MORINGA LEAF SIMPLICIA ENCHANCED HEALTH STATUS AND IMMUNE RESPONSE OF TILAPIA AGAINST Edwardsiella tarda INFECTION

The present study evaluated the effects of curcumin on growth, immune and antioxidant response in tilapia (Oreochromis niloticus). An optimum dose of curcumin was investigated by feeding four different levels of this compound in combination with three different regimes of stocking density (12 treatments). Fish were reared at three densities; low density (LD = 1.50 kg/m3), medium density (MD = 3.00 kg/m3), and high density (HD = 4.50 kg/m3). Each treatment was fed with four different levels of dietary supplementation of curcumin (C0 = 0 mg/kg, C1 = 50 mg/kg, C2 = 100 mg/kg, and C3 = 150 mg/kg) for 60 days. Each treatment has three replicates (n = 50/replicate in LD, 100/replicate in MD, 150/ replicate in HD). Although better growth was observed in MD, however treatments at all densities fed with C1 diet showed improved growth as compared to other diets. Chemical composition of fish and activity of amylase, lipase and protease in all treatments were noted to be similar. Levels of antioxidant enzymes (catalase, superoxide dismutase and glutathione peroxidase) and cortisol in MD and HD treatments were similar to those in LD treatment. However, fish fed with C1 diet in each density treatment showed the lowest values of antioxidant enzymes. Similarly, the levels of malondialdehyde were noted to be similar in MD and HD treatments as compared to that in LD. Its levels were lower in fish fed with C1 and C3 diets in all density treatments. Expression of pro-opiomelanocortin-α (POMC-α), Somatostatins-1 (SST-1) and Interleukin 1-β (IL-1β) did not increase in MD and HD treatments in response to high stocking density when compared with LD treatment. The lowest levels of these genes were noted in fish fed with C2 and C3 diets in all treatments. In conclusion, supplementation of curcumin in diet of tilapia improved growth and antioxidant response in tilapia. optimum dose of curcumin for tilapia culture is 50 mg/kg at the density of 3.00 kg/m3which might be further investigated for intensive culture.

The study investigated the impact of N-acetyl cysteine on growth, immune response, and antioxidant activity in tilapia (Oreochromis niloticus). Fish were reared at three densities (1.50, 3.00, and 4.50 kg/m3) with four levels of N-acetyl cysteine supplementation (0, 2, 4, and 6 mg/kg) over 60 days. Better growth was observed at low density, but at all densities, fish fed the highest N-acetyl cysteine level (6 mg/kg) showed improved growth. Chemical composition of fish and activity of amylase, lipase and protease in all treatments were noted to be insignificant. The levels of antioxidant enzymes (catalase, superoxide dismutase and glutathione peroxidase) and cortisol in HD treatments were high as compared to LD and MD treatment. However, fish fed with N3 diet in each density treatment showed the lowest level of antioxidant enzymes as well as cortisol. Similarly, the levels of malondialdehyde were noted to be high at HD treatments as compared to that in LD and MD. Its levels were lower in fish fed with N3 diets in all density treatments. Expression of somatostatins-1 did not increase in MD and HD treatments in response to high stocking density when compared with LD treatment. However, pro-opiomelanocortin-α level was reduced after N3 diet in HD treatment and interleukin 1-β expression increased after N3 supplement in HD treatment. In conclusion, N-acetyl cysteine supplementation improved growth and antioxidant response in tilapia. The most optimum dose of N-acetyl cysteine was noted to be 6 mg/kg at high stocking, suggesting the potential role of this nutraceutical in tilapia intensive culture.

