Postpandemic rebound of adeno-associated virus type 2 (AAV2) infections temporally associated with an outbreak of unexplained severe acute hepatitis in children in the United Kingdom

Over 1000 cases of unexplained severe acute hepatitis in children have been reported to date worldwide. An association with adeno-associated virus type 2 (AAV2) infection, a human parvovirus, prompted us to investigate the epidemiology of AAV in the United Kingdom. Three hundred pediatric respiratory samples collected before (April 03, 2009–April 03, 2013) and during (April 03, 2022) the COVID-19 pandemic were obtained. Wastewater samples were collected from 50 locations in London (August 2021–March 2022). Samples were tested for AAV using real-time polymerase chain reaction followed by sequencing. Selected adenovirus (AdV)-positive samples were also sequenced. The detection frequency of AAV2 was a sevenfold higher in 2022 samples compared with 2009–2013 samples (10% vs. 1.4%) and highest in AdV-positive samples compared with negatives (10/37, 27% vs. 5/94, 5.3%, respectively). AAV2-positive samples displayed high genetic diversity. AAV2 sequences were either very low or absent in wastewater collected in 2021 but increased in January 2022 and peaked in March 2022. AAV2 was detected in children in association with AdV of species C, with a highest frequency in 2022. Our findings are consistent with the expansion of the population of children unexposed to AAV2, leading to greater spread of the virus once distancing restrictions were lifted

Attenuated Induction of the Unfolded Protein Response in Adult Human Primary Astrocytes in Response to Recurrent Low Glucose

This is the final version. Available on open access from Frontiers Media via the DOI in this record. The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found in the article/Supplementary Material.AIMS/HYPOTHESIS: Recurrent hypoglycaemia (RH) is a major side-effect of intensive insulin therapy for people with diabetes. Changes in hypoglycaemia sensing by the brain contribute to the development of impaired counterregulatory responses to and awareness of hypoglycaemia. Little is known about the intrinsic changes in human astrocytes in response to acute and recurrent low glucose (RLG) exposure. METHODS: Human primary astrocytes (HPA) were exposed to zero, one, three or four bouts of low glucose (0.1 mmol/l) for three hours per day for four days to mimic RH. On the fourth day, DNA and RNA were collected. Differential gene expression and ontology analyses were performed using DESeq2 and GOseq, respectively. DNA methylation was assessed using the Infinium MethylationEPIC BeadChip platform. RESULTS: 24 differentially expressed genes (DEGs) were detected (after correction for multiple comparisons). One bout of low glucose exposure had the largest effect on gene expression. Pathway analyses revealed that endoplasmic-reticulum (ER) stress-related genes such as HSPA5, XBP1, and MANF, involved in the unfolded protein response (UPR), were all significantly increased following low glucose (LG) exposure, which was diminished following RLG. There was little correlation between differentially methylated positions and changes in gene expression yet the number of bouts of LG exposure produced distinct methylation signatures. CONCLUSIONS/INTERPRETATION: These data suggest that exposure of human astrocytes to transient LG triggers activation of genes involved in the UPR linked to endoplasmic reticulum (ER) stress. Following RLG, the activation of UPR related genes was diminished, suggesting attenuated ER stress. This may be a consequence of a successful metabolic adaptation, as previously reported, that better preserves intracellular energy levels and a reduced necessity for the UPR.Diabetes UKJDRF postdoctoral fellowshipMedical Research Council (MRC)Wellcome TrustBiotechnology & Biological Sciences Research Council (BBSRC)Novo Nordisk UK Research FoundationMary Kinross Charitable TrustEuropean Foundation for the Study of Diabetes/Novo Nordisk Programm

