Insecticidal action of mammalian galectin-1-transfected Arabidopsis thaliana.

BACKGROUND: Galectins (GALs) are a family of mammalian sugar-binding proteins specific for ß-galactosides. Our previous studies have shown that the larval development of the diamondback moth (Plutella xylostella) is significantly disturbed when fed with recombinant mammalian galectin 1 (GAL1) derived from Escherichia coli. To further explore its applicability, two GAL1-overexpressed Arabidopsis [GAL1-Arabidopsis (whole plant) and GAL1-Arabidopsis-vas (vascular bundle-specific)] lines were established for insecticidal activity and mechanism studies. RESULTS: The expression level of GAL1 in transgenic Arabidopsis is 1-0.5% (GAL1-Arabidopsis) and 0.08-0.01% (GAL1-Arabidopsis-vas) of total leaf soluble protein. Survival, body weight, and food consumption significantly decreased in a time-dependent manner in P. xylostella larvae (with chewing mouthparts) fed on GAL1-Arabidopsis. The mortality of Kolla paulula (with piercing-sucking mouthparts and xylem feeder) fed on GAL1-Arabidopsis-vas was also significantly higher than that fed on wild-type Arabidopsis (WT-Arabidopsis), but was lower than that fed on GAL1-Arabidopsis. The histochemical structure and results of immunostaining suggested that the binding of GAL1 to the midgut epithelium of P. xylostella fed on GAL1-Arabidopsis was dose- and time-dependent. Ultrastructural studies further showed the disruption of microvilli, abnormalities in epithelial cells, and fragments of the peritrophic membrane (PM) in P. xylostella larvae fed on GAL1-Arabidopsis. CONCLUSION: The insecticidal mechanism of GAL1 involves interference with PM integrity and suggests that GAL1 is a potential candidate for bioinsecticide development. © 2024 Society of Chemical Industry.

Application of nZnO supported with nanoclay for improving shrimp immunity.

This study discloses the nanoscale silicate platelet-supported nZnO (ZnONSP) applied as novel feed additives in aquaculture. The preparation of the nanohybrid (ZnO/NSP = 15/85, w/w) was characterized by UV-visible spectroscopy, powder X-ray diffraction and transmission electron microscope. The effects of ZnONSP on growth, zinc accumulation, stress response, immunity and resistance to Vibrio alginolyticus in white shrimp (Penaeus vannamei) were \demonstrated. To evaluate the safety of ZnONSP, shrimps (2.0 ± 0.3 g) were fed with ZnONSP containing diets (200, 400 and 800 mg/kg) for 56 days. Dietary ZnONSP did not affect the weight gain, specific growth rate, feed conversion ratio, survival rate, zinc accumulation, and the expression of heat shock protein 70 in tested shrimps. To examine the immunomodulatory effect of ZnONSP, shrimps (16.6 ± 2.4 g) were fed with the same experimental diets for 28 days. Dietary ZnONSP improved the immune responses of haemocyte in tested shrimps, including phagocytic rate, phagocytic index, respiratory burst, and phenoloxidase activity, and upregulated the expression of several genes, including lipopolysaccharide, ß-1,3-glucan binding protein, peroxinectin, penaeidin 2/3/4, lysozyme, crustin, anti-lipopolysaccharide factor, superoxide dismutase, glutathione peroxidase, clotting protein and α-2-macroglobulin. In the challenge experiment, shrimps (17.2 ± 1.8 g) were fed with ZnONSP containing diets (400 and 800 mg/kg) for 7 days and then infected with Vibrio alginolyticus. Notably, white shrimps that received ZnONSP (800 mg/kg) showed significantly improved Vibrio resistance, with a survival rate of 71.4 % at the end of 7-day observation. In conclusion, this study discovers that ZnONSP is a new type of immunomodulatory supplement that are effective on enhancing innate cellular and humoral immunities, and disease resistance in white shrimp.

Effect of nanoclay supported nanosilver on the growth inhibition of aquatic pathogens and immunomodulatory effect in Penaeusvannamei.

