miRador: a fast and precise tool for the prediction of plant miRNAs.

Plant microRNAs (miRNAs) are short, noncoding RNA molecules that restrict gene expression via posttranscriptional regulation and function in several essential pathways, including development, growth, and stress responses. Accurately identifying miRNAs in populations of small RNA sequencing libraries is a computationally intensive process that has resulted in the misidentification of inaccurately annotated miRNA sequences. In recent years, criteria for miRNA annotation have been refined with the aim to reduce these misannotations. Here, we describe miRador, a miRNA identification tool that utilizes the most up-to-date, community-established criteria for accurate identification of miRNAs in plants. We combined target prediction and Parallel Analysis of RNA Ends (PARE) data to assess the precision of the miRNAs identified by miRador. We compared miRador to other commonly used miRNA prediction tools and found that miRador is at least as precise as other prediction tools while being substantially faster than other tools. miRador should be broadly useful for the plant community to identify and annotate miRNAs in plant genomes.

The evolutionary history of small RNAs in Solanaceae.

The Solanaceae or "nightshade" family is an economically important group with remarkable diversity. To gain a better understanding of how the unique biology of the Solanaceae relates to the family's small RNA (sRNA) genomic landscape, we downloaded over 255 publicly available sRNA data sets that comprise over 2.6 billion reads of sequence data. We applied a suite of computational tools to predict and annotate two major sRNA classes: (1) microRNAs (miRNAs), typically 20- to 22-nucleotide (nt) RNAs generated from a hairpin precursor and functioning in gene silencing and (2) short interfering RNAs (siRNAs), including 24-nt heterochromatic siRNAs typically functioning to repress repetitive regions of the genome via RNA-directed DNA methylation, as well as secondary phased siRNAs and trans-acting siRNAs generated via miRNA-directed cleavage of a polymerase II-derived RNA precursor. Our analyses described thousands of sRNA loci, including poorly understood clusters of 22-nt siRNAs that accumulate during viral infection. The birth, death, expansion, and contraction of these sRNA loci are dynamic evolutionary processes that characterize the Solanaceae family. These analyses indicate that individuals within the same genus share similar sRNA landscapes, whereas comparisons between distinct genera within the Solanaceae reveal relatively few commonalities.

A semi-supervised approach to unobtrusively predict abnormality in breathing patterns using hydraulic bed sensor data in older adults aging in place.

Shortness of breath is often considered a repercussion of aging in older adults, as respiratory illnesses like COPD1 or respiratory illnesses due to heart-related issues are often misdiagnosed, under-diagnosed or ignored at early stages. Continuous health monitoring using ambient sensors has the potential to ameliorate this problem for older adults at aging-in-place facilities. In this paper, we leverage continuous respiratory health data collected by using ambient hydraulic bed sensors installed in the apartments of older adults in aging-in-place Americare facilities to find data-adaptive indicators related to shortness of breath. We used unlabeled data collected unobtrusively over the span of three years from a COPD-diagnosed individual and used data mining to label the data. These labeled data are then used to train a predictive model to make future predictions in older adults related to shortness of breath abnormality. To pick the continuous changes in respiratory health we make predictions for shorter time windows (60-s). Hence, to summarize each day's predictions we propose an abnormal breathing index (ABI) in this paper. To showcase the trajectory of the shortness of breath abnormality over time (in terms of days), we also propose trend analysis on the ABI quarterly and incrementally. We have evaluated six individual cases retrospectively to highlight the potential and use cases of our approach.

Holistic approach to waste mobil oil bioremediation: Valorizing waste through biosurfactant production for soil restoration.

The combustion of mobil oil leads to the emission of toxic compounds in the environment. In this study, the aromatic and aliphatic hydrocarbon fractions present in a waste mobil oil collected from automobile market were comprehensively identified and their toxicity was evaluated using wheat grain. Lysinibacillus sphaericus strain IITR51 isolated and characterized previously could degrade 30-80% of both aliphatic and aromatic hydrocarbons in liquid culture. Interestingly, the strain IITR51 produced 627 mg/L of rhamnolipid biosurfactant by utilizing 3% (v/v) of waste mobil oil in the presence of 1.5% glycerol as additional carbon source. In a soil microcosm study by employing strain IITR51, 50-86% of 3-6 ring aromatic hydrocarbons and 63-98% of aliphatic hydrocarbons (C8 to C22) were degraded. Addition of 60 µg/mL rhamnolipid biosurfactant enhanced the degradation of both aliphatic and aromatic hydrocarbons from 76.88% to 61.21%-94.11% and 78.27% respectively. The degradation of mobil oil components improved the soil physico-chemical properties and increased soil fertility to 64% as evident by the phytotoxicity assessments. The findings indicate that strain IITR51 with degradation capability coupled with biosurfactant production could be a candidate for restoring hydrocarbon contaminated soils.

