Ultrasensitive Immunosensor Based on Langmuir-Blodgett Deposited Ordered Graphene Assemblies for Dengue Detection.

In this manuscript partially reduced graphene oxide (RGO) nanosheet-based electrodes have been utilized for quantification of the NS1 protein and subsequently for dengue detection. NS1 is the biomarker found circulating in the body of dengue-infected persons on or after first day of the appearance of disease symptoms. Graphene oxide (GO) has been synthesized using a modified Hummer's method, and its ordered nanostructured films have been electrophoretically deposited on indium tin oxide (ITO)-coated glass substrates using Langmuir-Blodgett (LB) deposition. Deposited LB films of GO have been reduced with hydrazine vapors to obtain RGO-coated ITO electrodes. NS1 antibodies have been grafted onto the ordered thin films using covalent linking, and the bioelectrodes have been utilized for the specific detection of NS1 antigen. The electrochemical performance of the fabricated bioelectrodes for NS1 antigen detection has been explored in standard and spiked sera samples. The limit of detection for the standard samples and spiked serum samples is found to be 0.069 ng mL-1 and 0.081 ng mL-1, respectively, with a sensitivity of 8.41 and 36.75 Ω per ng mL, respectively, in the detection range of 101 to 107 ng mL-1.

Protein-conjugated quantum dots interface: binding kinetics and label-free lipid detection.

We propose a label-free biosensor platform to investigate the binding kinetics using antigen-antibody interaction via electrochemical and surface plasmon resonance (SPR) techniques. The L-cysteine in situ capped cadmium sulfide (CdS; size < 7 nm) quantum dots (QDs) self-assembled on gold (Au) coated glass electrode have been covalently functionalized with apolipoprotein B-100 antibodies (AAB). This protein conjugated QDs-based electrode (AAB/CysCdS/Au) has been used to detect lipid (low density lipoprotein, LDL) biomolecules. The electrochemical impedimetric response of the AAB/CysCdS/Au biosensor shows higher sensitivity (32.8 kΩ µM(-1)/cm(2)) in the detection range, 5-120 mg/dL. Besides this, efforts have been made to investigate the kinetics of antigen-antibody interactions at the CysCdS surface. The label-free SPR response of AAB/CysCdS/Au biosensor exhibits highly specific interaction to protein (LDL) with association constant of 33.4 kM(-1) s(-1) indicating higher affinity toward LDL biomolecules and a dissociation constant of 0.896 ms(-1). The results of these studies prove the efficacy of the CysCdS-Au platform as a high throughput compact biosensing device for investigating biomolecular interactions.

Improved estimation of population variance in stratified successive sampling using calibrated weights under non-response.

This paper introduces a new method to estimate the population variance of a study variable in stratified successive sampling over two occasions, while accounting for random non-response. The method uses a logarithmic type estimator and leverages information from a highly positively correlated auxiliary variable. The paper also presents calibrated weights for the new estimator and examines its properties through numerical and simulation studies. The results indicate that the suggested estimator is more effective than the standard estimator for estimating the population variance.

A general class of improved population variance estimators under non-sampling errors using calibrated weights in stratified sampling.

This paper proposes a new calibration estimator for population variance within a stratified two-phase sampling design. It takes into account random non-response and measurement errors, specifically applying this method to estimate the variance in Gas turbine exhaust pressure data. The study integrates additional information from two highly positively correlated auxiliary variables to develop a general class of estimators tailored for the stratified two-phase sampling scheme. The properties of these estimators, in terms of their biases and mean square errors, have been thoroughly examined and extensively analyzed through numerical and simulation studies. Furthermore, the calibrated weights of the strata are derived. The proposed estimators outperform the natural estimator of population variance. Finally, suitable recommendations have been made for survey statisticians intending to apply these findings to real-life problems.

Comparison of vertical and horizontal flow planted and unplanted subsurface flow wetlands treating municipal wastewater.

