An aptamer based competition assay for protein detection using CNT activated gold-interdigitated capacitor arrays

An aptamer can specifically bind to its target molecule, or hybridize with its complementary strand. A target bound aptamer complex has difficulty to hybridize with its complementary strand. It is possible to determine the concentration of target based on affinity separation system for the protein detection. Here, we exploited this property using C-reactive protein (CRP) specific RNA aptamers as probes that were immobilized by physical adsorption on carbon nanotubes (CNTs) activated gold interdigitated electrodes of capacitors. The selective binding ability of RNA aptamer with its target molecule was determined by change in capacitance after allowing competitive binding with CRP and complementary RNA (cRNA) strands in pure form and co-mixtures (CRP:cRNA=0:1, 1:0, 1:1, 1:2 and 2:1). The sensor showed significant capacitance change with pure forms of CRP/cRNA while responses reduced considerably in presence of CRP:cRNA in co-mixtures (1:1 and 1:2) because of the binding competition. At a critical CRP:cRNA ratio of 2:1, the capacitance response was dramatically lost because of the dissociation of adsorbed aptamers from the sensor surface to bind when excess CRP. Binding assays showed that the immobilized aptamers had strong affinity for cRNA (K(d)=1.98 μM) and CRP molecules (K(d)=2.4 μM) in pure forms, but low affinity for CRP:cRNA ratio of 2:1 (K(d)=8.58 μM). The dynamic detection range for CRP was determined to be 1-8 μM (0.58-4.6 μg/capacitor). The approach described in this study is a sensitive label-free method to detect proteins based on affinity separation of target molecules that can potentially be used for probing molecular interactions.     

Identification and functional expression of a family of nicotinic acetylcholine receptor subunits in the central nervous system of the mollusc Lymnaea stagnalis

We described a family of nicotinic acetylcholine receptor (nAChR) subunits underlying cholinergic transmission in the central nervous system (CNS) of the mollusc Lymnaea stagnalis. By using degenerate PCR cloning, we identified 12 subunits that display a high sequence similarity to nAChR subunits, of which 10 are of the alpha-type, 1 is of the beta-type, and 1 was not classified because of insufficient sequence information. Heterologous expression of identified subunits confirms their capacity to form functional receptors responding to acetylcholine. The alpha-type subunits can be divided into groups that appear to underlie cation-conducting (excitatory) and anion-conducting (inhibitory) channels involved in synaptic cholinergic transmission. The expression of the Lymnaea nAChR subunits, assessed by real time quantitative PCR and in situ hybridization, indicates that it is localized to neurons and widespread in the CNS, with the number and localization of expressing neurons differing considerably between subunit types. At least 10% of the CNS neurons showed detectable nAChR subunit expression. In addition, cholinergic neurons, as indicated by the expression of the vesicular ACh transporter, comprise approximately 10% of the neurons in all ganglia. Together, our data suggested a prominent role for fast cholinergic transmission in the Lymnaea CNS by using a number of neuronal nAChR subtypes comparable with vertebrate species but with a functional complexity that may be much higher.     

The role of prophage-like elements in the diversity of Salmonella enterica serovars

The Salmonella enterica serovar Typhi CT18 (S.Typhi) chromosome harbours seven distinct prophage-like elements, some of which may encode functional bacteriophages. In silico analyses were used to investigate these regions in S.Typhi CT18, and ultimately compare these integrated bacteriophages against 40 other Salmonella isolates using DNA microarray technology. S.Typhi CT18 contains prophages that show similarity to the lambda, Mu, P2 and P4 bacteriophage families. When compared to other S.Typhi isolates, these elements were generally conserved, supporting a clonal origin of this serovar. However, distinct variation was detected within a broad range of Salmonella serovars; many of the prophage regions are predicted to be specific to S.Typhi. Some of the P2 family prophage analysed have the potential to carry non-essential "cargo" genes within the hyper-variable tail region, an observation that suggests that these bacteriophage may confer a level of specialisation on their host. Lysogenic bacteriophages therefore play a crucial role in the generation of genetic diversity within S.enterica.     