Effect of COVID-19 lockdown on lifestyle habits and self-care practices of diabetic and hypertensive patients in rural Shimla and Udaipur - Findings from the HealthRise India program.

Background: COVID-19 pandemic has increased the risk of mortality among patients with noncommunicable diseases. Maintaining a good metabolic control, lifestyle modification along with improved self-care practices are not only associated with less severe COVID-19 infections but also with a high recovery rate. Objectives: This research article explores the changes in lifestyle habits, self-care practices, and metabolic control among patients enrolled in the HealthRise program. The study compares behavioral changes, before COVID-19 pandemic and during COVID-19 pandemic, between intervention and control arms in Shimla and Udaipur. Methods: A quasi-experimental study design was employed for program implementation in select villages of Shimla district, and Udaipur district. A total of 459 patients from Shimla and 309 patients from Udaipur with diabetes mellitus or hypertension or with both were enrolled and followed for 1 year. Results: Metabolic control in Shimla intervention arm was 2.6 times higher than in control arm (P = 0.001) before COVID-19 pandemic. During COVID-19 pandemic, Odds of metabolic control in Shimla intervention was 1.5 times higher when compared with control arm (P = 0.03). In Udaipur, metabolic control before COVID-19 pandemic was comparable between control and intervention arms. During the pandemic, metabolic control in intervention arm of Udaipur was 5 times higher when compared to the control arm ((P = 0.001). Conclusion: Participants exposed to support, appreciate, learn, and transfer-community life competence process (SALT-CLCP) intervention maintained metabolic control during the COVID-19 pandemic with improved behavioral and self-care practices. Community-based interventions such as SALT-CLCP method bring ownership and empower community in achieving the better health outcomes.

Pulsed laser assisted high-throughput intracellular delivery in hanging drop based three dimensional cancer spheroids.

Targeted intracellular delivery of biomolecules and therapeutic cargo enables the controlled manipulation of cellular processes. Laser-based optoporation has emerged as a versatile, non-invasive technique that employs light-based transient physical disruption of the cell membrane and achieves high transfection efficiency with low cell damage. Testing of the delivery efficiency of optoporation-based techniques has been conducted on single cells in monolayers, but its applicability in three-dimensional (3D) cell clusters/spheroids has not been explored. Cancer cells grown as 3D tumor spheroids are widely used in anti-cancer drug screening and can be potentially employed for testing delivery efficiency. Towards this goal, we demonstrated the optoporation-based high-throughput intracellular delivery of a model fluorescent cargo (propidium iodide, PI) within 3D SiHa human cervical cancer spheroids. To enable this technique, nano-spiked core-shell gold-coated polystyrene nanoparticles (ns-AuNPs) with a high surface-to-volume ratio were fabricated. ns-AuNPs exhibited high electric field enhancement and highly localized heating at an excitation wavelength of 680 nm. ns-AuNPs were co-incubated with cancer cells within hanging droplets to enable the rapid aggregation and assembly of spheroids. Nanosecond pulsed-laser excitation at the optimized values of laser fluence (45 mJ cm-2), pulse frequency (10 Hz), laser exposure time (30 s), and ns-AuNP concentration (5 × 1010 particles per ml) resulted in the successful delivery of PI dye into cancer cells. This technique ensured high delivery efficiency (89.6 ± 2.8%) while maintaining high cellular viability (97.4 ± 0.4%), thereby validating the applicability of this technique for intracellular delivery. The optoporation-based strategy can enable high-throughput single cell manipulation, is scalable towards larger 3D tissue constructs, and may provide translational benefits for the delivery of anti-cancer therapeutics to tumors.

