through sedimentation/concentration, Baermann, Harada-Mori and McMaster's, with a subgroup that also included Agar plate culture method (APC). Sensitivity was calculated against a composite reference standard. Of the 5625 samples collected, 944 qualified for this analysis, with a prevalence of 11.14% for A. lumbricoides, 8.16% for hookworm, 1.38% for T. trichiura, and 6.36% for S. stercoralis. The sedimentation/concentration method was the most sensitive for A. lumbricoides (96%), compared to the McMaster method, with a sensitivity of 62%. Similarly, for hookworms, sedimentation/concentration was more sensitive than McMaster's, Harada-Mori, and Baermann with sensitivities of 87%, 70%, 43%, and 13%, respectively. Most of these infections were of light intensity. For S. stercoralis, Baermann and sedimentation/concentration methods were the most sensitive, with 70% and 62% respectively, while Harada-Mori was the least sensitive. In a subset of 389 samples also analyzed by the APC, Baermann was more sensitive than APC for detecting S. stercoralis, and both methods were superior to Harada-Mori. Parasitological methods, mostly when used combined, offer adequate opportunities for the diagnosis of STH in clinical and public health laboratories. The incorporation of S. stercoralis into the control strategies of the World Health Organization requires rethinking the current diagnostic approach used for surveys. With sedimentation/concentration and Baermann appearing as the most sensitive methods for this species. Further studies, including implementation assessments, should help in identifying the most adequate and feasible all-STH diagnostic approach.

Haematology and plasma biochemistry reference intervals of Española, San Cristobal and Eastern Santa Cruz Galapagos tortoise species.

Normal reference intervals (RI) of hematologic and biochemical parameters are important for assessing and monitoring the health status of captive and free-living chelonians; however, such information is not available for most wildlife species. Giant Galapagos tortoises are one of the most iconic animals on earth and health information can make an important contribution to their conservation and management. This study provides formal RI of haematology and plasma biochemistry parameters and describes cell morphology along with morphometrics of free-living Eastern Santa Cruz (Chelonoidis donfaustoi), Española (Chelonoidis hoodensis) and San Cristóbal tortoises (Chelonoidis chathamensis). We explored differences in blood parameters between sexes, across the tortoise species in this study and with previously published RI of the Western Santa Cruz tortoise (Chelonoidis porteri). Biochemistry parameters of both Santa Cruz species were overall more similar to each other than to Española and San Cristobal tortoises. This research constitutes the first RI for these three Galapagos tortoise species and may be of value for advising captive-breeding and conservation plans. We recommend further research to establish RI in additional tortoise species so we may better understand and interpret haematology and biochemistry parameters as a valuable conservation tool for species of this critically endangered taxon.

Assessment of reference genes for qRT-PCR normalization to elucidate host response to African swine fever infection.

Viral infection disrupts the normal regulation of the host gene's expression. In order to normalise the expression of dysregulated host genes upon virus infection, analysis of stable reference housekeeping genes using quantitative real-time-PCR (qRT-PCR) is necessary. In the present study, healthy and African swine fever virus (ASFV) infected porcine tissues were assessed for the expression stability of five widely used housekeeping genes (HPRT1, B2M, 18 S rRNA, PGK1 and H3F3A) as reference genes using standard algorithm. Total RNA from each tissue sample (lymph node, spleen, kidney, heart and liver) from healthy and ASFV-infected pigs was extracted and subsequently cDNA was synthesized, and subjected to qRT-PCR. Stability analysis of reference genes expression was performed using the Comparative delta CT, geNorm, BestKeeper and NormFinder algorithm available at RefFinder for the different groups. Direct Cycle threshold (CT) values of samples were used as an input for the web-based tool RefFinder. HPRT1 in spleen, 18 S rRNA in liver and kidney and H3F3A in heart and lymph nodes were found to be stable in the individual healthy tissue group (group A). The majority of the ASFV-infected organs (liver, kidney, heart, lymph node) exhibited H3F3A as stable reference gene with the exception of the ASFV-infected spleen, where HPRT1 was found to be the stable gene (group B). HPRT1 was found to be stable in all combinations of all CT values of both healthy and ASFV-infected porcine tissues (group C). Of five different reference genes investigated for their stability in qPCR analysis, the present study revealed that the 18 S rRNA, H3F3A and HPRT1 genes were optimal reference genes in healthy and ASFV-infected different porcine tissue samples. The study revealed the stable reference genes found in healthy as well as ASF-infected pigs and these reference genes identified through this study will form the baseline data which will be very useful in future investigations on gene expression in ASFV-infected pigs.

