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Dried blood spots (DBSs) are formed by collecting a small sample of blood on specialized filter paper and allowing it to dry naturally. Various domains of life sciences and drug research extensively use DBSs as a sampling technique. The "Hematocrit (Ht) effect" affects assay bias, and several strategies have been put forth to deal with it, including the correction of quantified concentrations using an appropriate correction factor. The approach was previously applied, following the utilization of an image processing algorithm developed in Matlab® to derive a reliable equation correlating DBS areas to Ht% values. The present work looks more closely at the application of image analysis to the evaluation of Ht in DBS samples. Utilizing image analysis software, DBS samples with known Ht values were processed. Preparation of cards has followed a previously developed protocol for the appropriate formation of uniform area DBSs, irrespective of Ht. The resulting areas showed close resemblance to the respective theoretical areas calculated by applying the correlation equation. Following that, the equation was utilized to determine the Ht values for each sample, and a comprehensive comparison of measured versus calculated Ht was carried out using various statistical approaches for method comparison. The results demonstrated a strong correlation, suggesting the method's viability in estimating Ht for unknown DBS samples. [ABSTRACT FROM AUTHOR]

Background Detecting total cholesterol in dried blood spots could aid in identifying individuals with a high likelihood of familial hypercholesterolemia and could be used as a screening measure. This study aims to assess the diagnostic accuracy of dried blood spots on Whatman 903 paper cards using a manual enzymatic technique. Methods: A total of 394 samples were collected as serum and dried blood spots were compared. Cholesterol was determined in serum using the automated reference method, while cholesterol on paper was measured using a manual enzymatic method. Within- and between-day diagnostic variability were analyzed. The correlation between both methods was assessed using Passing–Bablok regression and Bland–Altman plot. Internal validation of our correlation formula was performed on 149 samples, along with external validation of the formula proposed by Corso et al. Results: The within- and between-day coefficient of variation was found to be lower than 10.14% and 14.09%, respectively. Passing–Bablok regression indicated a precision of 0.803 and an accuracy of 0.96. Internal validation precision was measured at 0.716. The resulting positive and negative predicted values were 0.77 and 0.92, respectively, vs. 0.46 and 0.96 from the external formula. Conclusions: Total cholesterol analysis in dried blood spots demonstrates high precision and reproducibility. This method reliably enables the incorporation of this biological marker into neonatal screening for familial hypercholesterolemia detection. [ABSTRACT FROM AUTHOR]

Dried blood spots (DBS) have proven to be a powerful sampling and storage method for newborn screening and many other applications. However, DBS methods have not yet been optimized for broad-spectrum targeted metabolomic analysis. In this study, we developed a robust, DBS-based, broad-spectrum, targeted metabolomic method that was able to measure over 400 metabolites from a 6.3 mm punch from standard Whatman 903TM filter paper cards. The effects of blood spot volumes, hematocrit, vacutainer chemistry, extraction methods, carryover, and comparability with plasma and fingerstick capillary blood samples were analyzed. The stability of over 400 metabolites stored under varying conditions over one year was also tested. No significant impacts of blood volume and hematocrit variations were observed when the spotted blood volume was over 60 µL and the hematocrit was between 31% and 50%. The median area under the curve (AUC) of metabolites in the DBS metabolome declined by 40% in the first 3 months and then did not decline further for at least 1 year. All originally detectable metabolites remained within detectable limits. The optimal storage conditions for metabolomic analysis were −80 °C with desiccants and without an O2 scavenger. The method was clinically validated for its potential utility in the diagnosis of the mitochondrial disease mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes (MELAS). Our method provides a convenient alternative to freezing, storing, and shipping liquid blood samples for comparative metabolomic studies. [ABSTRACT FROM AUTHOR]

