Contrasting somatic mutation patterns in aging human neurons and oligodendrocytes.

Characterizing somatic mutations in the brain is important for disentangling the complex mechanisms of aging, yet little is known about mutational patterns in different brain cell types. Here, we performed whole-genome sequencing (WGS) of 86 single oligodendrocytes, 20 mixed glia, and 56 single neurons from neurotypical individuals spanning 0.4-104 years of age and identified >92,000 somatic single-nucleotide variants (sSNVs) and small insertions/deletions (indels). Although both cell types accumulate somatic mutations linearly with age, oligodendrocytes accumulated sSNVs 81% faster than neurons and indels 28% slower than neurons. Correlation of mutations with single-nucleus RNA profiles and chromatin accessibility from the same brains revealed that oligodendrocyte mutations are enriched in inactive genomic regions and are distributed across the genome similarly to mutations in brain cancers. In contrast, neuronal mutations are enriched in open, transcriptionally active chromatin. These stark differences suggest an assortment of active mutagenic processes in oligodendrocytes and neurons.

Antagonistic Inflammatory Phenotypes Dictate Tumor Fate and Response to Immune Checkpoint Blockade.

Inflammation can support or restrain cancer progression and the response to therapy. Here, we searched for primary regulators of cancer-inhibitory inflammation through deep profiling of inflammatory tumor microenvironments (TMEs) linked to immune-dependent control in mice. We found that early intratumoral accumulation of interferon gamma (IFN-γ)-producing natural killer (NK) cells induced a profound remodeling of the TME and unleashed cytotoxic T cell (CTL)-mediated tumor eradication. Mechanistically, tumor-derived prostaglandin E2 (PGE2) acted selectively on EP2 and EP4 receptors on NK cells, hampered the TME switch, and enabled immune evasion. Analysis of patient datasets across human cancers revealed distinct inflammatory TME phenotypes resembling those associated with cancer immune control versus escape in mice. This allowed us to generate a gene-expression signature that integrated opposing inflammatory factors and predicted patient survival and response to immune checkpoint blockade. Our findings identify features of the tumor inflammatory milieu associated with immune control of cancer and establish a strategy to predict immunotherapy outcomes.

Recommendations for accelerating open preprint peer review to improve the culture of science.

Peer review is an important part of the scientific process, but traditional peer review at journals is coming under increased scrutiny for its inefficiency and lack of transparency. As preprints become more widely used and accepted, they raise the possibility of rethinking the peer-review process. Preprints are enabling new forms of peer review that have the potential to be more thorough, inclusive, and collegial than traditional journal peer review, and to thus fundamentally shift the culture of peer review toward constructive collaboration. In this Consensus View, we make a call to action to stakeholders in the community to accelerate the growing momentum of preprint sharing and provide recommendations to empower researchers to provide open and constructive peer review for preprints.

Reversal of C9orf72 mutation-induced transcriptional dysregulation and pathology in cultured human neurons by allele-specific excision.

Efforts to genetically reverse C9orf72 pathology have been hampered by our incomplete understanding of the regulation of this complex locus. We generated five different genomic excisions at the C9orf72 locus in a patient-derived induced pluripotent stem cell (iPSC) line and a non-diseased wild-type (WT) line (11 total isogenic lines), and examined gene expression and pathological hallmarks of C9 frontotemporal dementia/amyotrophic lateral sclerosis in motor neurons differentiated from these lines. Comparing the excisions in these isogenic series removed the confounding effects of different genomic backgrounds and allowed us to probe the effects of specific genomic changes. A coding single nucleotide polymorphism in the patient cell line allowed us to distinguish transcripts from the normal vs. mutant allele. Using digital droplet PCR (ddPCR), we determined that transcription from the mutant allele is upregulated at least 10-fold, and that sense transcription is independently regulated from each allele. Surprisingly, excision of the WT allele increased pathologic dipeptide repeat poly-GP expression from the mutant allele. Importantly, a single allele was sufficient to supply a normal amount of protein, suggesting that the C9orf72 gene is haplo-sufficient in induced motor neurons. Excision of the mutant repeat expansion reverted all pathology (RNA abnormalities, dipeptide repeat production, and TDP-43 pathology) and improved electrophysiological function, whereas silencing sense expression did not eliminate all dipeptide repeat proteins, presumably because of the antisense expression. These data increase our understanding of C9orf72 gene regulation and inform gene therapy approaches, including antisense oligonucleotides (ASOs) and CRISPR gene editing.