In the search for design criteria for constructed wetlands (CWs) in Nepal a semi-scale experimental setup including horizontal flow (HF) and vertical flow (VF) CWs was developed. This paper compares the performance of HF and VF wetlands, and planted with unplanted beds. The experimental setup consists of two units of HF and VF beds of size 6 m × 2 m × 0.6 m and 6 m × 2 m × 0.8 m (length × width × depth) respectively. For both HF and VF systems, one unit was planted with Phragmites karka (local reed) and one was not planted. The systems were fed with wastewater drawn from the grit chamber of a municipal wastewater treatment plant. The media consisted of river gravel. In the first phase of the experiment the hydraulic loading rate (HLR) was varied in steps; 0.2, 0.08, 0.04 m(3)/m(2)/d and the percent removal increase with decrease in HLR for all beds and parameters except for total phosphorus. In the second phase the loading rate of 0.04 m(3)/m(2)/d was run for 7 months. In both parts of the experiment the planted beds performed better than the unplanted beds and the VF better than the HF beds. To meet Nepalese discharge standards HF beds are sufficient, but to meet stricter requirements a combination of HF and VF beds are recommended.

Comparative pathology, molecular pathogenicity, immunological features, and genetic characterization of three highly pathogenic human coronaviruses (MERS-CoV, SARS-CoV, and SARS-CoV-2).

The last two decades have witnessed the emergence of three deadly coronaviruses (CoVs) in humans: severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory syndrome coronavirus (MERS-CoV), and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). There are still no reliable and efficient therapeutics to manage the devastating consequences of these CoVs. Of these, SARS-CoV-2, the cause of the currently ongoing coronavirus disease 2019 (COVID-19) pandemic, has posed great global health concerns. The COVID-19 pandemic has resulted in an unprecedented crisis with devastating socio-economic and health impacts worldwide. This highlights the fact that CoVs continue to evolve and have the genetic flexibility to become highly pathogenic in humans and other mammals. SARS-CoV-2 carries a high genetic homology to the previously identified CoV (SARS-CoV), and the immunological and pathogenic characteristics of SARS-CoV-2, SARS-CoV, and MERS contain key similarities and differences that can guide therapy and management. This review presents salient and updated information on comparative pathology, molecular pathogenicity, immunological features, and genetic characterization of SARS-CoV, MERS-CoV, and SARS-CoV-2; this can help in the design of more effective vaccines and therapeutics for countering these pathogenic CoVs.

Nanostructured conducting polymer based reagentless capacitive immunosensor.

Nanostructured polyaniline (PANI) film electrophoretically fabricated onto indium-tin-oxide (ITO) coated glass plate has been utilized for development of an immunosensor based on capacitance change of a parallel plate capacitor (PPC) by covalently immobilizing anti-human IgG (Anti-HIgG) using N-ethyl-N'-(3-dimethylaminopropyl) carbodiimide and N-hydroxysuccinimide chemistry. These fabricated PANI/ITO and Anti-HIgG/PANI/ITO plates have been characterized using scanning electron microscopy, cyclic voltammetry, differential pulse voltammetry and Fourier transform infra-red studies. The capacitance measurements indicate that dielectric medium of this biologically modified PPC (Anti-HIgG/PANI/ITO) is sensitive to HIgG in 5 - 5 x 10(5) ng mL(-1) range and has lower detection limit of 1.87 ng mL(-1). The observed results reveal that this Anti-HIgG modified PPC can be used as a robust, easy-to-use, reagentless, sensitive and selective immunosensor for estimation of human IgG.

Polyaniline-carboxymethyl cellulose nanocomposite for cholesterol detection.

Cholesterol oxidase (ChOx) has been covalently immobilized onto polyaniline-carboxymethyl cellulose (PANI-CMC) nanocomposite film deposited onto indium-tin-oxide (ITO) coated glass plate using glutaraldehyde as a cross-linker. Fourier transform infrared (FTIR) spectroscopic and electrochemical studies have been used to characterize the PANI-CMC/ITO nanocomposite electrode and ChOx/PANI-CMC/ITO bioelectrode. Scanning electron microscopy (SEM) studies reveal the formation of PANI-CMC nanocomposite fibers of size approximately 150 nm in diameter. The ChOx/PANI-CMC/ITO bioelectrode exhibits linearity as 0.5-22 mM, detection limit as 1.31 mM, sensitivity as 0.14 mA/mM cm2, response time as 10 s and shelf-life of about 10 weeks when bioelectrode is stored at 4 degrees C. The low value of Michaelis-Menten constant (K(m)) obtained as 2.71 mM reveals high affinity of immobilized ChOx for PANI-CMC/ITO nanocomposite electrode.