Dried blood spots (DBSs), a micro-sampling technique whereby a drop of blood is collected on filter paper has multiple advantages over conventional blood sampling regarding the sampling itself, as well as transportation and storage. This is the first paper describing the development and validation of a method for the determination of 23 mycotoxins and phase I metabolites in DBSs from pigs and broiler chickens using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The targeted mycotoxins belong to groups for which the occurrence in feed is regulated by the European Union, namely, aflatoxins, ochratoxin A and several Fusarium mycotoxins, and to two groups of unregulated mycotoxins, namely Alternaria mycotoxins and Fusarium mycotoxins (enniatins and beauvericin). The impact of blood haematocrit, DBS sampling volume and size of the analysed DBS disk on the validation results was assessed. No effects of variation in size of the analysed disk, haematocrit and spotted blood volume were observed for most mycotoxins, except for the aflatoxins and β-zearalanol (BZAL) at the lowest haematocrit (26%) level and for the enniatins (ENNs) at the lowest volume (40 µL). The developed method was transferred to an LC-high resolution mass spectrometry instrument to determine phase II metabolites. Then, the DBS technique was applied in a proof-of-concept toxicokinetic study including a comparison with LC-MS/MS data from plasma obtained with conventional venous blood sampling. A strong correlation (r > 0.947) was observed between plasma and DBS concentrations. Finally, DBSs were also applied in a pilot exposure assessment study to test their applicability under field conditions. [ABSTRACT FROM AUTHOR]

Objectives: Conduction of pharmacokinetic (PK) study in pediatric patients is challenging due to blood sampling limits. The dried blood spots (DBS) method represents a potential matrix for microsampling in support of PK studies in children. Herein, we used the Capitainer® qDBS device to develop a DBS method that can collect an exact 10 µL volume of blood on a paper card. This DBS method was developed to simultaneously quantify the concentrations of eight antibiotics, including sulbactam, tazobactam, ampicillin, meropenem, cefotaxime, cefoperazone, piperacillin, and metronidazole using ultra-high performance liquid chromatography–tandem mass spectrometry (UHPLC–MS/MS). Methods: The prepared DBS samples were extracted in methanol containing acetaminophen as the internal standard at 20 °C on a block bath shaker at 500 rpm for 30 min. The extracted antibiotics were eluted on an Acquity UPLC HSS T3 column (2.1 × 50 mm, 1.8 µm) using gradient elution with a total chromatographic run time of 6.5 min. The precursor and product ions of the analytes were detected by use of the multiple reaction monitoring (MRM) mode. Results: No interfering peaks at the respective retention times of the analytes were observed in DBS samples. The lower limits of quantification (LLOQ) for the antibiotics were between 0.25 and 2.0 μg/mL, and satisfactory accuracies (intra/inter-assay bias −16.7 to +13.6%) and precisions (intra/inter-assay coefficient of variations 1.5–15.6%) were obtained for the analytes. As a proof of concept, the method was applied to DBS samples obtained from neonatal patients treated with ampicillin and piperacillin/sulbactam. Conclusions: The DBS method is simple and robust, and it can be used in children with limited blood sampling. [ABSTRACT FROM AUTHOR]

During the lipidomic analysis of red blood cell membranes, the distribution and percentage ratios of the fatty acids are measured. Since fatty acids are the key constituents of cell membranes, by evaluating their quantities it possible to understand the general health of the cells and to obtain health indicators of the whole organism. However, because the analysis is precise, it is necessary to ensure that the blood does not undergo significant variations between the point of collection and analysis. The composition of the blood may vary dramatically weeks after collection, hence, here an attempt is made to stabilize these complex matrixes using antioxidants deposited on the paper cards on which the blood itself is deposited. [ABSTRACT FROM AUTHOR]

Certain micronutrients exhibit immunomodulatory effects. However, no intervention has yet investigated the effect of individualized supplementation on the severity of upper respiratory tract infections (URIs). Therefore, we investigated whether a personalized supplementation moderates the incidence and severity of URI. Selenium, zinc, and vitamin D were measured in dried blood spots from 59 healthy participants. Accordingly, a personalized supplement was provided with or without the respective micronutrients. We used WURSS-21 questionnaires to assess the disease status. The blood values converged during the intervention and micronutrients no longer differed between treated and untreated volunteers at the end of the intervention period. The incidence and severity of the illness did not significantly differ between the groups. However, when analyzing the WURSS-21 scores by the intention to treat, the initially randomized treatment arm revealed a significantly higher score than the placebo arm. Upon acute administration, individualized combinations of selenium, zinc and vitamin D do not reduce the number, or contribute to a milder course of URIs. Therefore, supplementation in acute infectious situations seems questionable. Further studies must address the habitual diet in more detail, to better understand the impact of individual micronutrient status on the prevention of URI. [ABSTRACT FROM AUTHOR]

