Phagocytic Activity Of Hemocytes Research Articles

Salvia miltiorrhiza polysaccharide promotes the health of crayfish (Procambarus clarkii) by promoting hemocyte phagocytosis, protecting hepatopancreas and enhancing intestinal barrier function

Plant polysaccharides as immunomodulators are considered one of the effective measures to reduce antibiotic therapy in aquaculture. The immunomodulatory function of Salvia miltiorrhiza polysaccharides (SMP) has been demonstrated and begun to be applied in vertebrates, but its potential effect on crustaceans is unclear. In this study, crayfish (Procambarus clarkii) was fed with 0 %, 0.3 %, 0.7 %, 1.1 %, and 1.5 % SMP for 4 weeks to investigate the effects of SMP on hemocytes phagocytosis, hepatopancreatic function, and intestinal barrier function. The results revealed that hemocyte phagocytic activity was increased in all SMP groups. During the process of hemocytes phagocytic recognition and formation of phagosomes and phagolysosomes, the mRNA expression levels of mas, hem, rab3, ctsb, and lamp-1 were up-regulated mainly in the 0.3 % SMP group. During the clearance phase of phagocytosis, respiratory burst activity, ROS level, T-SOD, CAT, GST, and LZM activities were mainly increased in the 1.5 % SMP group. Hepatopancreas AKP and GOT activity were no significant change in all SMP groups. ACP activity was significantly enhanced in the 1.1 % SMP group. The GPT activity of 0.3–0.7 % SMP group was significantly decreased. The 0.7 % SMP group had the highest intestinal fold height. The highest index values of OTUs, Ace, Chao, and Shannon were in the 0.3 % SMP group. The dietary addition of 0.3 % SMP led to a tendency of increased relative abundance of Firmicutes and Bacteroidota at the phylum level, while the relative abundance of Proteobacteria at the phylum level decreased. In conclusion, dietary SMP could promote crayfish health by enhancing phagocytosis, protecting hepatopancreas and enhancing intestinal barrier function. This study contributes to the theoretical foundation for exploring the potential application of plant polysaccharides in crustaceans.

Hemolymph exosomes inhibit Spiroplasma eriocheiris infection by promoting Tetraspanin-mediated hemocyte phagocytosis in crab.

Exosomes released from infected cells are thought to play an important role in the dissemination of pathogens, as well as in host-derived immune molecules during infection. As an intracellular pathogen, Spiroplasma eriocheiris is harmful to multiple crustaceans. However, the immune mechanism of exosomes during Spiroplasma infection has not been investigated. Here, we found exosomes derived from S. eriocheiris-infected crabs could facilitate phagocytosis and apoptosis of hemocytes, resulting in increased crab survival and suppression of Spiroplasma intracellular replication. Proteomic analysis revealed the altered abundance of EsTetraspanin may confer resistance to S. eriocheiris, possibly by mediating hemocyte phagocytosis in Eriocheir sinensis. Specifically, knockdown of EsTetraspanin in E. sinensis increased susceptibility to S. eriocheiris infection and displayed compromised phagocytic ability, whereas overexpression of EsTetraspanin in Drosophila S2 cells inhibited S. eriocheiris infection. Further, it was confirmed that intramuscular injection of recombinant LEL domain of EsTetraspanin reduced the mortality of S. eriocheiris-infected crabs. Blockade with anti-EsTetraspanin serum could exacerbate S. eriocheiris invasion of hemocytes and impair hemocyte phagocytic activity. Taken together, our findings prove for the first time that exosomes modulate phagocytosis to resist pathogenic infection in invertebrates, which is proposed to be mediated by exosomal Tetraspanin, supporting the development of preventative strategies against Spiroplasma infection.

Dietary effect of botanical blend (Phyto AquaNity™) on growth, immunity and survival of Pacific White shrimps challenged against WSSV