Development of AhMITE1 markers through genome-wide analysis in peanut (Arachis hypogaea L.).

OBJECTIVE: In peanut, the DNA polymorphism is very low despite enormous phenotypic variations. This limits the use of genomics-assisted breeding to enhance peanut productivity. This study aimed to develop and validate new AhMITE1 and cleaved amplified polymorphic sequences (CAPS) markers. RESULTS: In total, 2957 new AhMITE1 markers were developed in addition to identifying 465 already reported markers from the whole genome re-sequencing data (WGRS) of 33 diverse genotypes of peanut. The B sub-genome (1620) showed more number of markers than the A sub-genome (1337). Distribution also varied among the chromosomes of both the sub-genomes. Further, 52.6% of the markers were from genic regions; where 31.0% were from intronic regions and 5.2% were from exonic regions. Of the 343 randomly selected markers, 82.2% showed amplification validation, with up to 35.5% polymorphism. From the SNPs on the A03, B01, B02 and B03 chromosomes, 11,730 snip-SNPs (potential CAPS sites) were identified, and 500 CAPS markers were developed from chromosome A03. Of these markers, 30.0% showed validation and high polymorphism. This study demonstrated the potential of the WGRS data to develop AhMITE1 and CAPS markers, which showed high level of validation and polymorphism. These marker resources will be useful for various genetic studies and mapping in peanut.

Langmuir-Blodgett Nanoassemblies of the MoS2-Au Composite at the Air-Water Interface for Dengue Detection.

We report Langmuir-Blodgett (LB) films of molybdenum disulphide (MoS2) and gold nanoparticles (AuNP) composite being utilized as a biosensing platform for dengue detection. The LB films of the MoS2-AuNP composite have been transferred from the air-water interface to the indium tin oxide-coated glass substrate under optimized conditions. Further, antibodies specific to dengue NS1 antigen were immobilized onto these LB films. The fabricated immunosensor has been explored for NS1 antigen detection in standard samples as well as in spiked sera samples using electrochemical impedance spectroscopy. The NS1 antigen is present in the blood of infected persons from day one of the onset of clinical symptoms in primary dengue infection. The limit of detection for the standard and the spiked samples is found to be 1.67 and 1.19 ng mL-1, respectively, which is suitable for clinical applications, as NS1 antigen levels in patient's sera range from 0.04 to 2 µg mL-1 in primary infection and from 0.01 to 2 µg mL-1 in secondary infection.

Two cases of eyelid tuberculosis - An uncommon presentation of ocular tuberculosis.

Mycobacterium tuberculosis apart from being the causative agent of pulmonary tuberculosis is also notorious to cause tuberculosis at various sites in the human body and ocular tuberculosis is one of the extra pulmonary manifestations of this organism. The most common presentation of ocular tuberculosis is anterior uveitis or choroiditis caused by hematogenous infection or hypersensitivity after another organ infection. Eyelid involvement by tuberculosis is most of the times secondary to orbital involvement and often seen in the form of drainage sinus. Isolated eyelid tuberculosis is however uncommon. Here we report two such cases of eyelid tuberculosis in different age groups; first case in a young female and second case of an old aged female with different presentation. Fortunately both of them responded well to the antitubercular treatment.

Application of electrochemically prepared polypyrrole-polyvinyl sulphonate films to DNA biosensor.