Inborn errors of metabolism and diabetes share common derangements in analytes of metabolic networks that are tested for in newborn screening, usually performed 48–72 h after birth. There is limited research examining the metabolic imprint of diabetes on newborn screening results. This paper aims to demonstrate the links between diabetes, biochemical genetics and newborn screening in investigating disease pathophysiology in diabetes, provide possible reasons for the lack of research in diabetes in newborn screening and offer recommendations on potential research areas. We performed a systematic search of the available literature from 1 April 1998 to 31 December 2018 involving newborn screening and diabetes using OVID, MEDLINE, Cochrane and the PROSPERO register, utilizing a modified extraction tool adapted from Cochrane. Eight studies were included after screening 1312 records. Five studies reanalyzed dried blood spots (DBS) on filter paper cards, and three studies utilized pre-existing results. The results of these studies and how they relate to cord blood studies, the use of cord blood versus newborn screening dried blood spots as a sample and considerations on newborn screening and diabetes research is further discussed. The timing of sampling of newborn screening allows insight into neonatal physiology in a catabolic state with minimal maternal and placental influence. This, combined with the wide coverage of newborn screening worldwide, may aid in our understanding of the origins of diabetes. [ABSTRACT FROM AUTHOR]

Dried blood spot (DBS) microsampling is extensively employed in newborn screening (NBS) and neonatal studies. However, the impact of variable neonatal hematocrit (Ht) values on the results can be a source of analytical error, and the use of fixed Ht for calibration (Htcal) is not representative of all neonatal subpopulations. A computational approach based on neonatal demographics was developed and implemented in R® language to propose a strategy using correction factors to address the Ht effect in neonatal DBS partial-spot assays. A rational "tolerance level" was proposed for the Ht effect contribution to the total analytical error and a safe Ht range for neonatal samples, where the correction of concentrations can be omitted. Furthermore, an "alert zone" for a false positive or negative result in NBS was proposed, where the Ht effect has to be considered. Results point toward the use of Htcal values closely representative of populations under analysis and an acceptable level of percentage relative error can be attributed to the Ht effect, diminishing the probability of correction. Overall, the impact of the Ht effect on neonatal studies is important and future work may further investigate this parameter, correlated to other clinical variables potentially affecting results. [ABSTRACT FROM AUTHOR]

The aim of this study was to evaluate the difference in drug exposure of rifampicin in native versus non-native Paraguayan populations using dried blood spots (DBS) samples collected utilizing a limited sampling strategy. This was a prospective pharmacokinetic study that enrolled hospitalized tuberculosis (TB) patients from both native and non-native populations receiving oral rifampicin 10 mg/kg once-daily dosing. Steady-state DBS samples were collected at 2, 4, and 6 h after intake of rifampicin. The area under the time concentration curve 0–24 h (AUC0–24) was calculated using a Bayesian population PK model. Rifampicin AUC0–24 < 38.7 mg*h/L was considered as low. The probability of target attainment (PTA) was calculated using AUC0–24/MIC > 271 as a target and estimated MIC values of 0.125 and 0.25 mg/L. In total, 50 patients were included. Native patients (n = 30) showed comparable drug exposure to the non-natives (n = 20), median AUC0–24 24.7 (17.1–29.5 IQR) and 21.6 (15.0–35.4 IQR) mg*h/L (p = 0.66), respectively. Among total patients, only 16% (n = 8) had a rifampicin AUC0–24 > 38.7 mg*h/L. Furthermore, PTA analysis showed that only 12 (24%) of the patients met a target AUC0–24 /MIC ≥ 271, assuming an MIC of 0.125 mg/L, which plummeted to 0% at a wild-type MIC of 0.25 mg/L. We successfully used DBS and limited sampling for the AUC0–24 estimation of rifampicin. Currently, our group, the EUSAT-RCS consortium, is preparing a prospective multinational, multicenter phase IIb clinical trial evaluating the safety and efficacy of high-dose rifampicin (35 mg/kg) in adult subjects using the DBS technique for AUC0–24 estimation. [ABSTRACT FROM AUTHOR]