Systematic review on the implementation of metrological assurance systems for medical devices in Latin America.

Background: Metrology plays a crucial role in small healthcare service businesses to ensure the quality of products and services. While legal metrology in healthcare exists in some regions, it lacks harmonization. In other countries, there is limited presence of metrology in medical and biomedical engineering. We aimed to evaluate the implementation of metrological assurance systems for medical devices in Latin America. Methods: A systematic review was conducted following PRISMA 2020 guidelines and registered with PROSPERO (CRD42022359284). Searches were performed across 13 databases from October 30th to November 3rd, 2022. The search equation was "(((quality assurance) AND (metrology)) AND (medical devices))." A total of 7,789 documents were identified, of which only 16 met the inclusion criteria. Results: The majority of studies (75%) were conducted in Colombia, with a significant portion being undergraduate theses. The primary normative references used in the analyzed studies were ISO 10012 and ISO 17025, with the majority (68.75%) relying on national legislation for their approach. One study in Colombia referenced eight standards, and one in Brazil analyzed user involvement in medical device management. Among the included studies, 56.25% were conducted in healthcare institutions, mainly clinics. Most studies provided implementation guidelines, with ISO 10012 being prominent, alongside ISO 17025, which implicitly addresses ISO 9001 elements. Global bias was low across all studies. Conclusion: Our results underscore the importance of metrological assurance in managing medical devices in Latin America. The utilization of international standards and national legislation illustrates the diverse approaches adopted by different institutions. Future research should focus on optimizing metrological practices to enhance quality and safety in healthcare.

RPLP0/TBP are the most stable reference genes for human dental pulp stem cells under osteogenic differentiation.

BACKGROUND: Validation of the reference gene (RG) stability during experimental analyses is essential for correct quantitative real-time polymerase chain reaction (RT-qPCR) data normalisation. Commonly, in an unreliable way, several studies use genes involved in essential cellular functions [glyceraldehyde-3-phosphate dehydrogenase (GAPDH), 18S rRNA, and ß-actin] without paying attention to whether they are suitable for such experimental conditions or the reason for choosing such genes. Furthermore, such studies use only one gene when Minimum Information for Publication of Quantitative Real-Time PCR Experiments guidelines recommend two or more genes. It impacts the credibility of these studies and causes distortions in the gene expression findings. For tissue engineering, the accuracy of gene expression drives the best experimental or therapeutical approaches. AIM: To verify the most stable RG during osteogenic differentiation of human dental pulp stem cells (DPSCs) by RT-qPCR. METHODS: We cultivated DPSCs under two conditions: Undifferentiated and osteogenic differentiation, both for 35 d. We evaluated the gene expression of 10 candidates for RGs [ribosomal protein, large, P0 (RPLP0), TATA-binding protein (TBP), GAPDH, actin beta (ACTB), tubulin (TUB), aminolevulinic acid synthase 1 (ALAS1), tyrosine 3-monooxygenase/tryptophan 5-monooxygenase activation protein, zeta (YWHAZ), eukaryotic translational elongation factor 1 alpha (EF1a), succinate dehydrogenase complex, subunit A, flavoprotein (SDHA), and beta-2-microglobulin (B2M)] every 7 d (1, 7, 14, 21, 28, and 35 d) by RT-qPCR. The data were analysed by the four main algorithms, ΔCt method, geNorm, NormFinder, and BestKeeper and ranked by the RefFinder method. We subdivided the samples into eight subgroups. RESULTS: All of the data sets from clonogenic and osteogenic samples were analysed using the RefFinder algorithm. The final ranking showed RPLP0/TBP as the two most stable RGs and TUB/B2M as the two least stable RGs. Either the ΔCt method or NormFinder analysis showed TBP/RPLP0 as the two most stable genes. However, geNorm analysis showed RPLP0/EF1α in the first place. These algorithms' two least stable RGs were B2M/GAPDH. For BestKeeper, ALAS1 was ranked as the most stable RG, and SDHA as the least stable RG. The pair RPLP0/TBP was detected in most subgroups as the most stable RGs, following the RefFinfer ranking. CONCLUSION: For the first time, we show that RPLP0/TBP are the most stable RGs, whereas TUB/B2M are unstable RGs for long-term osteogenic differentiation of human DPSCs in traditional monolayers.

Quantitative Assessment of the Inadequate Intake of Macronutrients, Minerals, and Vitamins Associated with Ultra-Processed Food Consumption.