Deletion 22q13.3 syndrome

The deletion 22q13.3 syndrome (deletion 22q13 syndrome or Phelan-McDermid syndrome) is a chromosome microdeletion syndrome characterized by neonatal hypotonia, global developmental delay, normal to accelerated growth, absent to severely delayed speech, and minor dysmorphic features. The deletion occurs with equal frequency in males and females and has been reported in mosaic and non-mosaic forms. Due to lack of clinical recognition and often insufficient laboratory testing, the syndrome is under-diagnosed and its true incidence remains unknown. Common physical traits include long eye lashes, large or unusual ears, relatively large hands, dysplastic toenails, full brow, dolicocephaly, full cheeks, bulbous nose, and pointed chin. Behavior is autistic-like with decreased perception of pain and habitual chewing or mouthing. The loss of 22q13.3 can result from simple deletion, translocation, ring chromosome formation and less common structural changes affecting the long arm of chromosome 22, specifically the region containing the SHANK3 gene. The diagnosis of deletion 22q13 syndrome should be considered in all cases of hypotonia of unknown etiology and in individuals with absent speech. Although the deletion can sometimes be detected by high resolution chromosome analysis, fluorescence in situ hybridization (FISH) or array comparative genomic hybridization (CGH) is recommended for confirmation. Differential diagnosis includes syndromes associated with hypotonia, developmental delay, speech delay and/or autistic-like affect (Prader-Willi, Angelman, Williams, Smith-Magenis, Fragile X, Sotos, FG, trichorhinophalangeal and velocardiofacial syndromes, autism spectrum disorders, cerebral palsy). Genetic counseling is recommended and parental laboratory studies should be considered to identify cryptic rearrangements and detect parental mosaicism. Prenatal diagnosis should be offered for future pregnancies in those families with inherited rearrangements. Individuals with deletion 22q13 should have routine examinations by the primary care physician as well as genetic evaluations with referral to specialists if neurological, gastrointestinal, renal, or other systemic problems are suspected. Affected individuals benefit from early intervention programs, intense occupational and communication therapies, adaptive exercise and sport programs, and other therapies to strengthen their muscles and increase their communication skills. No apparent life-threatening organic abnormalities accompany the diagnosis of deletion 22q13

Genetic analysis of D-xylose metabolism by endophytic yeast strains of Rhodotorula graminis and Rhodotorula mucilaginosa

Two novel endophytic yeast strains, WP1 and PTD3, isolated from within the stems of poplar (Populus) trees, were genetically characterized with respect to their xylose metabolism genes. These two strains, belonging to the species Rhodotorula graminis and R. mucilaginosa, respectively, utilize both hexose and pentose sugars, including the common plant pentose sugar, D-xylose. The xylose reductase (XYL1) and xylitol dehydrogenase (XYL2) genes were cloned and characterized. The derived amino acid sequences of xylose reductase (XR) and xylose dehydrogenase (XDH) were 32%∼41% homologous to those of Pichia stipitis and Candida. spp., two species known to utilize xylose. The derived XR and XDH sequences of WP1 and PTD3 had higher homology (73% and 69% identity) with each other. WP1 and PTD3 were grown in single sugar and mixed sugar media to analyze the XYL1 and XYL2 gene regulation mechanisms. Our results revealed that for both strains, the gene expression is induced by D-xylose, and that in PTD3 the expression was not repressed by glucose in the presence of xylose

The Gaia–ESO Survey: dynamical models of flattened, rotating globular clusters

We present a family of self-consistent axisymmetric rotating globular cluster models which are fitted to spectroscopic data for NGC 362, NGC 1851, NGC 2808, NGC 4372, NGC 5927 and NGC 6752 to provide constraints on their physical and kinematic properties, including their rotation signals. They are constructed by flattening Modified Plummer profiles, which have the same asymptotic behaviour as classical Plummer models, but can provide better fits to young clusters due to a slower turnover in the density profile. The models are in dynamical equilibrium as they depend solely on the action variables. We employ a fully Bayesian scheme to investigate the uncertainty in our model parameters (including mass-to-light ratios and inclination angles) and evaluate the Bayesian evidence ratio for rotating to non-rotating models. We find convincing levels of rotation only in NGC 2808. In the other clusters, there is just a hint of rotation (in particular, NGC 4372 and NGC 5927), as the data quality does not allow us to draw strong conclusions. Where rotation is present, we find that it is confined to the central regions, within radii of R ≤ 2rh. As part of this work, we have developed a novel q-Gaussian basis expansion of the line-of-sight velocity distributions, from which general models can be constructed via interpolation on the basis coefficients.This work was partly supported by the European Union FP7 programme through ERC grant number 320360 and by the Leverhulme Trust through grant RPG-2012-541. We acknowledge the support from INAF and Ministero dell’ Istruzione, dell’ Università’ e della Ricerca (MIUR) in the form of the grant ‘Premiale VLT 2012’

Growth disrupting mutations in epigenetic regulatory molecules are associated with abnormalities of epigenetic aging.