The present study was performed to assess the effect of a commercial product Phyto AquaNity™ (Artepillin C and curcuminoids) supplementation on growth performance, immune parameters, and disease resistance of whiteleg shrimps (Litopenaeus vannamei) against White Spot Syndrome Virus (WSSV) infection. Shrimps of initial weight of 0.9–1 g were fed Phyto AquaNity™ at three dosages (0.0, 0.5 and 1.0 g kg−1) over 42 days. After the feeding period, growth performance parameters were significantly improved in the group of 1.0 g kg−1. The highest specific growth rate and feed efficiency were observed in the group 1.0 g kg−1 (P < 0.05). At the end of the feeding period, all groups were challenged against WSSV by recording daily mortalities over 14 days and measuring immune parameters. Shrimps fed with Phyto AquaNity™ (1.0 g kg−1) showed a significant reduction of mortalities with the lowest rate compared to other groups (0.5 and 0.0 g kg−1). Moreover, this dosage enhanced all immunological parameters in challenging conditions such as total hemocyte counts, granular cell, hyaline cell, phenoloxidase activity and phagocytic activity (p > 0.05). Analysis of lysozyme gene expression was improved in groups fed with Phyto AquaNity™ and the lowest value was observed for the positive group (challenged & without Phyto AquaNity™ supplement). Overall, 1.0 g kg−1 of Phyto AquaNity™ showed better results in improving growth performance, immune responses, and survival in shrimps challenged with WSSV.

The Influence of Adding Api-Api Mangrove Leaf Extract as an Immunostimulant on Vaname Shrimp Against Vibriosis Disease Caused by Vibrio parahaemolyticus Bacteria

Vanname shrimp cultivation is one of the most developed aquacultures in Indonesia and makes it one of the commodities with the highest foreign exchange inputs for the Indonesian state from the fisheries sector. Disease attack is one of the main causes of failure in vaname shrimp culture, especially disease attacks caused by Vibrio parahaemolyticus bacteria. Avicennia alba mangrove leaf extract is known to have active compounds that can act as antibacterial. This research was conducted by mixing Avicennia alba mangrove leaf extract with doses of 150 ppm (VA), 250 ppm (VB) and 350 ppm (VC) into commercial feed by re-pelletting. Positive control (KV+) treatment used commercial vitamins added to the feed. The vaname shrimp used was 60 days old with an average initial weight of 8.9 grams. The aquariums used were 15 pieces with a size of 60 x 30 x 40 cm and a volume of 54 liters of water. The white shrimp challenge test with Vibrio parahaemolyticus bacteria with a concentration of 106 Cfu/ml as much as 0.2 ml. The clinical symptoms of white vaname shrimp include pale white hepatopancreas, blackened tail, grippy swimming legs, necrosis in some parts of the body, necrosis of the tail, blackened and necrotic rectum, broken intestines that are partially brown and starting to turn white, empty intestines, presence of white feces. appears in the aquarium, decreased appetite, as well as not actively swimming. The results showed that Avicennia alba leaf extract had an effect on absolute weight growth (W), growth rate (GR), specific growth rate (SGR), survival rate (SR), total haemocyte count (THC) and phagocytic activity of white shrimp. The dose of 350 ppm VC treatment was the most influential dose as an immunostimulant in this study.

Biofloc system supplemented by Pseudoalteromonas piscicida 1Ub protects the Pacific white shrimp Penaeus vannamei from Vibrio parahaemolyticus infection

This study aimed to evaluate the supplementation of the probiotic Pseudoalteromonas piscicida 1Ub to the biofloc system as an ecofriendly strategy for protecting white shrimp (Penaeus vannamei) from Vibrio parahaemolyticus infection. Shrimp with an average body weight of (0.50 ± 0.09) g were reared in 30 glass jars with a working volume of 2.5 L at a density of 20 ind/L. Shrimp were reared for 5 d for each treatment, which included the biofloc system without and with 106 colony forming unit (CFU) per mL probiotic. The regular clear water system was used as control. All treatment groups were challenged with 103, 105, and 107 CFU/mL V. parahaemolyticus. For the negative control, shrimp were reared without V. parahaemolyticus. The results showed that the density of V. parahaemolyticus cocultured with P. piscicida 1Ub decreased and the density of V. parahaemolyticus in rearing water and shrimp body in the probiotic-treated group was lower than that in the control group (P < 0.05). The survival and immune response (total hemocyte count, phagocytic activity, respiratory burst, phenoloxidase, and superoxide dismutase) of shrimp in the probiotic group was higher than that in the positive control (P < 0.05). Moreover, supplementing the biofloc system with the probiotic could protect shrimp hepatopancreas from damage caused by V. parahaemolyticus, regardless of bacterial density. Thus, the supplementation of the probiotic P. piscicida 1Ub in the biofloc system could significantly protect and increase the resistance of shrimp to V. parahaemolyticus infection.

