Improving synthesis and binding affinities of nucleic acid aptamers and their therapeutics and diagnostic applications.

Nucleic acid aptamers have captivated the attention of analytical and medicinal scientists globally due to their several advantages as recognition molecules over conventional antibodies because of their small size, simple and inexpensive synthesis, broad target range, and high stability in varied environmental conditions. These recognition molecules can be chemically modified to make them resistant to nuclease action in blood serum, reduce rapid renel clearance, improve the target affinity and selectivity, and make them amenable to chemically conjugate with a support system that facilitates their selective applications. This review focuses on the development of efficient aptamer candidates and their application in clinical diagnosis and therapeutic applications. Significant advances have been made in aptamer-based diagnosis of infectious and non-infectious diseases. Collaterally, the progress made in therapeutic applications of aptamers is encouraging, as evident from their use in diagnosing cancer, neurodegenerative diseases, microbial infection, and in imaging. This review also updates the progress on clinical trials of many aptamer-based products of commercial interests. The key development and critical issues on the subject have been summarized in the concluding remarks.

Siderophore of plant growth promoting rhizobacterium origin reduces reactive oxygen species mediated injury in Solanum spp. caused by fungal pathogens.

AIMS: The study aims to explore antifungal properties of bacillibactin siderophore produced by the plant growth-promoting rhizobacterium (PGPR) Bacillus subtilis against fungal phytopathogens Alternaria porri and Fusarium equiseti isolated from Solanum lycopersicum and Solanum melongena plants. METHODS AND RESULTS: Alternaria porri and F. equiseti were isolated from infected plants of eggplant and tomato, respectively. A plate assay was employed to assess the effect of bacillibactin against the phytopathogens. The antifungal potential of the PGPR was evaluated by estimation of dry fungal biomass, visualization of cellular deformity using compound and scanning electron microscopy, antioxidative enzyme assay and analysis of membrane damage via using lipid peroxidation. Inductively coupled plasma atomic emission spectroscopy (ICP-AES) analysis was employed to investigate changes in intracellular iron content. The impact of bacillibactin on pathogenesis was evaluated by infecting detached leaves of S. lycopersicum and S. melongena plants with both the pathogens and treating the infected leaves with bacillibactin. Leaves were further investigated for ROS accumulation, extent of necrosis and cell death. Our findings revealed significant damage to the hyphal structure of A. porri and F. equiseti following treatment with bacillibactin. Biomass reduction, elevated antioxidative enzyme levels, and membrane damage further substantiated the inhibitory effects of the siderophore on fungal growth. ICP-AES analysis indicates an increase in intracellular iron content suggesting enhanced iron uptake facilitated by bacillibactin. Moreover, application of 1500 µg ml-1 bacillibactin on infected leaves demonstrated a substantial inhibition of ROS accumulation, necrosis, and cell death upon bacillibactin treatment. CONCLUSIONS: This study confirms the potent antagonistic activity of bacillibactin against both the phytopathogens A. porri and F. equiseti growth, supporting its potential as a promising biological control agent for fungal plant diseases. Bacillibactin-induced morphological, physiological, and biochemical alterations in the isolated fungi and pathogen-infected leaves highlight the prospects of bacillibactin as an effective and sustainable solution to mitigate economic losses associated with fungal infections in vegetable crops.

Geophysical and geostatistical assessment of groundwater and soil quality using GIS, VES, and PCA techniques in the Jaipur region of Western India.

