16S rRNA metagenomic profiling of red amaranth grown organically with different composts and soils.

In recent years organic food is gaining popularity as it is believed to promote better human health and improve soil sustainability, but there are apprehensions about pathogens in organic produces. This study was designed to understand the effect of different composts and soils on the status of the microbiome present in organically grown leafy vegetables. 16S rRNA metagenomic profiling of the leaves was done, and data were analyzed. It was found that by adding composts, the OTU of the microbiome in the organic produce was higher than in the conventional produce. The beneficial genera identified across the samples included plant growth promoters (Achromobacter, Paenibacillus, Pseudomonas, Sphingobacterium) and probiotics (Lactobacillus), which were higher in the organic produce. Some pathogenic genera, viz., plant pathogenic bacteria (Cellvibrio, Georgenia) and human pathogenic bacteria (Corynebacterium, Acinetobacter, Streptococcus, Streptomyces) were also found but with relatively low counts in the organic produce. Thus, the present study highlights that organic produce has lesser pathogen contamination than the conventional produce. KEY POINTS: • 16S rRNA metagenomics profiling done for organic red amaranth cultivar • Microbial richness varied with respect to the soil and compost type used • The ratio of beneficial to pathogenic genera improves with the addition of compost.

Optimizing the input feature sets and machine learning algorithms for reliable and accurate estimation of continuous, cuffless blood pressure.

The advent of mobile devices, wearables and digital healthcare has unleashed a demand for accurate, reliable, and non-interventional ways to measure continuous blood pressure (BP). Many consumer products claim to measure BP with a cuffless device, but their lack of accuracy and reliability limit clinical adoption. Here, we demonstrate how multimodal feature datasets, comprising: (i) pulse arrival time (PAT); (ii) pulse wave morphology (PWM), and (iii) demographic data, can be combined with optimized Machine Learning (ML) algorithms to estimate Systolic BP (SBP), Diastolic BP (DBP) and Mean Arterial Pressure (MAP) within a 5 mmHg bias of the gold standard Intra-Arterial BP, well within the acceptable limits of the IEC/ANSI 80601-2-30 (2018) standard. Furthermore, DBP's calculated using 126 datasets collected from 31 hemodynamically compromised patients had a standard deviation within 8 mmHg, while SBP's and MAP's exceeded these limits. Using ANOVA and Levene's test for error means and standard deviations, we found significant differences in the various ML algorithms but found no significant differences amongst the multimodal feature datasets. Optimized ML algorithms and key multimodal features obtained from larger real-world data (RWD) sets could enable more reliable and accurate estimation of continuous BP in cuffless devices, accelerating wider clinical adoption.

Oral gabapentin for the treatment of postoperative pain after photorefractive keratectomy.

PURPOSE: To evaluate oral gabapentin for postoperative pain after photorefractive keratectomy (PRK). DESIGN: Prospective, nonrandomized clinical trial. METHODS: In additional to a standard regimen of topical antibiotics, topical steroids, and topical tetracaine as required, all PRK patients at our laser vision center were treated after surgery for pain for a two-month period with Percocet (oxycodone/acetaminophen) [Endo Pharmaceuticals; Chadds Ford, Pennsylvania, USA] 5 mg/325 mg as required for three days (control group). Patients completed a pain assessment survey using a faces pain scale (from zero through 6) on the evening of surgery and each subsequent morning and evening until postoperative day 3. A successive cohort of patients received Neurontin (gabapentin) [Pfizer, New York, New York, USA] 300 mg thrice daily (first dose administered two hours or more before the procedure) as an oral pain medication for three days, and the same survey data were collected. RESULTS: Data were collected on 141 patients in each cohort. Successful pain management score (defined as faces zero through 2 on the scale) differences did not reach statistical significance between the two cohorts except on the morning of the second postoperative day, when gabapentin was superior. On all postoperative days, patients in the oxycodone/acetaminophen cohort used significantly less tetracaine eye drops as required. The percent of patients rating overall pain experience as better than expected was 35% and 36%, those rating pain experience as about what was expected was 50% and 49%, and those rating pain experience as worse than expected was 15% and 15% in the oxycodone/acetaminophen and gabapentin cohorts, respectively. CONCLUSIONS: We found no difference in overall subjective pain management ratings between gabapentin and oxycodone/acetaminophen for postoperative PRK pain, although gabapentin was associated with significantly more frequent use of anesthetic eye drops as required.