Immunological detection assays for recombinant Shiga toxin & Shigella dysenteriae.

Background & objectives: : Shiga toxin (Stx) is produced by Shigella dysenteriae, a Gram-negative, facultative anaerobic bacillus that causes shigellosis, haemolytic uraemic syndrome (HUS) and Reiter's syndrome. The detection methods for shiga toxin needs to be rapid, accurate, reliable and must be extensively evaluated under field conditions. The aim of this study was to develop rapid, sensitive and specific detection method for Stx. Methods: : Mice and rabbits were immunized with purified recombinant Shiga toxin B (rStxB). Using these antibodies dot ELISA, sandwich ELISA and flow through assay were developed. Results: : The high-titre antibodies specifically reacted with purified rStxB. Dot-ELISA, sandwich ELISA and flow-through assay were developed and standardized that could detect StxB with limit of detection (LOD) of 9.75, 9.7 ng/ml and 0.46 µg/cassette, respectively. Interpretation & conclusions: : The rStxB was used to produce antibodies to avoid handling of pathogen. The Flow through assay 'developed was specific, rapid and field amenable.

Postexposure Recovery and Analysis of Biological Agent in a Simulated Biothreat Scenario Using Tandem Mass Spectrometry.

Some pathogens and toxins have the potential to be used as weapons of mass destruction and instigate population-based fear. Rapid, sensitive, and unambiguous identification of biothreat agents is of paramount importance for confirmation of the event and to mitigate the direct and indirect damages to public health and resources. Although there are several potential dissemination scenarios to describe an attack with a biological weapon, artificially generated bioaerosol is of the greatest concern from a bioterrorism or warfare perspective, potentially capable of causing mass destruction to a civilian or military population by inhalation of toxic bioaerosol. The present investigation proposes methodologies for recovery of biological agent followed by an off-site unambiguous detection using tandem mass spectrometry, in a postattack situation. We envisaged a biothreat scenario wherein the polydispersed bioaerosol is disseminated in bulk over any geographical setting. The larger particles (>5 µm in diameter) of bioaerosol settle and bind to various surfaces depending on the geographical setting. Recovery of agent was optimized from foliage, sand, and glass in a simulated biothreat scenario using bovine serum albumin (BSA). The recovered agents were shown to be amenable to detection by a downstream tandem MS analysis. Applicability of the proposed methodology was demonstrated in validation experiments for the recovery and detection of toxin and bacterial agents. The use of cleaner matrices (foliage, exposed smooth surfaces, sand) is recommended for retrospective verification of agent in a biothreat scenario.

Can community health workers play a greater role in increasing access to medical abortion services? A qualitative study.

BACKGROUND: Despite being legally available in India since 1971, barriers to safe and legal abortion remain, and unsafe and/or illegal abortion continues to be a problem. Community health workers have been involved in improving access to health information and care for maternal and child health in resource poor settings, but their role in facilitating accurate information about and access to safe abortion has been relatively unexplored. A qualitative study was conducted in Rajasthan, India to study acceptability, perspectives and preferences of women and community health workers, regarding the involvement of community health workers in medical abortion referrals. METHODS: In-depth interviews were conducted with 24 women seeking early medical abortion at legal abortion facilities or presenting at these facilities for a follow-up assessment after medical abortion. Ten community health workers who were trained to assess eligibility for early medical abortion and/or to assess whether women needed a follow-up visit after early medical abortion were also interviewed. The transcripts were coded using ATLAS-ti 7 (version 7.1.4) in the local language and reports were generated for all the codes, emerging themes were identified and the findings were analysed. RESULTS: Community health workers (CHWs) were willing to play a role in assessing eligibility for medical abortion and in identifying women who are in need of follow-up care after early medical abortion, when provided with appropriate training, regular supplies and job aids. Women however had apprehensions about contacting CHWs in relation to abortions. Important barriers that prevented women from seeking information and assistance from community health workers were fear of breach of confidentiality and a perception that they would be pressurised to undergo sterilisation. CONCLUSIONS: Our findings support a potential for greater role of CHWs in making safe abortion information and services accessible to women, while highlighting the need to address women's concerns about approaching CHWs in case of unwanted pregnancy. Further intervention research would be needed to shed light on the effectiveness of role of CHWs in facilitating access to safe abortion and to outline specific components in a programme setting. TRIAL REGISTRATION: Not applicable.