In present study, geophysical and geostatistical variability of ground water and agricultural soil investigated in the Jaipur region of Rajasthan (Western India) by applying the geographic information system (GIS), vertical electrical sounding (VES) ,and statistical analysis. Ground water and soil samples collected from different sites from the selected study area and variation pattern of quality parameters were assessed. A contour map analysis of distribution of metals and other contaminants in the samples was conducted using GIS. Maximum concentration of metals recorded in the soil samples in order of Fe, 11.25 mg kg-1 > Mn, 8.6 mg kg-1 > Zn, 7.2 mg kg-1 > Cu, 0.455 mg kg-1; however, maximum concentration of metals in the ground water samples was found as Zn, 2.64 mg L-1 > Cu, 0.86 mg L-1 > Fe, 0.39 mg L-1 > Mn, 0.18 mg L-1 > Pb, 0.065 mg L-1 > Ni, 0.016 mg L-1. Observed data emphasis variability in groundwater and soil quality parameter by PCA technique indicated 84.60% and 66.98% of variance, respectively. Soil quality index (SQI) value was observed as 0.482 indicating that 46% of soil sampling sites deteriorated and shown poor quality. Similarly, water quality index (WQI) value indicates good water quality at the sampling sites TW1, TW8, TW10, and TW12; however, TW3, TW4, TW6, TW19, TW20, and TW22 sites showed very poor water quality. The present study concludes that overexploitation of groundwater and unregulated discharge of wastewater leads to depletion of water and soil quality. Further, applying geographical and geostatistical techniques in assessing water and soil quality could be more effective tools in environmental monitoring and management for environmental and health safety.

Single-stage chromatographic clarification of Chinese Hamster Ovary cell harvest reduces cost of protein production.

A single-stage clarification was developed using a single-use chromatographic clarification device (CCD) to recover a recombinant protein from Chinese Hamster Ovary (CHO) harvest cell culture fluid (HCCF). Clarification of a CHO HCCF is a complex and costly process, involving multiple stages of centrifugation and/or depth filtration to remove cells and debris and to reduce process-related impurities such as host cell protein (HCP), nucleic acids, and lipids. When using depth filtration, the filter train consists of multiple filters of varying ratios, layers, pore sizes, and adsorptive properties. The depth filters, in combination with a 0.2-micron membrane filter, clarify the HCCF based on size-exclusion, adsorptive, and charge-based mechanisms, and provide robust bioburden control. Each stage of the clarification process requires time, labor, and utilities, with product loss at each step. Here, use of the 3M™ Harvest RC Chromatographic Clarifier, a single-stage CCD, is identified as an alternative strategy to a three-stage filtration train. The CCD results in less overall filter area, less volume for flushing, and higher yield. Using bioprocess cost modeling, the single-stage clarification process was compared to a three-stage filtration process. By compressing the CHO HCCF clarification to a single chromatographic stage, the overall cost of the clarification process was reduced by 17%-30%, depending on bioreactor scale. The main drivers for the cost reduction were reduced total filtration area, labor, time, and utilities. The benefits of the single-stage harvest process extended throughout the downstream process, resulting in a 25% relative increase in cumulative yield with comparable impurity clearance.

Association of neutrophil gelatinase associated lipocalin, ischemia modified albumin with uric acid in the etiopathogenesis of preeclampsia.

OBJECTIVES: Preeclampsia is a multisystem illness that manifests in the third trimester of pregnancy after 20 weeks of gestation and is marked by proteinuria and hypertension (PE). Changes in lifestyle, such as eating a high-calorie diet and delaying delivery, have raised the likelihood of developing PE. Eclampsia, abrupt renal failure, thromboembolic episodes leading to cardiac and brain problems, pulmonary embolism, and coagulopathy associated with HELLP syndrome are a few of the complications that might follow preeclampsia in pregnant moms. The objects of this study is to estimate and correlate the levels of NGAL (neutrophil gelatinase associated lipocalin), IMA (ischemia modified albumin) and Uric acid in prreclampsia. METHODS: 40 diagnosed cases of preeclampsia and 40 healthy age and gestational age matched healthy controls were included in the study. Blood samples were collected from them and serum NGAL, IMA and Uric acid levels were estimated. Estimation of NGAL (neutrophil gelatinase associated lipocalin), IMA (ischemia modified albumin) was done by commercially available ELISA kits standard spectrophotometry methods in autoanalyzer Mind ray BS300 using commercially available kits. RESULTS: The parameters of NGAL and IMA were significantly increased in patients with PE (p<0.001) when compared with the healthy control subjects. γ-glutamyl transferases and OPN were found in patients with ALD (p<0.001) when compared with the control subjects. OPN showed significant positive correlations with AST (r=0.76, p<0.001), ALT (r=0.64 p<0.001), ALP (r=0.68, p<0.001), and GGT (r=0.61, p<0.001). CONCLUSIONS: The current study focuses on the roles of NGAL and IMA, two sensitive markers of kidney injury that are particularly useful in identifying widespread endothelial dysfunction. As a result, the pattern of elevated NGAL and IMA levels can be useful for diagnosis.

