Banana (
Musa acuminata Colla AAA) peel extracts obtained in this work had a high capacity to scavenge 2,2-diphenyl-1-picrylhydrazyl (DPPH
) and 2,2′-azino-bis(3-ethylbenzothiazoline)-6-sulfonic acid ...(ABTS
+
) free radicals, and they were also good lipid peroxidation inhibitors. Acetone:water extracts were considerably more effective (compared with methanol, ethanol, acetone, water, methanol:water or ethanol:water) at inhibiting the peroxidation of lipids in the β-carotene/linoleic acid system or scavenging free radicals. However, aqueous extracts had a high capacity to protect lipids from oxidation in the thiobarbituric acid reactive substances (TBARS) test, as well as in the β-carotene bleaching assay. In addition, acetone:water most efficiently extracted all extractable components (54
±
4%), phenolic compounds (3.3
±
0.8%), and anthocyanin compounds (434
±
97
μg cyanidin 3-glucoside equivalents/100
g freeze-dried banana peel). Banana peel contained large amounts of dopamine and L-dopa, catecholamines with a significant antioxidant activity. However, ascorbic acid, tocopherols or phytosterols were not detected in the different extracts. The antioxidant activity of banana peel extracts from different cultivars was similar. However, the impact of extraction time or temperature should be studied in greater depth.
The carotid body (CB) is a prototypical acute oxygen (O2)‐sensing organ that mediates reflex hyperventilation and increased cardiac output in response to hypoxaemia. CB overactivation, secondary to ...the repeated stimulation produced by the recurrent episodes of intermittent hypoxia, is believed to contribute to the pathogenesis of sympathetic hyperactivity present in sleep apnoea patients. Although CB functional plasticity induced by chronic intermittent hypoxia (CIH) has been demonstrated, the underlying mechanisms are not fully elucidated. Here, we show that CIH induces a small increase in CB volume and rearrangement of cell types in the CB, characterized by a mobilization of immature quiescent neuroblasts, which enter a process of differentiation into mature, O2‐sensing and neuron‐like, chemoreceptor glomus cells. Prospective isolation of individual cell classes has allowed us to show that maturation of CB neuroblasts is paralleled by an upregulation in the expression of specific glomus cell genes involved in acute O2‐sensing. CIH enhances mitochondrial responsiveness to hypoxia in maturing neuroblasts as well as in glomus cells. These data provide novel perspectives on the pathogenesis of CB‐mediated sympathetic overflow that may lead to the development of new pharmacological strategies of potential applicability in sleep apnoea patients.
Key points
Obstructive sleep apnoea is a frequent condition in the human population that predisposes to severe cardiovascular and metabolic alterations.
Activation of the carotid body, the main arterial oxygen‐sensing chemoreceptor, by repeated episodes of hypoxaemia induces exacerbation of the carotid body‐mediated chemoreflex and contributes to sympathetic overflow characteristic of sleep apnoea patients.
In rats, chronic intermittent hypoxaemia induces fast neurogenesis in the carotid body with rapid activation of neuroblasts, which enter a process of proliferation and maturation into O2‐sensing chemoreceptor glomus cells.
Maturing carotid body neuroblasts and glomus cells exposed to chronic intermittent hypoxia upregulate genes involved in acute O2 sensing and enhance mitochondrial responsiveness to hypoxia.
These findings provide novel perspectives on the pathogenesis of carotid body‐mediated sympathetic hyperactivation. Pharmacological modulation of carotid body fast neurogenesis could help to ameliorate the deleterious effects of chronic intermittent hypoxaemia in sleep apnoea patients.
figure legend Changes in the cellular composition of adult rat carotid body (CB) stem cell niche induced by chronic intermittent hypoxia (CIH). Cell populations are identified and sorted by flow cytometry to study their morphological and functional properties. CIH activates progenitors of the neuronal lineage and results in the maturation of CB neuroblasts into O2‐sensitive glomus cells.
•This study compared RT-qPCR sensitivity for SARS-CoV-2 detection.•False negatives were in the range of 2–39.8%.•The most sensitive solution (97.9% 92.8–99.7) targeted the SARS-CoV-2 E-gene.
The ...ongoing COVID-19 pandemic continues to impose demands on diagnostic screening. In anticipation that the recurrence of outbreaks and the measures for lifting the lockdown worldwide may cause supply chain issues over the coming months, this study assessed the sensitivity of a number of one-step retrotranscription and quantitative polymerase chain reaction (RT-qPCR) solutions to detect SARS-CoV-2.
