Net-Shaped DNA Nanostructures Designed for Rapid/Sensitive Detection and Potential Inhibition of the SARS-CoV-2 Virus.

We present a net-shaped DNA nanostructure (called "DNA Net" herein) design strategy for selective recognition and high-affinity capture of intact SARS-CoV-2 virions through spatial pattern-matching and multivalent interactions between the aptamers (targeting wild-type spike-RBD) positioned on the DNA Net and the trimeric spike glycoproteins displayed on the viral outer surface. Carrying a designer nanoswitch, the DNA Net-aptamers release fluorescence signals upon virus binding that are easily read with a handheld fluorimeter for a rapid (in 10 min), simple (mix-and-read), sensitive (PCR equivalent), room temperature compatible, and inexpensive (∼$1.26/test) COVID-19 test assay. The DNA Net-aptamers also impede authentic wild-type SARS-CoV-2 infection in cell culture with a near 1 × 103-fold enhancement of the monomeric aptamer. Furthermore, our DNA Net design principle and strategy can be customized to tackle other life-threatening and economically influential viruses like influenza and HIV, whose surfaces carry class-I viral envelope glycoproteins like the SARS-CoV-2 spikes in trimeric forms.

Determining the effect of climate and profession on dry eye disease: A prevalence study among young males in north, north-west and central India.

Background: Ocular tear film and the ocular surface together represent a dynamic playground of various protective factors against environmental influencers. Surrounding environment and geoclimatic milieu are the pertinent and continuous influencing factors. Prevalence study of Dry eye disease (DED) in various climatic conditions and professions provides an insight into this peculiar aspect of DED and human geography. Methods: This was a hospital based, cross-sectional, comparative, analytical observational study. 1840 young soldiers between 25 and 45 years of age were studied. Divided in five climate groups based on Köppen-Geiger climate classification system the subjects were sub-grouped into professions exposed to known aggravating environmental factors. The ocular surface was examined for signs and symptoms of DED and its prevalence and risk ratio for different climates and professions were calculated. Results: Highest prevalence of DED was found in cold desert climate and among visual display unit (VDU) users. Least prevalence was seen in highland type of climate and among people involved in professional exposed to high temperature. Prevalence of DED based upon the signs and ocular surface disease index (OSDI) was calculated and stratified with risk ratio for climatic conditions and professions. Conclusion: Different climatic conditions and professions had different prevalence of signs, symptoms of OSD and DED which indicated its influence on prevalence of the disease.

Homologous pairing in short double-stranded DNA-grafted colloidal microspheres.

Homologous pairing (HP), i.e., the pairing of similar or identical double-stranded DNA, is an insufficiently understood fundamental biological process. HP is now understood to also occur without protein mediation, but crucial mechanistic details remain poorly established. Unfortunately, systematic studies of sequence dependence are not practical due to the enormous number of nucleotide permutations and multiple possible conformations involved in existing biophysical strategies even when using as few as 150 basepairs. Here, we show that HP can occur in DNA as short as 18 basepairs in a colloidal microparticle-based system. Exemplary systematic studies include resolving opposing reports of the impact of % AT composition, validating the impact of nucleotide order and triplet framework and revealing isotropic bendability to be crucial for HP. These studies are enabled by statistical analysis of crystal size and fraction within coexisting fluid-crystal phases of double-stranded DNA-grafted colloidal microspheres, where crystallization is predicated by HP.

Enhanced translation expands the endo-lysosome size and promotes antigen presentation during phagocyte activation.