Hybrid of nanosilver and nanoscale silicate platelet (AgNSP) is a safe, non-toxic nanomaterial which has been applied in medical use due to its strong antibacterial activity. The application of AgNSP in aquaculture was first proposed in the present study by evaluating the in vitro antibacterial activities against four aquatic pathogens, in vitro effects toward shrimp haemocytes as well as the immune responses and disease resistance in Penaeus vannamei fed with AgNSP for 7 days. For evaluating the antibacterial activities of AgNSP in culture medium, the minimum bactericidal concentration (MBC) values against Aeromonas hydrophila, Edwardsiella tarda, Vibrio alginolyticus and Vibrio parahaemolyticus were 100, 15, 625 and 625 mg/L, respectively. Moreover, the inhibition of pathogen growth over a period of 48 h could be achieved by the appropriate treatment of AgNSP in culturing water. In freshwater containing bacterial size of 103 and 106 CFU/mL, the effective doses of AgNSP against A. hydrophila were 12.5 and 450 mg/L, respectively while the effective doses against E. tarda were 0.2 and 50 mg/L, respectively. In seawater with same bacterial size, the effective doses against V. alginolyticus were 150 and 2000 mg/L, respectively while the effective doses against V. parahaemolyticus were 40 and 1500 mg/L, respectively. For the in vitro immune tests, the superoxide anion production and phenoloxidase activity in haemocytes were elevated after in vitro incubation with 0.5-10 mg/L of AgNSP. In the assessment of dietary supplemental effects of AgNSP (2 g/kg), no negative effect on the survival was found at the end of 7 day feeding trail. In addition, the gene expression of superoxide dismutase, lysozyme and glutathione peroxidase were up-regulated in haemocytes taken from shrimps received AgNSP. The following challenge test against Vibrio alginolyticus showed that the survival of shrimp fed with AgNSP was higher than that of shrimp fed with control diet (p = 0.083). Dietary AgNSP improved the Vibrio resistance of shrimp by increasing 22.7% of survival rate. Therefore, AgNSP could potentially be used as a feed additive in shrimp culture.

Accelerating the Layup Sequences Design of Composite Laminates via Theory-Guided Machine Learning Models.

Experimental and numerical investigations are presented for a theory-guided machine learning (ML) model that combines the Hashin failure theory (HFT) and the classical lamination theory (CLT) to optimize and accelerate the design of composite laminates. A finite element simulation with the incorporation of the HFT and CLT were used to generate the training dataset. Instead of directly mapping the relationship between the ply angles of the laminate and its strength and stiffness, a multi-layer interconnected neural network (NN) system was built following the logical sequence of composite theories. With the forward prediction by the NN system and the inverse optimization by genetic algorithm (GA), a benchmark case of designing a composite tube subjected to the combined loads of bending and torsion was studied. The ML models successfully provided the optimal layup sequences and the required fiber modulus according to the preset design targets. Additionally, it shows that the machine learning models, with the guidance of composite theories, realize a faster optimization process and requires less training data than models with direct simple NNs. Such results imply the importance of domain knowledge in helping improve the ML applications in engineering problems.

Bioaccumulation of arsenic and immunotoxic effect in white shrimp (Penaeus vannamei) exposed to trivalent arsenic.

Trivalent arsenic (As (III)) contamination in the marine environment can produce adverse effects in crustaceans. The present study investigated the chronic toxicity of As (III) in white shrimp (Penaeus vannamei) by analyzing the tissue bioaccumulation and non-specific immune responses. Shrimps were exposed to 0 (control), 50, 500, and 2500 µg/L of As (III) for 21 days. The results showed that the hepatopancreas was the main tissue of arsenic accumulation in white shrimp. The cumulative concentration of total arsenic and inorganic arsenic but not arsenobetaine was positively correlated with the exposure concentration. In vitro As (III) treatment (0-2500 µg/L) with haemocytes isolated from healthy shrimp did not cause the cytotoxicity, but this arsenic treatments inhibited the phagocytic rate and O2- production. Moreover, the decrease of total haemocyte count and the inhibition of phagocytic rate, phagocytic index, O2- production and phenoloxidase activity were observed in white shrimp under the exposure of As (III) over a period of 21 days. This study revealed that chronic As (III) stress could disturb arsenic metabolism and immune responses in P. vannamei.

Effect of Methylmercury Exposure on Bioaccumulation and Nonspecific Immune Respsonses in Hybrid Grouper Epinephelus fuscoguttatus × Epinephelus lanceolatus.