7,8-Dihydroxy-4-methylcoumarin induces apoptosis of human lung adenocarcinoma cells by ROS-independent mitochondrial pathway through partial inhibition of ERK/MAPK signaling.

Coumarins have attracted intense interest in recent years because they have been identified from natural sources, especially green plants and have diverse pharmacological properties. In this study, we investigated whether 7,8-dihydroxy-4-methylcoumarin (DHMC) caused apoptosis in A549 human non-small cell lung carcinoma cells (NSCLC) and, if so, by what mechanisms. Here, we show that, in A549 human NSCLC cells, DHMC induces apoptosis through mitochondria-mediated caspase-dependent pathway. Although an increase in the levels of reactive oxygen species (ROS) was observed, pre-treatment with antioxidant showed no protective effect against DHMC-induced apoptosis. In addition, our immunoblot data revealed that DHMC treatment led to down-regulation of Bcl-xl, Bax, p21, Cox-2, p53 and upregulation of c-Myc. Results in the present study for the first time suggest that DHMC induces apoptosis in human lung A549 cells through partial inhibition of ERK/MAPK signaling.

PEI-alginate nanocomposites as efficient in vitro gene transfection agents.

The positive charge on PEI was partially shielded by forming ionic nanocomposites with a polysaccharide, alginic acid, in aqueous solution, bypassing tedious chemical synthesis. The content of alginic acid was varied systematically to obtain a series of nanocomposites. The nanocomposites were first characterized by assessing the surface charge (zeta potential), size (DLS) and morphology (AFM) followed by evaluation for their DNA interaction ability, cytotoxicity and transfection efficiency on various cell lines. The transfection efficiency of PEI-alginate (6.26%) nanocomposites improved dramatically (2-16-fold over native PEI) in all the cell lines studied. However, a decrease in transfection efficiency was observed on deviating from this optimal concentration of alginic acid in nanocomposites. Cytotoxicity of PEI-alginate/DNA complexes was nearly abolished on increasing the concentration of alginic acid in nanocomposites. PEI-alginate (6.26%) nanocomposites also delivered SiRNAs efficiently into mammalian cells, resulting in 80% suppression of GFP expression. The cellular uptake and endosomal escape of PEI-alginate nanocomposites and PEI were found to follow a similar route when transfection was carried out in presence of chloroquine, bafilomycin A1, cytochalasin B and methyl-beta-cyclodextrin. The results demonstrate a versatile vector that can be used for efficient cytoplasmic delivery of a broad range of nucleic acids.

Unifying themes in DNA replication: reconciling single molecule kinetic studies with structural data on DNA polymerases.

Structural data suggest that DNA polymerases, from at least three different families, employ common strategies for carrying out DNA replication. Universal features include a large conformational change in the enzyme-template complex and a conserved active-site geometry that imposes a sharp kink at the 5 end of the template strand. Recent single molecule experiments have shown that stretching the DNA template markedly alters the rate of DNA synthesis catalyzed by these motor enzymes. From these data, it was previously inferred that T7 DNA polymerase and two related enzymes convert two or four (depending on the enzyme) single-stranded (ss) template bases to double helix geometry in the polymerase active site during each catalytic cycle. We discuss structural data on related DNA polymerases, which suggest that only one (ss) template base is contracted to dsDNA geometry during the rate-limiting step of each replication cycle. Previous interpretations relied upon the global stretching curves for DNA polymers alone (with no reference to the enzyme or the structure of the transition state). In contrast, we present a structurally guided model that presumes the force dependence of the replication rate is governed chiefly by local interactions in the immediate vicinity of the enzyme s active site. Our analysis reconciles single molecule kinetic studies with structural data on DNA polymerases.