Direction-oriented fiber guiding with a tunable tri-layer-3D scaffold for periodontal regeneration.

Multilayered scaffolds mimicking mechanical and biological host tissue architectures are the current prerequisites for successful tissue regeneration. We propose our tunable tri-layered scaffold, designed to represent the native periodontium for potential regenerative applications. The fused deposition modeling platform is used to fabricate the novel movable three-layered polylactic acid scaffold mimicking in vivo cementum, periodontal ligament, and alveolar bone layers. The scaffold is further provided with multiple angulated fibers, offering directional guidance and facilitating the anchorage dependence on cell adhesion. Additionally, surface modifications of the scaffold were made by incorporating coatings like collagen and different concentrations of gelatin methacryloyl to enrich the cell adhesion and proliferation. The surface characterization of our designed scaffolds was performed using tribological studies, atomic force microscopy, contact angle measurement, scanning electron microscopy, and micro-computed tomography. Furthermore, the material characterization of this scaffold was investigated by attenuated total reflectance-Fourier transformed infrared spectroscopy. The scaffold's mechanical characterization, such as strength and compression modulus, was demonstrated by compression testing. The L929 mouse fibroblast cells and MG63 human osteosarcoma cells have been cultured on the scaffold. The scaffold's superior biocompatibility was evaluated using fluorescence dye with fluorescence microscopy, scanning electron microscopy, in vitro wound healing assay, MTT assay, and flow cytometry. The mineralization capability of the scaffolds was also studied. In conclusion, our study demonstrated the construction of a multilayered movable scaffold, which is highly biocompatible and most suitable for various downstream applications such as periodontium and in situ tissue regeneration of complex, multilayered tissues.

Advances in single-cell long-read sequencing technologies.

With an increase in accuracy and throughput of long-read sequencing technologies, they are rapidly being assimilated into the single-cell sequencing pipelines. For transcriptome sequencing, these techniques provide RNA isoform-level information in addition to the gene expression profiles. Long-read sequencing technologies not only help in uncovering complex patterns of cell-type specific splicing, but also offer unprecedented insights into the origin of cellular complexity and thus potentially new avenues for drug development. Additionally, single-cell long-read DNA sequencing enables high-quality assemblies, structural variant detection, haplotype phasing, resolving high-complexity regions, and characterization of epigenetic modifications. Given that significant progress has primarily occurred in single-cell RNA isoform sequencing (scRiso-seq), this review will delve into these advancements in depth and highlight the practical considerations and operational challenges, particularly pertaining to downstream analysis. We also aim to offer a concise introduction to complementary technologies for single-cell sequencing of the genome, epigenome and epitranscriptome. We conclude by identifying certain key areas of innovation that may drive these technologies further and foster more widespread application in biomedical science.

Laser-Induced Intracellular Delivery: Exploiting Gold-Coated Spiky Polymeric Nanoparticles and Gold Nanorods under Near-Infrared Pulses for Single-Cell Nano-Photon-Poration.

This study explores the potential of laser-induced nano-photon-poration as a non-invasive technique for the intracellular delivery of micro/macromolecules at the single-cell level. This research proposes the utilization of gold-coated spiky polymeric nanoparticles (Au-PNPs) and gold nanorods (GNRs) to achieve efficient intracellular micro/macromolecule delivery at the single-cell level. By shifting the operating wavelength towards the near-infrared (NIR) range, the intracellular delivery efficiency and viability of Au-PNP-mediated photon-poration are compared to those using GNR-mediated intracellular delivery. Employing Au-PNPs as mediators in conjunction with nanosecond-pulsed lasers, a highly efficient intracellular delivery, while preserving high cell viability, is demonstrated. Laser pulses directed at Au-PNPs generate over a hundred hot spots per particle through plasmon resonance, facilitating the formation of photothermal vapor nanobubbles (PVNBs). These PVNBs create transient pores, enabling the gentle transfer of cargo from the extracellular to the intracellular milieu, without inducing deleterious effects in the cells. The optimization of wavelengths in the NIR region, coupled with low laser fluence (27 mJ/cm2) and nanoparticle concentrations (34 µg/mL), achieves outstanding delivery efficiencies (96%) and maintains high cell viability (up to 99%) across the various cell types, including cancer and neuronal cells. Importantly, sustained high cell viability (90-95%) is observed even 48 h post laser exposure. This innovative development holds considerable promise for diverse applications, encompassing drug delivery, gene therapy, and regenerative medicine. This study underscores the efficiency and versatility of the proposed technique, positioning it as a valuable tool for advancing intracellular delivery strategies in biomedical applications.

