Subsequent Bioassay Research Articles

Identification and characterization of ATP-binding cassette transporters involved in chlorantraniliprole tolerance of model insect Drosophila melanogaster and agricultural pest Spodoptera frugiperda

ATP-binding cassette (ABC) transporter family is one of the largest transporter families, which plays an important role in insecticide tolerance. In this study, we found that the ABC transporter inhibitor verapamil could significantly enhance the toxicity of chlorantraniliprole (CHL) to the model insect Drosophila melanogaster. Forty-six ABC transporter genes of D. melanogaster were knocked down through the daughterless-GAL4 (Da-GAL4) strain. The subsequent bioassay result showed that D. melanogaster with DmCG5772, DmCG1494, and DmCG5853 genes silencing significantly increased mortality after CHL treatment. Based on the genome of the fall armyworm (FAW), three genes with the best hits were identified, and SfABCA1 (XM_035576510.2) and SfABCG10 (XM_035577893.2) were successfully cloned. Spatiotemporal expression pattern analysis showed that SfABCA1 and SfABCG10 were both highly expressed in adult and pupal stages. Hemolymph was also a tissue with high expression of these two genes. LC10 dose of CHL could induce the expression levels of SfABCA1 and SfABCG10, with SfABCG10 upregulated 8-fold after 48 h of CHL treatment. Furthermore, overexpression of SfABCA1 and SfABCG10 increased the viability of Sf9 cell under CHL treatment. Our findings indicate that SfABCA1 and SfABCG10 might associate with the tolerance of CHL in S. frugiperda. These results are not only helpful in understanding the role of ABC transporters in CHL tolerance of other agricultural pests, but also lay a theoretical foundation for delaying the development of CHL resistance in pest management.

Toxicity Assessment of Tannery Sludge Extract During Early Seedling Growth in Brassica nigra, Vigna radiata, Vigna mungo, Raphanus sativus and Capsicum frutescens

Tannery sludge (TS) contains a number of useful components like Organic carbon, Organic Matter, PO43-, Ca2+, Mg2+, and other nutrients essential for plant growth. However, presence of water-soluble toxic metals in TS may cause phytotoxicity. Primary objective of this work was to adopt a bio-chemical strategy for assessing the potential consequences of agricultural application of tannery sludge, via sludge characterization and subsequent bioassay investigations. Heavy metal concentration in the TS was found to be in the order of: Cr (19576±2065.52) > Pb (16.98±2.54) > Cu (1.76±0.20) > Ni (0.98±0.00) > Cd (0.19±0.00) mg kg-1. Further, bioassay tests were performed in terms of germination index (GI), relative seed germination (RSG) and relative root growth (RRG) on Brassica nigra, Vigna radiata, Vigna mungo, Raphanus sativus, and Capsicum frutescens. RSG and RRG were found to be more than 80% whereas, GI was > 66% in all the tested crops exposed to TS extract at 1:5 and 1:10 (w/v) dilutions. Results revealed that the toxicants present in the TS were failed to cause toxicity at early seedling growth and may be used as a soil conditioner after appropriate dilution. However, long term field trails are essential before agricultural utilization.

A chemical approach to extend flower longevity of Japanese morning glory via inhibition of master senescence regulator EPHEMERAL1.

Petal senescence in flowering plants is a type of programmed cell death with highly regulated onset and progression. A NAM/ATAF1,2/CUC2 transcription factor, EPHEMERAL1 (EPH1), has been identified as a key regulator of petal senescence in Japanese morning glory (Ipomoea nil). Here we used a novel chemical approach to delay petal senescence in Japanese morning glory by inhibiting the DNA-binding activity of EPH1. A cell-free high-throughput screening system and subsequent bioassays found two tetrafluorophthalimide-based compounds, Everlastin1 and Everlastin2, that inhibited the EPH1-DNA interaction and delayed petal senescence. The inhibitory mechanism was due to the suppression of EPH1 dimerization. RNA-sequencing analysis revealed that the chemical treatment strongly suppressed the expression of programmed cell death- and autophagy-related genes. These results suggest that a chemical approach targeting a transcription factor can regulate petal senescence.