Monitoring cortisol replacement therapy in congenital adrenal hyperplasia (CAH) patients is vital to avoid serious adverse events such as adrenal crises due to cortisol underexposure or metabolic consequences due to cortisol overexposure. The less invasive dried blood spot (DBS) sampling is an advantageous alternative to traditional plasma sampling, especially in pediatric patients. However, target concentrations for important disease biomarkers such as 17α-hydroxyprogesterone (17-OHP) are unknown using DBS. Therefore, a modeling and simulation framework, including a pharmacokinetic/pharmacodynamic model linking plasma cortisol concentrations to DBS 17-OHP concentrations, was used to derive a target morning DBS 17-OHP concentration range of 2–8 nmol/L in pediatric CAH patients. Since either capillary or venous DBS sampling is becoming more common in the clinics, the clinical applicability of this work was shown by demonstrating the comparability of capillary and venous cortisol and 17-OHP concentrations collected by DBS sampling, using a Bland-Altman and Passing-Bablok analysis. The derived target morning DBS 17-OHP concentration range is a first step towards providing improved therapy monitoring using DBS sampling and adjusting hydrocortisone (synthetic cortisol) dosing in children with CAH. In the future, this framework can be used to assess further research questions, e.g., target replacement ranges for the entire day. [ABSTRACT FROM AUTHOR]

Fabry disease (FD) is an X-linked lysosomal storage disorder where impaired α-galactosidase A enzyme activity leads to the intracellular accumulation of undegraded glycosphingolipids, including globotriaosylsphingosine (lyso-Gb3) and related analogues. Lyso-Gb3 and related analogues are useful biomarkers for screening and should be routinely monitored for longitudinal patient evaluation. In recent years, a growing interest has emerged in the analysis of FD biomarkers in dried blood spots (DBSs), considering the several advantages compared to venipuncture as a technique for collecting whole-blood specimens. The focus of this study was to devise and validate a UHPLC-MS/MS method for the analysis of lyso-Gb3 and related analogues in DBSs to facilitate sample collection and shipment to reference laboratories. The assay was devised in conventional DBS collection cards and in Capitainer®B blood collection devices using both capillary and venous blood specimens from 12 healthy controls and 20 patients affected with FD. The measured biomarker concentrations were similar in capillary and venous blood specimens. The hematocrit (Hct) did not affect the correlation between plasma and DBS measurements in our cohort (Hct range: 34.3–52.2%). This UHPLC-MS/MS method using DBS would facilitate high-risk screening and the follow-up and monitoring of patients affected with FD. [ABSTRACT FROM AUTHOR]

Biological bases of autism spectrum disorder (ASD) include both genetic and epigenetic causes. Patients with ASD show anomalies in the profile of certain plasma amino acids, including neuroactive amino acids. Monitoring plasma amino acids may be relevant for patient care and interventions. We evaluated the plasma amino acid profile in samples extracted from dry blood spots by electrospray ionization-tandem mass spectrometry. Fourteen amino acids and eleven amino acid ratios were examined in patients with ASD and intellectual disability (ID), and neurotypical control subjects (TD). The amino acid profile in the ASD group showed reduced levels of ornithine (p = 0.008), phenylalanine (p = 0.042) and tyrosine (p = 0.013). The statistically significant amino acid ratios were Leu+Val/Phe+Tyr (p = 0.002), Tyr/Leu (p = 0.007) and Val/Phe (p = 0.028), such differences remaining significant only in the comparison between ASD and TD. Finally, a positive correlation emerged between the score of the restricted and repetitive behavior on ADOS-2 and the citrulline levels in the ASD group (p = 0.0047). To conclude, patients with ASD may show a distinguishable metabolic profile useful for studying their metabolic pathways in order to develop screening tests and targeted therapies. [ABSTRACT FROM AUTHOR]

