Genomic Epidemiology of SARS-CoV-2 in Pakistan

COVID-19 has swept globally and Pakistan is no exception. To investigate the initial introductions and transmissions of the SARS-CoV-2 in Pakistan, we performed the largest genomic epidemiology study of COVID-19 in Pakistan and generated 150 complete SARS-CoV-2 genome sequences from samples collected from March 16 to June 1, 2020. We identified a total of 347 mutated positions, 31 of which were over-represented in Pakistan. Meanwhile, we found over 1000 intra-host single-nucleotide variants (iSNVs). Several of them occurred concurrently, indicating possible interactions among them or coevolution. Some of the high-frequency iSNVs in Pakistan were not observed in the global population, suggesting strong purifying selections. The genomic epidemiology revealed five distinctive spreading clusters. The largest cluster consisted of 74 viruses which were derived from different geographic locations of Pakistan and formed a deep hierarchical structure, indicating an extensive and persistent nation-wide transmission of the virus that was probably attributed to a signature mutation (G8371T in ORF1ab) of this cluster. Furthermore, 28 putative international introductions were identified, several of which are consistent with the epidemiological investigations. In all, this study has inferred the possible pathways of introductions and transmissions of SARS-CoV-2 in Pakistan, which could aid ongoing and future viral surveillance and COVID-19 control.     

Differential gene expression analysis of RAGE-S100A6 complex for target selection and the design of novel inhibitors for anticancer drug discovery

Receptor for advanced glycation end products (RAGE), a member of the immunoglobulin family, interactions with its ligands trigger downstream signaling and induce an inflammatory response linked to diabetes, inflammation, carcinogenesis, cardiovascular disease, and a variety of other human disorders. The interaction of RAGE and S100A6 has been associated with a variety of malignancies. For the control of RAGE-related illnesses, there is a great demand for more specialized drug options. To identify the most effective target for combating human malignancies associated with RAGE-S100A6 complex, we conducted single and differential gene expression analyses of S100A6 and RAGE, comparing normal and malignant tissues. Further, a structure-based virtual screening was conducted using the ZINC15 database. The chosen compounds were then subjected to a molecular docking investigation on the RAGE active site region, recognized by the various cancer-related RAGE ligands. An optimized RAGE structure was screened against a library of drug-like molecules. The screening results suggested that three promising compounds were presented as the top acceptable drug-like molecules with a high binding affinity at the RAGE V-domain catalytic region. We depicted that these compounds may be potential RAGE inhibitors and could be used to produce a successful medication against human cancer and other RAGE-related diseases based on their various assorted parameters, binding energy, hydrogen bonding, ADMET characteristics, etc. MD simulation on a time scale of 50 ns was used to test the stability of the RAGE-inhibitor complexes. Therefore, targeting RAGE and its ligands using these drug-like molecules may be an effective therapeutic approach.     

An Immunohistochemical Analysis to Validate the Rationale behind the Enhanced Immunogenicity of D-Ribosylated Low Density Lipo-Protein

Advanced glycation end products (AGEs) are thought to contribute to the abnormal lipoprotein profiles and increased risk of cardiovascular disease in patients with diabetes and renal failure. D-ribose is one of the naturally occurring pentose monosaccharide present in all living cells and is a key component of numerous biomolecules involved in many important metabolic pathways. Formation of D-ribose derived glycated low density lipoprotein (LDL) has been previously demonstrated but no studies have been performed to assess the immune complex deposition in the kidney of rabbits immunized with glycated LDL. In this study, LDL was glycated with D-ribose, and it was further used as an immunogen for immunizing NZW female rabbits. The results showed that female rabbits immunized with D-ribose modified LDL induced antibodies as detected by direct binding and competitive ELISA. The modified LDL was found to be highly immunogenic eliciting high titer immunogen-specific antibodies, while the native forms were moderately immunogenic. The induced antibodies from modified LDL exhibited wide range of heterogeneity in recognizing various proteins and amino acids conformers. Furthermore, our histopathological results illustrated the deposits of immune complex in glomerular basement membrane in rabbits immunized with D-ribose-LDL.     

