Cinnamaldehyde Inhibits Postharvest Gray Mold on Pepper Fruits via Inhibiting Fungal Growth and Triggering Fruit Defense.

Gray mold infected with Botrytis cinerea frequently appears on fruits and vegetables throughout the supply chain after harvest, leading to economic losses. Biological control of postharvest disease with phytochemicals is a promising approach. CA (cinnamaldehyde) is a natural phytochemical with medicinal and antimicrobial activity. This study evaluated the effect of CA in controlling B. cinerea on fresh pepper fruit. CA inhibited B. cinerea growth in vitro significantly in a dose- (0.1-0.8 mM) and time-dependent (6-48 h) manner, with an EC50 (median effective concentration) of 0.5 mM. CA induced the collapse and breakdown of the mycelia. CA induced lipid peroxidation resulting from ROS (reactive oxygen species) accumulation in mycelia, further leading to cell leakage, evidenced by increased conductivity in mycelia. CA induced mycelial glycerol accumulation, resulting in osmotic stress possibly. CA inhibited sporulation and spore germination resulting from ROS accumulation and cell death observed in spores. Spraying CA at 0.5 mM induced a defense response in fresh pepper fruits, such as the accumulation of defense metabolites (flavonoid and total phenols) and an increase in the activity of defense enzymes (PAL, phenylalanine ammonia lyase; PPO, polyphenol oxidase; POD, peroxidase). As CA is a type of environmentally friendly compound, this study provides significant data on the activity of CA in the biocontrol of postharvest gray mold in peppers.

Berberine Suppresses Lung Metastasis of Cancer via Inhibiting Endothelial Transforming Growth Factor Beta Receptor 1.

This study investigated the effects of berberine (BBR) on pancreatic cancer (PC) lung metastasis and explored the underlying mechanisms, using a BALB/C-nu/nu nude mouse model injected with PC cells (AsPC-1). Intragastric administration of BBR dose-dependently improves survival of mice intravenously injected with AsPC-1 cells, and reduces lung metastasis. Especially, BBR significantly reduces lung infiltration of circulating tumor cells (CTCs) 24 h after AsPC-1 cells injection. In vitro, tumor cells (TCs) trigger endothelial barrier disruption and promote trans-endothelial migration of CFSE-labeled TCs. BBR treatment effectively ameliorates TC-induced endothelial disruption, an effect that is diminished by inhibiting transforming growth factor-ß receptor 1 (TGFBR1). Blocking TGFBR1 blunts the anti-metastatic effect of BBR in vivo. Mechanistically, BBR binds to the intercellular portion of TGFBR1, suppresses its enzyme activities, and protects endothelial barrier disruption by TCs which express higher levels of TGF-ß1. Hence, BBR might be a promising drug for reducing PC lung metastasis in clinical practice.

Ecosystem impact and dietary exposure of polychlorinated biphenyls (PCBs) and heavy metals in Chinese mitten crabs (Eriocheir sinensis) and their farming areas in Jiangsu, China.

This study investigated the presence of 18 dioxin-like and non-dioxin-like polychlorinated biphenyls (dl- and ndl-PCBs), heavy metals (Cd, Hg, Pb, and As) in Chinese mitten crabs (Eriocheir sinensis) and their distribution in Jiangsu, China. Risk assessment and source apportionment were employed for evaluating the eco-toxicological impact and human exposure. It was found that the compositions of PCBs varied spatially, suggesting different sources of pollutants, whilst PCB 28, 105, 114, and 126 were consistently found in all sample types, suggesting a common pollution source remained, and the bio-accumulation process was in effect. The total PCBs in sediment were found much higher than in water, and brown meat had the highest and most diverse PCB congeners among all tissues. The presence of heavy metals was found in all samples in descending order of As>Cd>Pb>Hg and in the order of shell>brown meat>white meat>gill for crabs. The results of risk assessment indicated that the potential carcinogenic and non-carcinogenic risks were within the acceptable range for long-term consumption of the crabs overall. However, the highest toxic equivalent (TEQ), carcinogenic, and non-carcinogenic risks were all recorded in Location C, where dl-PCB 126, 169, and As contributed to the majority of the risks. The ecological risk posed by all HMs was low, but cases of serious point source pollution have been found in the investigated regions, and risks caused by Cd individually should raise concerns. Source apportionment study revealed that the contaminants mostly originated from anthropogenic activities. Natural deposition and transportation played an important role as well.

Establishment of a sandwich enzyme-linked immunosorbent assay for specific detection of Bacillus thuringiensis (Bt) Cry1Ab toxin utilizing a monoclonal antibody produced with a novel hapten designed with molecular model.

Cry1Ab toxin is commonly expressed in genetically modified crops in order to control chewing pests. At present, the detection method with enzyme-linked immunosorbent assay (ELISA) based on monoclonal antibody cannot specifically detect Cry1Ab toxin for Cry1Ab's amino acid sequence and spatial structure are highly similar to Cry1Ac toxin. In this study, based on molecular design, a novel hapten polypeptide was synthesized and conjugated to keyhole limpet hemocyanin (KLH). Then, through animal immunization with this antigen, a monoclonal antibody named 2C12, showing high affinity to Cry1Ab and having no cross reaction with Cry1Ac, was produced. The equilibrium dissociation constant (K D) value of Cry1Ab toxin with MAb 2C12 was 1.947 × 10-8 M. Based on this specific monoclonal antibody, a sandwich enzyme-linked immunosorbent assay (DAS-ELISA) was developed for the specific determination of Cry1Ab toxin and the LOD and LOQ values were determined as 0.47 ± 0.11 and 2.43 ± 0.19 ng mL-1, respectively. The average recoveries of Cry1Ab from spiked rice leaf and rice flour samples ranged from 75 to 115%, with coefficient of variation (CV) less than 8.6% within the quantitation range (2.5-100 ng mL-1), showing good accuracy for the quantitative detection of Cry1Ab toxin in agricultural samples. In conclusion, this study provides a new approach for the production of high specific antibody and the newly developed DAS-ELISA is a useful method for Cry1Ab monitoring in agriculture products. Graphical Abstract Establishment of a DAS-ELISA for the specific detecting of Bacillus thuringiensis (Bt) Cry1Ab toxin.