A novel ilarvirus protein CP-RT is expressed via stop codon readthrough and suppresses RDR6-dependent RNA silencing.

Ilarviruses are a relatively understudied but important group of plant RNA viruses that includes a number of crop pathogens. Their genomes comprise three RNA segments encoding two replicase subunits, movement protein, coat protein (CP), and (in some ilarvirus subgroups) a protein that suppresses RNA silencing. Here we report that, in many ilarviruses, RNA3 encodes an additional protein (termed CP-RT) as a result of ribosomal readthrough of the CP stop codon into a short downstream readthrough (RT) ORF. Using asparagus virus 2 as a model, we find that CP-RT is expressed in planta where it functions as a weak suppressor of RNA silencing. CP-RT expression is essential for persistent systemic infection in leaves and shoot apical meristem. CP-RT function is dependent on a putative zinc-finger motif within RT. Replacing the asparagus virus 2 RT with the RT of an ilarvirus from a different subgroup restored the ability to establish persistent infection. These findings open up a new avenue for research on ilarvirus silencing suppression, persistent meristem invasion and vertical transmission.

Transitions in development - an interview with Rashmi Priya.

Rashmi Priya is a Group Leader at The Francis Crick Institute in London, UK. Her research combines genetic, cell biological and biophysical approaches to understand the complex morphogenetic events of organogenesis, using the zebrafish heart as a model system. We met Rashmi at the Crick to learn how she got started as a researcher, and to discuss the challenges of starting a lab in the middle of a global pandemic.

An interview with Debby Silver.

Debby Silver is an Associate Professor at Duke University Medical Center and the Duke Institute for Brain Sciences, in Durham, North Carolina. Debby's research focusses on mammalian cortical neurogenesis, and on how RNA metabolism controls cell behaviors in the developing brain. Earlier this year, Debby became an Academic Editor at Development, and we met over Zoom to discuss her career path and research interests, as well as her motivation for joining the Development team.

Promotion of order Bunyavirales to class Bunyaviricetes to accommodate a rapidly increasing number of related polyploviricotine viruses.

Prior to 2017, the family Bunyaviridae included five genera of arthropod and rodent viruses with tri-segmented negative-sense RNA genomes related to the Bunyamwera virus. In 2017, the International Committee on Taxonomy of Viruses (ICTV) promoted the family to order Bunyavirales and subsequently greatly expanded its composition by adding multiple families for non-segmented to polysegmented viruses of animals, fungi, plants, and protists. The continued and accelerated discovery of bunyavirals highlighted that an order would not suffice to depict the evolutionary relationships of these viruses. Thus, in April 2024, the order was promoted to class Bunyaviricetes. This class currently includes two major orders, Elliovirales (Cruliviridae, Fimoviridae, Hantaviridae, Peribunyaviridae, Phasmaviridae, Tospoviridae, and Tulasviridae) and Hareavirales (Arenaviridae, Discoviridae, Konkoviridae, Leishbuviridae, Mypoviridae, Nairoviridae, Phenuiviridae, and Wupedeviridae), for hundreds of viruses, many of which are pathogenic for humans and other animals, plants, and fungi.

Identification of RNA Virus-Derived RdRp Sequences in Publicly Available Transcriptomic Data Sets.

RNA viruses are abundant and highly diverse and infect all or most eukaryotic organisms. However, only a tiny fraction of the number and diversity of RNA virus species have been catalogued. To cost-effectively expand the diversity of known RNA virus sequences, we mined publicly available transcriptomic data sets. We developed 77 family-level Hidden Markov Model profiles for the viral RNA-dependent RNA polymerase (RdRp)-the only universal "hallmark" gene of RNA viruses. By using these to search the National Center for Biotechnology Information Transcriptome Shotgun Assembly database, we identified 5,867 contigs encoding RNA virus RdRps or fragments thereof and analyzed their diversity, taxonomic classification, phylogeny, and host associations. Our study expands the known diversity of RNA viruses, and the 77 curated RdRp Profile Hidden Markov Models provide a useful resource for the virus discovery community.