Level of non-conventional lipid parameters and its comparative analysis with TSH in subclinical hypothyroidism.

OBJECTIVES: The objective of this study is to estimate lipid parameters in subclinical hypothyroidism and correlate it with TSH. METHODS: Forty newly diagnosed cases of subclinical hypothyroidism and Forty age and gender-matched healthy controls were recruited for the study. Blood samples were collected from them and serum lipid profile (i.e. HDL, LDL, TG, serum total cholesterol) of the subjects was estimated by standard photometric methods in a fully auto-analyzer (MINDRAY BS-300) using commercially available kits and VLDL cholesterol was calculated using the Friedewald's formula. While serum Ox-LDL, Lipoprotein A, Apolipoprotein A1 and Apo B were estimated by using commercial kit based on enzyme-linked immmunosorbent assay. RESULTS: The parameters such as Oxidized low-density lipoprotein (Ox-LDL), lipoprotein (a), apolipoprotein A1, apolipoprotein B and small dense lipoprotein (sd LDL) were significantly increased in subclinical hypothyroid cases when compared with the control subjects (p<0.0001). In present study results showed significant positive correlations of serum thyroid stimulating hormone (TSH) with Ox-LDL (r=0.85, p<0.01), sd LDL (r=0.71, p<0.01). CONCLUSIONS: The present study focuses on the role of Ox-LDL, sd-LDL Lipoprotein A, Apolipoprotein A1 and Apo B that are sensitive indicators of atherogenic dyslipidemia in subclinical hypothyroidism and can serve as a better & novel risk factor for CAD.

Association of inflammatory biomarker abnormalities with mortality in COVID-19: a meta-analysis.

Background: COVID-19 outbreak has engulfed different parts of the world, affecting more than 163 million people and causing more than 3 million deaths worldwide due to human transmission. Thus, it has become critical to identify the risk factors and laboratory parameters to identify patients who have high chances of worsening clinical symptoms or poor clinical outcomes. Therefore, the study aims to identify inflammatory markers that can help identify patients at increased risk for progression to critical illness, thus decreasing the risk of any mortality. Our study focussed on the predictive utility of C-reactive protein, Interleukin-6, D-dimer and Procalcitonin in assisting the management of COVID-19 patients with adverse clinical effects. Through literature search in electronic databases, we included the retrospective studies that evaluated the biomarkers among confirmed COVID-19 patients before initiation of treatment and who had a definite outcome (dead or discharged). Biomarkers were expressed in standardized difference in mean value, calculated based on study sizes and mean values between survivors and non-survivors considered the effect size. We carried out a meta-regression analysis to identify the causes of the heterogeneity between the studies. Results: Number of studies eligible for C-reactive protein, D-dimer and Interleukin-6 markers were eight, seven and four, respectively. Using random effect model revealed that the overall effect size with 95% confidence interval (CI) for C-reactive protein, D-dimer and Interleukin-6 were 1.45 (0.79-2.12) milligrams/litre, 1.12 (0.64-1.59) micrograms/millilitre Fibrinogen Equivalent Units and 1.34 (0.43-2.24) picograms/millilitre respectively was statistically significant (P < 0.05) inferring that the mean scores of these marker were significantly higher among the non-survivors compared to the survivors. Two studies were eligible for Procalcitonin marker and there was no heterogeniety (I 2-statistics = 0) between these studies. Therefore, fixed-effect model revealed that the overall effect size (95% CI) for Procalcitonin was 0.75 (0.30-1.21) Nanograms/millilitre was also high among non-survivors. Conclusions: The study found that serum levels of C-reactive protein, Interleukin-6 and D-dimer showed significant elevation in non-survivors compared to survivors. Raised inflammatory markers aid in the risk stratification of COVID-19 patients and their proper management.

