An ON-type direction-selective ganglion cell in primate retina.

To maintain a stable and clear image of the world, our eyes reflexively follow the direction in which a visual scene is moving. Such gaze-stabilization mechanisms reduce image blur as we move in the environment. In non-primate mammals, this behaviour is initiated by retinal output neurons called ON-type direction-selective ganglion cells (ON-DSGCs), which detect the direction of image motion and transmit signals to brainstem nuclei that drive compensatory eye movements1. However, ON-DSGCs have not yet been identified in the retina of primates, raising the possibility that this reflex is mediated by cortical visual areas. Here we mined single-cell RNA transcriptomic data from primate retina to identify a candidate ON-DSGC. We then combined two-photon calcium imaging, molecular identification and morphological analysis to reveal a population of ON-DSGCs in the macaque retina. The morphology, molecular signature and GABA (γ-aminobutyric acid)-dependent mechanisms that underlie direction selectivity in primate ON-DSGCs are highly conserved with those in other mammals. We further identify a candidate ON-DSGC in human retina. The presence of ON-DSGCs in primates highlights the need to examine the contribution of subcortical retinal mechanisms to normal and aberrant gaze stabilization in the developing and mature visual system.

Multi-omics atlas of combinatorial abiotic stress responses in wheat.

Field-grown crops rarely experience growth conditions in which yield can be maximized. Environmental stresses occur in combination, with advancements in crop tolerance further complicated by its polygenic nature. Strategic targeting of causal genes is required to meet future crop production needs. Here, we employed a systems biology approach in wheat (Triticum aestivum L.) to investigate physio-metabolic adjustments and transcriptome reprogramming involved in acclimations to heat, drought, salinity and all combinations therein. A significant shift in magnitude and complexity of plant response was evident across stress scenarios based on the agronomic losses, increased proline concentrations and 8.7-fold increase in unique differentially expressed transcripts (DETs) observed under the triple stress condition. Transcriptome data from all stress treatments were assembled into an online, open access eFP browser for visualizing gene expression during abiotic stress. Weighted gene co-expression network analysis revealed 152 hub genes of which 32% contained the ethylene-responsive element binding factor-associated amphiphilic repression (EAR) transcriptional repression motif. Cross-referencing against the 31 DETs common to all stress treatments isolated TaWRKY33 as a leading candidate for greater plant tolerance to combinatorial stresses. Integration of our findings with available literature on gene functional characterization allowed us to further suggest flexible gene combinations for future adaptive gene stacking in wheat. Our approach demonstrates the strength of robust multi-omics-based data resources for gene discovery in complex environmental conditions. Accessibility of such datasets will promote cross-validation of candidate genes across studies and aid in accelerating causal gene validation for crop resiliency.

Distinct Phenotypic Consequences of Pathogenic Mutants Associated with Late-Onset Retinal Degeneration.

A pathologic feature of late-onset retinal degeneration caused by the S163R mutation in C1q-tumor necrosis factor-5 (C1QTNF5) is the presence of unusually thick deposits between the retinal pigmented epithelium (RPE) and the vascular choroid, considered a hallmark of this disease. Following its specific expression in mouse RPE, the S163R mutant exhibits a reversed polarized distribution relative to the apically secreted wild-type C1QTNF5, and forms widespread, prominent deposits that gradually increase in size with aging. The current study shows that S163R deposits expand to a considerable thickness through a progressive increase in the basolateral RPE membrane, substantially raising the total RPE height, and enabling their clear imaging as a distinct hyporeflective layer by noninvasive optical coherence tomography in advanced age animals. This phenotype bears a striking resemblance to ocular pathology previously documented in patients harboring the S163R mutation. Therefore, a similar viral vector-based gene delivery approach was used to also investigate the behavior of P188T and G216C, two novel pathogenic C1QTNF5 mutants recently reported in patients for which histopathologic data are lacking. Both mutants primarily impacted the RPE/photoreceptor interface and did not generate basal laminar deposits. Distinct distribution patterns and phenotypic consequences of C1QTNF5 mutants were observed in vivo, which suggested that multiple pathobiological mechanisms contribute to RPE dysfunction and vision loss in this disorder.

