Excavating SARS-coronavirus 2 genome for epitope-based subunit vaccine synthesis using immunoinformatics approach.

Since the outbreak of severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2) in December 2019 in China, there has been an upsurge in the number of deaths and infected individuals throughout the world, thereby leading to the World Health Organization declaration of a pandemic. Since no specific therapy is currently available for the same, the present study was aimed to explore the SARS-CoV-2 genome for the identification of immunogenic regions using immunoinformatics approach. A series of computational tools were applied in a systematic way to identify the epitopes that could be utilized in vaccine development. The screened-out epitopes were passed through several immune filters, such as promiscuousity, conservancy, antigenicity, nonallergenicity, population coverage, nonhomologous to human proteins, and affinity with human leukocyte antigen alleles, to screen out the best possible ones. Further, a construct comprising 11 CD4, 12 CD8, 3 B cell, and 3 interferon-γ epitopes, along with an adjuvant ß-defensin, was designed in silico, resulting in the formation of a multiepitope vaccine. The in silico immune simulation and population coverage analysis of the vaccine sequence showed its capacity to elicit cellular, humoral, and innate immune cells and to cover up a worldwide population of more than 97%. Further, the interaction analysis of the vaccine construct with Toll-like receptor 3 (immune receptor) was carried out by docking and dynamics simulations, revealing high affinity, constancy, and pliability between the two. The overall findings suggest that the vaccine may be highly effective, and is therefore required to be tested in the lab settings to evaluate its efficacy.

Identification of T cell and B cell epitopes against Indian HCV-genotype-3a for vaccine development- An in silico analysis.

Hepatitis C virus (HCV) infects almost 150 million people and is a leading cause of liver disease worldwide. It has been classified into seven genotypes; the most common genotype affecting Indian population is genotype 3 (60-70%). Currently there is no vaccine for any genotype of HCV. In order to develop peptide based vaccine against HCV, it is important to identify the conservancy in the circulating genotypes, along with the Human Leucocyte Antigen (HLA) alleles in the target population. The present study aims to identify conserved CD4 and CD8 T cells and B cell epitopes against Indian HCV-genotype-3a using an in silico analysis. In the present study, 28 promiscuous CD4 T cell epitopes and some CD8 epitopes were identified. The NS4 region was predicted to be the most antigenic with maximum number of conserved and promiscuous CD4 T cell epitopes and CD8 T cell epitopes having strong and intermediate affinity towards a number of HLA alleles prevalent in Indian population. Additionally, some linear B cell epitopes were also identified, which could generate neutralizing antibodies. In order to ascertain the binding pattern of the identified epitopes with HLA alleles, molecular docking analysis was carried out. The authors suggest further experimental validation to investigate the immunogenicity of the identified epitopes.

Nasal intubation: A comprehensive review.

Nasal intubation technique was first described in 1902 by Kuhn. The others pioneering the nasal intubation techniques were Macewen, Rosenberg, Meltzer and Auer, and Elsberg. It is the most common method used for giving anesthesia in oral surgeries as it provides a good field for surgeons to operate. The anatomy behind nasal intubation is necessary to know as it gives an idea about the pathway of the endotracheal tube and complications encountered during nasotracheal intubation. Various techniques can be used to intubate the patient by nasal route and all of them have their own associated complications which are discussed in this article. Various complications may arise while doing nasotracheal intubation but a thorough knowledge of the anatomy and physics behind the procedure can help reduce such complications and manage appropriately. It is important for an anesthesiologist to be well versed with the basics of nasotracheal intubation and advances in the techniques. A thorough knowledge of the anatomy and the advent of newer devices have abolished the negative effect of blindness of the procedure.

In silico study to predict promiscuous peptides for immunodiagnosis of cystic echinococcosis.