Secondary metabolites from marine fungus Penicillium chrysogenum VH17 and their antimicrobial and cytotoxic potential.

One new compound, methyl 3-((1-((2-carbamoylphenyl)amino)-1-oxopropan-2-yl)amino)-3-oxopropanoate (1), along with 9 known secondary metabolites (2-10) were isolated and elucidated chemical structures from the methanol extract of the marine-derived fungus Penicillium chrysogenum VH17. Subsequent bioassays showed the antimicrobial and cytotoxic potential of the isolated compounds. All compounds 1-10 displayed antimicrobial effects against at least one tested reference microorganism with MIC values ranging from 32 to 256µg mL-1. Furthermore, compound 4 exhibited significant cytotoxicity against all tested cell lines, HepG2, A549, and MCF7 with IC50 values of 29.43±1.37, 33.02±1.53, and 36.72±1.88 µM, respectively, whereas compound 3 exhibited weak cytotoxicity against MCF7 and HepG2 cell lines with IC50 values of 87.17±6.31 and 97.32±5.66 µM, respectively.

Passive sampler housing and sorbent type determine aquatic micropollutant adsorption and subsequent bioassay responses

The combination of integrative passive sampling and bioassays is a promising approach for monitoring the toxicity of polar organic contaminants in aquatic environments. However, the design of integrative passive samplers can affect the accumulation of compounds and therewith the bioassay responses. The present study aimed to determine the effects of sampler housing and sorbent type on the number of chemical features accumulated in polar passive samplers and the subsequent bioassay responses to extracts of these samplers. To this end, four integrative passive sampler configurations, resulting from the combination of polar organic chemical integrative sampler (POCIS) and Speedisk housings with hydrophilic-lipophilic balance and hydrophilic divinylbenzene sorbents, were simultaneously exposed at reference and contaminated surface water locations. The passive sampler extracts were subjected to chemical non-target screening and a battery of five bioassays. Extracts from POCIS contained a higher number of chemical features and caused higher bioassay responses in 91% of cases, while the two sorbents accumulated similar numbers of features and caused equally frequent but different bioassay responses. Hence, the passive sampler design critically affected the number of accumulated polar organic contaminants as well as their toxicity, highlighting the importance of passive sampler design for effect-based water quality assessment.

Discovery of Novel Anti-Resistance AR Antagonists Guided by Funnel Metadynamics Simulation.

Androgen receptor (AR) antagonists are widely used for the treatment of prostate cancer (PCa), but their therapeutic efficacy is usually compromised by the rapid emergence of drug resistance. However, the lack of the detailed interaction between AR and its antagonists poses a major obstacle to the design of novel AR antagonists. Here, funnel metadynamics is employed to elucidate the inherent regulation mechanisms of three AR antagonists (hydroxyflutamide, enzalutamide, and darolutamide) on AR. For the first time it is observed that the binding of antagonists significantly disturbed the C-terminus of AR helix-11, thereby disrupting the specific internal hydrophobic contacts of AR-LBD and correspondingly the communication between AR ligand binding pocket (AR-LBP), activation function 2 (AF2), and binding function 3 (BF3). The subsequent bioassays verified the necessity of the hydrophobic contacts for AR function. Furthermore, it is found that darolutamide, a newly approved AR antagonist capable of fighting almost all reported drug resistant AR mutants, can induce antagonistic binding structure. Subsequently, docking-based virtual screening toward the dominant binding conformation of AR for darolutamide is conducted, and three novel AR antagonists with favorable binding affinity and strong capability to combat drug resistance are identified by in vitro bioassays. This work provides a novel rational strategy for the development of anti-resistant AR antagonists.

Reproducible growth of Brachypodium in EcoFAB 2.0 reveals that nitrogen form and starvation modulate root exudation.