The mechanisms that govern organelle adaptation and remodelling remain poorly defined. The endo-lysosomal system degrades cargo from various routes, including endocytosis, phagocytosis, and autophagy. For phagocytes, endosomes and lysosomes (endo-lysosomes) are kingpin organelles because they are essential to kill pathogens and process and present antigens. During phagocyte activation, endo-lysosomes undergo a morphological transformation, going from a collection of dozens of globular structures to a tubular network in a process that requires the phosphatidylinositol-3-kinase-AKT-mechanistic target of rapamycin (mTOR) signalling pathway. Here, we show that the endo-lysosomal system undergoes an expansion in volume and holding capacity during phagocyte activation within 2 h of lipopolysaccharides (LPS) stimulation. Endo-lysosomal expansion was paralleled by an increase in lysosomal protein levels, but this was unexpectedly largely independent of the transcription factor EB (TFEB) and transcription factor E3 (TFE3), which are known to scale up lysosome biogenesis. Instead, we demonstrate a hitherto unappreciated mechanism of acute organelle expansion via mTOR Complex 1 (mTORC1)-dependent increase in translation, which appears to be mediated by both S6Ks and 4E-BPs. Moreover, we show that stimulation of RAW 264.7 macrophage cell line with LPS alters translation of a subset but not all of mRNAs encoding endo-lysosomal proteins, thereby suggesting that endo-lysosome expansion is accompanied by functional remodelling. Importantly, mTORC1-dependent increase in translation activity was necessary for efficient and rapid antigen presentation by dendritic cells. Collectively, we identified a previously unknown and functionally relevant mechanism for endo-lysosome expansion that relies on mTORC1-dependent translation to stimulate endo-lysosome biogenesis in response to an infection signal.

Overcoming the limitations of COVID-19 diagnostics with nanostructures, nucleic acid engineering, and additive manufacturing.

The COVID-19 pandemic revealed fundamental limitations in the current model for infectious disease diagnosis and serology, based upon complex assay workflows, laboratory-based instrumentation, and expensive materials for managing samples and reagents. The lengthy time delays required to obtain test results, the high cost of gold-standard PCR tests, and poor sensitivity of rapid point-of-care tests contributed directly to society's inability to efficiently identify COVID-19-positive individuals for quarantine, which in turn continues to impact return to normal activities throughout the economy. Over the past year, enormous resources have been invested to develop more effective rapid tests and laboratory tests with greater throughput, yet the vast majority of engineering and chemistry approaches are merely incremental improvements to existing methods for nucleic acid amplification, lateral flow test strips, and enzymatic amplification assays for protein-based biomarkers. Meanwhile, widespread commercial availability of new test kits continues to be hampered by the cost and time required to develop single-use disposable microfluidic plastic cartridges manufactured by injection molding. Through development of novel technologies for sensitive, selective, rapid, and robust viral detection and more efficient approaches for scalable manufacturing of microfluidic devices, we can be much better prepared for future management of infectious pathogen outbreaks. Here, we describe how photonic metamaterials, graphene nanomaterials, designer DNA nanostructures, and polymers amenable to scalable additive manufacturing are being applied towards overcoming the fundamental limitations of currently dominant COVID-19 diagnostic approaches. In this paper, we review how several distinct classes of nanomaterials and nanochemistry enable simple assay workflows, high sensitivity, inexpensive instrumentation, point-of-care sample-to-answer virus diagnosis, and rapidly scaled manufacturing.

Virtual Screening of Alkaloid and Terpenoid Inhibitors of SMT Expressed in Naegleria sp.

The pathogenic form of thermophilic Naegleria sp. i.e., Naegleria fowleri, also known as brain eating amoeba, causes primary amoebic encephalitis (PAM) with a >97% fatality rate. To date, there are no specific drugs identified to treat this disease specifically. The present antimicrobial combinatorial chemotherapy is hard on many patients, especially children. Interestingly, Naegleria fowleri has complex lipid biosynthesis pathways like other protists and also has a strong preference to utilize absorbed host lipids for generating energy. The ergosterol biosynthesis pathway provides a unique drug target opportunity, as some of the key enzymes involved in this pathway are absent in humans. Sterol 24-C Methyltransferase (SMT) is one such enzyme that is not found in humans. To select novel inhibitors for this enzyme, alkaloids and terpenoids inhibitors were screened and tested against two isozymes of SMT identified in N. gruberi (non-pathogenic) as well as its homolog found in yeast, i.e., ERG6. Five natural product derived inhibitors i.e., Cyclopamine, Chelerythrine, Berberine, Tanshinone 2A, and Catharanthine have been identified as potential drug candidates based on multiple criteria including binding affinity, ADME scores, absorption, and, most importantly, its ability to cross the blood brain barrier. This study provides multiple leads for future drug exploration against Naegleria fowleri.

