Overcoming microbial resuscitation using stable ultrafine gold nanosystems.

While ultrafine gold nanosystems (UGNs) are being extensively studied for their antimicrobial activities, hitherto, no report is available on their propensity towards mitigating bacterial resuscitation-a potential factor contributing to the antimicrobial resistance. The investigations herein with two categories of gold nanosystems-modulated for their stability and surface accessibility through glutathione capping-have provided insights into overcoming resuscitation. Additionally, the study cautions that even moderate resistance development in bacteria exposed to nanosystems can result in significant cross-resistance against conventional antibiotics.

Probing the Effects of Antimicrobial-Lysozyme Derivatization on Enzymatic Degradation of Poly(ε-caprolactone) Film and Fiber.

Surface derivatization is essential for incorporating unique functionalities into biodegradable polymers. Nonetheless, its precise effects on enzymatic biodegradation still lack comprehensive understanding. In this study, a facile solution-based method is employed to surface derivatize poly(ε-caprolactone) films and electrospun fibers with lysozyme, aiming to impart antimicrobial properties and examine the impact on enzymatic degradation. The derivatized films and fibers have shown high antibacterial efficacy against Escherichia coli and Staphylococcus aureus. Through gravimetric analysis, it is observed that the degradation rate experiences a slight decrease upon lysozyme derivatization. However, this reduction is effectively countered by the inclusion of Tween-20, as affirmed by isothermal titration calorimetry. Comparing films and fibers, the latter undergoes degradation at a more accelerated pace, coupled with a rapid decline in molecular weight. This study provides valuable insights into the factors influencing the degradation of surface-derivatized biopolymers through electrospinning, offering a simple strategy to mitigate biomaterial-associated infections.

Direct experimental evidence for the boronic acid-mediated bridging of DNA hybrids.

The use of terminal deoxynucleotidyl transferase for the first time in a mechanistic exploration-through colorimetric sensing and isothermal titration calorimetric studies-has provided direct experimental evidence of a boronic acid moiety bridging two DNA duplexes via the 3' hydroxyl groups, offering new opportunities and insights into the domain of DNA (nano)biotechnology.

Boronic acid chemistry for fluorescence-based quantitative DNA sensing.

Contrary to the long-standing opinion of boronic acids being typically reactive with 1,2- and 1,3-diols and hence not suitable for quantitative sensing of DNA containing only a mono-ol unit, this proof-of-concept study has successfully shown the feasibility to quantitatively detect DNA in the concentration range of 5 to 50 nM plausibly through boronic acid-mediated bridging of two DNA double helices via the 3' hydroxy groups, which opens up new avenues in the realm of oligonucleotide biochemistry.

Fluorescence-based simultaneous dual oligo sensing of HCV genotypes 1 and 3 using magnetite nanoparticles.

Nucleic acid tests (NATs) have gained an important position in biosensing in the context of the increasing need to meet the stringent requirements for accurate diagnosis of infectious diseases with high sensitivity and selectivity. Recently, the development of new strategies towards multiplex detection of analytes in a single assay is gaining impetus since such an approach would lead to high throughput analysis, leading to substantial benefits in terms of time, infrastructure, labor, and cost. In this work, we demonstrate a facile fluorescence-based simultaneous dual oligo sensing of genotypes 1 and 3 by employing two target sequences (36-mers each) derived from the NS4B and NS5A regions of HCV genome, respectively. A set of 18-mer amine-tagged probes and another set of 18-mer fluorescently-labeled probes that were complementary to each half of the 36-mer target sequences were designed. The amine-tagged probes were immobilized over aldehyde-derivatized magnetite nanoparticles (NPs) via imine bond formation, which was characterized using X-ray photoelectron spectroscopy (XPS) and energy dispersive spectroscopy (EDS) mapping techniques. The successful hybridization between the two probes with their target followed by magnetic removal of the NPs from the solution enabled quantitative analysis of the target by measuring the fluorescence intensity of the residual concentration of the fluorescently-tagged probe. In this manner, the targets corresponding to genotypes 1 and 3 were simultaneously detected with the detection limit in the range of 10-15 nM. The current strategy can potentially be amalgamated with existing nanotechnology-based techniques towards multiplex oligo sensing of several pathogens.

Water-soluble caffeic acid-dopamine acid-base complex exhibits enhanced bactericidal, antioxidant, and anticancer properties.

Despite the highly potent biological characteristics, the poor water-solubility of caffeic acid (CA) limits its applications in various domains. Here, we present a facile approach, wherein CA has been treated with dopamine hydrochloride (Dopa.HCl) to obtain a water-soluble acid-base complex, which does not possess any covalent bond between the individual components and thus retains their nativity. Simple mixing of CA and Dopa.HCl did not provide water solubility to CA, but the complex became readily soluble in water when the mineral acid was scavenged using sodium bicarbonate. The obtained CA-Dopa complex had been characterized using FT-IR, 1H NMR, 13C NMR, 2D 1H-1H NOESY NMR, XPS, and DSC techniques. The complex was found to exhibit excellent bactericidal, antibiofilm, antioxidant, and anticancer properties in the physiologically relevant pH range of 5.5 to 7.5. The results have revealed the high potential of the simple acid-base complex of CA in diverse domains.