Temporal changes in sleep quality and knee function following primary total knee arthroplasty: a prospective study.

PURPOSE: Several patient-reported outcome measures (PROMs) have been used to assess improvement in the quality of life following total knee arthroplasty (TKA). However, there is paucity of studies evaluating the sleep quality and knee function following TKA. The primary aim of our study was to evaluate the sleep quality and knee function in primary TKA patients using the Pittsburgh Sleep Quality Index (PSQI) and Knee Society Score (KSS), respectively. The secondary aim was to assess the correlation between the two outcome measures over the course of first post-operative year following TKA. METHODS: One hundred sixty-eight patients (female-140/male-28) with mean age of 64.63 years (± 7.50) who underwent 168 primary unilateral TKA using a cemented posterior-stabilised implant without patella resurfacing between June 2018 and October 2018 were included in the study. Global PSQI and KSS were recorded pre-operatively and post-operatively weekly up to six weeks and at one year. Body mass index (BMI) and Charlson comorbidity index (CCI) were recorded during pre-operative assessment. RESULTS: Mean(± SD) BMI and CCI were 28.45(± 4.64) and 2.48(± 0.93), respectively. Pre-operative global PSQI of 1.98(± 0.97) increased to 13.48(± 3.36) in the first post-operative week (p < 0.001) and remained high during all the six weeks following TKA (p < 0.001), whereas at the first post-operative year, it reduced to 2.10(± 1.15) (p = 0.15). Pre-operative KSS of 52.00(± 9.98) increased to 71.67(± 6.58) and 85.49(± 4.67) at 6 weeks and the first post-operative year respectively (p < 0.001). Pre-operative global PSQI had moderate correlation with pre-operative KSS (r = 0.39) (p < 0.001). Strong correlation was noted between pre-operative global PSQI and six week post-operative KSS (r = 0.47) (p < 0.001). Low correlation was noted between pre-operative global PSQI and KSS at the first post-operative year (r = 0.10, p = 0.19) following TKA. Multiple regression analysis revealed age, CCI, and pre-operative range of motion as independent predictors of global PSQI. CONCLUSIONS: Patients undergoing TKA experience changes in sleep quality but report an overall improvement in knee function during the first post-operative year. Sleep quality has moderate to strong correlation with knee function in the early post-operative period beyond which there is a low correlation with knee function thereby suggesting a transient phenomenon. Hence patients undergoing TKA can be appropriately counselled regarding the variation in sleep quality in the post-operative period and reassured accordingly.

Tomato T2 ribonuclease LE is involved in the response to pathogens.

T2 ribonucleases (RNases) are RNA-degrading enzymes that function in various cellular processes, mostly via RNA metabolism. T2 RNase-encoding genes have been identified in various organisms, from bacteria to mammals, and are most diverse in plants. The existence of T2 RNase genes in almost every organism suggests an important biological function that has been conserved through evolution. In plants, T2 RNases are suggested to be involved in phosphate scavenging and recycling, and are implicated in defence responses to pathogens. We investigated the function of the tomato T2 RNase LE, known to be induced by phosphate deficiency and wounding. The possible involvement of LE in pathogen responses was examined. Expression analysis showed LE induction during fungal infection and by stimuli known to be associated with pathogen inoculation, including oxalic acid and hydrogen peroxide. Analysis of LE-suppressed transgenic tomato lines revealed higher susceptibility to oxalic acid, a cell death-inducing factor, compared to the wild type. This elevated sensitivity of LE-suppressed lines was evidenced by visual signs of necrosis, and increased ion leakage and reactive oxygen species levels, indicating acceleration of cell death. Challenge of the LE-suppressed lines with the necrotrophic pathogen Botrytis cinerea resulted in accelerated development of disease symptoms compared to the wild type, associated with suppressed expression of pathogenesis-related marker genes. The results suggest a role for plant endogenous T2 RNases in antifungal activity.