The role of accurate identification of vulnerable youth in vocational education and training systems for improved employability: Insights from experimental data.

The role of vocational education and training system (VET) in addressing the economic and social needs of vulnerable youth from low-income households can be significant. It enables economic empowerment, providing them with a pathway to sustainable employment opportunities; improving their overall well-being and sense of personal identity. This article presents qualitative and quantitative data elucidating different elements of employability issues faced by such youth. It distinguishes and divulges a vulnerable group out of a larger population thereby making a strong case for identifying and addressing their needs. The approach is therefore not a 'one size fits all' training method. Students from urban Mumbai and New Delhi were mobilized from multiple channels such as self-help groups (SHGs), the National Institute of Open School (NIOS), distance education institutes, local government colleges, night schools and through direct community outreach. After carefully matching for demographic and economic characteristics, a group of 387 students in the age bracket of 18 to 24 years were selected and interviewed. This first set of data was generated for a range of personal, economic, and household characteristics. Data manifests structural barriers, poor human capital endowments and exclusion. To gain further insight into characteristics and to design a targeted intervention for a sub-group of 130 students from the total population, a second dataset is generated through a questionnaire and interview. Of this, two equal groups - an experiment group and a comparison group are formed as part of quasi-research. The third type of data is generated using a 5-point Likert scale questionnaire and personal discussions. A total of 2600 responses from the experiment (trained and skilled) and comparison (not trained) groups provides a basis for comparison of scores between the two groups pre-and post-intervention. The entire data collection process is practical, straightforward, and simple. Easy to explain - the dataset can be leveraged to generate evidence-based insights, and informed decisions on resource allocation, program design and strategies to mitigate risk factors. The multifaceted approach of data collection can be adapted to accurately identify vulnerable youth and create a newer framework for skill development and re-skilling. It can be used to develop measurement tools for employability by those involved in VET and in their efforts to create viable employment opportunities for high-potential yet disadvantaged youth.

Magnitude and Impact of Workplace Violence Against Obstetric Healthcare Personnel: A Multicentre Cross-Sectional Study.

Background: Workplace violence is defined by the World Health Organization (WHO) as incidents where staff is abused, threatened or assaulted in work settings. In emergency predominated branch like obstetrics, there is a need to study the magnitude and impact of violence against healthcare workers (HCW). Materials and Methodology: This cross-sectional study was conducted in the Department of Obstetrics at 2 centres in Lucknow district, for a period of 6 months. The study population included trainee residents, senior residents, nursing staff and consultants. Standard definitions from the WHO were used to define the types of violence. The validated questionnaire was designed in English with 25 questions to understand the incidence of workplace violence, prevention policy, reporting and follow-ups of incidents and impact of violence. Results: With a response rate of 90%, 274 HCW participated in the study. In total, 172 HCW (62.7%) either faced physical or verbal assault. In 70% of incidents, patient, their relatives or public were perpetrators of violence, and the rest 30% incidents were by colleagues or management. Majority of the incidents were in emergency areas. Only 22% of the abused reported to the concerned authorities. At least 123 (71.5%) HCW were extremely dissatisfied with the action taken. Action was taken against only 9.8% of the perpetrators. None of the respondents received any training to handle workplace violence. Conclusion: There is an alarming high prevalence of workplace violence by patients and colleagues. Adequate training to handle these incidents, improvement of working environment and unconditional support from management will bring a positive work experience. Supplementary Information: The online version contains supplementary material available at 10.1007/s13224-023-01809-0.

Functionally gradient three-dimensional graphene foam-based polymeric scaffolds for multilayered tissue regeneration.