Understanding plant-microbe interactions requires examination of root exudation under nutrient stress using standardized and reproducible experimental systems. We grew Brachypodium distachyon hydroponically in fabricated ecosystem devices (EcoFAB 2.0) under three inorganic nitrogen forms (nitrate, ammonium, and ammonium nitrate), followed by nitrogen starvation. Analyses of exudates with liquid chromatography-tandem mass spectrometry, biomass, medium pH, and nitrogen uptake showed EcoFAB 2.0's low intratreatment data variability. Furthermore, the three inorganic nitrogen forms caused differential exudation, generalized by abundant amino acids-peptides and alkaloids. Comparatively, nitrogen deficiency decreased nitrogen-containing compounds but increased shikimates-phenylpropanoids. Subsequent bioassays with two shikimates-phenylpropanoids (shikimic and p-coumaric acids) on soil bacteria or Brachypodium seedlings revealed their distinct capacity to regulate both bacterial and plant growth. Our results suggest that (i) Brachypodium alters exudation in response to nitrogen status, which can affect rhizobacterial growth, and (ii) EcoFAB 2.0 is a valuable standardized plant research tool.

Identification of maturation-inducing steroid in sturgeons via comprehensive analyses of steroids produced during oocyte maturation

Although 17α, 20β-dihydroxy-4-pregnen-3-one (DHP) and 17α, 20β, 21-trihydroxy-4-pregnen-3-one (20β-S) have been identified as maturation-inducing steroids (MIS) in several teleosts, to date, no MISs have been identified in sturgeons. As it remains possible that an unidentified steroid is an MIS in sturgeons, this study aimed to identify a sturgeon MIS via comprehensive analyses and maturation-inducing (MI) assay of C21 steroids. In vivo and in vitro comprehensive analyses of C21 steroids revealed that serum DHP concentrations were rapidly elevated in the oocyte maturation phase and the DHP production level was notably high among C21 steroids. MI assay indicated that the MI activity of DHP, 17α-hydroxyprogesterone (17OHP), a precursor of DHP, 17α, 20α-dihydroxy-4-pregnen-3-one (αDHP), and 20β-S was high among C21 steroids, but the MI activity of these steroids were similar. In the C21 steroids produced in ovarian follicles during oocyte maturation, 17OHP, αDHP, and unidentified compounds had a low production level, and 20β-S was suggested to be metabolized from DHP after oocyte maturation. Against this background, this study concluded that DHP is a steroid that possesses strong MI activity and is highly produced during oocyte maturation. Although this study could not identify an MIS in sturgeons by fractionation of plasma and subsequent bio assay, it was suggested that DHP is a major MIS in sturgeons.

Transcriptome analysis revealed that short-term stress in Blattella germanica to β-cypermethrin can reshape the phenotype of resistance adaptation

Previous studies on insect resistance have primarily focused on resistance monitoring and the molecular mechanisms involved, while overlooking the process of phenotype formation induced by insecticide stress. In this study, we compared the expression profiles of a beta-cypermethrin (β-CYP) resistant strain (R) and a susceptible strain (S) of Blattella germanica after β-CYP induction using transcriptome sequencing. In the short-term stress experiment, we identified a total of 792 and 622 differentially expressed genes (DEGs) in the S and R strains. Additionally, 893 DEGs were identified in the long-term adaptation experiment. To validate the RNA-Seq data, we performed qRT-PCR on eleven selected DEGs, and the results were consistent with the transcriptome sequencing data. These DEGs exhibited down-regulation in the short-term stress group and up-regulation in the long-term adaptation group. Among the validated DEGs, CUO8 and Cyp4g19 were identified and subjected to knockdown using RNA interference. Subsequent insecticide bioassays revealed that the mortality rate of cockroaches treated with β-CYP increased by 69.3% and 66.7% after silencing the CUO8 and Cyp4g19 genes (P<0.05). Furthermore, the silencing of CUO8 resulted in a significant thinning of the cuticle by 59.3% and 53.4% (P<0.05), as observed through transmission electron microscopy and eosin staining, in the S and R strains, respectively. Overall, our findings demonstrate that the phenotypic plasticity in response to short-term stress can reshape the adaptive mechanisms of genetic variation during prolonged exposure to insecticides. And the identified resistance-related genes, CUO8 and Cyp4g19, could serve as potential targets for controlling these pest populations.