Physiological and transcriptomic analyses reveal critical immune responses to hypoxia and sulfide in the haemolymph of clam Tegillarca granosa

Hypoxia and sulfide are inducing potential damage to aquatic organisms. However, the effects of hypoxia and sulfide on their immune systems and molecular mechanisms are not fully understood. In the present study, the clam Tegillarca granosa was exposed to hypoxia alone or in combination with sulfide (0.1, 0.5 mM) to investigate the physiological and transcriptomic responses in haemolymph. The IBR analysis revealed that moderate sulfide stimulated immune responses via increasing the total hemocyte counts, phagocytic activity, antibacterial activity, and antioxidant activity. The transcriptomic analysis revealed many critical signaling pathways (Toll and Imd, FoxO, NLR) and biological processes (antimicrobial/antibacterial peptide, interferon, interleukin, leukocyte, lymphocyte, mitophagy) involved in the immunostimulation. Our results would offer insights into the sulfide-tolerant molecular mechanisms in this species and provide a useful tool for assessing the integrated biological impacts of hypoxia and sulfide on shellfish.

Impact assessment of metals realeased by aluminium-based galvanic anode on the physiology of the abalone Haliotis tuberculata in controlled conditions.

To protect metal structures immersed in the sea from corrosion, the galvanic anode cathodic protection system (GACP) is often applied. However, this association leads to continuous oxidation of the galvanic anode and therefore to a release of a metal cocktail in the forms of ions or oxy-hydroxides. Therefore, the main objective of our study was to investigate the toxicity of elements released from the dissolution of an aluminium-based galvanic anode (∼95% Al, ∼5% Zn, <0.1% for In, Cu, Cd, Mn, Fe) on a grazing gastropod, the abalone Haliotis tuberculata. The present study was carried out in complement to other research currently in submission. Gastropods were exposed for 16 weeks (12 weeks of exposure and 4 weeks of decontamination phase) to 6 conditions including a control, 4 concentrations based on total aluminium level (86, 425, 1096 and 3549 µg L-1) and a trophic control, corresponding to abalones placed in non-contaminated natural seawater but fed with contaminated algae. The effects of metals on growth, glycogen levels, brix index of hemolymph, MDA levels in digestive gland and gills, hemocyte phagocytic activity, ROS production, lysosomal system and the progress of gametogenesis were investigated throughout the entire exposure allowing the realization of kinetics. The results revealed that the aluminium-based anode does not seem to have an effect on the health status of the individuals for environmentally realistic concentrations. However, in extreme conditions strong effects were reported on the growth, immune system and reproduction of abalone.

The immunostimulatory effects of Theobroma cacao L. pod husk extract via injection and dietary administrations on Macrobrachium rosenbergii and its resistance against Lactococcus garvieae

The extract from Theobroma cacao L, pod husk served as the immunostimulant to enhance the immunity and resistance against Lactococcus garvieae of Macrobrachium rosenbergii. In this study, we employed the injection method and dietary administration method to determine the effect of cocoa pod husk (CPH) on M. rosenbergii. The non-specific immune parameters and disease resistance were evaluated after the prawn injected with 1 μg prawn−1 CPH extract (C1), 3 μg prawn−1 CPH extract (C3), and 5 μg prawn−1 CPH extract (C5) for 1, 3, and 7 days. The results showed a significant increase of total haemocyte count (THC), differential haemocyte count (DHC), phenoloxidase (PO) activity, respiratory bursts (RBs), and phagocytic activity and clearance efficiency to L. garvieae. The non-specific immune parameters, physiological parameters, and disease resistance and growth performance were evaluated after the prawn fed with 1 g kg−1 CPH extract diet (CD1), 3 g kg−1 CPH extract diet (CD3) and 5 g kg−1 CPH extract diet (CD5). The results showed a significant increase in all immune parameters and showed a significant decrease in physiological parameters. No significant difference was observed in growth performance of prawn fed with the CPH containing diet. Both injection and dietary method showed a significant increase in disease resistance against to L. garvieae. We therefore recommend that CPH extract can be used as a immunostimulant for prawn by dietary administration to regulate immune responses, and carbohydrate metabolism lead to enhance resistance against pathogen.

