A Single-Cell Atlas of In Vivo Mammalian Chromatin Accessibility.

We applied a combinatorial indexing assay, sci-ATAC-seq, to profile genome-wide chromatin accessibility in ∼100,000 single cells from 13 adult mouse tissues. We identify 85 distinct patterns of chromatin accessibility, most of which can be assigned to cell types, and ∼400,000 differentially accessible elements. We use these data to link regulatory elements to their target genes, to define the transcription factor grammar specifying each cell type, and to discover in vivo correlates of heterogeneity in accessibility within cell types. We develop a technique for mapping single cell gene expression data to single-cell chromatin accessibility data, facilitating the comparison of atlases. By intersecting mouse chromatin accessibility with human genome-wide association summary statistics, we identify cell-type-specific enrichments of the heritability signal for hundreds of complex traits. These data define the in vivo landscape of the regulatory genome for common mammalian cell types at single-cell resolution.

Photosynthate transfer from an autotrophic orchid to conspecific heterotrophic protocorms through a common mycorrhizal network.

The minute 'dust seeds' of some terrestrial orchids preferentially germinate and develop as mycoheterotrophic protocorms near conspecific adult plants. Here we test the hypothesis that mycorrhizal mycelial connections provide a direct pathway for transfer of recent photosynthate from conspecific green orchids to achlorophyllous protocorms. Mycelial networks of Ceratobasidium cornigerum connecting green Dactylorhiza fuchsii plants with developing achlorophyllous protocorms of the same species were established on oatmeal or water agar before the shoots of green plants were exposed to 14CO2. After incubation for 48 h, the pattern of distribution of fixed carbon was visualised in intact entire autotrophic/protocorm systems using digital autoradiography and quantified in protocorms by liquid scintillation counting. Both methods of analysis revealed accumulation of 14C above background levels in protocorms, confirming that autotrophic plants supply carbon to juveniles via common mycorrhizal networks. Despite some accumulation of plant-fixed carbon in the fungal mycelium grown on oatmeal agar, a greater amount of carbon was transferred to protocorms growing on water agar, indicating that the polarity of transfer may be influenced by sink strength. We suggest this transfer pathway may contribute significantly to the pattern and processes determining localised orchid establishment in nature, and that 'parental nurture' via common mycelial networks may be involved in these processes.

Condensin II mutation causes T-cell lymphoma through tissue-specific genome instability.

Chromosomal instability is a hallmark of cancer, but mitotic regulators are rarely mutated in tumors. Mutations in the condensin complexes, which restructure chromosomes to facilitate segregation during mitosis, are significantly enriched in cancer genomes, but experimental evidence implicating condensin dysfunction in tumorigenesis is lacking. We report that mice inheriting missense mutations in a condensin II subunit (Caph2nes) develop T-cell lymphoma. Before tumors develop, we found that the same Caph2 mutation impairs ploidy maintenance to a different extent in different hematopoietic cell types, with ploidy most severely perturbed at the CD4+CD8+ T-cell stage from which tumors initiate. Premalignant CD4+CD8+ T cells show persistent catenations during chromosome segregation, triggering DNA damage in diploid daughter cells and elevated ploidy. Genome sequencing revealed that Caph2 single-mutant tumors are near diploid but carry deletions spanning tumor suppressor genes, whereas P53 inactivation allowed Caph2 mutant cells with whole-chromosome gains and structural rearrangements to form highly aggressive disease. Together, our data challenge the view that mitotic chromosome formation is an invariant process during development and provide evidence that defective mitotic chromosome structure can promote tumorigenesis.

Prevalence and Risk Factors of Constipation in Trauma Patients: A Prospective Cohort Study.

