Direct nanopore RNA sequencing of umbra-like virus-infected plants reveals long non-coding RNAs, specific cleavage sites, D-RNAs, foldback RNAs, and temporal- and tissue-specific profiles.

The traditional view of plus (+)-strand RNA virus transcriptomes is that infected cells contain a limited variety of viral RNAs, such as full-length (+)-strand genomic RNA(s), (-)-strand replication intermediate(s), 3' co-terminal subgenomic RNA(s), and viral recombinant defective (D)-RNAs. To ascertain the full complement of viral RNAs associated with the simplest plant viruses, long-read direct RNA nanopore sequencing was used to perform transcriptomic analyses of two related umbra-like viruses: citrus yellow vein-associated virus (CY1) from citrus and CY2 from hemp. Analysis of different timepoints/tissues in CY1- and CY2-infected Nicotiana benthamiana plants and CY2-infected hemp revealed: (i) three 5' co-terminal RNAs of 281 nt, 442 nt and 671 nt, each generated by a different mechanism; (ii) D-RNA populations containing the 671 fragment at their 5'ends; (iii) many full-length genomic RNAs and D-RNAs with identical 3'end 61 nt truncations; (iv) virtually all (-)-strand reads missing 3 nt at their 3' termini; (v) (±) foldback RNAs comprising about one-third of all (-)-strand reads and (vi) a higher proportion of full-length gRNAs in roots than in leaves, suggesting that roots may be functioning as a gRNA reservoir. These findings suggest that viral transcriptomes are much more complex than previously thought.

Recovery from social isolation requires dopamine in males, but not the autism-related gene nlg3 in either sex.

Social isolation causes profound changes in social behaviour in a variety of species. However, the genetic and molecular mechanisms modulating behavioural responses to social isolation and social recovery remain to be elucidated. Here, we quantified the behavioural response of vinegar flies to social isolation using two distinct protocols (social space preference and sociability, the spontaneous tendencies to form groups). We found that social isolation increased social space and reduced sociability. These effects of social isolation were reversible and could be reduced after 3 days of group housing. Flies with a loss of function of neuroligin3 (orthologue of autism-related neuroligin genes) with known increased social space in a socially enriched environment were still able to recover from social isolation. We also show that dopamine (DA) is needed for a response to social isolation and recovery in males but not in females. Furthermore, only in males, DA levels are reduced after isolation and are not recovered after group housing. Finally, in socially enriched flies mutant for neuroligin3, DA levels are reduced in males, but not in females. We propose a model to explain how DA and neuroligin3 are involved in the behavioural response to social isolation and its recovery in a dynamic and sex-specific manner.

Expanding the Plant Virome: Umbra-Like Viruses Use Host Proteins for Movement.

Before the very recent discovery of umbra-like viruses (ULVs), the signature defining feature of all plant RNA viruses was the encoding of specialized RNA-binding movement proteins (MPs) for transiting their RNA genomes through gated plasmodesmata to establish systemic infections. The vast majority of ULVs share umbravirus-like RNA-dependent RNA polymerases and 3'-terminal structures, but they differ by not encoding cell-to-cell and long-distance MPs and by not relying on a helper virus for trans-encapsidation and plant-to-plant transmission. The recent finding that two groups of ULVs do not necessarily encode MPs is expanding our understanding of the minimum requirements for modern plant RNA viruses. ULV CY1 from citrus uses host protein PHLOEM PROTEIN 2 (PP2) for systemic movement, and related ULVs encode a capsid protein, thereby providing an explanation for the lack of helper viruses present in many ULV-infected plants. ULVs thus resemble the first viruses that infected plants, which were likely deposited from feeding organisms and would have similarly required the use of host proteins such as PP2 to exit initially infected cells.

Mitigation of TDP-43 toxic phenotype by an RGNEF fragment in amyotrophic lateral sclerosis models.

Aggregation of the RNA-binding protein TAR DNA binding protein (TDP-43) is a hallmark of TDP-proteinopathies including amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). As TDP-43 aggregation and dysregulation are causative of neuronal death, there is a special interest in targeting this protein as a therapeutic approach. Previously, we found that TDP-43 extensively co-aggregated with the dual function protein GEF (guanine exchange factor) and RNA-binding protein rho guanine nucleotide exchange factor (RGNEF) in ALS patients. Here, we show that an N-terminal fragment of RGNEF (NF242) interacts directly with the RNA recognition motifs of TDP-43 competing with RNA and that the IPT/TIG domain of NF242 is essential for this interaction. Genetic expression of NF242 in a fruit fly ALS model overexpressing TDP-43 suppressed the neuropathological phenotype increasing lifespan, abolishing motor defects and preventing neurodegeneration. Intracerebroventricular injections of AAV9/NF242 in a severe TDP-43 murine model (rNLS8) improved lifespan and motor phenotype, and decreased neuroinflammation markers. Our results demonstrate an innovative way to target TDP-43 proteinopathies using a protein fragment with a strong affinity for TDP-43 aggregates and a mechanism that includes competition with RNA sequestration, suggesting a promising therapeutic strategy for TDP-43 proteinopathies such as ALS and FTD.