Six different RT-qPCR alternatives were evaluated for SARS-CoV-2/COVID-19 diagnosis based on standard RNA extractions. The one with best sensitivity was also assessed with direct nasopharyngeal swab viral transmission medium (VTM) heating; thus overcoming the RNA extraction step.
A wide variability in the sensitivity of RT-qPCR solutions was found that was associated with a range of false negatives from 2% (0.3–7.9%) to 39.8% (30.2–50.2%). Direct preheating of VTM combined with the best solution provided a sensitivity of 72.5% (62.5–81.0%), in the range of some of the solutions based on standard RNA extractions.
Sensitivity limitations of currently used RT-qPCR solutions were found. These results will help to calibrate the impact of false negative diagnoses of COVID-19, and to detect and control new SARS-CoV-2 outbreaks and community transmissions.
•High-throughput screening of SARS-CoV-2 by RT-qPCR without previous RNA extraction.•Heated nasopharyngeal swab transport medium as template in the RT-qPCR test.•Fast and safety management of ...SARS-CoV-2 samples in the diagnostic process.
The gold-standard COVID-19 diagnosis relies on detecting SARS-CoV-2 using RNA purification and one-step retrotranscription and quantitative PCR (RT-qPCR). Based on the urgent need for high-throughput screening, we tested the performance of three alternative, simple and affordable protocols to rapidly detect SARS-CoV-2, bypassing the long and tedious RNA extraction step and reducing the time to viral detection.
We evaluated three methods based on direct nasopharyngeal swab viral transmission medium (VTM) heating before the RT-qPCR: a) direct without additives; b) in a formamide-EDTA (FAE) buffer, c) in a RNAsnapTM buffer.
Although with a delay in cycle threshold compared to the gold-standard, we found consistent results in nasopharyngeal swab samples that were subject to a direct 70°C incubation for 10 min.
Our findings provide valuable options to overcome any supply chain issue and help to increase the throughput of diagnostic tests, thereby complementing standard diagnosis.
Most causal variants of Mendelian diseases are exonic. Whole‐exome sequencing (WES) has become the diagnostic gold standard, but causative variant prioritization constitutes a bottleneck. Here we ...assessed an in‐house sample‐to‐sequence pipeline and benchmarked free prioritization tools for germline causal variants from WES data. WES of 61 unselected patients with a known genetic disease cause was obtained. Variant prioritizations were performed by diverse tools and recorded to obtain a diagnostic yield when the causal variant was present in the first, fifth, and 10th top rankings. A fraction of causal variants was not captured by WES (8.2%) or did not pass the quality control criteria (13.1%). Most of the applications inspected were unavailable or had technical limitations, leaving nine tools for complete evaluation. Exomiser performed best in the top first rankings, while LIRICAL led in the top fifth rankings. Based on the more conservative top 10th rankings, Xrare had the highest diagnostic yield, followed by a three‐way tie among Exomiser, LIRICAL, and PhenIX, then followed by AMELIE, TAPES, Phen‐Gen, AIVar, and VarNote‐PAT. Xrare, Exomiser, LIRICAL, and PhenIX are the most efficient options for variant prioritization in real patient WES data.
The mitochondrial genome (mtDNA) is of interest for a range of fields including evolutionary, forensic, and medical genetics. Human mitogenomes can be classified into evolutionary related haplogroups ...that provide ancestral information and pedigree relationships. Because of this and the advent of high-throughput sequencing (HTS) technology, there is a diversity of bioinformatic tools for haplogroup classification. We present a benchmarking of the 11 most salient tools for human mtDNA classification using empirical whole-genome (WGS) and whole-exome (WES) short-read sequencing data from 36 unrelated donors. We also assessed the best performing tool in third-generation long noisy read WGS data obtained with nanopore technology for a subset of the donors. We found that, for short-read WGS, most of the tools exhibit high accuracy for haplogroup classification irrespective of the input file used for the analysis. However, for short-read WES, Haplocheck and MixEmt were the most accurate tools. Based on the performance shown for WGS and WES, and the accompanying qualitative assessment, Haplocheck stands out as the most complete tool. For third-generation HTS data, we also showed that Haplocheck was able to accurately retrieve mtDNA haplogroups for all samples assessed, although only after following assembly-based approaches (either based on a referenced-based assembly or a hybrid de novo assembly). Taken together, our results provide guidance for researchers to select the most suitable tool to conduct the mtDNA analyses from HTS data.