Dysregulation of schizophrenia-associated genes and genome-wide hypomethylation in neurons overexpressing DNMT1.

Aim: To study the effects of DNMT1 overexpression on transcript levels of genes dysregulated in schizophrenia and on genome-wide methylation patterns. Materials & methods: Transcriptome and DNA methylome comparisons were made between R1 (wild-type) and Dnmt1tet/tet mouse embryonic stem cells and neurons overexpressing DNMT1. Genes dysregulated in both Dnmt1tet/tet cells and schizophrenia patients were studied further. Results & conclusions: About 50% of dysregulated genes in patients also showed altered transcript levels in Tet/Tet neurons in a DNA methylation-independent manner. These neurons unexpectedly showed genome-wide hypomethylation, increased transcript levels of Tet1 and Apobec 1-3 genes and increased activity and copy number of LINE-1 elements. The observed similarities between Tet/Tet neurons and schizophrenia brain samples reinforce DNMT1 overexpression as a risk factor.

Lay abstract DNMT1 controls cytosine methylation, which is often associated with reduced gene expression. Increased levels of DNMT1 is a risk factor for schizophrenia but information on the affected genes is limited. In this study, ∼50% of genes with altered levels of messenger RNAs in schizophrenia patients were also altered in neurons with increased DNMT1. Surprisingly, the neurons with higher DNMT1 levels showed genome-wide decrease in methylation. These findings uncover a new type of gene dysregulation that is independent of DNMT1's catalytic activity.

Lactobacillus amylovorus derived lipase-mediated silver derivatization over poly(ε-caprolactone) towards antimicrobial coatings.

Owing to the probiotic origin, lipases-derived from the Lactobacilli sp. are considered to be promising biomaterials for in vivo applications. On a different note, poly(ε-caprolactone) (PCL)-an FDA-approved polymer for implantable applications-lacks inherent antimicrobial property, because of which suitable modifications are required to render it with bactericidal activity. Here, we employ Lactobacillus amylovorous derived lipase to surface derivatize the PCL films with silver that is a highly efficient inorganic broad-spectrum antimicrobial substance. Two different surface functionalization strategies have been employed over the alkaline hydrolyzed PCL films towards this purpose: In the first strategy, lipase-capped silver nanoparticles (Ag NPs) have been synthesized in a first step, which have been covalently immobilized over the activated carboxylic groups on the PCL film surface in a subsequent step. In the second strategy, the lipase was covalently immobilized over the activated carboxylic groups of the PCL film surface in the first step, over which silver was deposited in the second step using the dip-coating method. While the characterization study using X-ray photoelectron spectroscopy (XPS) has revealed the successful derivatization of silver over the PCL film, the surface characterization using field-emission scanning electron microscopy (FE-SEM) study has shown a distinct morphological change with higher silver loading in both strategies. The antimicrobial studies employing E. coli have revealed 100 % inhibition in the bacterial growth in 4-6 h with the Ag NPs-immobilized PCL films as opposed to >8 h with those prepared through the dip-coating method. Additionally, the cytotoxicity assay using mouse fibroblast cells has shown that the PCL films immobilized with lipase-capped Ag NPs exhibit high cell compatibility, similar to that of pristine PCL film, and thereby making it suitable for in vivo applications.

Functional Analysis of DNMT1 SNPs (rs2228611 and rs2114724) Associated with Schizophrenia.

A recent study showed the association of minor alleles of rs2228611 (T allele) and rs2114724 (T allele) of DNMT1 with schizophrenia (SZ) and suggested their effects on splicing of the transcripts. We performed a replication study using 310 controls and 304 SZ patients and confirmed the association of the homozygous minor allele genotypes with SZ (P = 0.04 for rs2114724 and P = 0.007 for rs2228611). This significant association persisted after Bonferroni correction when the previously published data of 301 controls and 325 patients were also considered (P ≤ 0.0002). In addition, we found that the proportion of male patients with homozygous minor alleles at rs2114724 was significantly higher than that of females (P = 0.002). When haplotype analysis of both loci was performed, we observed a significant association of T/T-T/T and T/T-C/T (P = 0.04) haplotypes with SZ. To gain insights into the functional effects of the two SNPs on the levels of DNMT1 transcripts, quantitative real-time PCR experiments were performed using peripheral blood monocytes from 10 individuals each with T/T-T/T (homozygous minor allele), C/T-C/T (heterozygous), and C/C-C/C (homozygous major allele) haplotypes. Independently, the levels of DNMT1 protein were also compared in three individuals each by immunofluorescence. These results suggest that neither DNMT1 transcript nor the protein levels were significantly different in the peripheral blood monocytes among the individuals studied for the three groups. Taken together, our results confirm that the two minor alleles in homozygosity are associated with SZ but with no discernible effects on transcript or protein levels of DNMT1 in the peripheral blood monocytes of the small number of samples tested.