Evaluation of Insecticidal Performances of Jayanti Plant (Sesbania sesban) for Integrated Control of Cabbage Caterpillar (Plutella xylostella)

The study on the use of local natural materials as a source of insecticides is intended, among other things, to reduce farmers' dependency on using synthetic chemicals. This research was conducted to obtain the most selective insecticidal active ingredient from Sesbania sesban plant for controlling cabbage caterpillars, choose a target mode of action that has a safe impact on the environment, and determine insecticidal stability during storage of the material prior to application and its stability in water after application. The dried powder of S. sesban leaves was extracted in stages using a series of solvents, namely hexane, DCM, ethanol and water, respectively. Each of the extract fractions produced was tested for its lethal toxicity against Plutella xylostella larvae and Diadegma semiclausum imago. Each of mortality data were processed by probit analysis to produce LC50(s) to determine their insecticidal selectivity. Subsequent bioassays were carried out using extract-water of S. sesban leaves and the data were processed by probit analysis to determine their anti-ovipositor, ovicidal powers and anti-feedant properties against P. xylostella. The insecticidal stability of S. sesban was also studied according to variations in the storage time of the simplicia before extraction, the storage time of the extract before dissolution, and its stability in water during application. The data was processed using ANOVA. The results show that the insecticidal activity of the ethanol extract fraction of S. sesban leaves for controlling cabbage caterpillars was very selective, namely very toxic to P. xylostella but very less toxic to D. semiclausum. Against P. xylostella, S. sesban leaf extract has also been proven to significantly prevent oviposition, inhibit egg hatching and feeding activity. The insecticidal power of S. sesban did not decrease significantly during storage of the simplicia for less than three months and during storage of the extract for less than seven days. However, the toxicity of the extract solution decreased drastically after 24 hours of application

A 2-component blend of coconut oil-derived fatty acids as an oviposition deterrent against Drosophila suzukii (Drosophilidae: Diptera).

Coconut free fatty acid (CFFA), a mixture of 8 fatty acids derived from coconut oil, is an effective repellent and deterrent against a broad array of hematophagous insects. In this study, we evaluated the oviposition deterrent activity of CFFA on spotted-wing drosophila (SWD; Drosophila suzukii), a destructive invasive pest of berries and cherries, and identified bioactive key-deterrent compounds. In laboratory 2-choice tests, CFFA deterred SWD oviposition in a dose-dependent manner with the greatest reduction (99%) observed at a 20-mg dose compared with solvent control. In a field test, raspberries treated with 20-mg CFFA received 64% fewer SWD eggs than raspberries treated with the solvent control. In subsequent laboratory bioassays, 2 of CFFA components, caprylic and capric acids, significantly reduced SWD oviposition by themselves, while 6 other components had no effect. In choice and no-choice assays, we found that a blend of caprylic acid and capric acid, at equivalent concentrations and ratio as in CFFA, was as effective as CFFA, while caprylic acid or capric acid individually were not as effective as the 2-component blend or CFFA at equivalent concentrations, indicating the 2 compounds as the key oviposition deterrent components for SWD. The blend was also as effective as CFFA for other nontarget drosophilid species in the field. Given that CFFA compounds are generally regarded as safe for humans, CFFA and its bioactive components have potential application in sustainably reducing SWD damage in commercial fruit operations, thereby reducing the sole reliance on insecticides.

Characterization and cytotoxity analysis of acuthy chestnut (Couepia edulis Prance) for use in herbal therapy

The Brazilian Amazon rainforest is rich in natural resources, which are widely used in areas such as food and health, especially for local populations. Couepia edulis Prance is a tree from the Amazon, consisting of fruits called agouti nut, which its use as a herbal medicine is widespread in the Amazon region. That said, the aim of this article was to verify the cytotoxicity of Agouti nut oil. For this, the nutritional characteristics, physical-chemical and bioactive properties of agouti nut oil were verified, for subsequent cytotoxicity bioassay. In the centesimal composition it was identified: Caloric Value 658.84± 0.02; Proteins 10.01 ± 0.03; Ash 2.34 ± 0.04; Lipids 61.20 ± 0.15; Carbohydrates 17.00 ± 0.20; Fibers 29.74 ± 0.64. In the fatty acid profile: arachidic (0.42%), palmitoleic (1.02%), stearic (7.11%), linoleic (14%), palmitic (29.20%), oleic (35.01%). In the cytotoxicity assay at different extract concentrations (100; 50; 6.25 and 1.56 µg.mL-1) cell viability of 78.60 to 90.20% was identified. We conclude that the almond and the oil of the Agouti nut at the nutritional and physical-chemical level were within the food standards for consumption, with a high content of total fibers.