Differential proteomics reveals multiple components in retrogradely transported axoplasm after nerve injury

Information on axonal damage is conveyed to neuronal cell bodies by a number of signaling modalities, including the post-translational modification of axoplasmic proteins. Retrograde transport of a subset of such proteins is thought to induce or enhance a regenerative response in the cell body. Here we report the use of a differential 2D-PAGE approach to identify injury-correlated retrogradely transported proteins in nerves of the mollusk Lymnaea. A comprehensive series of gels at different pI ranges allowed resolution of approximately 4000 spots by silver staining, and 172 of these were found to differ between lesioned versus control nerves. Mass spectrometric sequencing of 134 differential spots allowed their assignment to over 40 different proteins, some belonging to a vesicular ensemble blocked by the lesion and others comprising an up-regulated ensemble highly enriched in calpain cleavage products of an intermediate filament termed RGP51 (retrograde protein of 51 kDa). Inhibition of RGP51 expression by RNA interference inhibits regenerative outgrowth of adult Lymnaea neurons in culture. These results implicate regulated proteolysis in the formation of retrograde injury signaling complexes after nerve lesion and suggest that this signaling modality utilizes a wide range of protein components.     

In vitro propagation and the acclimatization effect on the synthesis of 2-hydroxy-4-methoxy benzaldehyde in Decalepis hamiltonii Wight and Arn.

The present study reports a high frequency in vitro propagation protocol through apical bud sprouting and basal organogenic nodule formation in shoot tip explants of Decalepis hamiltonii, an endemic and endangered medicinal liana. Among different combinations of plant growth regulators (PGRs) and growth additives, maximum of 8.20 shoots per explant with mean shoot length of 6.54 cm were induced on Murashige and Skoog’s medium (MS) supplemented with 5.0 µM 6-benzyladenine (BA) + 0.5 µM indole-3-acetic acid (IAA) + 30.0 µM adenine sulphate (ADS) through apical bud sprouting. On single cytokinin treatment explants did not exhibit good multiplication but showed nodulation (N₁) from the basal cut end similar to cytokinin–auxin combination (N₂). Between two types of nodular tissues, N₂ was proved to be better for maximum shoot regeneration (15.40 shoots per explant) and shoot length (4.56 cm) when cultured on MS medium supplemented with 5.0 µM BA, 0.5 µM IAA, 30.0 µM ADS and 1.0 µM gibberellic acid (GA₃). Microshoots were efficiently rooted on half-strength MS medium supplemented with 2.5 μM α-naphthalene acetic acid (NAA). After successful acclimatization in Soilrite, 95.10 % plantlets were survived in field conditions. Histological investigation proved useful in ascertaining the callogenic nature of the regenerating nodular tissue formed at the basal cut end of shoot tip explant. Acclimatized plantlets were studied for the estimation of chlorophyll and carotenoid content as well as the net photosynthetic rate (PN) during subsequent days of transfer to ex vitro condition. Moreover, acclimatization had a significant effect on biomass production and the synthesis of 2-hydroxy-4-methoxy benzaldehyde (2HMB). Maximum fresh weight (3.78 gm/plant), dry weight (0.39 gm/plant) of roots and 2HMB content (15.94 µg/ml of extract) were noticed after 8 weeks of acclimatization.     

Impaired Autophagy Flux is Associated with Proinflammatory Microglia Activation Following Japanese Encephalitis Virus Infection

Neurochemical Research - Role of autophagy in Japanese encephalitis viral (JEV) infection is not well known. In the present study, we reported the role of autophagy flux in microglia activation,...     