Does the addition of convex uniplanar screws in hybrid constructs improve 3D surgical correction in thoracic adolescent idiopathic scoliosis posterior fusion?

Purpose: Hybrid techniques using thoracic sublaminar bands have proved their efficacy in adolescent idiopathic scoliosis posterior fusion, but clinical axial correction sometimes remained disappointing. One solution found was "the frame technique" and the second alternative was the replacement of the convex sublaminar bands by periapical uniplanar screws. The goal of this study was to compare clinical and radiological outcomes of both techniques in a consecutive cohort of adolescent idiopathic scoliosis patients. Methods: All patients undergoing primary posterior fusion for thoracic adolescent idiopathic scoliosis between January 2017 and March 2020 were included. Two groups were compared: Group 1 with thoracic sublaminar bands only and Group 2 with periapical uniplanar screws. All patients underwent standing stereoradiographs. The main frontal, sagittal, and axial (apical vertebra rotation) radiological parameters of interest were analyzed. Functional outcomes were assessed using the Scoliosis Research Society 30 score. Results: A total of 147 adolescents were included (Group 1, n = 73 and Group 2, n = 74 patients). In the frontal plane, a greater reduction index was observed in Group 2 (68% versus 62%, p < 0.001) as well as a better apical axial correction (67.8% versus 46.6%, p = 0.03). The number of thoracoplasty performed was reduced (6.7% versus 20.5%, p = 0.02) in Group 2, with a significant decrease in the rate of mechanical complication. No significant loss of correction was observed during follow-up in any of the group. Conclusion: The adjunction of convex uniplanar screws at the periapical levels improved the three-dimensional surgical correction of thoracic adolescent idiopathic scoliosis treated with hybrid constructs. Level of evidence: level III, retrospective comparative study.

Umbravirus-like RNA viruses are capable of independent systemic plant infection in the absence of encoded movement proteins.

The signature feature of all plant viruses is the encoding of movement proteins (MPs) that supports the movement of the viral genome into adjacent cells and through the vascular system. The recent discovery of umbravirus-like viruses (ULVs), some of which only encode replication-associated proteins, suggested that they, as with umbraviruses that lack encoded capsid proteins (CPs) and silencing suppressors, would require association with a helper virus to complete an infection cycle. We examined the infection properties of 2 ULVs: citrus yellow vein associated virus 1 (CY1), which only encodes replication proteins, and closely related CY2 from hemp, which encodes an additional protein (ORF5CY2) that was assumed to be an MP. We report that both CY1 and CY2 can independently infect the model plant Nicotiana benthamiana in a phloem-limited fashion when delivered by agroinfiltration. Unlike encoded MPs, ORF5CY2 was dispensable for infection of CY2, but was associated with faster symptom development. Examination of ORF5CY2 revealed features more similar to luteoviruses/poleroviruses/sobemovirus CPs than to 30K class MPs, which all share a similar single jelly-roll domain. In addition, only CY2-infected plants contained virus-like particles (VLPs) associated with CY2 RNA and ORF5CY2. CY1 RNA and a defective (D)-RNA that arises during infection interacted with host protein phloem protein 2 (PP2) in vitro and in vivo, and formed a high molecular weight complex with sap proteins in vitro that was partially resistant to RNase treatment. When CY1 was used as a virus-induced gene silencing (VIGS) vector to target PP2 transcripts, CY1 accumulation was reduced in systemic leaves, supporting the usage of PP2 for systemic movement. ULVs are therefore the first plant viruses encoding replication and CPs but no MPs, and whose systemic movement relies on a host MP. This explains the lack of discernable helper viruses in many ULV-infected plants and evokes comparisons with the initial viruses transferred into plants that must have similarly required host proteins for movement.

Ten years of hand hygiene excellence: a summary of outcomes, and a comparison of indicators, from award-winning hospitals worldwide.