Bacterial secretion systems contribute to rapid tissue decay in button mushroom soft rot disease.

The soft rot pathogen Janthinobacterium agaricidamnosum causes devastating damage to button mushrooms (Agaricus bisporus), one of the most cultivated and commercially relevant mushrooms. We previously discovered that this pathogen releases the membrane-disrupting lipopeptide jagaricin. This bacterial toxin, however, could not solely explain the rapid decay of mushroom fruiting bodies, indicating that J. agaricidamnosum implements a more sophisticated infection strategy. In this study, we show that secretion systems play a crucial role in soft rot disease. By mining the genome of J. agaricidamnosum, we identified gene clusters encoding a type I (T1SS), a type II (T2SS), a type III (T3SS), and two type VI secretion systems (T6SSs). We targeted the T2SS and T3SS for gene inactivation studies, and subsequent bioassays implicated both in soft rot disease. Furthermore, through a combination of comparative secretome analysis and activity-guided fractionation, we identified a number of secreted lytic enzymes responsible for mushroom damage. Our findings regarding the contribution of secretion systems to the disease process expand the current knowledge of bacterial soft rot pathogens and represent a significant stride toward identifying targets for their disarmament with secretion system inhibitors. IMPORTANCE The button mushroom (Agaricus bisporus) is the most popular edible mushroom in the Western world. However, mushroom crops can fall victim to serious bacterial diseases that are a major threat to the mushroom industry, among them being soft rot disease caused by Janthinobacterium agaricidamnosum. Here, we show that the rapid dissolution of mushroom fruiting bodies after bacterial invasion is due to degradative enzymes and putative effector proteins secreted via the type II secretion system (T2SS) and the type III secretion system (T3SS), respectively. The ability to degrade mushroom tissue is significantly attenuated in secretion-deficient mutants, which establishes that secretion systems are key factors in mushroom soft rot disease. This insight is of both ecological and agricultural relevance by shedding light on the disease processes behind a pathogenic bacterial-fungal interaction which, in turn, serves as a starting point for the development of secretion system inhibitors to control disease progression.

Imidacloprid: Impact on Africanized Apis mellifera L. (Hymenoptera: Apidae) workers and honey contamination

Apis mellifera L. (Hymenoptera: Apidae) is fundamental in the production chain, ensuring food diversity through the ecosystem service of pollination. The aim of this work was to evaluate the impact of imidacloprid, orally, topically, and by contact, on A. mellifera workers and to verify the presence of this active ingredient in honey. Toxicity levels were verified by bioassays. In bioassay 1, the levels correspond to the percentages of 100, 10, 1, 0.1, and 0.01% of the recommended concentration for field application of the commercial product Nortox® (active ingredient imidacloprid), with which we obtained the mean lethal concentration (LC50) in 48 h for A. mellifera, determining the concentration ranges to be used in the subsequent bioassays. Bioassays 2 and 3 followed the guidelines of the Organization for Economic Cooperation and Development, which specify the LC50 (48 h). In bioassay 4, the LC50 (48 h) and the survival rate of bees for a period of 120 h were determined by contact with a surface contaminated with imidacloprid, and in bioassay 5, the interference of the insecticide with the flight behavior of bees was evaluated. Honey samples were collected in agroecological and conventional georeferenced apiaries and traces of the imidacloprid were detected by means of high-performance liquid chromatography (HPLC-UV) with extraction by SPE C18. Bee survival was directly affected by the concentration and exposure time, as well behavioral performance, demonstrating the residual effect of imidacloprid on A. mellifera workers. Honey samples from a conventional apiary showed detection above the maximum residue limits (MRL) allowed by the European Union (0.05 μg mL−1), but samples from other apiaries showed no traces of this insecticide. Imidacloprid affects the survival rate and behavior of Africanized A. mellifera and honey quality.