Background: Cystic echinococcosis (CE), caused by Echinococcus granulosus, is a major zoonotic disease that causes significant human morbidity and mortality. This cosmopolitan disease is difficult to diagnose, treat, and control. So far, crude extracts of hydatid cyst fluid containing antigen B or antigen 5 have been used as the primary antigenic source for its immunodiagnosis. The main issue is that it reacts with sera from people infected with other helminths. There is currently no standard, specific, or sensitive test for disease diagnosis, and no human vaccine has been reported. Aims and Objectives: Considering the need for efficient immunization and/or immunodiagnosis, six E. granulosus antigens, antigen 5, antigen B, heat shock proteins such as Hsp-8 and Hsp-90, phosphoenolpyruvate carboxykinase, and tetraspanin-1, were chosen. Materials and Methods: Using various in silico tools, T cell and B cell epitopes (promiscuous peptides) were predicted by targeting antigen 5, antigen B, heat shock proteins such as Hsp-8 and Hsp-90, phosphoenolpyruvate carboxykinase, and tetraspanin-1. Results: There are twelve promiscuous peptides with overlapping human leukocyte antigen (HLA) class-I, class-II, and conformational B cell epitopes. Such immunodominant peptides could be useful as subunit vaccines. Furthermore, six peptides specific for E. granulosus were also discovered, which may prove to be important markers in the diagnosis of CE, potentially preventing misdiagnosis and mismanagement. Conclusion: These epitopes may be the most important vaccine targets in E. granulosus because they have the most promiscuous peptides and B cell epitopes, as well as the highest affinity for different alleles, as determined by docking scores. However, additional research using in vitro and in vivo models is undertaken.

Analysis of conformational stability of interacting residues in protein binding interfaces.

After approximately 60 years of work, the protein folding problem has recently seen rapid advancement thanks to the inventions of AlphaFold and RoseTTAFold, which are machine-learning algorithms capable of reliably predicting protein structures from their sequences. A key component in their success was the inclusion of pairwise interaction information between residues. As research focus shifts towards developing algorithms to design and engineer binding proteins, it is likely that knowledge of interaction features at protein interfaces can improve predictions. Here, 574 protein complexes were analyzed to identify the stability features of their pairwise interactions, revealing that interactions between pre-stabilized residues are a selected feature in protein binding interfaces. In a retrospective analysis of 475 de novo designed binding proteins with an experimental success rate of 19%, inclusion of pairwise interaction pre-stabilization parameters increased the frequency of identifying experimentally successful binders to 40%.

MutDock: A computational docking approach for fixed-backbone protein scaffold design.

Despite the successes of antibodies as therapeutic binding proteins, they still face production and design challenges. Alternative binding scaffolds of smaller size have been developed to overcome these issues. A subset of these alternative scaffolds recognizes target molecules through mutations to a set of surface resides, which does not alter their backbone structures. While the computational design of antibodies for target epitopes has been explored in depth, the same has not been done for alternative scaffolds. The commonly used dock-and-mutate approach for binding proteins, including antibodies, is limited because it uses a constant sequence and structure representation of the scaffold. Docking fixed-backbone scaffolds with a varied group of surface amino acids increases the chances of identifying superior starting poses that can be improved with subsequent mutations. In this work, we have developed MutDock, a novel computational approach that simultaneously docks and mutates fixed backbone scaffolds for binding a target epitope by identifying a minimum number of hydrogen bonds. The approach is broadly divided into two steps. The first step uses pairwise distance alignment of hydrogen bond-forming areas of scaffold residues and compatible epitope atoms. This step considers both native and mutated rotamers of scaffold residues. The second step mutates clashing variable interface residues and thermodynamically unfavorable residues to create additional strong interactions. MutDock was used to dock two scaffolds, namely, Affibodies and DARPins, with ten randomly selected antigens. The energies of the docked poses were minimized and binding energies were compared with docked poses from ZDOCK and HADDOCK. The top MutDock poses consisted of higher and comparable binding energies than the top ZDOCK and HADDOCK poses, respectively. This work contributes to the discovery of novel binders based on smaller-sized, fixed-backbone protein scaffolds.

Machine Learning Based Lens-Free Shadow Imaging Technique for Field-Portable Cytometry.