Correction: pUL21 is a viral phosphatase adaptor that promotes herpes simplex virus replication and spread.

[This corrects the article DOI: 10.1371/journal.ppat.1009824.].

Moving an exercise referral scheme to remote delivery during the Covid-19 pandemic: an observational study examining the impact on uptake, adherence, and costs.

BACKGROUND: Exercise Referral Schemes (ERSs) have been implemented across Western nations to stimulate an increase in adult physical activity but evidence of their effectiveness and cost-effectiveness is equivocal. Poor ERS uptake and adherence can have a negative impact on effectiveness and cost-effectiveness and, if patterned by socio-demographic factors, can also introduce or widen health inequalities. Different modes of ERS delivery have the potential to reduce costs and enhance uptake and adherence. The primary aim of this study was to examine the effect of different programmes of ERS delivery on scheme uptake and adherence. Secondary aims were to examine the effect of socio-demographic factors on scheme uptake and adherence, and the impact of delivery mode on the expected resource and corresponding costs of delivering core parts of the programme. METHODS: This was an observational cohort study with cost analysis. Routine monitoring data covering a three-year period (2019-2021) from one large UK ERS (number of patients = 28,917) were analysed. During this period three different programmes of delivery were operated in succession: standard (all sessions delivered face-to-face at a designated physical location), hybrid (sessions initially delivered face-to-face and then switched to remote delivery in response to the Covid-19 pandemic), and modified (sessions delivered face-to-face, remotely, or a combination of the two, as determined on a case-by-case basis according to Covid-19 risk and personal preferences). Multi-level binary logistic and linear regression were performed to examine the effect of programme of delivery and socio-demographic characteristics on uptake and adherence. Cost data were sourced from regional-level coordinators and through NERS audits supplied by national-level NERS managers and summarised using descriptive statistics. RESULTS: There was no effect of programme of delivery on scheme uptake. In comparison to those on the standard programme (who attended a mean of 23.1 exercise sessions) those on the modified programme had higher adherence (mean attendance of 25.7 sessions) while those on the hybrid programme had lower adherence (mean attendance of 19.4 sessions). Being older, or coming from an area of lower deprivation, increased the likelihood of uptake and adherence. Being female increased the chance of uptake but was associated with lower adherence. Patients referred to the programme from secondary care were more likely to take up the programme than those referred from primary care for prevention purposes, however their attendance at exercise sessions was lower. The estimated cost per person for face-to-face delivery of a typical 16-week cycle of the scheme was £65.42. The same cycle of the scheme delivered virtually (outside of a pandemic context) was estimated to cost £201.71 per person. CONCLUSIONS: This study contributes new evidence concerning the effect of programme of delivery on ERS uptake and adherence and strengthens existing evidence concerning the effect of socio-economic factors. The findings direct the attention of ERS providers towards specific patient sub-groups who, if inequalities are to be addressed, require additional intervention to support uptake and adherence. At a time when providers may be considering alternative programmes of delivery, these findings challenge expectations that implementing virtual delivery will necessarily lead to cost savings.

The Impact of Morbidities Following Pediatric Cardiac Surgery on Family Functioning and Parent Quality of Life.