A CBL-interacting protein kinase AdCIPK5 confers salt and osmotic stress tolerance in transgenic tobacco.

CBL interacting protein kinases play important roles in adaptation to stress conditions. In the present study, we isolated a CBL-interacting protein kinase homolog (AdCIPK5) from a wild peanut (Arachis diogoi) with similarity to AtCIPK5 of Arabidopsis. Expression analyses in leaves of the wild peanut showed AdCIPK5 induction by exogenous signaling molecules including salicylic acid, abscisic acid and ethylene or abiotic stress factors like salt, PEG and sorbitol. The recombinant AdCIPK5-GFP protein was found to be localized to the nucleus, plasma membrane and cytoplasm. We overexpressed AdCIPK5 in tobacco plants and checked their level of tolerance to biotic and abiotic stresses. While wild type and transgenic plants displayed no significant differences to the treatment with the phytopathogen, Phytophthora parasitica pv nicotianae, the expression of AdCIPK5 increased salt and osmotic tolerance in transgenic plants. Analysis of different physiological parameters revealed that the transgenic plants maintained higher chlorophyll content and catalase activity with lower levels of H2O2 and MDA content during the abiotic stress conditions. AdCIPK5 overexpression also contributed to the maintenance of a higher the K+/Na+ ratio under salt stress. The enhanced tolerance of transgenic plants was associated with elevated expression of stress-related marker genes; NtERD10C, NtERD10D, NtNCED1, NtSus1, NtCAT and NtSOS1. Taken together, these results indicate that AdCIPK5 is a positive regulator of salt and osmotic stress tolerance.

Utilization of Scenedesmus obliquus Protein as a Replacement of the Commercially Available Fish Meal Under an Algal Refinery Approach.

The approach of algal refinery as a method to reduce the cost of algal biodiesel by co-production of various value-added chemicals is the most up-coming strategy suggested for the economic viability of microalgal biodiesel. This concept being relatively new and novel, abundant literature on the subject is not available although fragmented data on some feedstocks are present. The main objective of this research paper is to propose an algal refinery design through utilization of Scenedesmus obliquus biomass for production of various industrially important products. For this purpose, first a protocol was standardized for maximum extraction of protein from S. obliquus biomass. Then, different experiments were conducted for 90 days each to find out the optimum concentration of microalgal protein that can be substituted in the diets of freshwater fishes for their maximum growth. During these experiments eight different growth parameters and seven water quality parameters were tested. Results showed that the standard + whole microalgal biomass + extracted microalgal protein diet (25:25:50) was the best diet for maximum growth of the freshwater fishes. After conducting these experiments, a detailed sequential extraction process for maximum valorization of the S. obliquus biomass or in other words an algal refinery was designed. The detailed sequential process developed, yielded 0.06 g of ß-carotene, 10 g of protein, 38 g (43 mL) of biodiesel, 2 g of omega-3 fatty acid, 3 g (2.4 mL) of glycerol and 18 g (23 mL) of bioethanol from 1 Kg wet (≈100 g dry) S. obliquus biomass thus converting 70% of the test microalgal biomass into biodiesel and other value-added products by using an algal refinery approach.

A proteomic study of cysteine protease induced cell death in anthers of male sterile tobacco transgenic plants.