Apoptogenic effect of 7,8-diacetoxy-4-methylcoumarin and 7,8-diacetoxy-4-methylthiocoumarin in human lung adenocarcinoma cell line: role of NF-kappaB, Akt, ROS and MAP kinase pathway.

Coumarin (1,2-benzopyrone) is a naturally occurring fragrant compound found in a variety of plants and spices. Coumarins have attracted intense interest in recent years because of their diverse pharmacological activities. This study examines the antioxidant coumarin 7,8-diacetoxy-4-methylcoumarin (DAMC) and its thiocoumarin derivative 7,8-diacetoxy-4-methylthiocoumarin (DAMTC) for their effect on human non-small cell lung cancer A549 cells. Here we show that both DAMC and DAMTC not only inhibited cell proliferation, but also induced apoptosis with an IC(50) of 160 microg/ml as confirmed by morphological examination, annexin-V assay and flow cytometric analysis. Interestingly, it was observed that these two coumarin compounds exhibited little cytotoxicity towards peripheral blood mononuclear cells but induced apoptosis in malignant cells. DAMC/DAMTC treatment also resulted in pronounced release of apoptogenic cytochrome c from mitochondria to cytosol, alteration of mitochondrial membrane potential (DeltaPsi(m)), and activation of caspase-9 and caspase-3. Although an increase in the levels of reactive oxygen species (ROS) was observed, pre-treatment with antioxidant showed no protective effect against DAMC/DAMTC-induced apoptosis. Results of present study suggest that downregulation of Bcl-xl, Cox-2 and mitogen activated protein kinase pathway and upregulation of p53, Akt and NF-kappaB pathway are involved in the underlying molecular mechanism of apoptosis induction by DAMC and DAMTC in A549 cells.

Harnessing biological motors to engineer systems for nanoscale transport and assembly.

Living systems use biological nanomotors to build life's essential molecules--such as DNA and proteins--as well as to transport cargo inside cells with both spatial and temporal precision. Each motor is highly specialized and carries out a distinct function within the cell. Some have even evolved sophisticated mechanisms to ensure quality control during nanomanufacturing processes, whether to correct errors in biosynthesis or to detect and permit the repair of damaged transport highways. In general, these nanomotors consume chemical energy in order to undergo a series of shape changes that let them interact sequentially with other molecules. Here we review some of the many tasks that biomotors perform and analyse their underlying design principles from an engineering perspective. We also discuss experiments and strategies to integrate biomotors into synthetic environments for applications such as sensing, transport and assembly.

Tuning and switching a DNA polymerase motor with mechanical tension.

Recent single-molecule experiments reveal that mechanical tension on DNA can control both the speed and direction of the DNA polymerase motor. We present a theoretical description of this tension-induced "tuning" and "switching." The internal conformational states of the enzyme motor are represented as nodes, and the allowed transitions between states as links, of a biochemical network. The motor moves along the DNA by cycling through a given sequence of internal states. Tension and other external control parameters, particularly the ambient concentrations of enzyme, nucleotides, and pyrophosphates, couple into the internal conformational dynamics of the motor, thereby regulating the steady-state flux through the network. The network links are specified by bulk-phase kinetic data (in the absence of tension), and rudimentary models are used to describe the dependence on tension of key links. We find that this network analysis simulates well the chief results from single-molecule experiments including the tension-induced attenuation of polymerase activity, the onset of exonucleolysis at high tension, and insensitivity to large changes in concentration of the enzyme. A major dependence of the switching tension on the nucleotide concentration is also predicted.

Dependence of DNA polymerase replication rate on external forces: a model based on molecular dynamics simulations.

Molecular dynamics simulations are presented for a Thermus aquaticus (Taq) DNA polymerase I complex (consisting of the protein, the primer-template DNA strands, and the incoming nucleotide) subjected to external forces. The results obtained with a force applied to the DNA template strand provide insights into the effect of the tension on the activity of the enzyme. At forces below 30 pN a local model based on the parameters determined from the simulations, including the restricted motion of the DNA bases at the active site, yields a replication rate dependence on force in agreement with experiment. Simulations above 40 pN reveal large conformational changes in the enzyme-bound DNA that may have a role in the force-induced exonucleolysis observed experimentally.