We previously selected and defined nine important post-operative morbidities linked to paediatric cardiac surgery, and prospectively measured their incidence following 3090 consecutive operations. Our aim was to study the impact of these morbidities on family functioning and parental quality of life over 6 months in a subset of cases. As part of a prospective case matched study in five of the ten children's cardiac centers in the UK, we compared outcomes for parents of children who had a 'single morbidity', 'multiple morbidities', 'extracorporeal life support (ECLS)' or 'no morbidity'. Outcomes were evaluated using the PedsQL Family impact module (FIM) at 6 weeks and 6 months post-surgery. Outcomes were modelled using mixed effects regression, with adjustment for case mix and clustering within centers. We recruited 340 patients with morbidity (60% of eligible patients) and 326 with no morbidity over 21 months. In comparison to the reference group of 'no morbidity', after adjustment for case mix, at 6 weeks parent health-related quality of life (HRQoL) and total FIM sores were lower (worse) only for ECLS (p < 0.005), although a higher proportion of parents in both the ECLS and multi-morbidity groups had low/very low scores (p < .05). At 6 months, parent outcomes had improved for all groups but parent HRQoL and total score for ECLS remained lower than the 'no morbidity' group (p < .05) and a higher proportion of families had low or very low scores in the ECLS (70%) group (p < .01). Post-operative morbidities impact parent HRQoL and aspects of family functioning early after surgery, with this impact lessening by 6 months. Families of children who experience post-operative morbidities should be offered timely psychological support.

Sensitivity of Meloidogyne incognita and Rotylenchulus reniformis to cyclobutrifluram.

Cyclobutrifluram, a succinate dehydrogenase inhibitor fungicide, is being evaluated as a seed-applied nematicide in cotton and soybean to manage plant-parasitic nematodes. Currently, there is no information on the toxicity, ovicidal activity, nematode recovery, or effects on nematode infection for Meloidogyne incognita or Rotylenchulus reniformis after exposure to low concentrations of cyclobutrifluram. Nematode toxicity assays were performed in aqueous solutions of cyclobutrifluram, while root infection assays were conducted on tomato. Nematode paralysis was observed after 2 h of exposure to 0.5 µg/ml cyclobutrifluram for both nematode species. Based on an assay of nematode motility, the 2-hr EC50 value for M. incognita and R. reniformis was 0.48 and 1.07 µg/ml, respectively. In a comparable assay with a similar nematicide, continuous exposure to 0.5 µg/ml cyclobutrifluram for 24 h resulted in at least 45% more immotile nematodes for both species compared to those treated with 0.5 µg/ml fluopyram. Continuous exposure to concentrations >1.0 µg/ml suppressed hatching for both species compared to the water control. Nematode recovery from paralysis was greater than 80% for M. incognita and R. reniformis 24 h after nematodes were rinsed and removed from a 1-h treatment to their respective 2-hr EC50 concentrations. Nematode infection of tomato roots was reduced following a 1-h treatment with aqueous solutions of cyclobutrifluram, ranging from 0.12 to 0.48 µg/ml for M. incognita and 0.27 to 1.07 µg/ml for R. reniformis. Overall, the toxicity of cyclobutrifluram to these nematode species was greater than that of fluopyram and although the effects of cyclobutrifluram were reversible, low concentrations were effective at reducing the ability of both nematodes to infect tomato roots.

Prospective evaluation of acute neurological events after paediatric cardiac surgery.

OBJECTIVES: Children with CHD are at heightened risk of neurodevelopmental problems; however, the contribution of acute neurological events specifically linked to the perioperative period is unclear. AIMS: This secondary analysis aimed to quantify the incidence of acute neurological events in a UK paediatric cardiac surgery population, identify risk factors, and assess how acute neurological events impacted the early post-operative pathway. METHODS: Post-operative data were collected prospectively on 3090 consecutive cardiac surgeries between October 2015 and June 2017 in 5 centres. The primary outcome of analysis was acute neurological event, with secondary outcomes of 6-month survival and post-operative length of stay. Patient and procedure-related variables were described, and risk factors were statistically explored with logistic regression. RESULTS: Incidence of acute neurological events after paediatric cardiac surgery in our population occurred in 66 of 3090 (2.1%) consecutive cardiac operations. 52 events occurred with other morbidities including renal failure (21), re-operation (20), cardiac arrest (20), and extracorporeal life support (18). Independent risk factors for occurrence of acute neurological events were CHD complexity 1.9 (1.1-3.2), p = 0.025, longer operation times 2.7 (1.6-4.8), p < 0.0001, and urgent surgery 3.4 (1.8-6.3), p < 0.0001. Unadjusted comparison found that acute neurological event was linked to prolonged post-operative hospital stay (median 35 versus 9 days) and poorer 6-month survival (OR 13.0, 95% CI 7.2-23.8). CONCLUSION: Ascertainment of acute neurological events relates to local measurement policies and was rare in our population. The occurrence of acute neurological events remains a suitable post-operative metric to follow for quality assurance purposes.