Mercury (Hg) is a dangerous heavy metal that can accumulate in fish and is harmful when consumed by humans. This study investigated the bioaccumulation of mercury in the form of methylmercury (MeHg) and evaluated nonspecific immune responses such as phagocytic activity and superoxide anion (O2-) production in hybrid grouper (Epinephelus fuscoguttatus × E. lanceolatus). The hybrid grouper leukocytes were incubated with methylmercury chloride (CH3HgCl) at concentrations of 10-10,000 µg/L to determine cell viability, phagocytic activity, and O2- production in vitro. Subsequently, the grouper were exposed daily to CH3HgCl mixed in the experimental diets at concentrations of 0, 1, 5, and 10 mg/kg for 28 days. The bioaccumulation of MeHg in the liver, head kidney, and muscle tissue was measured, and the phagocytic activity and O2- production were evaluated. In vitro results indicated that cell viability was significantly lower than that of the control group at concentrations > 500 µg/L. The phagocytic rate and O2- production at concentrations ˃ 500 and ˃ 200 µg/L, respectively, were significantly lower than those of the control group. The dietary exposure demonstrated that MeHg accumulated more substantially in the liver and head kidney compared with the muscle tissue in the treatment groups. Moreover, the cumulative concentration significantly increased with higher concentrations and more days of exposure. The phagocytic rate and O2- production in the treatment groups were significantly lower than those in the control group from days 2 and 1, respectively. In conclusion, hybrid grouper accumulated significant MeHg in the liver and head kidney compared with the muscle tissue, and higher concentrations and more exposure days resulted in decreased cell viability, phagocytic activity, and O2- production.

Guava (Psidium guajava) leaf extract enhances immunity, growth, and resistance against Vibrio parahaemolyticus in white shrimp Penaeus vannamei.

This study investigated the effects of guava leaf extract (GLE) on immune responses, growth performance, and resistance to Vibrio parahaemolyticus in white shrimp (Penaeus vannamei). To examine the effect of GLE on the immune response of white shrimps, they were treated with various concentrations of GLE on hemocyte (in vitro) and were orally administered (in vivo) feed containing various concentrations of 0, 1, 5, and 10 g kg-1 GLE (control, GLE1, GLE5, and GLE10, respectively) for 28 days. Furthermore, their growth performance was evaluated for 56 days. In a separate experiment, the shrimps were challenged with V. parahaemolyticus injection after 7 days of culture. In vitro experiments indicated that GLE is nontoxic and can activate immune response. In vivo experiments revealed that the GLE5 led to the highest total hemocyte count, phenoloxidase activity, phagocytic activity, and superoxide anion production and the highest upregulation of lipopolysaccharide, ß-1,3-glucan-binding protein, peroxinectin, lysozyme, crustin, penaeidin 2, penaeidin 3, clotting protein, superoxide dismutase, and glutathione peroxidase. Moreover, better growth performance was observed in the GLE groups, with GLE5 exhibiting the highest specific growth rate, weight gain, and feed conversion rate. In addition, GLE5 enhanced resistance to V. parahaemolyticus, with a survival rate of 72.27%. In conclusion, GLE was found to be effective in enhancing nonspecific immune response and growth performance and in reducing V. parahaemolyticus infection in white shrimp.

Composition of nanoclay supported silver nanoparticles in furtherance of mitigating cytotoxicity and genotoxicity.

Silver nanoparticle (Ag-NP) is well known for its high antibacterial efficacy. However, its toxicity toward mammalian cells is still a concern in clinical applications. The aim of our study was to evaluate the composition effects of Ag-NP supported by silicate nanoplatelet (NSP) with respect to the cytotoxicity and genotoxicity, and was in reference to the poly (styrene-co-maleic anhydride)-supported Ag-NP (Ag-NP/SMA). The NSP at the geometric dimension of averaged 80 x 80 x 1 nm3 was prepared from the exfoliation of natural clays and used to support different weight ratio of Ag-NP. The supporting limitation of NSP on Ag-NP was below the weight ratio of 15/85 (Ag-NP to NSP), and the detached Ag-NP from the Ag-NP/NSP (30/70) and Ag-NP/SMA hybrids were observed by TEM. Ames test was performed to assess the mutagenic potential of different compositions of Ag-NP/NSP, only Ag-NP/NSP (30/70) and Ag-NP/SMA hybrids exhibited mutagenicity when the concentration was 1.09 ppm or higher. In viewing of cytotoxicity using MTT tests toward HaCaT cells, the IC30 of Ag-NP/NSP (1/99, 7/93 and 15/85) were 1416.7, 243.6, and 148.9 ppm respectively, while Ag-NP/SMA was 64.8 ppm. The IC30 of Ag-NP/NSP (1/99, 7/93 and 15/85) were at least 833, 78 and 7 folds higher than their corresponding minimum inhibitory concentrations (MIC) respectively, and whereas Ag-NP/SMA was 6.4 folds. The Ag-NP/NSP and Ag-NP/SMA hybrids had been further investigated for genotoxicity by chromosomal aberrations and in vivo micronucleus assay within the concentration at IC10 and IC30, only Ag-NP/SMA showed a higher frequency of chromosomal aberrations. Our findings indicated that the viability of utilizing the NSP to maintain Ag-NP for antimicrobial activity, and the high-surface area of NSP served as an excellent support for associating Ag-NP and consequently rendering the mitigation of the inherent toxicity of Ag-NP in clinical uses.