Influence of neck shaft angle of humerus in prosthesis design.

Background: A better understanding of the proximal humerus is essential for anatomical reconstruction of the glenohumeral joint during prosthetic replacement. The neck-shaft angle is critical for correct calcar screw positioning when fixing a proximal humeral fracture with a locking plate. It's essential for exact implant placement and treatment of any soft-tissue or bone pathology. Improper implants cause discomfort and post-operative complications, therefore understanding the humeral neck shaft angle is fundamental in the design and manufacturing of prostheses. This study looked into the necessity regional data of the humeral neck shaft angle (NSA) and its importance in shoulder prosthesis manufacturing. Method: This cross-sectional investigation was conducted on 300 dry cadaveric humeri of unknown gender and age that were free of damage or deformity. The Goniometer was used to measure the neck shaft angle. The data was analysed using SPSS software. The standard deviation and mean were calculated. The statistical difference between the right and left humeri was assessed using the students' t' test. Observations: The NSA of the humerus was 131.265.82° on average. The NSA mean values were substantially higher on the right side. A statistically significant difference between the right and left humeri was revealed by a P value of 0.001. Conclusion: Success of the shoulder arthroplasty demands anatomical reconstruction of the normal neck-shaft angle of the humerus. Racial variations in the morphometry of the neck-shaft angle needs to be considered in prosthesis design.

Carbon dioxide expanded liquid: an effective solvent for the extraction of quercetin from South African medicinal plants.

BACKGROUND: Quercetin is one of the most important bioflavonoids having positive effects on the biological processes and human health. Typically, it is extracted from plant matrices using conventional methods such as maceration, sonication, infusion, and Soxhlet extraction with high solvent consumption. Our study aimed to optimize the environmentally friendly carbon dioxide-based method for the extraction of quercetin from quince fruit with an emphasis on extraction yield, repeatability, and short extraction time. RESULTS: A two-step design of experiments was used for the optimization of the key parameters affecting physicochemical properties, including CO2/co-solvent ratio, co-solvent type, temperature, and pressure. Finally, gas expanded liquid combining CO2/ethanol/H2O in a ratio of 10/81/9 (v/v/v) provided the best extraction yield. Extraction temperature 66 °C and pressure 22.3 MPa were the most suitable conditions after careful optimization, although both parameters did not significantly affect the process. It was confirmed by experiments in various pressure and temperature conditions and statistical comparison of obtained data. The optimized extraction procedure at a flow rate of 3 mL/min took 30 min. The repeatability of the extraction method exhibited an RSD of 20.8%. CONCLUSIONS: The optimized procedure enabled very fast extraction in 30 min using environmentally friendly solvents and it was successfully applied to 16 different plant samples, including 14 bulbs and 2 fruits from South Africa. The quercetin content in extracts was quantified using ultra-high performance liquid chromatography (UHPLC) with tandem mass spectrometry. UHPLC hyphenated with high-resolution mass spectrometry was used to confirm chemical identity of quercetin in the analyzed samples. We quantified quercetin in 11 samples of all 16 tested plants. The quercetin was found in Agapanthus praecox from the Amaryllidaceae family and its presence in this specie was reported for the first time.

Fine-mapping of a putative glutathione S-transferase (GST) gene responsible for yellow seed colour in flax (Linum usitatissimum).