The Science Behind Nefertiti's Beauty: A Plastic Surgeon's Analysis.

Introduction The famous stucco limestone coated "Bust of Nefertiti" housed in the Neues Museum, Germany dated 1,345 BC is an icon of beauty. Sculpted around three millennia ago by Thutmose, the bust still emits a charm that leaves its audience spellbound. However, no one, to the best of author's knowledge, has analyzed this sculpture or its photographs objectively to determine if there is any scientific basis to its attractiveness. Materials and Methods High-resolution photographs of the bust were anthropometrically analyzed in frontal and right lateral profile views using neoclassical canons and Farkas' studies. Results The photographs of the bust exhibit many of the neoclassical canons and proportions of Farkas' studies exactly, while many of the remaining are very close to these measurements. A few measurements are out of range of what is considered acceptable these days; however, her overall appearance is pleasing. Conclusion Despite passage of more than three millennia, the proportions and parameters defining beautiful faces have largely remained unchanged.

Endoplasmic reticulum-plasma membrane contact sites integrate sterol and phospholipid regulation.

Tether proteins attach the endoplasmic reticulum (ER) to other cellular membranes, thereby creating contact sites that are proposed to form platforms for regulating lipid homeostasis and facilitating non-vesicular lipid exchange. Sterols are synthesized in the ER and transported by non-vesicular mechanisms to the plasma membrane (PM), where they represent almost half of all PM lipids and contribute critically to the barrier function of the PM. To determine whether contact sites are important for both sterol exchange between the ER and PM and intermembrane regulation of lipid metabolism, we generated Δ-super-tether (Δ-s-tether) yeast cells that lack six previously identified tethering proteins (yeast extended synatotagmin [E-Syt], vesicle-associated membrane protein [VAMP]-associated protein [VAP], and TMEM16-anoctamin homologues) as well as the presumptive tether Ice2. Despite the lack of ER-PM contacts in these cells, ER-PM sterol exchange is robust, indicating that the sterol transport machinery is either absent from or not uniquely located at contact sites. Unexpectedly, we found that the transport of exogenously supplied sterol to the ER occurs more slowly in Δ-s-tether cells than in wild-type (WT) cells. We pinpointed this defect to changes in sterol organization and transbilayer movement within the PM bilayer caused by phospholipid dysregulation, evinced by changes in the abundance and organization of PM lipids. Indeed, deletion of either OSH4, which encodes a sterol/phosphatidylinositol-4-phosphate (PI4P) exchange protein, or SAC1, which encodes a PI4P phosphatase, caused synthetic lethality in Δ-s-tether cells due to disruptions in redundant PI4P and phospholipid regulatory pathways. The growth defect of Δ-s-tether cells was rescued with an artificial "ER-PM staple," a tether assembled from unrelated non-yeast protein domains, indicating that endogenous tether proteins have nonspecific bridging functions. Finally, we discovered that sterols play a role in regulating ER-PM contact site formation. In sterol-depleted cells, levels of the yeast E-Syt tether Tcb3 were induced and ER-PM contact increased dramatically. These results support a model in which ER-PM contact sites provide a nexus for coordinating the complex interrelationship between sterols, sphingolipids, and phospholipids that maintain PM composition and integrity.

Methylene Chloride Burns: A Case Report and Review of Literature.