Quaternized Polydopamine Coatings for Anchoring Molecularly Dispersed Broad-Spectrum Antimicrobial Silver Salts.

Because of the broad-spectrum antimicrobial efficacy, silver-based coatings have emerged as the popular choice to apply over frequently touched surfaces for mitigating the spread of nosocomial infections. Despite the advancements through various coating strategies, clustering of the active component remains a bottleneck in achieving the molecular-scale dispersion of silver. To circumvent this, the current study takes advantage of the recent findings of quaternary ammonium moieties forming molecular complexes with silver salts that differ from the simple adduct between the individual components. Here we demonstrate the quaternization of oxidatively cross-linked polydopamine coatings over magnetite nanoparticles to anchor ionic silver at a molecular-scale dispersion. The silver-derivatized materials exhibit remarkable broad-spectrum antimicrobial properties against representative microbes like E. coli, S. aureus, and A. niger. Also, the study reveals the materials' antibiofilm efficacy (∼80-90%) against both bacteria. Further recyclability studies have proven the sustained bactericidal properties up to five cycles. The surface derivatization strategy has then been extended to cover glass slips that have also shown the retention of the bactericidal properties even after wiping 20 times with artificial sweat. The biocompatibility of the materials has been ascertained with treated water against the mouse fibroblast and human embryonic kidney cell lines. The current study offers insights in developing coatings with molecular-scale dispersion of ionic silver to achieve broad-spectrum antimicrobial properties in an atom-economical and sustainable manner.

Analysis of transcript levels of a few schizophrenia candidate genes in neurons from a transgenic mouse embryonic stem cell model overexpressing DNMT1.

Overexpression of DNA Methyltransferase I (DNMT1) is considered as one of the etiological factors for schizophrenia (SZ). However, information on genes subjected to dysregulation because of DNMT1 overexpression is limited. To test whether a larger group of SZ-associated genes are affected, we selected 15 genes reported to be dysregulated in patients (Gad1, Reln, Ank3, Cacna1c, Dkk3, As3mt, Ppp1r11, Smad5, Syn1, Wnt1, Pdgfra, Gsk3b, Cxcl12, Tcf4 and Fez1). Transcript levels of these genes were compared between neurons derived from Dnmt1tet/tet (Tet/Tet) mouse embryonic stem cells (ESCs) that overexpress DNMT1 with R1 (wild-type) neurons. Transcript levels of thirteen genes were significantly altered in Tet/Tet neurons of which, the dysregulation patterns of 11 were similar to patients. Transcript levels of eight out of these eleven were also significantly altered in Tet/Tet ESCs, but the dysregulation patterns of only five were similar to neurons. Comparative analyses among ESCs, embryoid bodies and neurons divided the 15 genes into four distinct groups with a majority showing developmental stage-specific patterns of dysregulation. Reduced Representational Bisulfite Sequencing data from neurons did not show any altered promoter DNA methylation for the dysregulated genes. Doxycycline treatment of Tet/Tet ESCs that eliminated DNMT1, reversed the direction of dysregulation of only four genes (Gad1, Dkk3, As3mt and Syn1). These results suggest that 1. Increased DNMT1 affected the levels of a majority of the transcripts studied, 2. Dysregulation appears to be independent of promoter methylation, 3. Effects of increased DNMT1 levels were reversible for only a subset of the genes studied, and 4. Increased DNMT1 levels may affect transcript levels of multiple schizophrenia-associated genes.

Schrodinger's scat: a critical review of the currently available tiger (Panthera Tigris) and leopard (Panthera pardus) specific primers in India, and a novel leopard specific primer.

BACKGROUND: Non-invasive sampling has opened avenues for the genetic study of elusive species, which has contributed significantly to their conservation. Where field based identity of non-invasive sample is ambiguous (e.g. carnivore scats), it is essential to establish identity of the species through molecular approaches. A cost effective procedure to ascertain species identity is to use species specific primers (SSP) for PCR amplification and subsequent resolution through agarose gel electrophoresis. However, SSPs if ill designed can often cross amplify non-target sympatric species. Herein we report the problem of cross amplification with currently published SSPs, which have been used in several recent scientific articles on tigers (Panthera tigris) and leopards (Panthera pardus) in India. Since these papers form pioneering research on which future work will be based, an early rectification is required so as to not propagate this error further. RESULTS: We conclusively show cross amplification of three of the four SSPs, in sympatric non-target species like tiger SSP amplifying leopard and striped hyena (Hyaena hyaena), and leopard SSP amplifying tiger, lion (Panthera leo persica) and clouded leopard (Neofelis nebulosa), with the same product size. We develop and test a non-cross-amplifying leopard specific primer pair within the mitochondrial cytochrome b region. We also standardize a duplex PCR method to screen tiger and leopard samples simultaneously in one PCR reaction to reduce cost and time. CONCLUSIONS: These findings suggest the importance of an often overlooked preliminary protocol of conclusive identification of species from non-invasive samples. The cross amplification of published primers in conspecifics suggests the need to revisit inferences drawn by earlier work.