OBJECTIVE: The brown seed coat colour of flax (Linum ustiatissimum) results from proanthocyanidin synthesis and accumulation. Glutathione S-transferases (GSTs), such as the TT19 protein in Arabidopsis, have been implicated in the transport of anthocyanidins during the synthesis of the brown proanthocyanidins. This study fine mapped the g allele responsible for yellow seed colour in S95407 and identified it as a putative mutated GST. RESULTS: We developed a Recombinant Inbred Line population with 320 lines descended from a cross between CDC Bethune (brown seed coat) and S95407 (yellow seed) and used molecular markers to fine map the G gene on Chromosome 6 (Chr 6). We used Next Generation Sequencing (NGS) to identify a putative GST was identified in this region and Sanger sequenced the gene from CDC Bethune, S95407 and other yellow seeded genotypes. The putative GST from S95407 had 13 SNPs encoding, including four non-synonymous amino acid changes, compared to the CDC Bethune reference sequence and the other genotypes. The GST encoded by Lus10019895 is a lambda-GST in contrast to the Arabidopsis TT19 which is a phi-GST.

Effect of storage on plant biostimulant and bioactive properties of freeze-dried Chlorella vulgaris biomass.

Microalgae are potential plant biostimulants and biocontrol agents. A major hurdle towards their commercialization is the production of large volumes of biomass at the correct time of year. Secondary metabolites are unstable and the "shelf-life" of bioactive microalgal biomass needs to be investigated. The aim of the study was to investigate the effects of storage conditions on freeze-dried microalgae to determine how long the biomass retained its growth promoting and bioactive properties under various temperature and light conditions. Chlorella vulgaris biomass was stored in the dark at - 70 °C, 10 °C, and 25 °C and in the light at 25 °C. Samples were tested every 3-4 months for 15 months. Storage time significantly influenced the rate of change in the bioactivity in the C. vulgaris biomass with storage temperature also having some effect. Rooting activity decreased in the mungbean rooting assay over time up to 12 months and then increased slightly. Antimicrobial activity increased against Staphylococcus aureus and Escherichia coli for up to 12 months and then declined. Antioxidant activity measured in the DPPH assay remained relatively stable for up to 12 months and then significantly decreased with longer storage. The change in bioactivity over time was attributed to the gradual breakdown of the rigid cell wall of C. vulgaris, thereby improving extraction efficiency but exposing the secondary metabolites to oxygen, thus quickening their degradation. Biomass produced for commercial purposes requires preliminary validation as the results of the present study showed that bioactive compounds are susceptible to degradation over time. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s10811-021-02596-9.

Intra-dural intercommunications between dorsal roots of adjacent spinal nerves and their clinical significance.

PURPOSE: The dorsal roots of adjacent spinal nerves are known to communicate with each other through rami communicantes. These intercommunications can cause deviations in the normal dermatomal organization which leads to errors during clinical decision-making. The objective of the study was to augment the existing knowledge of these communications which shall help minimize the diagnostic and therapeutic errors. METHODS: The present study examined thirty cadaveric spinal cord specimens to document the data of intra-dural, intercommunications between dorsal roots of adjacent spinal nerves. RESULTS: All the regions of the spinal cord exhibited the presence of intercommunications with variable frequency. The intercommunications were categorized into a total of nine groups based on their patterns. The levels of spinal cord exhibiting higher and lower frequencies of intercommunications were identified. CONCLUSION: This information will be useful during the clinical evaluation of patients with spinal cord pathologies or radiculopathies. The outcomes of rhizotomy can also be improved with knowledge of intercommunications.

Interactive effects of plant growth-promoting rhizobacteria and a seaweed extract on the growth and physiology of Allium cepa L. (onion).

Detrimental effects caused by the overuse of synthetic agrochemicals have led to the development of natural biostimulants such as seaweed extracts and plant growth-promoting rhizobacteria (PGPR) being used as an alternative, environmentally-friendly technology to improve crop growth and increase agricultural yields. The present study aimed to investigate the interactions between PGPR and a commercial seaweed extract on the growth and biochemical composition of onion (Allium cepa). A pot trial was conducted under greenhouse conditions where onion plants were treated individually with the two PGPR, namely Bacillus licheniformis (BL) and Pseudomonas fluorescens (PF) and a seaweed extract Kelpak® (KEL) and combinations of KEL + BL and KEL + PF. Growth and yield parameters were measured after 12 weeks. KEL-treated plants showed the best growth response and overcame the inhibitory effects of BL treatment. KEL-treated plants also had the highest chlorophyll content. PGPR application improved the mineral nutrition of onion with these plants having the highest mineral content in the leaves and bulb. All biostimulant treatments increased the endogenous cytokinin and auxin content with the highest concentrations generally detected in the PF-treated plants. These results suggest that co-application of different biostimulant classes with different modes of action could further increase crop productivity with an improvement in both growth and nutrition content being achieved in onion with the co-application of a seaweed extract and PGPR.