Effects of two fish-derived probiotics on growth performance, innate immune response, intestinal health, and disease resistance of Procambarus clarkii

Selection of the appropriate strain for particular aquatic animals is necessary for the desirable benefits of probiotics application in aquaculture, however, it is not very clear whether fish-derived probiotics could provide beneficial effects on invertebrates. In the present study, two fish-derived probiotics, Bacillus amyloliquefaciens, and Lactococcus lactis, were administrated as feed additives for crayfish (Procambarus clarkii) at concentrations of 106, 107, and 108 CFU/g for 28 days, respectively. Then their effects on the growth performance, innate immune enzyme activity, immune-related genes expression, haemocyte phagocytic activity, intestinal microbial diversity, and pathogen resistance of crayfish were evaluated. The results showed that the final weight, weight gain (WG), and specific growth rate (SGR) were significantly higher in 108 CFU/g B. coagulans and L. lactis additive groups. Compared to the control group, the activities of immune-related enzymes (acid phosphatase, alkaline phosphatase, glutathione peroxidase, and lysozyme) in the intestine were significantly increased in most of the probiotics added groups. In almost all experimental groups, the mRNA expression of two AMP genes (PcALF3 and PcAst) was enhanced in the intestinal of crayfish, and the expression level was proportional to the amounts of probiotics used. Moreover, the intestinal of crayfish in probiotics groups have better integrity and a thicker mucosal layer, and higher density granules in epithelial cells were also observed. According to the results of next-generation sequencing technologies, the diversity of intestinal microbiota in crayfish was also increased after supplementation with B. coagulans or L. lactis. Finally, the phagocytosis rate of haemocytes and pathogen resistance has also been improved. Taken together, our results suggested that fish-derived probiotics have beneficial effects on crayfish, and they can be treated as potential feed additives in practical applications.

The growth performance, antioxidant and immune responses, and disease resistance of Litopenaeus vannamei fed on diets supplemented with Indian ginseng (Withania somnifera)

In the current study, white-leg shrimp (Litopenaeus vannamei) were fed on diets containing varying doses of Withania somnifera aqueous extract (WSAE) at a rate of 0 (control), 0.5, 1.0, and 2.0 g/kg feed for 56 days. After the feeding trial, shrimps in all groups were challenged with the exposure to Vibrio harveyi for ten days during which animals' mortality was observed. It is noted that the dietary WSAE linearly and quadratically stimulated shrimp's growth indices particularly at the treatment of 2.0 g/kg feed. Compared to the control group, the WSAE-fed L. vannamei had significantly higher villi length, villi width, and absorption area particularly in the treatment of 2.0 g/kg feed. Furthermore, L. vannamei fed on WSAE-enriched diets consumed more feed and exhibited higher total proteolytic activity, lipase, and α-amylase activities as compared with the control group. The dietary WSAE at escalating levels linearly and quadratically enhanced the antioxidant activity (serum superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), total antioxidant capacity, and reduced glutathione) and the immune response (total hemocyte counts, total protein, lysozyme, and phagocytic activity). Similarly, the mRNA expression levels of cMn-SOD, CAT, and GPx genes were linearly and quadratically upregulated in the hepatopancreas of L. vannamei fed on WSAE-enriched diets (especially in the 2.0 g/kg feed treatment), while their lowest levels were significantly observed in the control group. On the other hand, malondialdehyde levels were significantly decreased in WSAE-supplemented shrimp groups, and its highest levels were observed in animals fed on the control diet. After the bacterial exposure, the survival rates of L. vannamei fed on 1.0 and 2.0 g WSAE/kg feed (61.3% and 66.7%, respectively) were higher than those in the control animals. Taken together, the results obtained herein indicate that inclusion of WSAE in diets of L. vannamei effectively enhanced the growth, antioxidant biomarkers, immune response, and resistance to the V. harveyi infection, particularly at the treatment of 2.0 g/kg feed.

Acute toxic effects of thiamethoxam on Chinese mitten crab Eriocheir sinensis.