Moringa oleifera Leaves' Extract Enhances Nonspecific Immune Responses, Resistance against Vibrio alginolyticus, and Growth in Whiteleg Shrimp (Penaeus vannamei).

Moringa is widely known as a plant with high medicinal properties. Therefore, moringa has a high potential for use as an immunostimulant in shrimp. This study investigated the effect of a moringa water extract on the immune response, resistance against V. alginolyticus, and growth performance of whiteleg shrimp. To perform the in vitro assay, hemocytes were incubated with different concentrations of the moringa extract. Furthermore, the moringa extract was incorporated at 0 (control), 1.25 g (ME1.25), 2.5 g (ME2.5), and 5.0 g (ME5.0) per kg of diet for the in vivo assay. During the rearing period, immune responses, namely the total hemocyte count (THC), phenoloxidase (PO) activity, phagocytosis activity, superoxide anion production, and immune-related gene expression were examined on days 0, 1, 2, 4, 7, 14, 21, and 28. Growth performance was measured 60 days after the feeding period. Furthermore, the shrimp were challenged with V. alginolyticus after being fed for different feeding durations. The results of the in vitro assay revealed that 100-250 ppm of the moringa extract enhanced the PO activity, phagocytic rate (PR), and superoxide anion production. The findings of the in vivo assay demonstrated that the THC, PO activity, PR, and immune-related gene expression, including alpha-2-macroglobulin, prophenoloxidase II, penaeidin2, penaeidin3, anti-lipopolysaccharide factor, crustin, lysozyme, superoxide dismutase, and clotting protein, were higher in the group of ME.25 and ME5.0 than in the control and ME1.25 at several time points. Growth performance was significantly increased (p < 0.05) in the ME2.5 group compared to the control group. Furthermore, the dietary ME2.5 resulted in a higher survival rate compared to that of the control group after challenging with V. alginolyticus, especially at ME2.5 administered for 4 and 7 days. This study indicated that the incorporation of the moringa extract at 2.5 g per kg of diet enhanced the immune response, the growth performance of the whiteleg shrimp, and the resistance against V. alginolyticus infection.

Effects of Phyllanthus amarus extract on nonspecific immune responses, growth, and resistance to Vibrio alginolyticus in white shrimp Litopenaeus vannamei.

This study investigates the effects of Phyllanthus amarus extract (PAE) on immune responses, growth, and resistance to Vibrio alginolyticus in white shrimp (Litopenaeus vannamei). In vitro PAE treatment did not alter the cell viability of haemocytes and significantly enhanced immune parameters such as phenoloxidase (PO) activity, phagocytic activity, and superoxide anion (O2-) production. We conducted two feeding trials to examine the effects of PAE on the growth, disease resistance, and innate immune parameters of white shrimp. In the first in vivo trial, shrimps (4.01 ± 0.03 g) were fed a diet containing 0 g (control), 10 g (PAE10), 20 g (PAE20), or 40 g (PAE40) of PAE per kilogram of feed for 56 days. After the feeding period, the PAE20 group showed a significantly higher weight gain and specific growth rate than shrimp fed the control diet. Furthermore, after challenge with V. alginolyticus, shrimp fed a diet containing PAE showed significantly higher survival than those fed the control diet. The second in vivo trial (28 days) was performed to identify the mechanisms of enhanced immunity in PAE-fed shrimp. Shrimp fed the PAE20 diet generally had the highest total haemocyte count, PO activity, phagocytic activity, and O2- production, followed by the PAE40 and PAE10 groups. Thus, our results suggest that administration of 20 g of PAE per kilogram of feed can enhance immunity, growth, and resistance to V. alginolyticus in white shrimp.