In this research, a UHPLC–MS/MS method was developed and validated for the determination of zonisamide in dried plasma spots (DPS) and dried blood spots (DBS). Detection of zonisamide and internal standard, 1-(2,3-dichlorphenyl)piperazine, was carried out in ESI+ mode by monitoring two MRM transitions per analyte. Total run time, less than 2.5 min, was achieved using Acquity UPLC BEH Amide (2.1 × 100 mm, 1.7 µm particle size) column with mobile phase comprising acetonitrile–water (85:15%, v/v) with 0.075% formic acid. The flow rate was 0.225 mL/min, the column temperature was 30 °C and the injection volume was 3 µL. Desolvation temperature, desolvation gas flow rate, ion source temperature and cone gas flow rate were set by the IntelliStart software tool in combination with tuning. All of the Guthrie cards were scanned, and DPS/DBS areas were determined by the image processing tool. The influence of hematocrit values (20–60%) on accuracy and precision was evaluated to determine the range within which method for DBSs is free from Hct or dependency is within acceptable limits. The validated method was applied to the determination of zonisamide levels in DPS and DBS samples obtained from patients confirming its suitability for clinical application. Finally, the distribution of zonisamide into the red blood cells was estimated by correlating its DPS and DBS levels. [ABSTRACT FROM AUTHOR]

Numerous sero-epidemiological studies have been initiated to investigate the spread and dynamics of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). To address the concomitant need for serological high-throughput assays, a bead-based multiplex serology assay, specific for SARS-CoV-2, had been developed. SARS-CoV-2 serolomics allows for measuring antibody responses to almost the entire SARS-CoV-2 proteome in up to 2000 serum samples per day. To enlarge the pool of eligible sample collection methods, we here test the compatibility of serolomics with dried blood spot (DBS)-derived eluates. Antibody levels of nine SARS-CoV-2 antigens, including the nucleocapsid (N) and receptor-binding domain of the spike protein (S1-RBD), were measured in 142 paired DBS and serum samples. The numeric correlation between the two sample types was high, with a Pearson's r of 0.88 for both S1-RBD and N and intraclass correlation coefficients of 0.93 and 0.92, respectively. Systematically reduced antibody levels in DBS eluates were compensated by lowering the cutoffs for seropositivity accordingly. This enabled the concordant classification of SARS-CoV-2 seropositivity, without loss in sensitivity. Antibody levels against accessory SARS-CoV-2 antigens also showed a high concordance, demonstrating that DBS-derived eluates are eligible for SARS-CoV-2 serolomics. DBS cards facilitate the collection of blood samples, as they obviate the need for medically trained personnel and can be shipped at room temperature. In combination with SARS-CoV-2 serolomics, DBS cards enable powerful sero-epidemiological studies, thus allowing for the monitoring of patients and epidemiological analyses in resource-poor settings. [ABSTRACT FROM AUTHOR]

Loiasis, caused by the filarial nematode Loa loa, is endemic in Central and West Africa. Loa loa has been associated with severe adverse reactions in high Loa-infected individuals receiving ivermectin during mass drug administration programs for the control of onchocerciasis and lymphatic filariasis. Diagnosis of loiasis still depends on microscopy in blood samples, but this is not effective for large-scale surveys. New diagnostics methods for loiasis are urgently needed. Previously, we developed a colorimetric high-sensitive and species-specific LAMP for Loa loa DNA detection. Here, we evaluate it in a set of 100 field-collected clinical samples stored as dried blood spots. In addition, Loa loa-LAMP was also evaluated in real-time testing and compared with microscopy and a specific PCR/nested PCR. A simple saponin/Chelex-based method was used to extract DNA. Colorimetric and real-time LAMP assays detected more samples with microscopy-confirmed Loa loa and Loa loa/Mansonella perstans mixed infections than PCR/nested-PCR. Samples with the highest Loa loa microfilariae counts were amplified faster in real-time LAMP assays. Our Loa loa-LAMP could be a promising molecular tool for the easy, rapid and accurate screening of patients for loiasis in endemic areas with low-resource settings. The real-time testing (feasible in a handheld device) could be very useful to rule out high-microfilariae loads in infected patients. [ABSTRACT FROM AUTHOR]

Clinical monitoring of pediatric HIV treatment remains a major challenge in settings where drug resistance genotyping is not routinely available. As a result, our understanding of drug resistance, and its impact on subsequent therapeutic regimens available in these settings, remains limited. We investigate the prevalence and correlates of HIV-1 drug resistance among 94 participants of the Ethiopia Pediatric HIV Cohort failing first-line combination antiretroviral therapy (cART) using dried blood spot-based genotyping. Overall, 81% (73/90) of successfully genotyped participants harbored resistance mutations, including 69% (62/90) who harbored resistance to both Nucleoside Reverse Transcriptase Inhibitors (NRTIs) and Non-nucleoside Reverse Transcriptase Inhibitors (NNRTIs). Strikingly, 42% of resistant participants harbored resistance to all four NRTIs recommended for second-line use in this setting, meaning that there are effectively no remaining cART options for these children. Longer cART duration and prior regimen changes were significantly associated with detection of drug resistance mutations. Replicate genotyping increased the breadth of drug resistance detected in 34% of cases, and thus is recommended for consideration when typing from blood spots. Implementation of timely drug resistance testing and access to newer antiretrovirals and drug classes are urgently needed to guide clinical decision-making and improve outcomes for HIV-infected children on first-line cART in Ethiopia. [ABSTRACT FROM AUTHOR]