•Assessment of the use of sample pooling in retrospective samples.•A pool size of five samples had the highest sensitivity.•Pooling prospective samples identified positive cases with cycle thresholds ...ranging from 16.7 to 39.4.
Limited testing capacity has characterized the ongoing coronavirus disease 2019 (COVID-19) pandemic in Spain, hampering timely control of outbreaks and opportunities to reduce the escalation of community transmission. This study investigated the potential to use sample pooling, followed by one-step retrotranscription and real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR) to increase testing capacity for severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2).
Various pool sizes (five, 10 and 15 samples) were evaluated prior to RNA extraction followed by standard RT-qPCR for the diagnosis of COVID-19. The pool size achieving reproducible results with individual sample testing was subsequently used to assess nasopharyngeal samples in a tertiary hospital in August 2020.
A pool size of five samples had higher sensitivity compared with pool sizes of 10 and 15 samples, showing a mean cycle threshold (Ct) shift of 3.5 standard deviation (SD) 2.2 between the pooled test and positive samples in the pool. Next, a pool size of five was used to test a total of 895 pools (4475 prospective samples) using two different RT-qPCR kits. The Real Accurate Quadruplex corona-plus PCR Kit (PathoFinder) reported the lowest mean Ct shift 2.2 (SD 2.4) between the pool and individual samples. This strategy enables detection of individual positive samples in positive pools with Ct of 16.7–39.4.
Grouping samples into pools of five for RT-qPCR resulted in an increase in SARS-CoV-2 testing capacity with minimal loss of sensitivity compared with testing each sample individually.
Despite asthma has a considerable genetic component, an important proportion of genetic risks remain unknown, especially for non-European populations. Canary Islanders have the largest African ...genetic ancestry observed among Southwestern Europeans and the highest asthma prevalence in Spain. Here we examined broad chromosomal regions previously associated with an excess of African genetic ancestry in Canary Islanders, with the aim of identifying novel risk variants associated with asthma susceptibility. In a two-stage cases-control study, we revealed a variant within HLA-DQB1 significantly associated with asthma risk (rs1049213, meta-analysis p = 1.30 × 10
, OR 95% CI = 1.74 1.41-2.13) previously associated with asthma and broad allergic phenotype. Subsequent fine-mapping analyses of classical HLA alleles revealed a novel allele significantly associated with asthma protection (HLA-DQA1*01:02, meta-analysis p = 3.98 × 10
, OR 95% CI = 0.64 0.50-0.82) that had been linked to infectious and autoimmune diseases, and peanut allergy. HLA haplotype analyses revealed a novel haplotype DQA1*01:02-DQB1*06:04 conferring asthma protection (meta-analysis p = 4.71 × 10
, OR 95% CI = 0.47 0.29- 0.73).
Diagnosis of Wilson disease (WD) is difficult and, as early detection may prevent all symptoms, it is essential to know the exact prevalence to evaluate the cost-efficacy of a screening program. As ...the number of WD patients was high in our population, we wished to estimate prevalence by determining the carrier frequency for clinically relevant ATP7B mutations.
To estimate prevalence, screening for the most prevalent mutation was performed in 1661 individuals with ancestry in Gran Canaria, and the frequency of other mutations was estimated from patient records. Alternatively, ATP7B mutations were detected from exomes and genomes from 851 individuals with Canarian ancestry, 236 from Gran Canaria, and a public Spanish exome database.
Estimated carrier frequencies in Gran Canaria ranged from 1 in 20 to 28, depending on the method used, resulting in prevalences of 1 case per 1547 to 3140 inhabitants. Alternatively, the estimated affected frequencies were 1 in 5985 to 7980 and 1 in 6278 to 16,510 in the archipelago or mainland Spain respectively.
The number of carriers predicts much higher prevalences than reported, suggesting that WD is underdiagnosed; specific mutations may remain unnoticed due to low penetrance or no signs of disease at all; regional prevalence rather than national prevalence should be considered in cost-efficacy models to approach preventive screening in the asymptomatic population and genetic screening strategies will have to deal with the genetic heterogeneity of ATP7B in the general population and in patients.
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