Reprint of: Implementation of depression management by ambulatory care pharmacists in the primary care setting.

BACKGROUND: In the United States, depression is one of the most common mental health disorders. Ambulatory care pharmacists play a critical role in assisting with medication and dosage selection, identifying and managing drug interactions and adverse effects, and increasing medication adherence. Existing data on depression management by ambulatory care pharmacists trained in primary care is limited and outdated. OBJECTIVES: This study provides insight into current practices for depression management by primary care pharmacy specialists within an academic health center and how pharmacist interventions may impact functional outcomes of depression. METHODS: This single-center, retrospective study analyzed 27 patients with a primary care physician within the health system who were seen by an ambulatory care pharmacist for depression. Subjects were excluded if they were under 18 years old, pregnant, or had a diagnosis of bipolar disorder, schizophrenia, schizoaffective disorder, or dementia. The primary outcome was characterization of pharmacist interventions for treatment of depression. Secondary outcomes included change in depressive symptoms, as measured by the patient health questionnaire (PHQ), characterization of adverse effects correlated with medications for depression, and utilization of pharmacogenomics testing and results. RESULTS: Of 27 patients seen by a pharmacist for depression management, 38 total interventions were made, with an average of 1.77 interventions per patient. The most common intervention was new medication initiation (32%). Average PHQ-9 scores dropped from 14.9 to 7.3 twelve weeks following the initial pharmacist visit. Only 6 patients reported adverse effects to a current antidepressant during their visit with the pharmacist, and only 2 of these cases warranted a change in therapy. Ten patients obtained pharmacogenomic testing with pharmacist facilitation. CONCLUSION: Pharmacists in the primary care setting are positioned to be an additional resource for depression management and can offer a wide variety of interventions to improve patient health.

Implementation of depression management by ambulatory care pharmacists in the primary care setting.

BACKGROUND: In the United States, depression is one of the most common mental health disorders. Ambulatory care pharmacists play a critical role in assisting with medication and dosage selection, identifying and managing drug interactions and adverse effects, and increasing medication adherence. Existing data on depression management by ambulatory care pharmacists trained in primary care is limited and outdated. OBJECTIVES: This study provides insight into current practices for depression management by primary care pharmacy specialists within an academic health center and how pharmacist interventions may impact functional outcomes of depression. METHODS: This single-center, retrospective study analyzed 27 patients with a primary care physician within the health system who were seen by an ambulatory care pharmacist for depression. Subjects were excluded if they were under 18 years old, pregnant, or had a diagnosis of bipolar disorder, schizophrenia, schizoaffective disorder, or dementia. The primary outcome was characterization of pharmacist interventions for treatment of depression. Secondary outcomes included change in depressive symptoms, as measured by the patient health questionnaire (PHQ), characterization of adverse effects correlated with medications for depression, and utilization of pharmacogenomics testing and results. RESULTS: Of 27 patients seen by a pharmacist for depression management, 38 total interventions were made, with an average of 1.77 interventions per patient. The most common intervention was new medication initiation (32%). Average PHQ-9 scores dropped from 14.9 to 7.3 twelve weeks following the initial pharmacist visit. Only 6 patients reported adverse effects to a current antidepressant during their visit with the pharmacist, and only 2 of these cases warranted a change in therapy. Ten patients obtained pharmacogenomic testing with pharmacist facilitation. CONCLUSION: Pharmacists in the primary care setting are positioned to be an additional resource for depression management and can offer a wide variety of interventions to improve patient health.

Characteristics of Human Immunodeficiency Virus (HIV) Seroconversions in a Large Prospective Implementation Cohort Study of Oral HIV Preexposure Prophylaxis in Men Who Have Sex with Men (EPIC-NSW).