Vital Parameters Assessments of Starvation Tolerance of <i>in vitro Populus alba</i> Culture

Populus alba is a large woody deciduous plant. The plant has been introduced to shooting, then multiplication of rooting on Murashige and Skoog (MS) medium. This work was designed to estimate the effect of two factors (low levels of 1-Naphthaleneacetic acid NAA and sucrose) on P. alba response resulting in 6 treatments compared to the control, with twelve measured responses. There was a significant difference in some measurements in morphology, like plantlets fresh-weight, shoot-, root-length, and leaf number. In the physiological measurements, there were significant differences in all the measured parameters. The low concentrations of sucrose and media composition/power (MS grams/L) led to starvation in plants; however, these conditions led to enhancement in some morphological and physiological parameters to overcome the starvation effect, compared to the control. The RAPD-PCR molecular marker (four decamers) was used to evaluate the new individuals’ genetic variation (instability), resulting in a total polymorphism percentage of 50.83%. It was formerly known that the plantlets were identical to each other and to the mother plant. In this study, however, the use of distinct media power, hormonal and sucrose levels resulted in molecular variation reflected in P. alba’s morphological and physiological responses.     

Effect of Exercise Interventions on Kainate Induced Status Epilepticus and Associated Co-morbidities; A Systematic Review and Meta-Analysis

Purpose

To conduct a systematic review and meta-analysis of studies testing the effect of exercise in Kainic-acid (KA) induced status-epilepticus (SE) and to quantify the efficacy of exercise strategies in the prognosis of SE and co-morbidities.

Methods

Two authors searched online databases (Pubmed and Web of Science) independently for studies testing the efficacy of exercise programs in KA-induced SE models. Reviewers autonomously extracted data on models used, exercise interventions and prognosis in all reported outcomes (behavioral, histological, biochemical and cognitive outcomes). All studies were summarized and relevant outcomes’ data were pooled by means of a meta-analysis.

Results

Among 14 selected studies; Quantitative analysis of studies with pre-SE exercise interventions showed significant reduction in mortality rate among 76 animals of four studies (RR?=?0.57, [95% CI 0.34, 0.95], p?=?0.03, I²?=?57%) and seizure rating score among three studies (n?=?56) with MD?=???1.04, [95% CI ??2.07, ??0.00], p?=?0.05, I²?=?71%. Three studies (n?=?62) presented with improved anti-oxidant enzymes’ profile (SMD?=?0.75, [95% CI 0.55, 2.31], p?=?0.0008, I²?=?44%) as a result of exercise intervention. Same intervention failed to show any significant measure for BDNF level and neuroprotection assessed through neuronal number in different brain areas with MD?=???1.22, [95% CI ??136.66, 134.22], p?=?0.99, I²?=?0% and SMD?=???0.05, [95% CI ??0.62, 0.52], p?=?0.86, I²?=?61% respectively. Qualitative review concluded in the reduction of median seizure score, depression and anxiety-like behaviors with improved cognitive performances in pre-SE exercised animals while improved memory and learning capabilities with increased neurogenesis were observed in post-SE exercised models.

Conclusions

Exercise before SE reduces behavioral seizures and oxidative stress with improvements in cognitive abilities. Post-SE exercise enhances learning and memory with neurogenesis in KA models. More extensive research on morphological and biochemical profiles is needed to explore underlying mechanisms.

     

Bacterial Thermotaxis by Speed Modulation

Naturally occurring gradients often extend over relatively long distances such that their steepness is too small for bacteria to detect. We studied the bacterial behavior in such thermal gradients. We find that bacteria migrate along shallow thermal gradients due to a change in their swimming speed resulting from the effect of temperature on the intracellular pH, which also depends on the chemical environment. When nutrients are scarce in the environment the bacteria''s intracellular pH decreases with temperature. As a result, the swimming speed of the bacteria decreases with temperature, which causes them to slowly drift toward the warm end of the thermal gradient. However, when serine is added to the medium at concentrations >300 μM, the intracellular pH increases causing the swimming speed to increase continuously with temperature, and the bacteria to drift toward the cold end of the temperature gradient. This directional migration is not a result of bacterial thermotaxis in the classical sense, because the steepness of the gradients applied is below the sensing threshold of bacteria. Nevertheless, our results show that the directional switch requires the presence of the bacterial sensing receptors. This seems to be due to the involvement of the receptors in regulating the intracellular pH.     