Effects of three medicinal herbs Bidens pilosa, Lonicera japonica, and Cyathula officinalis on growth and non-specific immune responses of cobia (Rachycentron canadum).

This study investigates the effects of three medicinal herbal extracts, namely Bidens pilosa (BPE), Lonicera japonica (LJE), and Cyathula officinalis (COE), on nonspecific immune parameters of cobia (Rachycentron canadum) in vitro and in vivo. During in vitro tests, BPE treatment increased reactive oxygen species (ROS) production in a dose-dependent manner in primary head kidney leukocytes. Similarly, ROS production rates were enhanced by LJE (50 and 100 mg/ml) and COE (100 mg/ml). This suggests that these three herbal extracts possess immunostimulating properties. We then conducted two feeding trials to examine the effects of these three herbal extracts on growth and innate immune parameters of cobia, and sought an optimal dietary supplementation proportion required for activating the non-specific immune responses. In the first trial, we supplemented the diet with 1, 5, or 10% of the individual extracts. After a ten-week feeding trial, no negative impacts on weight gain, feed conversion rate, and survival rate were observed in fish offered experimental diets. Further, ROS production, phagocytic capacity of the head kidney leukocytes, and serum lysozyme activity were enhanced by differing degrees in fish fed the herbal extracts compared to fish in the control group. A similar albumin/globulin ratio was seen between each experimental group and the control group regardless of the type and dose of herbal extract used, indicating these medicinal herbal extracts are safe for cobia. We then performed a 30-day feeding trial with lower extract concentrations (1, 3, and 5% of the diet) to identify dose responses in cobia at various time points so that we could establish a cost-effective manner of administering the three extracts for cobia. All BPE fed fish had higher ROS production compared to the control group, while phagocytosis rate and index were simultaneously raised in only the BPE30 group (3% BPE). Immune parameters such as ROS production, phagocytic rate, and serum lysozyme activity were triggered when fish received 30 g LJE per kg of feed. However, ROS production only increased in the LJE10 group (1% LJE) on day 30 and was not enhanced in the LJE50 group (5% LJE). Additionally, although the phagocytic rate and phagocytic index were induced in the LJE50 group, serum lysozyme activity was not elevated in this group (LJE50) at any time point examined. ROS production was greatly improved in all COE fed groups, but only the COE30 group (3% COE) showed prolonged enhanced phagocytic rate over the 30-day feeding trial.

ß-glucan alleviates the immunosuppressive effects of oxytetracycline on the non-specific immune responses and resistance against Vibrio alginolyticus infection in Epinephelus fuscoguttatus × Epinephelus lanceolatus hybrids.

This study was conducted to examine the combinatory effects of ß-glucan and oxytetracycline (OTC) on hybrid giant tiger groupers (Epinephelus fuscoguttatus × Epinephelus lanceolatus). In vitro tests, OTC significantly reduced superoxide anion production and phagocytic activity in primary head kidney leukocytes. However, this suppressive effect was alleviated by co-treatment with ß-glucan. Subsequently, feeding trials were performed to investigate the potential immunomodulatory effects of dietary ß-glucan alone or in combination with OTC on groupers. A total of 210 healthy groupers (368.00 ± 51.03 g) were divided into six groups. Group 1 was the control group, group 2 (BG) received 5 g ß-glucan per kg feed weight, groups 3-5 received 5 g/kg ß-glucan in combination with 10, 30, or 50 mg OTC/kg fish weight/day (groups M1, M2, and M3, respectively), and group 6 (O) received 50 mg OTC/kg fish weight/day. Fish were sampled to determine the innate immunity parameters and residual OTC levels in the muscle tissue during a 28-day feeding regimen. Residual OTC levels were considerably higher in groups M3 and O compared with the other groups, and peaked on day 14. This was followed by a slight decrease on day 28, despite a continuous supply of OTC. Notably, fish fed with OTC alone had significantly decreased phagocytic rates and superoxide anion production observed in head kidney leukocytes, as well as poorer protection against Vibrio alginolyticus infection. These immunosuppressive effects were not observed in the fish fed with ß-glucan in combination with a lower dose of OTC (group M2). Thus, these data suggest that the combination of dietary ß-glucan and OTC exerts synergistic immunostimulating effects that protect groupers from bacterial infection.