Manifestation of male sterility in plants is an important requirement for hybrid seed production. Tapetum cell layer of anther is a primary target for genetic manipulation for male sterility. In our previous report, the targeted expression of Arachis cysteine protease in tapetum led to premature degeneration of tapetal layer that resulted in complete male sterility in transgenic tobacco plants. To correlate cysteine protease mediated cell death of tapetum, transmission electron microscopy (TEM) and proteomic pattern of anthers of cysteine protease induced male sterile plant were compared with the untransformed control plant. TEM study revealed the abnormal growth of tapetal cells exhibiting excessive vacuolization that synchronized with irregular exine wall formation of the microspores. In anther proteome, a total 250 protein spots were detected that were reproducible and exhibited similar distribution pattern. Further, anther proteome of male sterile plant showed the significant upregulation (≥ 1.5) of 56 protein spots. Using Mass spectroscopy (MALDI TOF/TOF), we have identified 14 protein spots that were involved in several processes such as energy metabolism, protein synthesis, plastid protein, lipid metabolism, and cell wall assembly. Upregulation of patatin-like protein-2 homolog, carboxylesterase 17 and dicer like protein-4 in male sterile anthers that have been demonstrated to induce cell death, suggesting that cysteine protease mediated premature tapetal cell death might involve the lipid peroxidation pathway in coordination with gene silencing mechanism.

Dye Coupled Aptamer-Captured Enzyme Catalyzed Reaction for Detection of Pan Malaria and P. falciparum Species in Laboratory Settings and Instrument-Free Paper-Based Platform.

Malaria diagnosis methods offering species-specific information on the causative parasites, along with their flexibility to use in different resource settings, have great demand for precise treatment and management of the disease. Herein, we report the detection of pan malaria and P. falciparum species using a dye-based reaction catalyzed by the biomarker enzymes Plasmodium lactate dehydrogenase ( PLDH) and Plasmodium falciparum glutamate dehydrogenase ( PfGDH), respectively, through instrument-based and instrument-free approaches. For the detection, two ssDNA aptamers specific to the corresponding PLDH and PfGDH were used. The aptamer-captured enzymes were detected through a substrate-dependent reaction coupled with the conversion of resazurin (blue, ∼λ605nm) to resorufin (pink, ∼λ570nm) dye. The reaction was monitored by measuring the fluorescence intensity at λ660nm for resorufin, absorbance ratio (λ570nm/λ605nm), and change in color (blue to pink). The detection approach could be customized to a spectrophotometer-based method and an instrument-free device. For both the approaches, the biomarkers were captured from the serum samples with the help of aptamer-coated magnetic beads prior to the analysis to exclude potential interferences from the serum. In the instrument-free device, a medical syringe (5 mL) prefabricated with a magnet was used for in situ separation of the enzyme-captured beads from the reaction supernatant. The converted dye in the supernatant was then efficiently adsorbed over a DEAE cellulose-treated paper wick assembled in the syringe hose. The biomarkers could be detected by both qualitative and quantitative format following the color and pixel intensity, respectively, developed on the paper surface. The developed method and technique offered detection of the biomarkers within a clinically relevant dynamic range, with the limit of detection values in the picomolar level. Flexible detection capability, low cost, interference-free detections, and portable nature (for instrument-free devices) are the major advantages offered by the developed approaches.

Protein-Induced Fluorescence Enhancement Based Detection of Plasmodium falciparum Glutamate Dehydrogenase Using Carbon Dot Coupled Specific Aptamer.