Suppression subtractive hybridization coupled with microarray analysis to examine differential expression of genes in Japanese flounder Paralichthys olivaceus leucocytes during Edwardsiella tarda and viral hemorrhagic septicemia virus infection

Carvacrol, a major compound naturally present in oregano and thyme and its precursor, cymene, were tested in vitro for their antimicrobial activity againstEdwardsiella tarda, a fish pathogenic bacteria causing edwardsiellosis. When used alone, carvacrol, but not cymene, inhibited the bacterium with the minimal inhibitory concentration (MIC) of 20 ppm. However, cymene was shown to be able to enhance the inhibitory ability of carvacol indicated by the reduction of MIC to 5 ppm when used with 2.5 ppm of cymene. Based on mortalities in 2 weeks after intraperitoneal E. tarda injection, the median lethal dose (LD50) of E. tarda for tilapia (Oreochromis niloticus) was 5.0 × 102 CFU/g of fish. Fish diets supplemented with carvacrol and cymene were also tested for their protective effect against E. tarda infection in tilapia. The results showed that carvacrol (200 ppm) but not cymene (200 ppm) when fed prophylactically decreased the mortality in E. tarda-challenged tilapia. However, carvacrol at the same concentration could cause no mortality of E. tarda infected fish when used in conjunction with 200 ppm of cymene. Key words: Carvacrol, cymene, Edwardsiella tarda, tilapia.

Identification and functional analysis of dual-specificity phosphatases (DUSP) genes in Japanese flounder (Paralichthys olivaceus) against temperature and Edwardsiella tarda stress

Simple SummaryThe health of farmed marine fish, including the Japanese flounder (Paralichthys olivaceus), is severely compromised by temperature fluctuations and infections such as Edwardsiella tarda, resulting in significant economic losses. A previous study revealed that elevated temperature enhanced E. tarda dissemination in flounder tissues. However, the underlying mechanism has not been fully explained, especially the changes in protein level. This study utilized label-free proteomics to investigate the immune response of flounder to E. tarda infection under low (15 °C) and high (23 °C) temperature conditions. Multiple differentially abundant proteins (DAPs) were identified in each group. GO and KEGG analyses highlighted immune-related pathways and proteins, with key DAPs forming extensive interaction networks. Venn analysis revealed distinct responses: high temperature promoted endocytosis and complement activation, while low temperature increased histone levels and impaired RNA transport. These results enhance our understanding of how water temperature changes affect antibacterial immunity in fish.Fluctuating water temperatures and bacterial pathogens such as Edwardsiella tarda pose a serious threat to mariculture, resulting in significant economic losses within the flounder industry. A previous study revealed that elevated temperature enhanced E. tarda dissemination in flounder tissues. However, the underlying mechanism has not been fully explained, especially the changes in protein level. In this study, label-free proteomics was utilized to investigate the impact of high temperature (23 °C) and low temperature (15 °C) on flounder immune response to E. tarda infection. Our results identified 317 differentially abundant proteins (DAPs) in the low-temperature group (LI-LC) and 302 DAPs in the high-temperature group (HI-HC). GO and KEGG analyses of DAPs revealed numerous immune-related proteins and pathways. Twenty-six key DAPs in the LI-LC group and twenty-seven key DAPs in the HI-HC group were further identified and formed extensive interaction networks, respectively. Through the analysis of key immune-related DAPs that were specifically identified in both groups via Venn diagram analysis, we demonstrated that the endocytosis capacity and complement activity were enhanced in the HI-HC group, while histone abundance and RNA transport function were, respectively, increased and severely interfered with in the LI-LC group. These findings highlight a clear divergence in the immune response of flounder to E. tarda infection between 15 °C and 23 °C, providing valuable insights into how temperature variation influences antibacterial immunity in fish.