BACKGROUND: Hospitalized patients are well described as having a high prevalence of constipation. While the risks associated with constipation in trauma patients are well known, the prevalence rate is not. OBJECTIVE: This study aims to measure the prevalence of constipation and associated risk factors in trauma patients. METHODS: This study is a single-center analytic cross-sectional study on constipation in hospitalized trauma patients aged 18-65 years, admitted from January 2021 to July 2021 to the trauma service at The Royal Melbourne Hospital, a Level I major trauma and teaching hospital servicing the state of Victoria, Australia. Exclusion criteria include patients with traumatic brain injury, blunt or penetrating abdominal or spinal injuries, pregnancy, and gastrointestinal comorbidities. RESULTS: A total of N = 99 patients were studied, of which n = 78 (78.8%) were male with a median (interquartile range) age of 46 years (33-58). The overall prevalence of constipation was 76%. The univariate analysis demonstrated higher constipation rates in males and patients with multisystem injuries. However, in the multivariate analysis, mode of toileting and mobility were not associated with constipation after adjusting for confounding factors. CONCLUSION: This study demonstrated a high prevalence of constipation in all trauma patients. There is a strong association between the development of constipation in patients with multisystem injuries when compared to those with single system.

Diversity of viroids infecting grapevines in the South African Vitis germplasm collection.

Seven viroid species and one putative viroid species have been reported to infect grapevine namely, hop stunt viroid (HSVd), grapevine yellow speckle viroid 1 (GYSVd-1), grapevine yellow speckle viroid 2 (GYSVd-2), Australian grapevine viroid (AGVd), Japanese grapevine viroid (JGVd), grapevine latent viroid (GLVd), and citrus exocortis viroid (CEVd), as well as a grapevine hammerhead viroid-like RNA (GHVd), so far. In this study, RNA sequence (RNA-Seq) data, from 229 Vitis accessions from the field-maintained vineyard of the South African Vitis germplasm collection, were analysed to determine the diversity of the viroids present. Five of the seven known grapevine-infecting viroids and one putative grapevine-infecting viroid species were very commonly found, with 214 of the 229 samples containing at least one viroid species. HSVd, GYSVd-1, GYSVd-2, AGVd, and JGVd, as well as GHVd, were identified in the RNA-Seq data of the samples and confirmed using RT-PCR and Sanger sequencing. The HSVd sequences indicated the presence of two variants, with one showing multiple nucleotide insertions. AGVd and GYSVd-2 did not display significant sequence diversity, confirming past international studies. GYSVd-1 occurs as four major variants worldwide and representatives of all four variants were identified in this vineyard. This is the first report on the diversity of viroids infecting grapevine in South Africa and the first report of JGVd outside of Japan and GHVd in South Africa. Further studies are needed to fully assess the population and to identify potentially new viroid species.

Biases in the collection of blood alcohol data for adult major trauma patients in Victoria, Australia.

BACKGROUND: In-hospital alcohol testing provides an opportunity to implement prevention strategies for patients with high risk of experiencing repeated alcohol-related injuries. However, barriers to alcohol testing in emergency settings can prevent patients from being tested. In this study, we aimed to understand potential biases in current data on the completion of blood alcohol tests for major trauma patients at hospitals in Victoria, Australia. METHODS: Victorian State Trauma Registry data on all adult major trauma patients from 1 January 2018 to 31 December 2021 were used. Characteristics associated with having a blood alcohol test recorded in the registry were assessed using logistic regression models. RESULTS: This study included 14 221 major trauma patients, of which 4563 (32.1%) had a blood alcohol test recorded. Having a blood alcohol test completed was significantly associated with age, socioeconomic disadvantage level, preferred language, having pre-existing mental health or substance use conditions, smoking status, presenting during times associated with heavy community alcohol consumption, injury cause and intent, and Glasgow Coma Scale scores (p<0.05). Restricting analyses to patients from a trauma centre where blood alcohol testing was part of routine clinical care mitigated most biases. However, relative to patients injured while driving a motor vehicle/motorcycle, lower odds of testing were still observed for patients with injuries from flames/scalds/contact burns (adjusted OR (aOR)=0.33, 95% CI 0.18 to 0.61) and low falls (aOR=0.17, 95% CI 0.12 to 0.25). Higher odds of testing were associated with pre-existing mental health (aOR=1.39, 95% CI 1.02 to 1.89) or substance use conditions (aOR=2.33, 95% CI to 1.47-3.70), and living in a more disadvantaged area (most disadvantaged quintile relative to least disadvantaged quintile: aOR=2.30, 95% CI 1.52 to 3.48). CONCLUSION: Biases in the collection of blood alcohol data likely impact the surveillance of alcohol-related injuries. Routine alcohol testing after major trauma is needed to accurately inform epidemiology and the subsequent implementation of strategies for reducing alcohol-related injuries.