Physiological bioengineering of multilayered tissues requires an optimized geometric organization with comparable biomechanics. Currently, polymer-reinforced three-dimensional (3D) graphene foams (GFs) are gaining interest in tissue engineering due to their unique morphology, biocompatibility, and similarity to extracellular matrixes. However, the homogeneous reinforcement of single polymers throughout a GF matrix does not provide tissue-level organization. Therefore, a triple-layered structure is developed in a GF matrix to closely mimic native tissue structures of the periodontium of the teeth. The scaffold aims to overcome the issue of layer separation, which generally occurs in multilayered structures due to the poor integration of various layers. The 3D GF matrix was reinforced with a polycaprolactone (PCL), polyvinyl alcohol (PVA), and PCL-hydroxyapatite (HA) mixture, added sequentially, via spin coating, vacuum, and hot air drying. Later, PVA was dissolved to create a middle layer, mimicking the periodontal fibers, while the layers present on either side resembled cementum and alveolar bone, respectively. Scanning electron microscopy and micro-computed tomography revealed the structure of the scaffold with internal differential porosities. The nanoindentation and tensile testing demonstrated the closeness of mechanical properties to that of native tissues. The biocompatibility was assessed by the MTT assay with MG63 cells (human osteosarcoma cells) exhibiting high adhesion and proliferation rate inside the 3D architecture. Summing up, this scaffold has the potential for enhancing the regeneration of various multilayered tissues.

Protocol for generating embedding-free brain organoids enriched with oligodendrocytes.

In response to the scarcity of advanced in vitro models dedicated to human CNS white matter research, we present a protocol to generate neuroectoderm-derived embedding-free human brain organoids enriched with oligodendrocytes. We describe steps for neuroectoderm differentiation, development of neural spheroids, and their transferal to Matrigel. We then detail procedures for the development, maturation, and application of oligodendrocyte-enriched brain organoids. The presence of myelin-producing cells makes these organoids useful for studying human white matter diseases, such as leukodystrophy.

Stem cell therapy: a novel & futuristic treatment modality for disaster injuries.

Stem cell therapy hold the potential to meet the demand for transplant cells/tissues needed for treating damages resulting from both natural and man-made disasters. Pluripotency makes embryonic stem cells and induced pluripotent stem cells ideal for use, but their teratogenic character is a major hindrance. Therapeutic benefits of bone marrow transplantation are well known but characterizing the potentialities of haematopoietic and mesenchymal cells is essential. Haematopoietic stem cells (HSCs) have been used for treating both haematopoietic and non-haematopoietic disorders. Ease of isolation, in vitro expansion, and hypoimmunogenecity have brought mesenchymal stem cells (MSCs) into limelight. Though differentiation of MSCs into tissue-specific cells has been reported, differentiation-independent mechanisms seem to play a more significant role in tissue repair which need to be addressed further. The safety and feasibility of MSCs have been demonstrated in clinical trials, and their use in combination with HSC for radiation injury treatment seems to have extended benefit. Therefore, using stem cells for treatment of disaster injuries along with the conventional medical practice would likely accelerate the repair process and improve the quality of life of the victim.

Microfluidic platforms for single neuron analysis.

Single-neuron actions are the basis of brain function, as clinical sequelae, neuronal dysfunction or failure for most of the central nervous system (CNS) diseases and injuries can be identified via tracing single-neurons. The bulk analysis methods tend to miscue critical information by assessing the population-averaged outcomes. However, its primary requisite in neuroscience to analyze single-neurons and to understand dynamic interplay of neurons and their environment. Microfluidic systems enable precise control over nano-to femto-liter volumes via adjusting device geometry, surface characteristics, and flow-dynamics, thus facilitating a well-defined micro-environment with spatio-temporal control for single-neuron analysis. The microfluidic platform not only offers a comprehensive landscape to study brain cell diversity at the level of transcriptome, genome, and/or epigenome of individual cells but also has a substantial role in deciphering complex dynamics of brain development and brain-related disorders. In this review, we highlight recent advances of microfluidic devices for single-neuron analysis, i.e., single-neuron trapping, single-neuron dynamics, single-neuron proteomics, single-neuron transcriptomics, drug delivery at the single-neuron level, single axon guidance, and single-neuron differentiation. Moreover, we also emphasize limitations and future challenges of single-neuron analysis by focusing on key performances of throughput and multiparametric activity analysis on microfluidic platforms.