The Chinese soft-shelled turtle (Pelodiscus sinensis) is an important aquatic species in southern China that is threatened by many serious diseases. Edwardsiella tarda is one of the highly pathogenic bacteria that cause the white abdominal shell disease. Yet, little is known about the immune and metabolic responses of the Chinese soft-shelled turtle against E. tarda infection. In the paper, gene expression profiles in the turtle liver were obtained to study the immune responses and metabolic regulations induced by E. tarda infection using RNA sequencing. A total of 3908 differentially expressed unigenes between the experimental group and the control group were obtained by transcriptome analysis, among them, were the significantly upregulated unigenes and downregulated unigenes 2065 and 1922, respectively. Further annotation and analysis revealed that the DEGs were mainly enriched in complement and coagulation cascades, phagosome, and steroid hormone biosynthesis pathways, indicating that they were mainly associated with defense mechanisms in the turtle liver against E. tarda four days post infection. For the first time, we reported on the gene profile of anti-E. tarda response in the soft-shelled turtle, and our research might provide valuable data to support further study on anti-E. tarda defense mechanisms in turtles.

Discovery of immunoglobulin T in Nile tilapia (Oreochromis niloticus): A potential molecular marker to understand mucosal immunity in this species

Immunometabolic interplay in Edwardsiella tarda-infected crucian carp (Carassius auratus) and in vitro identification of the antimicrobial activity of apolipoprotein D (ApoD) by utilization of multiomics analyses