Chemical burns account for a small percentage of burns but contribute to significant number of burn-related mortalities. The major challenge posed by chemical burns is difficulty in correct depth estimation, as the damage continues to progress until they are effectively neutralized. Besides the most common etiology of chemical burns by alkalis and acids, there are many other unique causes of chemical burns. The author describes a case of chemical burns by heavy duty paint remover, the main composition of which is methylene chloride. Although several studies about methylene chloride poisoning are available in literature, only one case report of burns due to methylene chloride exists in literature. These burns are unique in their presentation. The author describes the presentation and challenges in management of this burn. Clinical trial registration Not applicable.

A PhotoClick cholesterol-based quantitative proteomics screen for cytoplasmic sterol-binding proteins in Saccharomyces cerevisiae.

Ergosterol is a prominent component of the yeast plasma membrane and essential for yeast cell viability. It is synthesized in the endoplasmic reticulum and transported to the plasma membrane by nonvesicular mechanisms requiring carrier proteins. Oxysterol-binding protein homologues and yeast StARkin proteins have been proposed to function as sterol carriers. Although many of these proteins are capable of transporting sterols between synthetic lipid vesicles in vitro, they are not essential for ergosterol transport in cells, indicating that they may be functionally redundant with each other or with additional-as yet unidentified-sterol carriers. To address this point, we hypothesized that sterol transport proteins are also sterol-binding proteins (SBPs), and used an in vitro chemoproteomic strategy to identify all cytosolic SBPs. We generated a cytosol fraction enriched in SBPs and captured the proteins with a photoreactive clickable cholesterol analogue. Quantitative proteomics of the captured proteins identified 342 putative SBPs. Analysis of these identified proteins based on their annotated function, reported drug phenotypes, interactions with proteins regulating lipid metabolism, gene ontology, and presence of mammalian orthologues revealed a subset of 62 characterized and nine uncharacterized candidates. Five of the uncharacterized proteins play a role in maintaining plasma membrane integrity as their absence affects the ability of cells to grow in the presence of nystatin or myriocin. We anticipate that the dataset reported here will be a comprehensive resource for functional analysis of sterol-binding/transport proteins and provide insights into novel aspects of non-vesicular sterol trafficking.

The "Safdarjung Method" of Estimation of Total Body Surface Area (TBSA) Burns in Major Amputees.

Accurate estimation of total body surface area (TBSA) burns is the cornerstone of initiating correct fluid therapy in burns. The current formulae are designed for patients with intact body. However, the authors having worked at a high-volume burns center (Safdarjung Hospital, New Delhi, India) have encountered patients with major amputations presenting with burns. The existing formulae are inaccurate for estimating TBSA in major amputees, leading to inaccurate fluid estimation in such cases. The authors have designed a novel method to estimate TBSA burns in patients with major amputations that involves a "correction factor" to account for the amputated parts.

A Giant Pseudoaneurysm Mimicking Retropharyngeal Abscess in a Child.

Pseudoaneurysms of the cervical internal carotid artery are rare and usually result from trauma, infection, or rarely spontaneously. They harbor potential risk of life-threatening hemorrhage and warrant immediate management. Endovascular treatment in the form of stent placement and coiling is a well established technique for dealing with both intracranial and extracranial pseudoaneurysms. We present a case of a child who presented with fever, neck swelling, dysphagia, and respiratory distress in emergency for which clinical diagnosis of a neck abscess was made. Intubation was not possible because of bulge in the retropharyngeal wall. An elective tracheotomy was planned with imaging of the neck. Imaging and careful planning to avoid incision and drainage in the initial presentation led us to diagnose a pseudoaneurysm and successful management. We present this case to highlight the importance of early recognition and timely meticulous management to avoid serious complications and morbidity in a child.

Morphogenesis checkpoint kinase Swe1 is the executor of lipolysis-dependent cell-cycle progression.