Residual Behaviors of Six Pesticides in Shiitake from Cultivation to Postharvest Drying Process and Risk Assessment.

The dissipation of six pesticides (carbendazim, thiabendazole, procymidone, bifenthrin, λ-cyhalothrin, and ß-cyfluthrin) in shiitakes from cultivation to postharvest drying process was investigated, and the dietary exposure risk was estimated thereafter. The field trial study indicates that the half-lives of carbendazim, thiabendazole, and procymidone were much shorter than those of bifenthrin, λ-cyhalothrin, and ß-cyfluthrin. Furthermore, the effects of two drying processes on the residues and processing factors (PFs) were investigated. The results showed that hot-air drying resulted in higher residues than sunlight exposure drying. Both drying processes led to pesticide residue concentration (with PF > 1), except for thiabendazole upon sunlight exposure treatment. The estimated daily intakes (EDIs) ranged from 0.06% of the acceptable daily intake (ADI) for thiabendazole to 42.43% of the ADI for procymidone. The results show that the six pesticide residues in dried shiitakes are still within acceptable levels for human consumption on the basis of a dietary risk assessment.

Production and Characterization of Monoclonal Antibody Broadly Recognizing Cry1 Toxins by Use of Designed Polypeptide as Hapten.

In this study, by use of synthesized polypeptides as haptens, a monoclonal antibody with broad recognition against seven major Cry1 toxins (Cry1Aa, Cry1Ab, Cry1Ac, Cry1B, Cry1C, Cry1E, and Cry1F) has been produced and characterized. First, by comparing the three-dimensional structures of seven Cry1 toxins, analyzing the conserved sequences, and considering the antigenicity and hydrophilicity, three polypeptides (T1, T2, and T3) have been chosen and coupled to keyhole limpet hemocyanin as immunogens for the generic monoclonal antibody (Mab) generation. Thereafter, a double antibody sandwich enzyme-linked immunosorbent assay method (DAS-ELISA) was developed for simultaneous determination of seven Cry1 toxins. The results revealed that the haptens T1, T2, and T3 had different effects in the production of antibodies. Among them, the obtained Mab (strain 2D3) generated by T2 can recognize seven Cry1 toxins simultaneously. Equilibrium dissociation constant (KD) values for seven Cry1 toxins with Mab 2D3 were 1.198 × 10(-8) M for Cry1Aa, 2.197 × 10(-8) M for Cry1Ab, 1.367 × 10(-8) M for Cry1Ac, 2.092 × 10(-8) M for Cry1B, 5.177 × 10(-8) M for Cry1C, 4.016 × 10(-8) M for Cry1E, and 3.497 × 10(-8) M for Cry1F. For 2D3-based DAS-ELISA, the limits of detection (LOD) and limits of quantification (LOQ) can reach 15 and 30 ng·mL(-1) for each Cry1 toxin, respectively. Our study is the first report of a broadly specific immunoassay for multidetermination of seven major Cry1 toxins, and it will provide a new idea and technical routes for development of multidetermination immunoassays.

Organophosphorus pesticides detection using broad-specific single-stranded DNA based fluorescence polarization aptamer assay.

An approach is developed to detect the organophosphorus pesticides via competitive binding to a recombinant broad-specificity DNA aptamer with a molecular beacon (MB), the binding of the MB to the aptamer results in the activation of a fluorescent signal, which can be measured for pesticide quantification. Aptamers selected via the Systematic Evolution of Ligands by Exponential Enrichment (SELEX) were structurally modified and truncated to narrow down the binding region of the target, which indicated that loops of the aptamer contributed different functions for different chemical recognition. Thereafter, a variant fused by two different minimum functional structures, was clarified with broad specificity and increased affinity. Further molecular docking and molecular dynamics simulations was conducted to understand the molecular interaction between DNA structure and chemicals. 3D modeling revealed a hot spot area formed by 3 binding sites, forces including hydrogen bonds and van der Waals interactions appear to play a significant role in enabling and stabilizing the binding of chemicals. Finally, an engineered aptamer based approach for the detection of organophosphorus pesticides was successfully applied in a test using a real sample, the limit of quantification (LOQ) for phorate, profenofos, isocarbophos, and omethoate reached 19.2, 13.4, 17.2, and 23.4 nM (0.005 mg L(-1)), respectively.

Selection of DNA aptamers that bind to four organophosphorus pesticides.

Single-stranded DNA (ssDNA) aptamers against four organophosphorus pesticides (phorate, profenofos, isocarbophos and omethoate) were simultaneously isolated from an immobilized random ssDNA library by systematic evolution of ligands by exponential enrichment (SELEX) technique. After 12 rounds of in vitro selection, five ssDNA aptamer candidates were selected and their binding affinities were identified by a novel method using a molecular beacon. Two of the five ssDNA sequences, SS2-55 and SS4-54, demonstrated higher affinities and specificities to the four organophosphorus pesticides. They were defined as broad-spectrum aptamers binding to four different targets and their simulated secondary structures showed highly distinct features with typical stem and loop structures. The dissociation constant of SS2-55 and SS4-54 binding to the four organophosphorus pesticides ranged from 0.8 to 2.5 µM. These aptamers offered application potential in the analysis and/or neutralization of the residues of the four organophosphorus pesticides.