BACKGROUND: Most human immunodeficiency virus (HIV) seroconversions in people who have initiated preexposure prophylaxis (PrEP) occur in the context of insufficient adherence. We describe participants who seroconverted after being dispensed PrEP in a large PrEP implementation study in Australia. METHODS: Expanded PrEP Implementation in Communities in New South Wales was an implementation study of daily oral PrEP in individuals aged ≥18 years at high risk for acquiring HIV. HIV seroconversions were defined as a positive HIV test by either antigen, antibody, or detectable HIV viral load after enrollment. Insufficient adherence, measured by dispensing logs or participant self-report, was defined as <4 PrEP doses per week. RESULTS: A total of 9596 participants were enrolled and dispensed PrEP between 1 March 2016 and 30 April 2018; 30 were diagnosed with HIV by 31 March 2019. The median (interquartile range [IQR]) age was 31 (25-38) years, all identified as male, 29 (97%) identified as gay or homosexual, and 20 (69%) lived in a postcode with a low concentration of gay male residents. The median (IQR) days from first PrEP dispensing to diagnosis was 409 (347-656). There was no evidence that participants who seroconverted had been sufficiently adherent to PrEP. Nineteen (63%) participants who seroconverted were diagnosed with chlamydia, gonorrhoea, syphilis, or new hepatitis C infection. One participant had resistance to emtricitabine (M184V mutation) at diagnosis. CONCLUSIONS: Participants who seroconverted were insufficiently adherent to PrEP despite being at high risk for acquiring HIV. Understanding the reasons for poor PrEP adherence in individuals who subsequently acquire HIV is critical to improving PrEP effectiveness.

Cryo-annealing of Photoreduced CdS Quantum Dot-Nitrogenase MoFe Protein Complexes Reveals the Kinetic Stability of the E4(2N2H) Intermediate.

A critical step in the mechanism of N2 reduction to 2NH3 catalyzed by the enzyme nitrogenase is the reaction of the four-electron/four-proton reduced intermediate state of the active-site FeMo-cofactor (E4(4H)). This state is a junction in the catalytic mechanism, either relaxing by the reaction of a metal bound Fe-hydride with a proton forming H2 or going forward with N2 binding coupled to the reductive elimination (re) of two Fe-hydrides as H2 to form the E4(2N2H) state. E4(2N2H) can relax to E4(4H) by the oxidative addition (oa) of H2 and release of N2 or can be further reduced in a series of catalytic steps to release 2NH3. If the H2 re/oa mechanism is correct, it requires that oa of H2 be associative with E4(2N2H). In this report, we have taken advantage of CdS quantum dots in complex with MoFe protein to achieve photodriven electron delivery in the frozen state, with cryo-annealing in the dark, to reveal details of the E-state species and to test the stability of E4(2N2H). Illumination of frozen CdS:MoFe protein complexes led to formation of a population of reduced intermediates. Electron paramagnetic resonance spectroscopy identified E-state signals including E2 and E4(2N2H), as well as signals suggesting the formation of E6 or E8. It is shown that in the frozen state when pN2 is much greater than pH2, the E4(2N2H) state is kinetically stable, with very limited forward or reverse reaction rates. These results establish that the oa of H2 to the E4(2N2H) state follows an associative reaction mechanism.

ICTV Virus Taxonomy Profile: Discoviridae 2023.

Discoviridae is a family of negative-sense RNA viruses with genomes of 6.2-9.7 kb that have been associated with fungi and stramenopiles. The discovirid genome consists of three monocistronic RNA segments with open reading frames (ORFs) that encode a nucleoprotein (NP), a nonstructural protein (Ns), and a large (L) protein containing an RNA-directed RNA polymerase (RdRP) domain. This is a summary of the International Committee on Taxonomy of Viruses (ICTV) Report on the family Discoviridae, which is available at ictv.global/report/discoviridae.

ICTV Virus Taxonomy Profile: Mypoviridae 2023.

Mypoviridae is a family of negative-sense RNA viruses with genomes of about 16.0 kb that have been found in myriapods. The mypovirid genome consists of three monocistronic RNA segments that encode a nucleoprotein (NP), a glycoprotein (GP), and a large (L) protein containing an RNA-directed RNA polymerase (RdRP) domain. This is a summary of the International Committee on Taxonomy of Viruses (ICTV) Report on the family Mypoviridae, which is available at: ictv.global/report/mypoviridae.