Medical Cannabis and Industrial Hemp Tissue Culture: Present Status and Future Potential.

In recent years high-THC (psychoactive) and low-THC (industrial hemp) type cannabis (Cannabis sativa L.) have gained immense attention in medical, food, and a plethora of other consumer product markets. Among the planting materials used for cultivation, tissue culture clones provide various advantages such as economies of scale, production of disease-free and true-to-type plants for reducing the risk of GMP-EuGMP level medical cannabis production, as well as the development and application of various technologies for genetic improvement. Various tissue culture methods have the potential application with cannabis for research, breeding, and novel trait development, as well as commercial mass propagation. Although tissue culture techniques for plant regeneration and micropropagation have been reported for different cannabis genotypes and explant sources, there are significant variations in the response of cultures and the morphogenic pathway. Methods for many high-yielding elite strains are still rudimentary, and protocols are not established. With a recent focus on sequencing and genomics in cannabis, genetic transformation systems are applied to medical cannabis and hemp for functional gene annotation via traditional and transient transformation methods to create novel phenotypes by gene expression modulation and to validate gene function. This review presents the current status of research focusing on different aspects of tissue culture, including micropropagation, transformation, and the regeneration of medicinal cannabis and industrial hemp transformants. Potential future tissue culture research strategies helping elite cannabis breeding and propagation are also presented.

Synthetic strigolactone (rac-GR24) alleviates the adverse effects of heat stress on seed germination and photosystem II function in lupine seedlings.

There is increasing experimental evidence that strigolactones, a class of carotenoid-derived sesquiterpenoid hormones, and their downstream signal components play a role in plant resilience to abiotic stress. Strigolactones positively influence plant coping mechanisms in response to abiotic stressors like drought and high salinity. In this study, we examined the effects of rac-GR24 (a synthetic strigolactone analog) and strigolactone inhibitors on the physiological and molecular responses associated with thermotolerance during seed germination and seedling development in Lupinus angustifolius under heat stress. Photosystem I & II functions were also evaluated via Chl a fluorescence transient analysis in heat stressed lupine seedlings. Our results suggest a putative role for GR24 in mediating tolerance to heat stress during seed germination and seedling development albeit these responses appeared independent of D14-mediated signalling. Seeds primed with GR24 had the highest of all germination indices, enhanced proline content and reduced peroxidation of lipids. GR24 also enhanced the activities of enzymes of the antioxidant and glyoxalase systems in lupine seedlings. The JIP-test indicated that GR24 conferred resistance to heat stress-induced damage to the oxygen evolution complex while also preventing the inactivation of PSII reaction centres thus ensuring PSII thermotolerance.

Drought-Induced Regulatory Cascades and Their Effects on the Nutritional Quality of Developing Potato Tubers.

Competition for scarce water resources and the continued effects of global warming exacerbate current constraints on potato crop production. While plants' response to drought in above-ground tissues has been well documented, the regulatory cascades and subsequent nutritive changes in developing tubers have been largely unexplored. Using the commercial Canadian cultivar "Vigor", plants were subjected to a gradual drought treatment under high tunnels causing a 4 °C increase in the canopy temperature. Tubers were sampled for RNAseq and metabolite analysis. Approximately 2600 genes and 3898 transcripts were differentially expressed by at least 4-fold in drought-stressed potato tubers, with 75% and 69% being down-regulated, respectively. A further 229 small RNAs were implicated in gene regulation during drought. Expression of several small RNA clusters negatively correlated with expression of their six target patatin genes, suggesting involvement in the regulation of storage proteins during drought. The comparison of protein homologues between Solanum tuberosum L. and Arabidopsis thaliana L. indicated that down-regulated genes were associated with phenylpropanoid and carotenoid biosynthesis. As is indicative of reduced flow through the phenylpropanoid pathway, phenylalanine accumulated in drought-stressed tubers. This suggests that there may be nutritive implications to drought stress occurring during the potato tuber bulking phase in sensitive cultivars.