The E3 ubiquitin ligase activity of RING1B is not essential for early mouse development.

Polycomb-repressive complex 1 (PRC1) and PRC2 maintain repression at many developmental genes in mouse embryonic stem cells and are required for early development. However, it is still unclear how they are targeted and how they function. We show that the ability of RING1B, a core component of PRC1, to ubiquitinate histone H2A is dispensable for early mouse embryonic development and much of the gene repression activity of PRC1. Our data support a model in which PRC1 and PRC2 reinforce each other's binding but suggest that the key functions of PRC1 lie beyond the enzymatic capabilities of RING1B.

Inorganic phosphorus nutrition in green-leaved terrestrial orchid seedlings.

BACKGROUND AND AIMS: Many terrestrial orchids have an obligate dependence on their mycorrhizal associations for nutrient acquisition, particularly during germination and early seedling growth. Though important in plant growth and development, phosphorus (P) nutrition studies in mixotrophic orchids have been limited to only a few orchid species and their fungal symbionts. For the first time, we demonstrate the role of a range of fungi in the acquisition and transport of inorganic P to four phylogenetically distinct green-leaved terrestrial orchid species (Diuris magnifica, Disa bracteata, Pterostylis sanguinea and Microtis media subsp. media) that naturally grow in P-impoverished soils. METHODS: Mycorrhizal P uptake and transfer to orchids was determined and visualized using agar microcosms with a diffusion barrier between P source (33P orthophosphate) and orchid seedlings, allowing extramatrical hyphae to reach the source. KEY RESULTS: Extramatrical hyphae of the studied orchid species were effective in capturing and transporting inorganic P into the plant. Following 7 d of exposure, between 0.5 % (D. bracteata) and 47 % (D. magnifica) of the P supplied was transported to the plants (at rates between 0.001 and 0.097 fmol h-1). This experimental approach was capable of distinguishing species based on their P-foraging efficiency, and highlighted the role that fungi play in P nutrition during early seedling development. CONCLUSIONS: Our study shows that orchids occurring naturally on P-impoverished soils can obtain significant amounts of inorganic P from their mycorrhizal partners, and significantly more uptake of P supplied than previously shown in other green-leaved orchids. These results provide support for differences in mycorrhiza-mediated P acquisition between orchid species and fungal symbionts in green-leaved orchids at the seedling stage. The plant-fungus combinations of this study also provide evidence for plant-mediated niche differentiation occurring, with ecological implications in P-limited systems.

Genomic characterization of a novel potyvirus infecting Barleria repens in South Africa.

The complete RNA genome sequence of a novel member of the genus Potyvirus infecting Barleria repens has been determined. A plant showing symptoms of virus-like leaf mosaic was sampled in the Western Cape province of South Africa, and the associated virus has been tentatively named "Barleria repens mottle virus" (BaRMoV). The genome of BaRMoV consists of 9,561 nucleotides and encodes a typical potyvirus polyprotein that is 3,105 amino acids long. Pairwise comparisons showed that the BaRMoV genome shared an average of 70% nucleotide sequence identity with that of yam mosaic virus (YMV), and the corresponding polyprotein shared an average of 76.6% amino acid sequence identity with that of YMV. Phylogenetic analysis of the BaRMoV polyprotein amino acid sequence and those of other extant potyviruses confirmed the relationship between BarMoV and YMV. This is the first time that a member of the family Potyviridae has been identified in a member of the species Barleria.