Germline mutations in fundamental epigenetic regulatory molecules including DNA methyltransferase 3 alpha (DNMT3A) are commonly associated with growth disorders, whereas somatic mutations are often associated with malignancy. We profiled genome-wide DNA methylation patterns in DNMT3A c.2312G > A; p.(Arg771Gln) carriers in a large Amish sibship with Tatton-Brown-Rahman syndrome (TBRS), their mosaic father, and 15 TBRS patients with distinct pathogenic de novo DNMT3A variants. This defined widespread DNA hypomethylation at specific genomic sites enriched at locations annotated as genes involved in morphogenesis, development, differentiation, and malignancy predisposition pathways. TBRS patients also displayed highly accelerated DNA methylation aging. These findings were most marked in a carrier of the AML-associated driver mutation p.Arg882Cys. Our studies additionally defined phenotype-related accelerated and decelerated epigenetic aging in two histone methyltransferase disorders: NSD1 Sotos syndrome overgrowth disorder and KMT2D Kabuki syndrome growth impairment. Together, our findings provide fundamental new insights into aberrant epigenetic mechanisms, the role of epigenetic machinery maintenance, and determinants of biological aging in these growth disorders

Novel epigenetic clock for fetal brain development predicts prenatal age for cellular stem cell models and derived neurons

Induced pluripotent stem cells (iPSCs) and their differentiated neurons (iPSC-neurons) are a widely used cellular model in the research of the central nervous system. However, it is unknown how well they capture age-associated processes, particularly given that pluripotent cells are only present during the earliest stages of mammalian development. Epigenetic clocks utilize coordinated age-associated changes in DNA methylation to make predictions that correlate strongly with chronological age. It has been shown that the induction of pluripotency rejuvenates predicted epigenetic age. As existing clocks are not optimized for the study of brain development, we developed the fetal brain clock (FBC), a bespoke epigenetic clock trained in human prenatal brain samples in order to investigate more precisely the epigenetic age of iPSCs and iPSC-neurons. The FBC was tested in two independent validation cohorts across a total of 194 samples, confirming that the FBC outperforms other established epigenetic clocks in fetal brain cohorts. We applied the FBC to DNA methylation data from iPSCs and embryonic stem cells and their derived neuronal precursor cells and neurons, finding that these cell types are epigenetically characterized as having an early fetal age. Furthermore, while differentiation from iPSCs to neurons significantly increases epigenetic age, iPSC-neurons are still predicted as being fetal. Together our findings reiterate the need to better understand the limitations of existing epigenetic clocks for answering biological research questions and highlight a limitation of iPSC-neurons as a cellular model of age-related diseases