A critical review on exploiting the pharmaceutical potential of plant endophytic fungi.

The escalating demand for secondary metabolites in international markets poses a severe threat to many plant species. An unscrupulous collection is also the immediate challenge to the survival of many unthreatened as well as vulnerable plants. Fungal endophytes have emerged in recent years as a promising substitute for sources of plant secondary metabolites. Many appealing secondary metabolites with potent antibacterial, antifungal, insecticidal, antioxidant, cytotoxic and anticancer properties have been discovered from endophytic fungi. Concerning their distinctive genetic and metabolic diversity and promising activities, they hold a plausible application in medicine and industry. However, there is little success in utilizing the pharmaceutical potential of fungal endophytes. Cutting-edge research is desirable to establish and bolster in vitro biosynthetic proficiency of fungal endophytes. Modern biotechnological techniques [such as multilocus sequence typing (MLST), metabolomics, metagenomics and next-generation sequencing (NGS) technologies] and bioinformatics approaches can fill a gap in fungal endophyte research. The present review focuses on how advanced chemical, biotechnological and computational molecular biology methods can be used for robust exploitation of bioactive compounds from these microorganisms.

WheatCRISPR: a web-based guide RNA design tool for CRISPR/Cas9-mediated genome editing in wheat.

BACKGROUND: CRISPR/Cas9 gene editing has become a revolutionary technique for crop improvement as it can facilitate fast and efficient genetic changes without the retention of transgene components in the final plant line. Lack of robust bioinformatics tools to facilitate the design of highly specific functional guide RNAs (gRNAs) and prediction of off-target sites in wheat is currently an obstacle to effective application of CRISPR technology to wheat improvement. DESCRIPTION: We have developed a web-based bioinformatics tool to design specific gRNAs for genome editing and transcriptional regulation of gene expression in wheat. A collaborative study between the Broad Institute and Microsoft Research used large-scale empirical evidence to devise algorithms (Doech et al., 2016, Nature Biotechnology 34, 184-191) for predicting the on-target activity and off-target potential of CRISPR/SpCas9 (Streptococcus pyogenes Cas9). We applied these prediction models to determine on-target specificity and potential off-target activity for individual gRNAs targeting specific loci in the wheat genome. The genome-wide gRNA mappings and the corresponding Doench scores predictive of the on-target and off-target activities were used to create a gRNA database which was used as a data source for the web application termed WheatCRISPR. CONCLUSION: The WheatCRISPR tool allows researchers to browse all possible gRNAs targeting a gene or sequence of interest and select effective gRNAs based on their predicted high on-target and low off-target activity scores, as well as other characteristics such as position within the targeted gene. It is publicly available at https://crispr.bioinfo.nrc.ca/WheatCrispr/ .

Role of Smoke Stimulatory and Inhibitory Biomolecules in Phytochrome-Regulated Seed Germination of Lactuca sativa.