Genomic characterization of grapevine viruses N and O: novel vitiviruses from South Africa.

A survey was performed on a Vitis cultivar collection in Stellenbosch, South Africa. Metaviromes were generated for each cultivar, using an RNAtag-seq workflow. Analysis of assembled contigs indicated the presence of two putatively novel members of the genus Vitivirus, provisionally named "grapevine virus N" (GVN) and "grapevine virus O" (GVO). Comparisons of amino acid sequences showed that GVN and GVO are most closely related to grapevine virus G and grapevine virus E, respectively. The incidence of these novel viruses within the sampling site was low, with GVO and GVN associated with only five and two cultivars, respectively, of the 229 sampled.

Genomic characterization of soybean blotchy mosaic virus, a cytorhabdovirus from South Africa.

Samples showing blotchy mottle symptoms were collected from soybeans in North-West province, South Africa. The assembly of high-throughput sequencing data from three samples yielded contigs of 13,426 to 13,435nt, which represent the first complete genome sequences of soybean blotchy mosaic virus (SbBMV). SbBMV shows a typical cytorhabdovirus gene organization (3'-N-P-P3-M-G-L-5'), with each putative gene product being most similar, but with only 49.1-71.1% sequence identity, to those of cucurbit cytorhabdovirus 1. Given the species demarcation thresholds for rhabdoviruses, SbBMV is thus a distinct member of the genus Cytorhabdovirus.

Genomic and phylogenetic characterization of wheat yellows virus, a novel tenuivirus infecting wheat in South Africa.

During the 2019 winter wheat season, symptoms of severe chlorosis and stunting were observed on wheat in the irrigation production areas of South Africa. RNAtag-seq data were generated for seven samples from KwaZulu-Natal province and one from Limpopo. Analysis of assembled contigs indicated the presence of a putatively novel member of the genus Tenuivirus, tentatively named "wheat yellows virus" (WhYV). The genome is made up of four segments, which are 8952, 3451, 2338, and 2045 nucleotides in length and code for a total of seven ORFs. Phylogenies of each segment (nucleotide) and the polymerase gene (amino acid), as well as amino acid sequence comparisons of each gene product, showed that WhYV is most closely related to rice stripe virus.

Stable integrant-specific differences in bimodal HIV-1 expression patterns revealed by high-throughput analysis.

HIV-1 gene expression is regulated by host and viral factors that interact with viral motifs and is influenced by proviral integration sites. Here, expression variation among integrants was followed for hundreds of individual proviral clones within polyclonal populations throughout successive rounds of virus and cultured cell replication, with limited findings using CD4+ cells from donor blood consistent with observations in immortalized cells. Tracking clonal behavior by proviral "zip codes" indicated that mutational inactivation during reverse transcription was rare, while clonal expansion and proviral expression states varied widely. By sorting for provirus expression using a GFP reporter in the nef open reading frame, distinct clone-specific variation in on/off proportions were observed that spanned three orders of magnitude. Tracking GFP phenotypes over time revealed that as cells divided, their progeny alternated between HIV transcriptional activity and non-activity. Despite these phenotypic oscillations, the overall GFP+ population within each clone was remarkably stable, with clones maintaining clone-specific equilibrium mixtures of GFP+ and GFP- cells. Integration sites were analyzed for correlations between genomic features and the epigenetic phenomena described here. Integrants inserted in the sense orientation of genes were more frequently found to be GFP negative than those in the antisense orientation, and clones with high GFP+ proportions were more distal to repressive H3K9me3 peaks than low GFP+ clones. Clones with low frequencies of GFP positivity appeared to expand more rapidly than clones for which most cells were GFP+, even though the tested proviruses were Vpr-. Thus, much of the increase in the GFP- population in these polyclonal pools over time reflected differential clonal expansion. Together, these results underscore the temporal and quantitative variability in HIV-1 gene expression among proviral clones that are conferred in the absence of metabolic or cell-type dependent variability, and shed light on cell-intrinsic layers of regulation that affect HIV-1 population dynamics.