The lens-free shadow imaging technique (LSIT) is a well-established technique for the characterization of microparticles and biological cells. Due to its simplicity and cost-effectiveness, various low-cost solutions have been developed, such as automatic analysis of complete blood count (CBC), cell viability, 2D cell morphology, 3D cell tomography, etc. The developed auto characterization algorithm so far for this custom-developed LSIT cytometer was based on the handcrafted features of the cell diffraction patterns from the LSIT cytometer, that were determined from our empirical findings on thousands of samples of individual cell types, which limit the system in terms of induction of a new cell type for auto classification or characterization. Further, its performance suffers from poor image (cell diffraction pattern) signatures due to their small signal or background noise. In this work, we address these issues by leveraging the artificial intelligence-powered auto signal enhancing scheme such as denoising autoencoder and adaptive cell characterization technique based on the transfer of learning in deep neural networks. The performance of our proposed method shows an increase in accuracy >98% along with the signal enhancement of >5 dB for most of the cell types, such as red blood cell (RBC) and white blood cell (WBC). Furthermore, the model is adaptive to learn new type of samples within a few learning iterations and able to successfully classify the newly introduced sample along with the existing other sample types.

A ricin-based peptide BRIP from Hordeum vulgare inhibits Mpro of SARS-CoV-2.

COVID-19 pandemic caused by SARS-CoV-2 led to the research aiming to find the inhibitors of this virus. Towards this world problem, an attempt was made to identify SARS-CoV-2 main protease (Mpro) inhibitory peptides from ricin domains. The ricin-based peptide from barley (BRIP) was able to inhibit Mpro in vitro with an IC50 of 0.52 nM. Its low and no cytotoxicity upto 50 µM suggested its therapeutic potential against SARS-CoV-2. The most favorable binding site on Mpro was identified by molecular docking and steered molecular dynamics (MD) simulations. The Mpro-BRIP interactions were further investigated by evaluating the trajectories for microsecond timescale MD simulations. The structural parameters of Mpro-BRIP complex were stable, and the presence of oppositely charged surfaces on the binding interface of BRIP and Mpro complex further contributed to the overall stability of the protein-peptide complex. Among the components of thermodynamic binding free energy, Van der Waals and electrostatic contributions were most favorable for complex formation. Our findings provide novel insight into the area of inhibitor development against COVID-19.

Effect of inclining strain on the crystal lattice along an extended series of lanthanide hydroxysulfates Ln(OH)SO4 (Ln = Pr-Yb, except Pm).

A series of trivalent lanthanide hydroxysulfates, Ln(OH)SO(4), (Ln = Pr through Yb, except radioactive Pm) has been synthesized via hydrothermal methods from Ln(2)(SO(4))(3)·8H(2)O by reaction with aqueous NaOH at 170 °C in Teflon lined Parr steel autoclaves, and were characterized by single crystal X-ray diffraction and FT-IR spectroscopy. Two types of arrangements were found in the solid state. The lighter (Ln = Pr-Nd, Sm-Gd) and heavier lanthanide(III) hydroxysulfates (Tb-Yb) are each isostructural. Both structure types exhibit the monoclinic space group P2(1)/n, but the unit cell content is doubled with two crystallographically distinct LnO(8) polyhedra for the heavier lanthanide compounds. The lighter complexes maintain the coordination number 9, forming a three-dimensional extended lattice. The heavier counterparts exhibit the coordination number 8, and arrange as infinite columns of two crystallographically different LnO(8) polyhedra, while extending along the "c" axis. These columns of LnO(8) polyhedra are surrounded and separated by six columns of sulfate ions, also elongating in the "c" direction. The rigid sulfate entities seem to obstruct the closing in of the lighter LnO(9) polyhedra, and show an inclining degree of torsion into the "ac" layers. The crystal lattice of the lighter 4f complexes can sufficiently withstand the tension buildup, caused by the decreasing Ln(3+) radius, up to Gd(OH)SO(4). The energy profile of this structural arrangement then seems to exceed levels at which this structure type is favorable. The lattice arrangement of the heavier Ln-analogues seems to offer a lower energy profile. This appears to be the preferred arrangement for the heavier lanthanide hydroxysulfates, whose crystal lattice exhibits more flexibility, as the coordination sphere of these analogues is less crowded. The IR absorbance frequencies of the hydroxide ligands correlate as a function of the Ln(3+) ionic radius. This corresponds well with the X-ray single crystal analysis data.