The biologically active molecules karrikinolide (KAR1) and trimethylbutenolide (TMB) present in wildfire smoke play a key role in regulating seed germination of many plant species. To elucidate the physiological mechanism by which smoke-water (SW), KAR1, and TMB regulate seed germination in photosensitive 'Grand Rapids' lettuce (Lactuca sativa), we investigated levels of the dormancy-inducing hormone abscisic acid (ABA), three auxin catabolites, and cytokinins (26 isoprenoid and four aromatic) in response to these compounds. Activity of the hydrolytic enzymes α-amylase and lipase along with stored food reserves (lipids, carbohydrate, starch, and protein) were also assessed. The smoke compounds precisely regulated ABA and hydrolytic enzymes under all light conditions. ABA levels under red (R) light were not significantly different in seeds treated with TMB or water. However, TMB-treated seeds showed significantly inhibited germination (33%) compared with water controls (100%). KAR1 significantly enhanced total isoprenoid cytokinins under dark conditions in comparison with other treatments; however, there was no significant effect under R light. Enhanced levels of indole-3-aspartic acid (an indicator of high indole-3-acetic acid accumulation, which inhibits lettuce seed germination) and absence of trans-zeatin and trans-zeatin riboside (the most active cytokinins) in TMB-treated seeds might be responsible for reduced germination under R light. Our results demonstrate that SW and KAR1 significantly promote lettuce seed germination by reducing levels of ABA and enhancing the activity of hydrolytic enzymes, which aids in mobilizing stored reserves. However, TMB inhibits germination by enhancing ABA levels and reducing the activity of hydrolytic enzymes.

Quantification of karrikins in smoke water using ultra-high performance liquid chromatography-tandem mass spectrometry.

BACKGROUND: Karrikins (KARs) are plant growth regulators that promote seed germination and the subsequent growth and development of seedlings of many plant species. In nature they are generated and released by combustion of plant material and promote the restoration of burned ecosystems. Smoke water can be artificially prepared as a saturated extract of all substances in smoke produced by burning plants, and it has various horticultural and agricultural applications. RESULTS: We have developed, validated and applied the first fast, specific and sensitive method, based on ultra-high performance liquid chromatography-tandem mass spectrometry, for quantifying KARs in smoke water. To assist these efforts and further analyses, standards of the main KARs (which are not commercially available) were synthesized. Due to the complex matrix of smoke waters, two quantification approaches (standard dilution with a structural KAR analogue and standard addition) were compared. The standard addition method allowed absolute quantification of KARs in six of eight smoke water samples of diverse origins and ages. CONCLUSIONS: Our findings reveal differences in both total and relative levels of KARs in smoke water, and indicate that differences in its KAR composition may be linked to variations in its bioactivity.

Synthesis of potent neuroprotective butenolides based on plant smoke derived 3,4,5-Trimethylfuran-2(5H)-one and 3-methyl-2H-furo[2,3-c]pyrone-2-one.

Smoke derived karrikinolide and trimethylbutenolide exerted neuroprotective effects against monoamine oxidase and acetylcholinesterase. Synthesis of potent analogs was achieved. Sulphur substitution in the bicyclic ring structure of KAR1 displayed the most encouraging activity returning IC50 values of 13.75 ±â€¯0.001 µM and 0.03 ±â€¯0.02 µM for monoamine oxidase A and B and 0.08 ±â€¯0.006 µM for acetylcholinesterase. Neuroprotective butenolides may be particularly useful in the treatment of depressive disorders, Alzheimer's and Parkinson's diseases.

Bioactive molecules derived from smoke and seaweed Ecklonia maxima showing phytohormone-like activity in Spinacia oleracea L.

Bioactive compounds such as karrikinolide (KAR1 from smoke) and eckol (from the seaweed Ecklonia maxima) show promising effects on several important crop plants. These plant growth-stimulating organic biomolecules, along with crude extracts (smoke-water and Kelpak® product prepared from Ecklonia maxima), were tested on spinach plants. Eckol sprayed at 10-6 M significantly increased all the growth and biochemical parameters examined compared to control spinach plants. All tested plant growth biostimulants significantly increased total chlorophyll, carotenoids and protein content of spinach leaves. The cytokinin profile of spinach plants was also determined. Cis-zeatin, dihydrozeatin and isopentenyladenine types of cytokinins were promoted by both smoke- and seaweed-based biostimulants. In comparison to the control plants, the level of free sinapic acid was greater in all spinach plants treated with these biostimulants. The application of these biostimulants can help spinach crop by improving growth, yield and nutritional quality; moreover, they are organic and cost-effective.