Insecticidal and residual effect of chlorfenapyr on different surfaces for the control of Sitophilusoryzae

The immediate and residual insecticidal effects of chlorfenapyr applied on concrete and metal surfaces were evaluated against Sitophilus oryzae adults in laboratory bioassays. The dose used was 0.016 mg active ingredient (a.i.)/cm2 which refers to the highest label dose in case of severe infestation. Treated surfaces were either kept in continuous darkness or exposed to light. A series of subsequent bioassays were conducted in the treated surfaces, starting from the day of application and continued for three consecutive months, with new insects each time. For each bioassay, mortality levels were recorded after exposure for 3, 7 and 14 days. At the same time, the dissipation of chlorfenapyr residues on the treated surfaces was estimated at different time intervals (0, 3, 7, 15, 30, 60, 90 and 120 days) after application using a high-performance liquid chromatography system with a UV-detector (HPLC-UV). According to the results, mortality rates of S. oryzae adults after direct exposure to chlorfenapyr-treated concrete surfaces were high, reaching 100% at the end of the first bioassay. However, we observed a gradual reduction in mortality with the increase of the storage (post application) interval. On the contrary, mortality on metal surfaces was high and reached in all cases 100% after 7 days of exposure, regardless of the storage interval. Regarding the residues of chlorfenapyr on the surfaces, they remained relatively constant on metal surfaces for the entire 120-day period in both illuminated and non-illuminated surfaces. On concrete, an initial 20–35% reduction was observed over the first 3 days, whereas chlorfenapyr residues remained constant for the rest of the period regardless of the presence of illumination. Our results indicate that chlorfenapyr is effective for the control of S. oryzae, but its residual efficacy is moderated by the time after application, the surface type and partially by the presence of illumination.

Molecular Networking-Guided Isolation of Cyclopentapeptides from the Hydrothermal Vent Sediment Derived Fungus Aspergillus pseudoviridinutans TW58-5 and Their Anti-inflammatory Effects.

Repetitive isolation of known compounds remains a major challenge in natural-product-based drug discovery. LC-MS/MS-based molecular networking has become a highly efficient strategy for the discovery of new natural products from complex mixtures. Herein, we report a molecular networking-guided isolation procedure, which resulted in the discovery of seven new cyclopentapeptides, namely, pseudoviridinutans A-F (1-7), from the marine-derived fungus Aspergillus pseudoviridinutans TW58-5. Compounds 1-7 feature a rare amino acid moiety, O,β-dimethyltyrosine, observed for the first time from a marine-derived fungus. The planar structures of 1-7 were elucidated by detailed analyses of IR, UV, HR ESI-Q-TOF MS, and 1D and 2D NMR spectroscopic data. Meanwhile, their absolute configurations were determined through a combination of Marfey's method and X-ray diffraction. Subsequent bioassay revealed the anti-inflammation potential of 1-7, especially 6, which inhibited the production of nitric oxide (NO), a vital inflammatory mediator, in LPS-induced murine macrophage RAW264.7 cells by regulating the expression level of NLRP3 and iNOS.

Silver/chiral pyrrolidinopyridine relay catalytic cycloisomerization/(2 + 3) cycloadditions of enynamides to asymmetrically synthesize bispirocyclopentenes as PDE1B inhibitors

Significant progress has been made in asymmetric synthesis through the use of transition metal catalysts combined with Lewis bases. However, the use of a dual catalytic system involving 4-aminopyridine and transition metal has received little attention. Here we show a metal/Lewis base relay catalytic system featuring silver acetate and a modified chiral pyrrolidinopyridine (PPY). It was successfully applied in the cycloisomerization/(2 + 3) cycloaddition reaction of enynamides. Bispirocyclopentene pyrazolone products could be efficiently synthesized in a stereoselective and economical manner (up to >19:1 dr, 99.5:0.5 er). Transformations of the product could access stereodivergent diastereoisomers and densely functionalized polycyclic derivatives. Mechanistic studies illustrated the relay catalytic model and the origin of the uncommon chemoselectivity. In subsequent bioassays, the products containing a privileged drug-like scaffold exhibited isoform-selective phosphodiesterase 1 (PDE1) inhibitory activity in vitro. The optimal lead compound displayed a good therapeutic effect for ameliorating pulmonary fibrosis via inhibiting PDE1 in vivo.