Reversion of the ELISPOT test after treatment in Gambian tuberculosis cases

BACKGROUND: New tools are required to improve tuberculosis (TB) diagnosis and treatment, including enhanced ability to compare new treatment strategies. The ELISPOT assay uses Mycobacterium tuberculosis-specific antigens to produce a precise quantitative readout of the immune response to pathogen. We hypothesized that TB patients in The Gambia would have reduced ELISPOT counts after successful treatment. METHODS: We recruited Gambian adults with sputum smear and culture positive tuberculosis for ELISPOT assay and HIV test, and followed them up one year later to repeat testing and document treatment outcome. We used ESAT-6, CFP-10 and Purified Protein Derivative (PPD) as stimulatory antigens. We confirmed the reliability of our assay in 23 volunteers through 2 tests one week apart, comparing within and between subject variation. RESULTS: We performed an ELISPOT test at diagnosis and 12 months later in 89 patients. At recruitment, 70/85 HIV-negative patients (82%) were ESAT-6 or CFP-10 (EC) ELISPOT positive, 77 (90%) were PPD ELISPOT positive. Eighty-two cases (96%) successfully completed treatment: 44 (55%; p < 0.001) were EC ELISPOT negative at 12 months, 17 (21%; p = 0.051) were PPD ELISPOT negative. Sixty (73%) cured cases had a CFP-10 ELISPOT count decrease, 64 (78%) had an ESAT-6 ELISPOT count decrease, 58 (70%) had a PPD ELISPOT count decrease. There was a mean decline of 25, 44 and 47 SFU/2 × 10(5 )cells for CFP-10, ESAT-6 and PPD respectively (p < 0.001 for all). Three of 4 HIV positive patients were cured, all 3 underwent ELISPOT reversion; all 4 not cured subjects (3 HIV-negative, 1 HIV positive) were ESAT-6, CFP-10 and PPD ELISPOT positive at 12 months. CONCLUSION: Successful tuberculosis treatment is accompanied by a significant reduction in the M. tuberculosis-specific antigen ELISPOT count. The ELISPOT has potential as a proxy measure of TB treatment outcome. Further investigation into the decay kinetics of T-cells with treatment is warranted

Biallelic SQSTM1 mutations in early-onset, variably progressive neurodegeneration.

OBJECTIVE: To characterize clinically and molecularly an early-onset, variably progressive neurodegenerative disorder characterized by a cerebellar syndrome with severe ataxia, gaze palsy, dyskinesia, dystonia, and cognitive decline affecting 11 individuals from 3 consanguineous families. METHODS: We used whole-exome sequencing (WES) (families 1 and 2) and a combined approach based on homozygosity mapping and WES (family 3). We performed in vitro studies to explore the effect of the nontruncating SQSTM1 mutation on protein function and the effect of impaired SQSTM1 function on autophagy. We analyzed the consequences of sqstm1 down-modulation on the structural integrity of the cerebellum in vivo using zebrafish as a model. RESULTS: We identified 3 homozygous inactivating variants, including a splice site substitution (c.301+2T>A) causing aberrant transcript processing and accelerated degradation of a resulting protein lacking exon 2, as well as 2 truncating changes (c.875_876insT and c.934_936delinsTGA). We show that loss of SQSTM1 causes impaired production of ubiquitin-positive protein aggregates in response to misfolded protein stress and decelerated autophagic flux. The consequences of sqstm1 down-modulation on the structural integrity of the cerebellum in zebrafish documented a variable but reproducible phenotype characterized by cerebellum anomalies ranging from depletion of axonal connections to complete atrophy. We provide a detailed clinical characterization of the disorder; the natural history is reported for 2 siblings who have been followed up for >20 years. CONCLUSIONS: This study offers an accurate clinical characterization of this recently recognized neurodegenerative disorder caused by biallelic inactivating mutations in SQSTM1 and links this phenotype to defective selective autophagy

Conserved and highly expressed tRNA derived fragments in zebrafish

Background: Small non-coding RNAs (sncRNAs) are a class of transcripts implicated in several eukaryotic regulatory mechanisms, namely gene silencing and chromatin regulation. Despite significant progress in their identification by next generation sequencing (NGS) we are still far from understanding their full diversity and functional repertoire. Results: Here we report the identification of tRNA derived fragments (tRFs) by NGS of the sncRNA fraction of zebrafish. The tRFs identified are 18–30 nt long, are derived from specific 5′ and 3′ processing of mature tRNAs and are differentially expressed during development and in differentiated tissues, suggesting that they are likely produced by specific processing rather than random degradation of tRNAs. We further show that a highly expressed tRF (5′tRF-ProCGG) is cleaved in vitro by Dicer and has silencing ability, indicating that it can enter the RNAi pathway. A computational analysis of zebrafish tRFs shows that they are conserved among vertebrates and mining of publicly available datasets reveals that some 5′tRFs are differentially expressed in disease conditions, namely during infection and colorectal cancer. Conclusions: tRFs constitute a class of conserved regulatory RNAs in vertebrates and may be involved in mechanisms of genome regulation and in some diseases. Keywords: tRNA derived fragments, Zebrafish, Small non coding RNAs, tRNAspublishe