Double stranded calf thymus deoxyribonucleic acid (DNA) was physisorbed onto polypyrrole-polyvinyl sulphonate (PPY-PVS) films electrochemically deposited onto indium-tin-oxide (ITO) coated glass plates. These DNA immobilized PPY-PVS films optimized for various conditions, such as polymerization potential, pH of buffer, DNA concentration and scan rate were characterized using Fourier-transform infrared (FT-IR) spectroscopy, atomic force microscopy (AFM) and cyclic voltammetry (CV) techniques, respectively. The amperometric response studies of these DNA/PPY-PVS electrodes were carried out as a function of 2-aminoantharcene (2-AA, 0.01-20 ppm) and o-chlorophenol (OCP, 0.1-30 ppm) concentration, respectively at 25 degrees C. The observed amperometric current arising due to oxidation of guanine in the DNA/PPY-PVS films decreased linearly with the increase in the concentration of 2-AA and OCP. It has been revealed that 10 ppm of 2-AA is sufficient to reduce the observed guanine oxidation peak current by approximately -95+/-10% as compared to the reported values. A 25 ppm of OCP was capable enough to reduce the guanine oxidation current to zero. These DNA/PPY-PVS electrodes were found to have a shelf life of about 4 months when stored at 25 degrees C.

Immobilization of urease on poly(N-vinyl carbazole)/stearic acid Langmuir-Blodgett films for application to urea biosensor.

Urease was immobilized in mixed monolayers of poly(N-vinyl carbazole) (PNVK) and stearic acid (SA) formed at an air-water interface. The monolayers were transferred onto indium-tin-oxide (ITO) coated glass plates using Langmuir-Blodgett (LB) film deposition technique. Urease immobilized on PNVK/SA LB films, characterized using FTIR and UV-visible spectroscopy, was found to exhibit increased stability over a wide pH (6.5-8.5) and temperature (25-50 degrees C) range. Potentiometric measurements on these urease electrodes were carried out using an ammonium ion analyzer. Two values for K(m)(app) were obtained at lower and higher concentrations of substrate urea.

Signal amplification by substrate recycling on polyaniline/lactate oxidase/lactate dehydrogenase bienzyme electrodes.

The bienzyme electrodes were fabricated by coimmobilization of lactate oxidase (LOD) and lactate dehydrogenase (LDH) onto electrochemically prepared polyaniline (PANI) films. These PANI/LOD/LDH bienzyme electrodes were shown to provide signal amplification by substrate recycling, making it possible to detect L-lactate at lower concentrations (0.1-1 mM). The PANI/LOD/LDH bienzyme electrodes were found to be stable for about 21 d at 4-10 degrees C.

Phonon dispersion in polyinosinic acid.

A study of the normal modes of vibration and their dispersion in polyinosinic acid [poly (I)] along the helix axis based on Urey-Bradley force field is reported. It leads to a better interpretation of Raman and FTIR spectra. A comparison of dispersion curves of poly (I) with poly (G) has been made. Characteristic features of dispersion curves, such as regions of high density-of-states, repulsion and character mixing are discussed. Predictive value of heat capacity as a function of temperature is reported.

Quantitative trait locus analysis and construction of consensus genetic map for foliar disease resistance based on two recombinant inbred line populations in cultivated groundnut (Arachis hypogaea L.).

Late leaf spot (LLS) and rust have the greatest impact on yield losses worldwide in groundnut (Arachis hypogaea L.). With the objective of identifying tightly linked markers to these diseases, a total of 3,097 simple sequence repeats (SSRs) were screened on the parents of two recombinant inbred line (RIL) populations, namely TAG 24 × GPBD 4 (RIL-4) and TG 26 × GPBD 4 (RIL-5), and segregation data were obtained for 209 marker loci for each of the mapping populations. Linkage map analysis of the 209 loci resulted in the mapping of 188 and 181 loci in RIL-4 and RIL-5 respectively. Using 143 markers common to the two maps, a consensus map with 225 SSR loci and total map distance of 1,152.9 cM was developed. Comprehensive quantitative trait locus (QTL) analysis detected a total of 28 QTL for LLS and 15 QTL for rust. A major QTL for LLS, namely QTL(LLS)01 (GM1573/GM1009-pPGPseq8D09), with 10.27-62.34% phenotypic variance explained (PVE) was detected in all the six environments in the RIL-4 population. In the case of rust resistance, in addition to marker IPAHM103 identified earlier, four new markers (GM2009, GM1536, GM2301 and GM2079) showed significant association with the major QTL (82.96% PVE). Localization of 42 QTL for LLS and rust on the consensus map identified two candidate genomic regions conferring resistance to LLS and rust. One region present on linkage group AhXV contained three QTL each for LLS (up to 67.98% PVE) and rust (up to 82.96% PVE). The second candidate genomic region contained the major QTL with up to 62.34% PVE for LLS. Molecular markers associated with the major QTL for resistance to LLS and rust can be deployed in molecular breeding for developing groundnut varieties with enhanced resistance to foliar diseases. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s11032-011-9661-z) contains supplementary material, which is available to authorized users.