Influence of Nutrition on Growth and Yield of G4 Mulberry Variety (Morus alba L.) and Bio-assay of Multivoltine Silkworm, Pure Mysore in Southern dry Zone of Karnataka, India

The experiment was carried out during 2019-20 at P-2 Pure Mysore Multivoltine Basic Seed Farm (BSF), National Silkworm Seed Organisation, Nagenahalli, Karnataka to know the effect of nutrient management on the growth and yield of G4 mulberry variety and its subsequent bioassay of multivoltine silkworm, Pure Mysore. The mulberry garden (three years old) with G4 variety planted in paired row system was used for the experiment with seven treatments and three replications. The growth parameters (average of 5 crops) viz., plant height, number of branches per plant, lowest number of leaf in 100g weight, weight of individual leaf, weight of 100 fresh leaf, leaf yield per plant and leaf yield per ha-1were significantly highest (134.6 cm, 14.0, 24.0, 4.18g, 384g, 800g and 55.55 Mt) with the application of 100 % RDF, poshan spray and application of vermicompost at5 t/ha/year. The results of the bio-assay (average of five rearings) also showed the superiority forweight of 10 full grown larvae (27.2 gm), single cocoon weight (1.27g), single shell weight (0.18g), shell ratio (14.25%), ERR (95.00%) pupation (92.00%), number of cocoons/kg (787) and yield per 100 dfls (52.10 kg) in the treatment having 100 % RDF, poshan spray and application of vermicompost of at5 t/ha than other treatments. Combined application of organic and inorganic sources of nutrients increased the productivity of the mulberry in G4 variety and subsequently better performance of Pure Mysore multivoltine seed cocoon parameters.

Quality control and purification of ready-to-use conjugated gold nanoparticles to ensure effectiveness in biosensing

Introduction: Gold nanoparticles (AuNPs) and their conjugates are used for many applications in the field of sensors. Literature lacks procedures able to separate, purify and characterize these species in native conditions without altering them while assuring a high throughput. This technological gap can be reduced by exploiting Asymmetrical Flow Field Flow Fractionation multidetection platforms (AF4 multidetection).Method: This work describes a complete set of strategies based on the AF4 system, from nanoparticle synthesis to separative method optimization to conjugates screening and characterization, achieving quantitative control and purification of ready-to-use conjugated Gold nanoparticles and ensuring effectiveness in biosensing.Results and Discussion: AF4-multidetection was used to study AuNPs with different types of surface coating [Poly ethylene glycol, (PEG) and Citrate], their binding behaviour with protein (Bovine serum albumin, BSA) and their stability after conjugation to BSA. A robust but flexible method was developed, able to be applied to different AuNPs and conjugating molecules. The morphology and conjugation mechanism of AuNPs-BSA conjugates were evaluated by combining online Multiangle light scattering (MALS) and offline Dynamic Light Scattering (DLS) measures, which provided an important feature for the quality control required to optimize bio-probe synthesis and subsequent bioassays.

Joint Application of Biological Techniques for the Remediation of Waste Contaminated with Hydrocarbons

Cleaning the oil industry's fuel storage and management facilities generates high levels of hazardous waste. This research aims to assess the use of biological bioremediation treatments, most commonly used for decontaminating soil, by applying them to hydrocarbon-contaminated waste. Turned pile composting using food-derived sludge as a co-substrate and the necessary proportion of bulking agent enabled the bioremediation of the initial mixture via the succession of microbial populations (PLFAs), with a 70% lower TPH concentration obtained 6 months after the start of the process. Subsequent bioassays using the composted material showed survival rates of over 80% with earthworms (Eisenia andrei) and a larger decrease in TPH in the joint treatment with earthworms and plants (Pennisetum clandestinum). The composting process reduces the concentration of hazardous organic compounds, allowing for the proper development of fauna and flora in the compost by improving the biodegradation rate.Graphical