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The cost of the purification process hinders the extensive use of cytosine phosphate guanosine-oligodeoxynucleotides (CpG-ODNs) for shrimp culture. Therefore, this study used a shuttle vector plasmid to carry 60 copies of CpG-ODN 1668 (pAD43-25_60CpG), which can replicate in Escherichia coli and Bacillus subtilis strain RIK1285. The first experiment used a reverse gavage procedure to deliver a substance (PBS [CK], pAD43-25 [P0], and pAD43-25_60CpG [P60], respectively) directly into the anterior midgut of Penaeus vannamei and transcriptome sequence analysis with a reference genome was performed to examine the expression of well-known immune-related genes. The results showed that the expression levels of immune-related genes in P60 group were significantly increased, particularly those associated with AMPs. In addition, using RT‒qPCR, the expression levels of AMP genes (LvALF, LvPEN-2, and LvPEN-3) in the P60 group may vary depending on the tissue and time point. The second experiment used dietary supplementation with three kinds of heat-killed B. subtilis (HKBS, HKBS-P0, and HKBS-P60) in 28 days of feeding experiments. The results showed that dietary supplementation with HKBS-P60 did not significantly improve shrimp growth performance and survival. However, on days 14 and 28 of the feeding regimens, alkaline phosphatase (AKP) and acid phosphatase (ACP) activity were considerably higher than in other treatments. In addition, following infection with Vibrio harveyi, AKP and ACP activity in the HKBS-P60 group was significantly higher than in other treatments, particularly at the early stage of bacterial infection. Moreover, HKBS-P60 was found to be better protected against V. harveyi infection with lower cumulative mortality (60%) compared to HKBS (90%) and HKBS-P0 (100%) at 7 days after infection. Overall, these findings confirmed that P60 could increase immunological responses in the shrimp midgut, and HKBS-P60 could be used as an effective tool to enhance the immune response and disease resistance in shrimp. [ABSTRACT FROM AUTHOR]

The supplementation of cytosine phosphate guanosine-oligodeoxynucleotides (CpG-ODNs) in feeds to enhance the immune responses of crustaceans has been studied. However, the immunomodulatory effects of CpG-ODNs alone on prawn hepatopancreas and midgut remain unclear. Hence, we delivered CpG-ODNs directly into the midgut of Macrobrachium rosenbergii by reverse gavage (RG). In this study, M. rosenbergii was fasted before receiving different doses (0, 2, 4, and 10 μg) of CpG-ODN 2006. Furthermore, the hemolymph, hepatopancreas, and midgut were sampled at 2, 6, 12, and 24 h after RG. The results showed that administration of CpG-ODN 2006 could boost total hemocyte count (THC), transglutaminase (TG), phenoloxidase (PO), lysozyme-like (LYS), and respiratory burst (RB) responses in the hemolymph. In addition, the expression levels of prophenoloxidase (MrProPO), crustin (MrCrs), peroxinectin (MrPxt), and superoxide dismutase (MrSOD) in the hepatopancreas and midgut were also enhanced, in a dose- and time-dependent manner. Overall, our results indicate that administration of 4 μg CpG-ODN 2006 alone by RG elicited the most potent immunological response in the hemolymph, hepatopancreas, and midgut of M. rosenbergii. [ABSTRACT FROM AUTHOR]

This study evaluated the inhibitory effects and adhesion properties of the isolated Bacillus safensis strain VQV8 (abbreviated as VQV8) against pathogenic Vibrio species under different culture conditions. VQV8 was cultured under specific conditions of temperature, pH, and NaCl (%) before testing its antibacterial activities using the dot-spotting method. In addition, a medium from agriculture by-products was screened for replacing Marine broth (MB) medium to culture VQV8. The bacteriostatic activities of VQV8 against pathogenic Vibrio bacteria were also determined. The adhesion properties of this bacterium were evaluated based on three main attributes: hydrophobicity, aggregation, and biofilm formation. The results indicated that VQV8 had the highest antibacterial activities against the tested Vibrio species when cultured in a medium containing 2% NaCl and incubation temperature ranged from 25 to 37 °C. However, the antibacterial activities against V. parahaemolyticus, V. haveyi, and V. alginolyticus were not found at 45 °C. Additionally, the pH from 6 to 9 did not demonstrate a significant effect on the antibacterial activities of VQV8. On the other hand, the use of pineapple peel extract (20–30%) medium demonstrated no significant difference in either growth or antibacterial activities of VQV8 compared to those found in MB medium. These results suggest the potential ability of this bacterial strain to strongly inhibit the growth of all tested Vibrio spp. High adhesion ability was also confirmed through the results of the hydrophobicity, aggregation, and biofilm formation assays. Thus, our results suggest that VQV8 could be used as a potential probiotic candidate for the prevention of Vibrio spp. infections in aquaculture. [ABSTRACT FROM AUTHOR]

The application of herbal medicine is one of the world's most ancient approaches to treating diseases. This study aimed to examine the immunomodulatory effect of Radix Scutellariae water extract (RSWE) on the Macrobrachium rosenbergii immunity by reverse gavage feeding (RGF). In this study, M. rosenbergii was administered RSWE by RGF to stimulate its immunity. Furthermore, total hemocyte count (THC), phenoloxidase (PO), respiratory burst (RB), transglutaminase (TG), and lysozyme activity in hemocytes were evaluated to determine the immunological responses. Meanwhile, midgut and hepatopancreas were isolated to observe immune-related genes (LGPB, PE, and proPO). The study employed four treatments (0, 1, 5, and 10 µg of RWSE), and each treatment consisted of 3 replications. All data were analyzed using SPSS 17.0 (one-way ANOVA and Duncan test) to evaluate the differences among treatments. The results showed that the 5 µg RSWE was the optimum dosage to enhance the immune responses and induce the expression of immune-related genes in the hepatopancreas and midgut of M. rosenbergii. Finally, the RGF method can be used to investigate the immunomodulatory effect of RSWE on prawn, and the immune response associated with each dose can be compared precisely. [ABSTRACT FROM AUTHOR]

Oligodeoxynucleotides (ODNs) with unmethylated CpG dinucleotides mimic the immunostimulatory activity of bacterial DNA in vertebrates through the recognition of cellular Toll-like receptor 9 (TLR9). We previously demonstrated that CpG motif, 5′-GACGTT-3′ (1668), and 5′-GTCGTT-3′ (2395) were highly immunostimulatory in cobia. In this study, motifs containing CpA, CpC, and CpT were used to evaluate their effects on cobia TLR9 (RCTLR9) gene expression. RCTLR9 expression increased significantly in response to CpT ODN in comparison with other ODN motifs. The optimal dosage of CpT ODN was 10 µg/fish. Furthermore, when the endosomal acidification was blocked using chloroquine immersion, the CpT ODN-induced expression of RCTLR9 in spleen and liver was inhibited in a dose-dependent manner. Our results demonstrated that ODNs containing motifs other than CpG, such as CpT ODN, can stimulate RCTLR9 expression, which is mediated, at least partially, by the interaction between CpT ODN and RCTLR9. [ABSTRACT FROM AUTHOR]