Genomic characterization of two novel viruses infecting Barleria cristata L. from the genera Orthotospovirus and Polerovirus.

Barleria cristata L. has become naturalized in South Africa, where it is commonly used as an ornamental. In 2019, plants of B. cristata showing putative viral symptoms were collected from two locations in Gauteng, South Africa. RNAtag-seq libraries were prepared and sequenced using an Illumina HiSeq 2500 platform. De novo assembly of the resulting data revealed the presence of a novel member of the family Tospoviridae associated with the plants from both locations, and this virus was given the tentative name "barleria chlorosis-associated virus". Segments L, M, and S have lengths of 8752, 4760, and 2906 nt, respectively. Additionally, one of the samples was associated with a novel polerovirus, provisionally named "barleria polerovirus 1", with a complete genome length of 6096 nt. This is the first study to show the association of viruses with a member of the genus Barleria.

Novel viruses associated with plants of the family Amaryllidaceae in South Africa.

Nineteen samples from members of the plant genera Agapanthus, Clivia, Hippeastrum, and Scadoxus were collected from gardens in the Gauteng and Western Cape provinces of South Africa. The plants displayed highly variable symptoms of viral disease, including chlorosis, necrosis, streaking, and ringspot. RNAtag-seq was used to characterize the associated viral populations. Plants of the genus Agapanthus were found to be associated with three novel viruses from the families Caulimoviridae, Closteroviridae, and Betaflexiviridae; plants of the genus Clivia were associated with novel members of the families Potyviridae and Betaflexiviridae; and plants of the genus Scadoxus were associated with a novel member of the family Tospoviridae. Nerine latent virus was associated with plants of the genera Agapanthus, Clivia, and Hippeastrum, while hippeastrum mosaic virus was associated exclusively with a Hippeastrum cultivar.

Delayed ileostomy closure increases the odds of Clostridium difficile infection.

AIM: A diverting ileostomy is typically performed to divert intestinal contents in high-risk colorectal anastomoses. Ileostomy closure is associated with high rates of postoperative Clostridium difficile infection (CDI). Risk factors for the development of CDI are unclear; however, a correlation has been observed with delayed closure. This study aimed to assess the odds of developing CDI in patients who had a delay to reversal of ileostomy, compared to those who had no delay. METHODS: A retrospective cohort study was conducted of patients undergoing reversal of ileostomy between 2010 and 2019 at a single tertiary centre. A delay to reversal of ileostomy was defined if the procedure was performed at >365 days following the index procedure. CDI was defined as the presence of Clostridium difficile toxin associated with diarrhoea. Univariable logistic regression analysis was performed to estimate odds of CDI for each covariable, comparing patients who had a delay to reversal of ileostomy with those who did not. Multivariable logistic regression analysis was used to adjust for the potential confounding effects of covariables. RESULTS: Of 195 patients, 11 (5.6%), developed postoperative CDI. Multivariable analysis showed that delay to reversal of ileostomy was associated with a nearly 7-fold increase in odds of CDI (OR = 6.95, CI: 1.06-81.6; p-value = 0.03). CONCLUSION: A delay to reversal of ileostomy of >365 days was associated with a higher incidence of CDI postoperatively. Careful consideration should be given to the timing of reversal and appropriate preoperative counselling of patients.

Elastic-plastic properties of mesoscale electrodeposited LIGA nickel alloy films: microscopy and mechanics.

The elastic-plastic properties of mesoscale electrodeposited LIGA Ni alloy specimens are investigated as a function of specimen size, strain rate, and material composition. Two material compositions are studied: a high-strength fine-grained Ni-Fe alloy and a high-ductility coarse-grained Ni-Co alloy. The specimens have thicknesses of approximately 200 µm and gauge widths ranging from 75 µm to 700 µm. Tensile tests are conducted at strain rates of 0.001/s and 1/s using tabletop loading apparatuses and digital image correlation (DIC) for strain measurement. For each test condition, the apparent Young's modulus, yield strength, ultimate tensile strength, and strain hardening exponent and strength coefficient are extracted from the tensile tests. The true strains to failure are also assessed from fractography. Size, rate, and composition effects are discussed. For most properties, the statistical scatter represented by the standard deviation exceeds the measurement uncertainty; the notable exceptions to these observations are the apparent Young's modulus and yield strength, where large measurement uncertainties are ascribed to common experimental factors and material microplasticity.