The information about toxic effects of thiamethoxam on non-target aquatic organisms is still incomplete. The semi-static toxicity test method was used to investigate the acute toxic effects of thiamethoxam on Eriocheir sinensis. The results showed that the median lethal concentration (LC50) of thiamethoxam to E. sinensis at 96h was 510μg/L, and the safety concentration (SC) was 51μg/L. After 96h exposure to thiamethoxam, the survival rates of crabs at concentrations of 0, 151.11, 226.67, 340, and 510μg/L were 100%, 76.19%, 64.29%, 61.91%, and 46.43%, respectively. A significant (P < 0.05) decrease of the number of hemocytes was observed in thiamethoxam groups. With the increase of thiamethoxam concentration, the phagocytic activity of hemocytes, the activities of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH-Px), total antioxidant capacity (T-AOC), and the activities of protease, amylase, and lipase of crabs increased firstly and then decreased, and the above indexes reached the maximum in 151.11μg/L thiamethoxam group. Collectively, a high concentration of thiamethoxam (510μg/L) had a great effect on the gene expression of immune metabolism-related factors in hepatopancreas and gill of crabs. These findings indicated that thiamethoxam exposure had the ability to impair immune and metabolic systems and resulted in the reduction of survival rate of crabs.

Protective effects of the biofloc system in Pacific white shrimp (Penaeus vannamei) culture against pathogenic Vibrio parahaemolyticus infection

The mitigation of vibriosis in shrimp culture could be accomplished by reducing the virulence of the pathogen or by increasing the immune response of the shrimp. This study aims to evaluate the application of a biofloc system in protecting Pacific white shrimp (Penaeus vannamei) from pathogenic Vibrio parahaemolyticus infection. Shrimp post-larvae (PL 20) with an average body weight of 0.041 ± 0.019 g were reared in an aquarium with a working volume of 33 L at a density of 3 L−1 for 21 days using two rearing systems, i.e., the biofloc system and the regular clear water system as control. In each system, the shrimp post-larvae were challenged by adding V. parahaemolyticus at different densities, 103, 105, and 107 CFU mL−1, while the negative control was performed by maintaining shrimp post-larvae in the clear water system without the addition of V. parahaemolyticus. The results of the in vitro experiment showed that the density and biofilm activity of V. parahaemolyticus reared in biofloc suspension was lower than that of the positive control (p < 0.05). The density of V. parahaemolyticus in shrimp rearing water was lower than that in the control at 105 CFU mL−1, especially on the 3rd day post-challenge, but there was no significant difference in the total presumptive Vibrio count between the biofloc treatment and the control. The survival, growth, and immune response parameters, such as total hemocyte count, phagocytic activity, respiratory burst, and phenoloxidase activity, of the shrimp, reared in the biofloc system were also higher than those of the positive control (p < 0.05), regardless of the density of V. parahaemolyticus. The present study demonstrated that the application of biofloc could significantly protect and increase the resistance of Pacific white shrimp against pathogenic V. parahaemolyticus infection.

Pectin from dry cacao pod husk mediates growth performance, immune resistance responses and carbohydrate metabolism of Litopenaeus vannamei through dietary administration

The sustainable uses of cacao, Theobroma cacao L, pod husk (CPH) produced from cacao bean industry bring environment great benefits. Botanical polysaccharides possessing a variety of physiological functions are therefore referred to as biological response modifiers, and those are also found in CPH. In this study, the weight gain rate (WGR), length gain rate (LGR), feeding efficiency (FE), and survival rate (SR) of shrimp, Litopenaeus vannamei, fed diets containing CPH pectin at 0 (CON), 1.0 (CPH1), 3.0 (CPH3) and 6.0 (CPH6) g/kg were calculated every 7 days within 56 days of feeding. The non-specific immune parameters, disease resistance, tolerance to hypothermal stress and stress indicators were also evaluated after the shrimp fed with CPH pectin containing diets. Shrimp that were fed with CPH3 after 56 days had higher WGR, LGR, and FE than those in CPH0. The SR significantly increased for shrimp challenged with Vibrio alginolyticus, and subjected to hypothermal (14 °C) stress when they were fed diets containing CPH3 and CPH6, and CPH6 after 14 or 28 days of feeding, respectively. Dietary CPH3 and/or CPH6 showed a significant increase of total hemocyte count (THC), differential hemocyte count (DHC), phenoloxidase (PO) activity, respiratory bursts (RBs), and phagocytic activity and clearance efficiency to V. alginolyticus at 14 and/or 28 days. Furthermore, shrimp fed with CPH pectin-contained diets after 14 and 28 days retarded secretion of dopamine (DA) and norepinephrine (NE), and decreased the level of glucose and lactate in plasma under cold stress. We therefore recommend that CPH pectin can be used as a physiomodulator for shrimp by dietary administration at 3.0 g/kg for 56 days to promote growth; and at 3.0–6.0 g/kg within an appropriate time frame (28 days) to regulate neuroendocrine and immune responses, and carbohydrate metabolism lead to enhance resistance against pathogen and hypothermal stress.