BACKGROUND: Hand hygiene is a crucial measure for the prevention of healthcare-associated infections (HAIs). The Hand Hygiene Excellence Award (HHEA) is an international programme acknowledging healthcare facilities for their leadership in implementing hand hygiene improvement programmes, including the World Health Organisation's Multimodal Improvement Strategy. This study aimed at summarising the results of the HHEA campaign between 2010 and 2021 and investigating the relationship between different hand hygiene parameters based on data from participating healthcare facilities. METHODS: A retrospective analysis was performed on datasets from HHEA forms, including data on hand hygiene compliance, alcohol-based handrub (ABHR) consumption, and Hand Hygiene Self-Assessment Framework (HHSAF) scores. Descriptive statistics were reported for each variable. The correlation between variables was inspected through Kendall's test, while possible non-linear relationships between hand hygiene compliance, ABHR consumption and HHSAF scores were sought through the Locally Estimated Scatterplot Smoothing or logistic regression models. A tree-structured partitioning model was developed to further confirm the obtained findings. RESULTS: Ninety-seven healthcare facilities from 28 countries in three world regions (Asia-Pacific, Europe, Latin America) were awarded the HHEA and thus included in the analysis. HHSAF scores indicated an advanced hand hygiene promotion level (median 445 points, IQR 395-480). System change (100 [95-100] points) and institutional safety climate (85 [70-95] points) showed the highest and lowest score, respectively. In most cases, hand hygiene compliance was above 70%, with heterogeneity between countries. ABHR consumption above 20 millilitres per patient-day (ml/PD) was widely reported, with overall increasing trends. HHSAF scores were positively correlated with hand hygiene compliance (τ = 0.211, p = 0.007). We observed a positive correlation between compliance rates and ABHR consumption (τ = 0.193, p < 0.001), although the average predicted consumption was stable around 55-60 ml/PD for compliance rates above 80-85%. Logistic regression and partitioning tree analyses revealed that higher HHSAF scores were more likely in the high-ABHR consumption group at cut-offs around 57-59 ml/PD. CONCLUSION: Ten years after its inception, the HHEA proves to be a valuable hand hygiene improvement programme in healthcare facilities worldwide. Consistent results were provided by the different hand hygiene indicators and the HHSAF score represents a valuable proxy measure of hand hygiene compliance.

Distal femoral osteotomy and patellar tendon advancement for the treatment of crouch gait in patients with bilateral spastic cerebral palsy.

BACKGROUND: Crouch gait, or flexed knee gait, represents a common gait pattern in patients with spastic bilateral cerebral palsy (CP). Distal femoral extension and/or shortening osteotomy (DFEO/DFSO) and patellar tendon advancement (PTA) can be considered as viable options when knee flexion contractures are involved. Better outcomes have been reported after a combination of both, independently of the presence of knee extensor lag. In this study, we evaluated the clinical and kinematic outcomes of these procedures. PATIENTS AND METHODS: We reviewed a cohort of 52 limbs (28 patients) who were treated for crouch gait by DFEO/DFSO alone (group 1, n = 15) or DFEO/DFSO + PTA (group 2, n = 37) as a part of single event multilevel surgery (SEMLS). The mean age at surgery was 14 years, and the mean follow-up time was 18 months. The physical examination data and three-dimensional standardized gait analysis were collected and analyzed before the surgery and postoperatively. RESULTS: Overall knee range of motion improved in all limbs. The knee flexion decreased significantly in both groups at initial, mid, and terminal stance. Hip flexion significantly decreased in mid-stance for limbs in group 2. Both clinical and gait parameters were most improved in limbs who underwent DFEO/DFSO + PTA. Increased pelvic tilt was observed in both groups after surgery. CONCLUSION: Although DFEO/DFSO alone was successful in correcting knee flexion contractures, PTA has helped to improve knee extensor lag and knee extension during gait. LEVEL OF EVIDENCE: Therapeutic level IV.

A Disintegrin And Metalloprotease 10 expression within the murine central nervous system.

A Disintegrin And Metalloprotease 10 (ADAM10), is able to control several important physiopathological processes through the shedding of a large number of protein substrates. Although ADAM10 plays a crucial role in the central nervous system (CNS) development and function, its protein distribution in the CNS has not been fully addressed. Here, we described the regional and cellular ADAM10 protein expression in C57BL/6 mice examined by immunofluorescence 1) throughout the adult mouse brain, cerebellum and spinal cord in vivo and 2) in different cell types as neurons, astrocytes, oligodendrocytes and microglia in vitro. We observed ADAM10 expression through the whole CNS, with a strong expression in the hippocampus, in the hypothalamus and in the cerebral and piriform cortex in the brain, in the Purkinje and in granular cell layers in the cerebellum and in the spinal cord to a lower extent. In vivo, ADAM10 protein expression was mainly found in neurons and in some oligodendroglial cell populations. However, in primary cultures we observed ADAM10 expression in neurons, oligodendrocytes, astrocytes and microglia. Interestingly, ADAM10 was not only found in the membrane but also in cytoplasmic vesicles and in the nucleus of primary cultured cells. Overall, this work highlights a wide distribution of ADAM10 throughout the CNS. The nuclear localization of ADAM10, probably due to its intracellular domain, emphasizes its role in cell signalling in physiological and pathological conditions. Further investigations are required to better elucidate the role of ADAM10 in glial cells.