Rice bran contains essential nutrients, antioxidants, and bioactives with anti-inflammatory and diarrheal protective properties important for infants. This 6-month randomized controlled trial investigated the effects of heat-stabilized rice bran supplementation during Malian infant weaning. Fifty healthy 6-month-old infants were randomized to a rice bran intervention (N = 25) or non-intervention control group (N = 25). Intervention infants received dose-escalating rice bran supplementation for 6 months (1–5 g/day). Monthly infant dried blood spot and anthropometric measurements were collected. Dried blood spot metabolite abundances were compared monthly according to diet for six months. Supplementation resulted in favorable weight-for-age and weight-for-length z-score changes. Non-targeted dried blood spot-based metabolomics identified 796 metabolites, of which 33% had significant fold differences between groups (7–12 months). Lipids and amino acids represented 70.6% of the metabolites identified. Rice bran supplementation during infant weaning significantly modulated the metabolites involved in antioxidant defenses and with neuroactive properties including reduced glutathione, glycine, glutamate, cysteinylglycine, tryptophan betaine, and choline. These findings support rice bran as a weaning ingredient to meet infant nutritional requirements and with the potential to reduce oxidative stress and improve cognitive outcomes. This study provides evidence for dried blood spots as a cost-effective tool to detect infant biomarkers of nutritional and metabolic status. [ABSTRACT FROM AUTHOR]

Background: Low vitamin D in pregnancy may impair the development of the fetal immune system and influence the risk of later development of rheumatoid arthritis (RA) in the offspring. The aim was to examine whether lower 25-hydroxyvitamin D3 (25(OH)D) concentrations at birth were associated with the risk of developing RA in early adulthood. Methods: This case-cohort study obtained data from Danish registers and biobanks. Cases included all individuals born during 1981–1996 and recorded in the Danish National Patient Register with a diagnosis of RA with age >18 years at first admission. The random comparison consisted of a subset of Danish children. Vitamin D concentrations were measured in newborn dried blood. In total, 805 RA cases and 2416 individuals from the subcohort were included in the final analysis. Weighted Cox regression was used to calculate hazard ratio (HR). Results: The median (interquartile rage (IQR)) 25(OH)D concentrations among cases were 24.9 nmol/L (IQR:15.4;36.9) and 23.9 nmol/L (IQR:13.6;36.4) among the subcohort. There was no indication of a lower risk of RA among individuals in the highest vitamin D quintile compared with the lowest (HRadj.:1.21 (0.90;1.63)). Conclusion: The risk of RA in early adulthood was not associated with vitamin D concentrations at birth. [ABSTRACT FROM AUTHOR]

Are free carnitine concentrations on newborn screening (NBS) 48–72 h after birth lower in patients who develop type 1 diabetes than in controls? A retrospective case-control study of patients with type 1 diabetes was conducted. NBS results of patients from a Sydney hospital were compared against matched controls from the same hospital (1:5). Multiple imputation was performed for estimating missing data (gestational age) using gender and birthweight. Conditional logistic regression was used to control for confounding and to generate parameter estimates (α = 0.05). The Hommel approach was used for post-hoc analyses. Results are reported as medians and interquartile ranges. A total of 159 patients were eligible (80 females). Antibodies were detectable in 86. Median age at diagnosis was 8 years. Free carnitine concentrations were lower in patients than controls (25.50 µmol/L;18.98–33.61 vs. 27.26; 21.22–34.86 respectively) (p = 0.018). Immunoreactive trypsinogen was higher in this group (20.24 µg/L;16.15–29–52 vs. 18.71; 13.96–26.92) (p = 0.045), which did not persist in the post-hoc analysis. Carnitine levels are lower and immunoreactive trypsinogen might be higher, within 2–3 days of birth and years before development of type 1 diabetes as compared to controls, although the differences were well within reference ranges and provide insight into the pathogenesis into neonatal onset of type 1 diabetes development rather than use as a diagnostic tool. Given trypsinogen's use for evaluation of new-onset type 1 diabetes, larger studies are warranted. [ABSTRACT FROM AUTHOR]