Cell growth and division requires the precise duplication of cellular DNA content but also of membranes and organelles. Knowledge about the cell-cycle-dependent regulation of membrane and storage lipid homeostasis is only rudimentary. Previous work from our laboratory has shown that the breakdown of triacylglycerols (TGs) is regulated in a cell-cycle-dependent manner, by activation of the Tgl4 lipase by the major cyclin-dependent kinase Cdc28. The lipases Tgl3 and Tgl4 are required for efficient cell-cycle progression during the G1/S (Gap1/replication phase) transition, at the onset of bud formation, and their absence leads to a cell-cycle delay. We now show that defective lipolysis activates the Swe1 morphogenesis checkpoint kinase that halts cell-cycle progression by phosphorylation of Cdc28 at tyrosine residue 19. Saturated long-chain fatty acids and phytosphingosine supplementation rescue the cell-cycle delay in the Tgl3/Tgl4 lipase-deficient strain, suggesting that Swe1 activity responds to imbalanced sphingolipid metabolism, in the absence of TG degradation. We propose a model by which TG-derived sphingolipids are required to activate the protein phosphatase 2A (PP2A(Cdc55)) to attenuate Swe1 phosphorylation and its inhibitory effect on Cdc28 at the G1/S transition of the cell cycle.

Translabyrinthine approach to internal auditory meatus: A retrospective study.

BACKGROUND: Ear and the lateral skull base surgery is challenging and yet fascinating for a Neuro-otologist. A thorough knowledge of the complex anatomy is indispensable for the surgeon in order to provide the best possible care. METHODS: The aim of the study was to highlight the present day indications for translabyrinthine approach to IAM from a Neuro-otologist perspective. RESULTS: There were a total of 7 patients who underwent Translabyrinthine approach at our centre. In the present study we have reported cases of Vestibular Schwannoma, Facial nerve schwannoma, Cholesteatoma involving the IAM, Meniere's disease with refractory vertigo which were managed via translabyrinthine approach. We also encountered, probably the first reported case, tuberculoma of the IAM which was clinical suspected to be vestibular schwannoma. CONCLUSION: The article presents different clinical situations where this approach can be suitably utilized and has been dealt with via a retrospective study encountered at our centre.

Regulation of Sphingolipid Biosynthesis by the Morphogenesis Checkpoint Kinase Swe1.

Sphingolipid (SL) biosynthesis is negatively regulated by the highly conserved endoplasmic reticulum-localized Orm family proteins. Defective SL synthesis in Saccharomyces cerevisiae leads to increased phosphorylation and inhibition of Orm proteins by the kinase Ypk1. Here we present evidence that the yeast morphogenesis checkpoint kinase, Swe1, regulates SL biosynthesis independent of the Ypk1 pathway. Deletion of the Swe1 kinase renders mutant cells sensitive to serine palmitoyltransferase inhibition due to impaired sphingoid long-chain base synthesis. Based on these data and previous results, we suggest that Swe1 kinase perceives alterations in SL homeostasis, activates SL synthesis, and may thus represent the missing regulatory link that controls the SL rheostat during the cell cycle.

Lipid topogenesis--35years on.

Glycerophospholipids are the principal fabric of cellular membranes. The pathways by which these lipids are synthesized were elucidated mainly through the work of Kennedy and colleagues in the late 1950s and early 1960s. Subsequently, attention turned to cell biological aspects of lipids: Where in the cell are lipids synthesized? How are lipids integrated into membranes to form a bilayer? How are they sorted and transported from their site of synthesis to other cellular destinations? These topics, collectively termed 'lipid topogenesis', were the subject of a review article in 1981 by Bell, Ballas and Coleman. We now assess what has been learned about early events of lipid topogenesis, i.e. "lipid synthesis, the integration of lipids into membranes, and lipid translocation across membranes", in the 35 years since the publication of this important review. We highlight the recent elucidation of the X-ray structures of key membrane enzymes of glycerophospholipid synthesis, progress on identifying lipid scramblase proteins needed to equilibrate lipids across membranes, and new complexities in the subcellular location and membrane topology of phosphatidylinositol synthesis revealed through a comparison of two unicellular model eukaryotes. This article is part of a Special Issue entitled: The cellular lipid landscape edited by Tim P. Levine and Anant K. Menon.