Immuno-informatics approach to design a multi-epitope vaccine to combat cytomegalovirus infection.

Human cytomegalovirus (HCMV) poses a serious public health problem causing morbidity and mortality in transplant recipients, immunocompromised patients, and congenitally infected newborns. Considering the recent reports of emergence of Ganciclovir drug resistance, vaccine development is the need of an hour. In the present study, a multi-epitope vaccine was constructed targeting the major hotspot- the pentavalent complex of glycoproteins (H/L/UL128-UL130-UL131) of HCMV, and other important target proteins- gB and pp65. The vaccine designed was composed of series of epitopes belonging to CD4, CD8 and B cells. As an immunobooster, the CpG motifs was linked to the vaccine which severed as an adjuvant. The affinity, stability and flexibility of the vaccine construct with the immune receptor- Toll like receptor -9 (TLR-9) was investigated by molecular docking and molecular dynamics simulations. The in-silico immune simulations of the vaccine sequence were also carried out to determine its ability to stimulate different immune components. Further, an in-silico cloning of the vaccine construct was performed to analyze the feasibility of its expression and translation efficiency in pET-28a (+) vector. The overall results obtained indicated the vaccine to be immunogenic, non-allergic, had high population coverage, high affinity and stability with the immune receptor, had efficient expression in host E. coli and was effective in stimulating different immune cell types like T helper, T cytotoxic, B cells, dendritic cells, macrophages and interleukins. The proposed vaccine construct is expected to be highly efficacious and needs to be carried forward by the vaccinologists to test its efficacy in lab settings.

COVID-19: the impetus for change-sustaining healthcare team communication in times of social distancing.

The COVID-19 pandemic has challenged existing healthcare systems and has made prevention of healthcare personnel exposure a high priority. Essential healthcare services, including multi-disciplinary team (MDT) meetings that make medical decisions, are expected to continue uninterrupted in this time of social distancing. There are a multitude of virtual platforms available to enable remote MDT meetings, and the pandemic has accelerated their arrival into daily healthcare practice. While we deal with a pandemic crisis, we have comprehensively reviewed and reported on the popular platforms and services available for this purpose. While each platform has its own unique features and drawbacks, it is essential to liaise with information technology departments and data governance teams to understand the optimal platforms for use within each healthcare setting. Level of evidence: Not ratable.

Designing a multi-epitope based vaccine to combat Kaposi Sarcoma utilizing immunoinformatics approach.

Kaposi's sarcoma-associated herpesvirus (KSHV) responsible for causing Kaposi sarcoma (KS), an opportunistic angioproliferative neoplasm is emerging rapidly. Despite this there is no permanent cure for this disease. The present study was aimed to design a multi-epitope based vaccine targeting the major glycoproteins of KSHV which plays an important role in the virus entry. After the application of rigorous immunoinformatics analysis and several immune filters, the multi-epitope vaccine was constructed, consisting of CD4, CD8 and IFN-γ inducing epitopes. Several physiochemical characteristics, allergenicity and antigenicity of the multi-epitope vaccine were analyzed in order to ensure its safety and immunogenicity. Further, the binding affinity and stability of the vaccine with Toll like receptor -9 (TLR-9) was analyzed by molecular docking and dynamics simulation studies. In addition, an in silico cloning was performed to ensure the expression and translation efficiency of the vaccine, utilizing pET-28a (+) vector. Such T-cell-based immunotherapies which leverage this mechanism could prove their potential against cancer. Further, the authors propose to test the present findings in the lab settings to ensure the safety, immunogenicity and efficacy of the presented vaccine which may help in controlling KSHV infection.

Clinical implications of scalp ictal EEG pattern in patients with temporal lobe epilepsy.