Studies indicate that ultra-processed food (UP) consumption correlates negatively with essential vitamin and mineral intake and positively with sodium and lipid intake. The objective of this study was to explore the relationship between UP consumption and deviations from nutritional guidelines. An observational, cross-sectional analytical study was conducted on a probability sample of manufacturing workers in the state of Rio Grande do Norte, Brazil. Food consumption was assessed with a 24 h recall survey, and nutrient intake inadequacies were calculated as the difference between individuals' intake of energy, macronutrients, minerals and vitamins, and the dietary reference intakes for individuals of the same sex and age group, and then analyzed for trends across the percentage contribution of UP to total energy intake with nonparametric multiple regression adjusted for covariates. The study included 921 workers from 33 industries, 55.9% male, with a mean age of 32 years. Overall, the study population exhibited deficits in energy, all macronutrients, and in some micronutrients. With increasing UP contribution to total energy intake, there is a trend towards a greater intake of energy (p < 0.001), total, saturated, monounsaturated, and trans fats (p < 0.001), n6-polyunsaturated fatty acids (p = 0.03), carbohydrates (p < 0.001), calcium (p = 0.008), and manganese (p < 0.001), thiamin (p < 0.001), and vitamin B6 (p = 0.01); however, this comes with a negative consequence in terms of reducing the protein consumption (p = 0.037), fiber (p = 0.035), copper (p = 0.033), and vitamin E (p = 0.002) intake. The results show that correcting energy and micronutrient deficiencies by increasing UP consumption can also lead to a decrease in diet quality.

RNA-seq validation: software for selection of reference and variable candidate genes for RT-qPCR.

BACKGROUND: Real-time quantitative PCR (RT-qPCR) is one of the most widely used gene expression analyses for validating RNA-seq data. This technique requires reference genes that are stable and highly expressed, at least across the different biological conditions present in the transcriptome. Reference and variable candidate gene selection is often neglected, leading to misinterpretation of the results. RESULTS: We developed a software named "Gene Selector for Validation" (GSV), which identifies the best reference and variable candidate genes for validation within a quantitative transcriptome. This tool also filters the candidate genes concerning the RT-qPCR assay detection limit. GSV was compared with other software using synthetic datasets and performed better, removing stable low-expression genes from the reference candidate list and creating the variable-expression validation list. GSV software was used on a real case, an Aedes aegypti transcriptome. The top GSV reference candidate genes were selected for RT-qPCR analysis, confirming that eiF1A and eiF3j were the most stable genes tested. The tool also confirmed that traditional mosquito reference genes were less stable in the analyzed samples, highlighting the possibility of inappropriate choices. A meta-transcriptome dataset with more than ninety thousand genes was also processed successfully. CONCLUSION: The GSV tool is a time and cost-effective tool that can be used to select reference and validation candidate genes from the biological conditions present in transcriptomic data.

Reference Ranges for Arterial Oxygen Saturation, Heart Rate, and Cerebral Oxygen Saturation during Immediate Postnatal Transition in Neonates Born Extremely or Very Preterm.

OBJECTIVE: To define percentile charts for arterial oxygen saturation (SpO2), heart rate (HR), and cerebral oxygen saturation (crSO2) during the first 15 minutes after birth in neonates born very or extremely preterm and with favorable outcome. STUDY DESIGN: We conducted a secondary-outcome analysis of neonates born preterm included in the Cerebral regional tissue Oxygen Saturation to Guide Oxygen Delivery in preterm neonates during immediate transition after birth III (COSGOD III) trial with visible cerebral oximetry measurements and with favorable outcome, defined as survival without cerebral injuries until term age. We excluded infants with inflammatory morbidities within the first week after birth. SpO2 was obtained by pulse oximetry, and electrocardiogram or pulse oximetry were used for measurement of HR. crSO2 was assessed with near-infrared spectroscopy. Measurements were performed during the first 15 minutes after birth. Percentile charts (10th to 90th centile) were defined for each minute. RESULTS: A total of 207 neonates born preterm with a gestational age of 29.7 (23.9-31.9) weeks and a birth weight of 1200 (378-2320) g were eligible for analyses. The 10th percentile of SpO2 at minute 2, 5, 10, and 15 was 32%, 52%, 83%, and 85%, respectively. The 10th percentile of HR at minute 2, 5, 10, and 15 was 70, 109, 126, and 134 beats/min, respectively. The 10th percentile of crSO2 at minute 2, 5, 20, and 15 was 15%, 27%, 59%, and 63%, respectively. CONCLUSIONS: This study provides new centile charts for SpO2, HR, and crSO2 for neonates born extremely or very preterm with favorable outcome. Implementing these centiles in guiding interventions during the stabilization process after birth might help to more accurately target oxygenation during postnatal transition period.