Crystal Structure of a Bivalent Antibody Fab Fragment

We determined the crystal structure to 1.8 Å resolution of the Fab fragment of an affinity-matured human monoclonal antibody (HC84.26.5D) that recognizes the E2 envelope glycoprotein of hepatitis C virus (HCV). Unlike conventional Fabs, which are monovalent monomers, Fab HC84.26.5D assembles into a bivalent domain-swapped dimer in which the two V_L/V_H modules are separated by ~25 Å. In solution, Fab HC84.26.5D exists predominantly as a dimer (~80%) in equilibrium with the monomeric form of the Fab (~20%). Dimerization is mediated entirely by deletion of a single residue, V_HSer113 (Kabat numbering), in the elbow region linking the V_H and C_H1 domains. In agreement with the crystal structure, dimeric Fab HC84.26.5D is able to bind two HCV E2 molecules in solution. This is only the second example of a domain-swapped Fab dimer from among >3000 Fab crystal structures determined to date. Moreover, the architecture of the doughnut-shaped Fab HC84.26.5D dimer is completely different from that of the previously reported Fab 2G12 dimer. We demonstrate that the highly identifiable shape of dimeric Fab HC84.26.5D makes it useful as a fiducial marker for single-particle cryoEM analysis of HCV E2. Bivalent domain-swapped Fab dimers engineered on the basis of HC84.26.5D may also serve as a means of doubling the effective size of conventional Fab–protein complexes for cryoEM.     

Muslims and evolution: a study of Pakistani physicians in the United States

This study investigated the views of Pakistani-American medical doctors regarding biological evolution. We used a mixed-methods approach, chiefly consisting of a short interview that presented evolution in the contexts of microbial, animal, and human evolution; evolution's acceptability or unacceptability to Muslims; and evolution's relevance to medicine. The participants were 23 doctors attending a convention in the United States. Fourteen participants accepted evolution, three rejected evolution, and six held other views. While a majority of participants indicated that they accepted evolution, a slightly smaller plurality accepted human evolution. A majority of participants, including some who did not wholly accept or reject evolution, thought that one could mutually accept evolution and also believe in Allah. Nearly every participant, including two who rejected evolution, thought that evolution was relevant to medicine. We find that participants assigned a plurality of meanings to the theory that depended on interactions between a participant’s perception of religion, science, medicine, and a host of other cultural influences. This study is the first of a collection of studies carried out by the authors, who collected data with the same instrument in five other countries with significant populations of Muslim doctors and medical students.     

Pharmacological properties of biogenically synthesized silver nanoparticles using endophyte Bacillus cereus extract of Berberis lyceum against oxidative stress and pathogenic multidrug-resistant bacteria

The emergence of multidrug resistance in pathogenic bacteria limits the utilization of available antibiotics. The development of alternate options to treat infectious diseases is the need of the day.The present study was aimed to synthesize, characterize and evaluate the bioactive properties of silver nanoparticles. Endophytic bacterium Bacillus cereus (MT193718) isolated from Berberis lycium was used to synthesize biocompatible silver nanoparticles. Antibacterial properties of AgNPs were evaluated against clinically isolated multidrug-resistant strains of Staphylococcus aureus, Pseudomonas aeruginosa, Acinetobacter baumannii and Klebsiella pneumoniae. AgNPs indicated significant antibacterial activity against S. aureus and K. pneumoniae fwith a zone of inhibition of 17 and 18 mm at a concentration of 1000 µg/ mL with minimum inhibitory concentration of 15.6 and 62.5 µg/mL respectively. Significant antioxidant activity with an IC50 value of 9.5 µg/mL was recorded. Biosynthesized AgNPs were found compatible with red blood cells at a concentration of 31.5 µg/ml with no clumping of erythrocytes. The study suggested that AgNPs synthesized by the endophytic bacterium Bacillus cereus are biologically active and can be used as antioxidant and antibacterial agents against drug-resistant bacteria.