Contrast-enhanced ultrasound assessed renal microvascular perfusion may predict postoperative renal complications following colorectal surgery.

Colorectal surgery is associated with an above-average mortality rate of approximately 15%. During surgery, maintenance of vital organ perfusion is essential in order to reduce postoperative mortality and morbidity, with renal perfusion of particular importance. Oesophageal Doppler monitors (ODM) are commonly used to try and provide accurate measures of fluid depletion during surgery; however, it is unclear to what extent they reflect organ perfusion. In addition, it is not known whether macro- and/ or microvascular perfusion indices are associated with renal complications following colorectal surgery. Thirty-two participants scheduled for colorectal surgery had three measures of macro- and microvascular renal blood flow via contrast enhanced ultrasound (CEUS), and simultaneous measures of cardiac output indices via ODM: (i) pre-operatively; (ii) intra-operatively at the mid-point of operation, and (iii) after the conclusion of surgery. The Postoperative Morbidity Survey (POMS) was used to assess postoperative complications. Intra-operatively, there was a significant correlation between renal microvascular flow (RT) and renal macrovascular flow (TTI) (ρ = 0.52; p = 0.003). Intra-operative TTI, but not RT, was associated with cardiac index (ρ = -0.50; p=0.0003). Intra-operative RT predicted increases in renal complications (OR 1.46; 95% CI 1.03-2.09) with good discrimination (C-statistic, 0.85). Complications were not predicted by TTI or ODM-derived indices. There was no relationship between RT and TTI before or after surgery. ODM measures of haemodynamic status do not correlate with renal microvascular blood flow, and as such are likely not suitable to determine vital organ perfusion. Only CEUS-derived measures of microvascular perfusion were predictive of postoperative renal complications.

Predicting gene expression in the human malaria parasite Plasmodium falciparum using histone modification, nucleosome positioning, and 3D localization features.

Empirical evidence suggests that the malaria parasite Plasmodium falciparum employs a broad range of mechanisms to regulate gene transcription throughout the organism's complex life cycle. To better understand this regulatory machinery, we assembled a rich collection of genomic and epigenomic data sets, including information about transcription factor (TF) binding motifs, patterns of covalent histone modifications, nucleosome occupancy, GC content, and global 3D genome architecture. We used these data to train machine learning models to discriminate between high-expression and low-expression genes, focusing on three distinct stages of the red blood cell phase of the Plasmodium life cycle. Our results highlight the importance of histone modifications and 3D chromatin architecture in Plasmodium transcriptional regulation and suggest that AP2 transcription factors may play a limited regulatory role, perhaps operating in conjunction with epigenetic factors.

Metaviromic analysis reveals coinfection of papaya in western Kenya with a unique strain of Moroccan watermelon mosaic virus and a novel member of the family Alphaflexiviridae.

Severe mottling symptoms were observed on Carica papaya L. in Koyonzo, Kakamega County, Kenya. Total RNA was sequenced via an RNAtag-seq workflow. Assembled contigs indicated the presence of a divergent strain of Moroccan watermelon mosaic virus (genus Potyvirus) with a complete genome length of 9,733 nt (GenBank accession no. MN418119). Additionally, the complete genome sequence of a novel member of the viral genus Allexivirus was determined (GenBank accession no. MN418120). The genome contains six open reading frames (ORFs) that show varying degrees of sequence similarity to members of the genus Allexivirus; however, it appears to lack an ORF encoding a nucleic-acid-binding homolog. The tentative name "papaya virus A" (PaVA) has been proposed for this virus.