The World Health Organization has established an elimination plan for hepatitis C virus (HCV) by 2030. In Sub-Saharan Africa (SSA) access to diagnostic tools is limited, and a number of genotype 4 subtypes have been shown to be resistant to some direct-acting antivirals (DAAs). This study aims to analyze diagnostic assays for HCV based on dried blood spots (DBS) specimens collected in Kinshasa and to characterize genetic diversity of the virus within a group of mainly HIV positive patients. HCV antibody detection was performed on 107 DBS samples with Vidas® anti-HCV and Elecsys anti-HCV II, and on 31 samples with INNO-LIA HCV. Twenty-six samples were subjected to molecular detection. NS3, NS5A, and NS5B regions from 11 HCV viremic patients were sequenced. HCV seroprevalence was 12.2% (72% with detectable HCV RNA). Both Elecsys Anti-HCV and INNO-LIA HCV were highly sensitive and specific, whereas Vidas® anti-HCV lacked full sensitivity and specificity when DBS sample was used. NS5B/NS5A/NS3 sequencing revealed exclusively GT4 isolates (50% subtype 4r, 30% 4c and 20% 4k). All 4r strains harbored NS5A resistance-associated substitutions (RAS) at positions 28, 30, and 31, but no NS3 RAS was detected. Elecsys Anti-HCV and INNO-LIA HCV are reliable methods to detect HCV antibodies using DBS. HCV subtype 4r was the most prevalent among our patients. RASs found in subtype 4r in NS5A region confer unknown susceptibility to DAA. [ABSTRACT FROM AUTHOR]

The review presents data from the last few years on bioanalytical methods used in therapeutic drug monitoring (TDM) of the 1st–3rd generation and the newest antiepileptic drug (AEDs) cenobamate in patients with various forms of seizures. Chemical classification, structure, mechanism of action, pharmacokinetic data and therapeutic ranges for total and free fractions and interactions were collected. The primary data on bioanalytical methods for AEDs determination included biological matrices, sample preparation, dried blood spot (DBS) analysis, column resolution, detection method, validation parameters, and clinical utility. In conclusion, the most frequently described method used in AED analysis is the LC-based technique (HPLC, UHPLC, USLC) combined with highly sensitive mass detection or fluorescence detection. However, less sensitive UV is also used. Capillary electrophoresis and gas chromatography have been rarely applied. Besides the precipitation of proteins or LLE, an automatic SPE is often a sample preparation method. Derivatization was also indicated to improve sensitivity and automate the analysis. The usefulness of the methods for TDM was also highlighted. [ABSTRACT FROM AUTHOR]

Pediatric human immunodeficiency virus (HIV) care in resource-limited settings remains a major challenge to achieving global HIV treatment and virologic suppression targets, in part because the administration of combination antiretroviral therapies (cART) is inherently complex in this population and because viral load and drug resistance genotyping are not routinely available in these settings. Children may also be at elevated risk of transmission of drug-resistant HIV as a result of suboptimal antiretroviral administration for prevention of mother-to-child transmission. We investigated the prevalence and the correlates of pretreatment HIV drug resistance (PDR) among HIV-infected, cART-naive children in Ethiopia. We observed an overall PDR rate of 14%, where all cases featured resistance to non-nucleoside reverse transcriptase inhibitors (NNRTIs): ~9% of participants harbored resistance solely to NNRTIs while ~5% harbored resistance to both NNRTIs and nucleoside reverse transcriptase inhibitors (NRTIs). No resistance to protease inhibitors was observed. No sociodemographic or clinical parameters were significantly associated with PDR, though limited statistical power is noted. The relatively high (14%) rate of NNRTI resistance in cART-naive children supports the use of non-NNRTI-based regimens in first-line pediatric treatment in Ethiopia and underscores the urgent need for access to additional antiretroviral classes in resource-limited settings. [ABSTRACT FROM AUTHOR]