Ascorbic Acid Regulates the Immunity, Anti-Oxidation and Apoptosis in Abalone Haliotis discus hannai Ino.

The present study was conducted to investigate the roles of ascorbic acid (AA) in immune response, anti-oxidation and apoptosis in abalone (Haliotis discus hannai Ino). Seven semi-purified diets with graded levels of AA (0, 50, 100, 200, 500, 1000 and 5000 mg/kg) were fed to abalone (initial weight: 12.01 ± 0.001 g, initial shell length: 48.44 ± 0.069 mm) for 100 days. The survival, weight gain rate and daily increment in shell length were not affected by dietary AA. The AA content in the gill, muscle and digestive glands of abalone was significantly increased by dietary AA. In terms of immunity, dietary AA significantly improved the total hemocyte count, respiratory burst and phagocytic activity in hemolymph, and lysozyme activity in cell-free hemolymph (CFH). In the digestive gland, the TLR-MyD88-dependent and TLR-MyD88-independent signaling pathways were suppressed by dietary AA supplementation. The mRNA levels of β-defensin and arginase-I in the digestive gland were significantly increased by dietary AA. In the gill, only the TLR-MyD88-dependent signaling pathway was depressed by dietary AA to reduce inflammation in abalone. The level of mytimacin 6 in the gill was significantly upregulated by dietary AA. After Vibrio parahaemolyticus infection, the TLR signaling pathway in the digestive gland was suppressed by dietary AA, which reduced inflammation in the abalone. In terms of anti-oxidation, superoxide dismutase, glutathione peroxidase and catalase activities, as well as total anti-oxidative capacity and reduced glutathione content in CFH, were all significantly upregulated. The malondialdehyde content was significantly downregulated by dietary AA. The anti-oxidative capacity was improved by triggering the Keap1-Nrf2 pathway in abalone. In terms of apoptosis, dietary AA could enhance the anti-apoptosis ability via the JNK-Bcl-2/Bax signaling cascade in abalone. To conclude, dietary AA was involved in regulating immunity, anti-oxidation and apoptosis in abalone.

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, β-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.

Suppression of a JAK-STAT pathway target gene Ftz-F1H enhances the resistance of shrimp to Vibrio parahaemolyticus

Bacterial diseases, especially those caused by Vibrio infection, are major constraints to shrimp aquaculture. The fushi tarazu (FTZ) transcription factor (Ftz-F1) is an orphan nuclear receptor now known to be essentially involved in embryonic development, but its role in immunity is still unknown. In the current study, a Ftz-F1 homologue (Ftz-F1H) was identified from Pacific white shrimp Litopenaeus vannamei and functionally studied in the context of antibacterial response. Ftz-F1H was a target gene of the JAK-STAT pathway and its product was mainly localized in the nucleus. The expression of Ftz-F1H was significantly up-regulated upon bacterial stimulation, indicating its involvement in antibacterial response. In Ftz-F1H-silenced shrimp, expression of a set of immune effector genes and the phagocytic activity of hemocytes were increased. Moreover, the mortality rate of Vibrio parahaemolyticus infected shrimp as well as the bacterial load in tissues was significantly decreased after silencing of Ftz-F1H. These suggested that Ftz-F1H functioned as an inhibitor of antibacterial response in shrimp. To our knowledge, it is the first report on the immune function of Ftz-F1 family in invertebrates. The shrimp Ftz-F1H gene could be potentially used as a therapeutic target for improving immune defense against bacterial infection.

Studies on biochemical and synergistic effects of immunosuppressive concentration of imidacloprid with Beauveria bassiana and Metarhizium anisopliae for enhancement of virulence against vector mosquito Culex quinquefasciatus