Quantitative trait locus analysis and construction of consensus genetic map for drought tolerance traits based on three recombinant inbred line populations in cultivated groundnut (Arachis hypogaea L.).

Groundnut (Arachis hypogaea L.) is an important food and cash crop grown mainly in semi-arid tropics (SAT) regions of the world where drought is the major constraint on productivity. With the aim of understanding the genetic basis and identification of quantitative trait loci (QTL) for drought tolerance, two new recombinant inbred line (RIL) mapping populations, namely ICGS 76 × CSMG 84-1 (RIL-2) and ICGS 44 × ICGS 76 (RIL-3), were used. After screening of 3,215 simple sequence repeat (SSR) markers on the parental genotypes of these populations, two new genetic maps were developed with 119 (RIL-2) and 82 (RIL-3) SSR loci. Together with these maps and the reference map with 191 SSR loci based on TAG 24 × ICGV 86031 (RIL-1), a consensus map was constructed with 293 SSR loci distributed over 20 linkage groups, spanning 2,840.8 cM. As all these three populations segregate for drought-tolerance-related traits, a comprehensive QTL analysis identified 153 main effect QTL (M-QTL) and 25 epistatic QTL (E-QTL) for drought-tolerance-related traits. Localization of these QTL on the consensus map provided 16 genomic regions that contained 125 QTL. A few key genomic regions were selected on the basis of the QTL identified in each region, and their expected role in drought adaptation is also discussed. Given that no major QTL for drought adaptation were identified, novel breeding approaches such as marker-assisted recurrent selection (MARS) and genomic selection (GS) approaches are likely to be the preferred approaches for introgression of a larger number of QTL in order to breed drought-tolerant groundnut genotypes. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s11032-011-9660-0) contains supplementary material, which is available to authorized users.

DNA biosensor for detection of Neisseria gonorrhoeae causing sexually transmitted disease.

A sequence-specific electrochemical sexually transmitted disease (STD) sensor based on self-assembled monolayer of thiolated DNA probe specific to target opa gene for detection of Gonorrhoea, a sexually transmitted disease has been fabricated. 6-Mercapto-1-hexanol (MCH) has been used as a blocking agent to facilitate oligos "stand" up at the surface, a configuration favoring subsequent DNA hybridization and to repel non-specific adsorption of undesired DNA. The results of differential pulse voltammetric studies of this STD sensor reveal low detection limit (1.0 × 10(-18)M) and a wide dynamic range (from 1.0 × 10(-6)M to 0.5 × 10(-18)M) arising due to the stable hybridization using methylene blue as an electro-active DNA hybridization indicator. The experimental results with genomic DNA, clinical patient sample of Neisseria gonorrhoeae, culture of non-N. gonorrhoeae Neisseria species (NgNS) and gram negative bacteria indicate that the fabricated sensor is specific to this STD.

Nucleic acid sensor for insecticide detection.

Nucleic acid sensor based on polyaniline (PANI) has been fabricated by covalently immobilizing double stranded calf thymus (dsCT) DNA onto perchlorate (ClO(-) (4))-doped PANI film deposited onto indium-tin-oxide (ITO) glass plate using 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC)/N-hydroxysuccinimide (NHS) chemistry. These dsCT-DNA-PANI-ClO(4)/ITO and PANI-ClO(4)/ITO electrodes have been characterized using square wave voltammetry, electrochemical impedance, scanning electron microscopy (SEM) and Fourier-transform-infrared (FTIR) measurements. This disposable dsCT-DNA-PANI-ClO(4)/ITO bioelectrode, stable for about 4 months, can be used to detect cypermethrin (0.005 ppm) and trichlorfon (0.01 ppm) in 30 and 60 s, respectively.