A novel 90-mer long ssDNA aptamer (NG3) covering a 40-mer random region targeting Plasmodium falciparum glutamate dehydrogenase ( PfGDH) developed through systematic evolution of ligands by exponential enrichment (SELEX) technique. The binding affinity of the aptamer to PfGDH discerned by circular dichroism (CD) was 0.5 ± 0.04 µM. The specificity of the aptamer toward the target was confirmed by gel electrophoresis and CD studies. The presence of two quadruplex forming regions, two big and four small stem loop structures with a δG of -7.99 kcal mol-1 for NG3 were deduced by computational studies. The spherical carbon dots (Cdots) of size 2-4 nm, synthesized by pyrolysis method using l-glutamate as a substrate were covalently linked to the amine modified aptamer. The Cdot with a band gap of 2.8 eV and a quantum yield of 34% produced fluorescence at ∼ λ410 nm when excited at λ320nm. The quantum yield of Cdot-aptamer assembly was increased up to 40% in the presence of the PfGDH in solution. A linear relationship with a dynamic range of 0.5 nM to 25 nM (R2 = 0.98) and a limit of detection (LOD) of 0.48 nM was observed between the fluorescence intensity of the Cdots-aptamer conjugate and the concentration of PfGDH. The method could detect PfGDH with an LOD of 2.85 nM in diluted serum sample. This novel simple, sensitive and specific protein induced fluorescence enhancement based detection of PfGDH has a great potential to develop as a method for malaria detection.

Molecular Approaches for Manipulating Male Sterility and Strategies for Fertility Restoration in Plants.

Usable pollination control systems have proven to be effective system for the development of hybrid crop varieties, which are important for optimal performance over varied environments and years. They also act as a biocontainment to check horizontal transgene flow. In the last two decades, many genetic manipulations involving genes controlling the production of cytotoxic products, conditional male sterility, altering metabolic processes, post-transcriptional gene silencing, RNA editing and chloroplast engineering methods have been used to develop a proper pollination control system. In this review article, we outline the approaches used for generating male sterile plants using an effective pollination control system to highlight the recent progress that occurred in this area. Furthermore, we propose possible future directions for biotechnological improvements that will allow the farmers to buy hybrid seed once for many generations in a cost-effective manner.

Aromatic Surfactant as Aggregating Agent for Aptamer-Gold Nanoparticle-Based Detection of Plasmodium Lactate Dehydrogenase.

A novel ssDNA aptamer (P38), with a 40 mer random region flanked by primer-binding sites on both sides, targeting Plasmodium falciparum lactate dehydrogenase (PfLDH) has been developed through systematic evolution of ligands by exponential enrichment (SELEX), including counter SELEX against human lactate dehydrogenase A and B (hLDH A and B). The 2D structure of P38 shows the presence of three stem loops with a δG of -9.18 kcal/mol. EMSA studies on P38 complexes with the increasing concentration of PfLDH revealed a dissociation constant of 0.35 µM. P38 has been exploited for the quantitative detection of PfLDH using cationic surfactant-mediated aggregation of gold nanoparticles (16-nm diameter) as an optical probe. Among the three different cationic surfactants, characterized by different hydrocarbon tail groups, benzalkonium chloride (BCK) was found to be most efficient with a limit of detection of 281 ± 11 pM. This BCK-based approach with the novel highly selective aptamer provides simple and sensitive detection of PfLDH in the clinically relevant range.

A novel zinc-binding alcohol dehydrogenase 2 from Arachis diogoi, expressed in resistance responses against late leaf spot pathogen, induces cell death when transexpressed in tobacco.

A novel zinc-binding alcohol dehydrogenase 2 (AdZADH2) was significantly upregulated in a wild peanut, Arachis diogoi treated with conidia of late leaf spot (LLS) pathogen, Phaeoisariopsis personata. This upregulation was not observed in a comparative analysis of cultivated peanut, which is highly susceptible to LLS. This zinc-binding alcohol dehydrogenase possessed a Rossmann fold containing NADB domain in addition to the MDR domain present in all previously characterized plant ADH genes/proteins. Transient over-expression of AdZADH2 under an estradiol inducible promoter (XVE) resulted in hypersensitive response (HR)-like cell death in tobacco leaf. However, the same level of cell death was not observed when the domains were transiently expressed individually. Cell death observed in tobacco was associated with overexpression of cell death related proteins, antioxidative enzymes such as SOD, CAT and APX and pathogenesis-related (PR) proteins. In A. diogoi, AdZADH2 expression was significantly upregulated in response to the plant signaling hormones salicylic acid, methyl jasmonate, and sodium nitroprusside.