OBJECTIVE: To determine the clinical implications of scalp ictal EEG pattern in patients with temporal lobe epilepsy (TLE). METHODS: Scalp EEG ictal patterns were retrospectively determined in 27 consecutive patients with medically refractory temporal lobe epilepsy who underwent phase-1 scalp video-EEG and phase-2 simultaneous scalp and intracranial video-EEG recordings for pre-surgical evaluation. RESULTS: Of the 192 temporal lobe seizures recorded during phase-1 and phase-2 scalp video-EEG studies, 124 (65%) seizures were associated with theta/alpha (5-9 Hz) ictal onset pattern, and 68 (35%) seizures were associated with delta (2-5 Hz) ictal onset pattern. Fourteen (52%) patients had exclusively theta/alpha ictal onset, 3 (11%) patients had exclusively delta ictal onset, and 10 (37%) patients had mixed theta/alpha and delta ictal onsets. MTLE was observed in 26 patients who had 124 seizures with theta/alpha ictal onset and 59 seizures with delta ictal onset. LTLE was observed in one patient who had 9 seizures with delta ictal onset. Scalp ictal EEG pattern was not significantly correlated with postsurgical seizure outcomes. CONCLUSIONS: Both scalp delta and theta/alpha ictal onset patterns can be commonly found in patients with MTLE. SIGNIFICANCE: Scalp delta ictal onset is not a unique EEG pattern for LTLE as commonly believed.

Identification of broadly reactive epitopes targeting major glycoproteins of Herpes simplex virus (HSV) 1 and 2 - An immunoinformatics analysis.

Infections due to both HSV-1 and HSV-2 constitute an enormous health burden worldwide. Development of vaccine against herpes infections is a WHO supported public health priority. The viral glycoproteins have always been the major hotspots for vaccine designing. The present study was aimed to identify the conserved T and B cell epitopes in the major glycoproteins of both HSV-1 and HSV-2 via rigorous computational approaches. Identification of promiscuous T cell epitopes is of utmost importance in vaccine designing as such epitopes are capable of binding to several allelic forms of HLA and could generate effective immune response in the host. The criteria designed for identification of T and B cell epitopes was that it should be conserved in both HSV-1 and 2, promiscuous, have high affinity towards HLA alleles, should be located on the surface of glycoproteins and not be present in the glycosylation sites. This study led to the identification of 17 HLA Class II and 26 HLA Class I T cell epitopes, 9 linear and some conformational B cell epitopes. The identified T cell epitopes were further subjected to molecular docking analysis to analyze their binding patterns. Altogether we have identified 4 most promising regions in glycoproteins (2-gB, 1-gD, 1-gH) of HSV-1 and 2 which are promiscuous to HLA Class II alleles and have overlapping HLA Class I and B cell epitopes, which could be very useful in generating both arms of immune response in the host i.e. adaptive as well as humoral immunity. Further the authors propose the cross-validation of the identified epitopes in experimental settings for confirming their immunogenicity to support the present findings.

Plaster Burn: Challenge to Plastic Surgeon.

BACKGROUND: The thermal burn is accidental and also is the hidden and worst complication of medical plaster application. This study evaluated clinical and aetiological profile and severity of plaster burns. METHODS: In Department of Burns, Plastic and Reconstructive Surgery from 1st August 2014 to 31st December 2015, six patients with plaster burn were assessed for total body surface area and depth of burn. The wounds were cultured and dressed with moist dressings daily till the surgical procedure and satisfactory healing. RESULTS: The youngest patient was 10 and oldest 65 years (mean age: 40.20±6.67 years, 4 males and two females with ratio of 2:1). Three patients sustained plaster burn injury accidentally at home and 3 developed burn after medical application of plaster. The hands burns were observed commonly in accidental plaster burns, while ankle was often involved in iatrogenic plaster burns. The iatrogenic burns were mostly deep in thickness varying 2nd to 4th degree, while accidental burns were often 2nd degree. Superficial 2nd degree burns were managed conservatively, and deep 2nd degree burns were skin grafted. Fourth degree burn was managed with reverse sural flap alone and another with vacuum-assisted closure followed by reverse sural flap and skin grafting. CONCLUSION: Plaster burn is still a challenge to plastic surgeon and it is advised for applying casts to utilize all available means to limit the amount of casting material provided. It can be accomplished without compromising the cast strength to minimize the risk of thermal injury when applying plaster or composite casts.