Palynomorph distribution in a mangrove ecosystem along environmental and salinity gradient: a tool for palaeoecological reconstruction

Pollen assemblages closely reflect the local vegetation that characterizes the salinity status, providing useful analogs for paleoecologxical reconstruction in regional deposits. Palynological evidences of surface sediments from the Coringa mangrove wetland were correlated with the physicochemical and sediment salinity records to observe the relationship between them. The statistical analysis of the data obtained here revealed a marked horizontal salinity gradient from north to south.. In this study, ordination (detrended correspondence analysis) of palynomorph groups has identified a salinity gradient of 1.1 to 3.0 PSU from the north to the south in the wetland. High palynomorph deposition and lower salinity are observed along channel margins due to the mixing of estuarine water during the rainy season. The mudflats along the transects show a lower diversity of plants in the pollen record and high total dissolved solids (TDS) than commonly found in the coastal wetlands. The presence of scrubby halophytic vegetation in the upper saltmarsh and oligohaline-freshwater vegetation in the low tidal saline marshes is a zonation pattern related to the localized influence of freshwater conditions. Palynomarine Index (PMI) reveals the highest freshwater/tidal inundation along Ramannapalem due to the presence of numerous riverine channels while the remaining part of the wetland has restricted tidal inundation leading to the conversion of mangrove forests to salt pans and paddy fields. Thus, the health and productivity of the mangrove ecosystem are also governed by hydrodynamics, catchment land use, water discharge in the channels, and tidal flushing.

     

Observations on the floristic,life-form,leaf-size spectra and habitat diversity of vegetation in the Bhimber hills of Kashmir Himalayas

Vegetation analysis provides the prerequisites to understand the overall community structure and function of any ecosystem and is a fundamental requirement for the precise evaluation of biodiversity. Although many studies have assessed floristic attributes of specific areas, there are still unexplored regions, as is the case of the mountain region in the Kashmir Himalayas. Current research highlighted the recent findings of the scientific characterization of floristic and ecological aspects on the forest flora found in the Bhimber hills, Pakistan. Floristically, a total of 93 species belonging to 80 genera in 41 families were recorded. The species distribution patterns across the families were disproportionate with half of the species contributed by 8 families and 25 families were monotypic. Based on the floristic analysis, Asteraceae was the largest family with 12% of species followed by Poaceae with (11%) species. PAST software, a multivariate ecological community analysis was used to classify the species similarities and differences among the different habitat types. According to the habitat wise distribution, 21% of species were growing in the natural forest habitat, while 15% of species were dispersed in highly distributed habitats along roadsides and 8% on pedestrians. In terms of functional diversity, the herbaceous growth form was dominant (58%). The biological spectrum revealed therophytes as the dominant life form as it indicates the disturbed habitat vegetation. The phytogeographical analysis revealed that the maximum (69%) species were native, while the minimum (31%) species were exotic. Thus, the study of these functional and habitat diversity patterns can significantly improve our understanding of the ecological aspects of the flora in the geographical location. This information may additionally be useful in devising management plans to ensure sustainable utilization and better management of forest landscapes in this Himalayan region.     

Thermophilic (55°C) and moderately hyperthermophilic (65°C) fermentation of poultry manure triggers release of high heavy metal concentrations leading to enhanced genotoxicity

Proper disposal of waste from poultry industries may be done by thermophilic anaerobic digestion. Particularly, thermophilic fermentation exhibits pivotal advantages. However, not only the accrued waste material (digestate) serves as organic fertilizer but also poses environmental problems when the pollutant release is poorly synchronized with the environmental demand. To minimize environmental risks, additional hyperthermophilic (moderately hyperthermophilic) treatment of animal by‐products is necessary, but concurrent release of pollutants is notably increased. To estimate the quantity and quality of pollutants released, we analyzed various organic compounds as well as 21 metals and tested their genotoxic/mutagenic potential by using the Allium cepa assay and the Ames test. As, Cd, Ni, and W were not detected or had low concentration (0.01–0.47 mg/kg) and were considered of little relevance. Co, Cr, Ni, Pb, and Mo were detected at concentrations up to 5.76 mg/kg. Fe, Mn, Zn, and Cu had the highest concentrations and were accumulated to inhibitory levels. Phenolic compounds were detected in negligible concentrations. We found that approximately 30% of onion root chromosomes were permanently damaged by high heavy metal concentrations released from the digestates. Higher temperatures and longer fermentation times fostered an increased release of pollutants above permissible limits.