In order to increase the virulence of two commonly used entomopathogens namely Beauveria bassiana and Metarhizium anisopliae for effective mosquito control programs, we developed new combined formulation by the use of immunosuppressive concentration of imidacloprid. Exposure of sublethal concentration (LC10 1.34 ppb) of imidacloprid to second instar larvae for 92 h that differently modulated the larval biochemical markers by decreasing the total protein (20.65% reduction), acetylcholinesterase (35.74% reduction) and increases the β-carboxylesterase (26.59% increase) and acid phosphatase (30.69% increase) levels while no significant difference was noticed in alkaline phosphatase of C. quinquefasciatus. Further we demonstrated that the imidacloprid exposure brings about reduction in phenoloxidase level (39.48% decrease) and nitric oxide production (37.64% decrease) in larval homogenate over control. The reduction in viability of hemocytes (15.37% decrease) and phagocytic activity of hemocytes (32.18% decrease) was noticed in imidacloprid treatment. Moreover in laboratory condition, exposure of 1 × 107 spores/ml of B. bassiana (984) and M. anisopliae (6060) alone for 92 h cumulative toxicity assay exhibited larval mortality of 36.47% and 47.64% respectively against C. quinquefasciatus. However in the synergistic experimental studies with LC10 of imidacloprid and 1 × 107 spores/ml of B. bassiana or M. anisopliae to the larvae for 92 h cumulative assay brought 60% and 50.59% more insecticidal activity than the respective entomopathogens alone. The substantial increase of larvicidal activity noticed in the synergistic test conditions against larvae of C. quinquefasciatus revealed that the inclusion of sublethal concentration was proved to be useful for effective larval control.

A Novel Forkhead Box Protein P (FoxP) From Litopenaeus vannamei Plays a Positive Role in Immune Response.

The forkhead box protein P (FoxP) family members have been known to be important for regulation of immune responses in vertebrates, but their roles in invertebrate immunity remain unclear. In this study, a novel FoxP gene (LvFoxP) was identified from Pacific white shrimp Litopenaeus vannamei and functionally studied in the context of immune response. Possessing a conserved FoxP coiled-coil domain and a forkhead domain, LvFoxP shared homology to vertebrate FoxP family members, in particular FoxP1. Expression of LvFoxP was detectable in all the examined tissues and could be up-regulated by immune challenge in gill and hemocytes. The LvFoxP protein was present in both the cytoplasm and nucleus of hemocytes and could be nuclear-translocated upon immune stimulation. Silencing of LvFoxP increased the susceptibility of shrimp to infections by Vibrio parahaemolyticus and white spot syndrome virus (WSSV) and down-regulated the expression of multiple components of NF-κB and JAK-STAT pathways and almost all the examined immune effector genes. Moreover, the phagocytic activity of hemocytes from LvFoxP-silenced shrimp against V. parahaemolyticus was decreased. These suggested that LvFoxP could play a positive role in immune response. The current study may provide novel insights into the immunity of invertebrates and the functional evolution of the FoxP family.

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 Yu-Ping-Feng polysaccharides (YPS) on the immune response, intestinal microbiota, disease resistance and growth performance of Litopenaeus vannamei

A 28-day feeding trial was conducted to investigate the effects of Yu-Ping-Feng polysaccharides (YPS) containing Astragalus polysaccharides (APS), Atractylodes macrocephala polysaccharides (AMP) and Saposhnikoviae polysaccharides (SPS) on the immune response, intestinal microbiota, disease resistance and growth performance of Litopenaeus vannamei. Seven hundred and twenty shrimp (3.04 ± 0.33 g) were fed the following diets: Control, YPS1 (0.13% APS + 0.0325% AMP + 0.0325% SPS), YPS2 (0.13% APS + 0.0325% AMP + 0.065% SPS) and YPS3 (0.13% APS + 0.0325% AMP+0.0975% SPS). After 14 and 28 days of feeding, the immune responses of hemocytes and intestine were measured. Intestinal microbiota and growth performance were measured after 28 days of feeding, after that, a 7-day challenge test against Vibrio harveyi was conducted. A significant (P < 0.05) increase of the total haemocyte count (THC), phagocytic activity, antibacterial activity and phenoloxidase (PO) activity was observed in shrimp fed YPS diets compared to the control. Also, dietary YPS supplementation particularly YPS3 group significantly increased the expressions of immune-related genes in the hemocytes and intestine. Regarding the intestinal microbiota, the microbial diversity and richness decreased and functional genes associated with short-chain fatty acids metabolism increased in YPS groups. After Vibrio harveyi challenge, the cumulative mortality in YPS groups was significantly lower than that of the control. Besides, dietary YPS had no significant effect on growth performance of shrimp (P > 0.05). The present results suggested that YPS could be considered as potential prebiotics for aquaculture farmed shrimp.