Background: Edwardsiella tarda has been report as etiology of gastroenteritis in both human and veterinary medicine, usually associated with exposure to aquatic environments in immunocompromised individuals. The present report describes a case of typhlocolitis in a cow in the west region of Santa Catarina state, Brazil. Case: After clinically evaluation of animal and proceeding euthanasia and realized the necropsy. Tissue samples were collected, and routinely processed for histological examination. Histopathological lesions were described as mild, moderate and severe. Tissue and swabs samples of small and large intestine were sent to standard microbiological culture processing. At necropsy, cattle presented severe dehydration and emaciation. Eye and vaginal mucosa were severely congested. The opening of the abdominal cavity revealed a great amount of greenish fluid and a large amount of fecal material, associated with diffuse severe peritonitis, evidenced by hyperemia and severe deposition of fibrin in the visceral and parietal peritoneal surface. In the serosa of the cecum, there were two points of rupture, observed in the proximal colon. The mucosa of cecum and colon were severely edematous, hyperemic, and presented diffusely distributed pinpoint round hemorrhages, as well as fibrinonecrotic material adhered to the surface. Histologically, in the mucosa of cecum and colon, moderate to severe diffuse inflammatory infiltrate of neutrophils, lymphocytes and plasma cells associated with multifocal severe necrosis were observed. Moderate diffuse fibrin deposition was evidenced in the submucosa and muscular, as well as multifocal moderate necrosis in the muscular layer. In the serous, severe diffuse inflammatory infiltrate of neutrophils associated with fibrin deposition and innumerous coccoid bacterial colonies were observed. The samples subjected to bacterial isolation showed growth of Edwardsiella tarda. All samples were negative for Salmonella spp. and Yersinia spp.Discussion: The final diagnosis was established through the association of clinical history, clinical signs, gross and histopathological lesions, as well as, bacterial isolation of the etiological agent, Edwardsiella tarda. In this case, it is conjectured that the reservoirs which cattle had access represented the source of infection. The fact that the animal was in the immediate postpartum period may have predisposed to the development of clinical disease due to immunosuppression. In domestic animals, Edwardsiella tarda has been reported in swine, and as a cause of septicemia in calves. Clinically, intestinal manifestations observed in edwardsiellosis in cattle are indistinguishable from several other conditions that cause diarrhea, such as infectious, nutritional or parasitic diseases. The main differential diagnoses are salmonellosis and yersiniosis due to the similarities regarding to gross and histopathological lesions in these cases compared to cases of edwardsiellosis. Salmonellosis is characterized by grey to yellowish, fetid diarrhea in which blood and mucus are oftentimes observed. At necropsy, catarrhal, hemorrhagic or fibrinous enteritis may be evidenced. The lesions initially are seen in the ileum. However, in the chronic stages of infection, foci of necrosis and ulceration may be noted mainly in the cecum and colon. Histologically, a fibrin layer associated with necrosis and mucosal ulceration can be observed in the small intestine and initial portion of large intestine. Inflammatory infiltrate composed predominantly by neutrophils, as well as fibrin thrombi in capillaries and venules are also observed. Lesions observed are similar that described in ulcerative colitis by E. tarda in human patients. In conclusion, Edwardsiella tarda can lead to a fatal typhlocolitis in cattle, being an important differential diagnosis in cases of acute diarrhea.Arya A.V., Rostom A., Dong W.F. & Flynn A.N. 2011. Crohn’s Disease Exacerbation Induced by Edwardsiella tarda Gastroenteritis. Gastroenterology. 5(3): 623-627.Engel J.J. & Martin T.L. 2006. Edwardsiella tarda as a cause of postdysenteric ulcerative colitis. International Journal Colarectal Disease. 21(2): 184-185.Ewing W.H., McWhorter A.C., Escobar M.R. & Lubin A.H. 1965. Edwardsiella, a new genus of enterobacteriaceae based on a new species, E. tarda. International Journal of Systematic and Evolutionary Microbiology. 15(1): 33-38.Gelberg H.B. 2013.Sistema Alimentar, Peritônio, Omento Mesentério e Cavidade Peritonial. In: McGavin M.D. & Zachary F.M. (Eds). Bases da patologia em veterinária. 2.ed. Rio de Janeiro: Elsevier, pp.378-382.Janda J.M. & Abbott S.L. 1993.Infections Associated with the Genus Edwardsiella: the role of Edwardsiella tarda in human disease. Clinical Infectious Diseases. 17(4): 742-748.Leung K.Y., Siame B.A., Tenkink B.J., Noort R.J. & Mok Y.K. 2012. Edwardsiella tarda – Virulence mechanisms of an emerging gastroenteritis pathogen. Microbes and Infection. 14(1): 26-34. Litton K.M. & Rogers B.A. 2016. Edwardsiella tarda Endocarditis Confirmed by Indium-111 White Blood Cell Scan: An Unusual Pathogen and Diagnostic Modality. Case Reports in Infectious Diseases. 2016:1-3. Magalhães H., Freitas M.A., Santos J.A. & Costa C.H.C. 1984. Septicemia por Edwardsiella tarda, em bezerro. Pesquisa Agropecuária Brasileira. 19(3): 367-370.Hirai Y., Ashata-Tago S., Ainoda Y., Fujita T. & Kikuchi K. 2015.Edwardsiella tarda bacteremia. A rare but fatal water – and foodborne infection: Review of the literature and clinical cases from a single centre. The Canadian Journal of Infectious Diseases & Medical Microbiology. 26(6): 313-318. Mikamo H., Ninomiya M., Sawamura H. & Tamaya T. 2003. Puerperal intrauterine infection caused by Edwardsiella tarda. Journal of Infection and Chemotherapy. 9(4): 341-343.Mohanti B.R. & Sahoo P.K. 2007. Edwardsiellosis in fish: a brief review. Journal of biosciences. 32(7): 1331-1344. Owens D.R., Nelson S.L. & Addinon J.B. 1974. Isolation of Edwardsiella tarda from Swine. Appllied microbiology. 27(4): 703-705.Park S.B., Aokil T. & Jung T.S. 2012. Pathogenesis of and strategies for preventing Edwardsiella tarda infection in fish. Veterinary Research. 43(1): 67.Riet-Correa F., Schild A.L., Méndez M.D.C. & Lemos R.A.A. 2007.Doenças de Ruminantes e Equídeos. 3.ed. Santa Maria: Pallotti, 998p. Slaven E.M., Lopez F.A., Hart S.M. & Sanders C.V. 2001. Myonecrosis Caused by Edwardsiella tarda: A Case Report and Case Series of Extraintestinal E. tarda Infections. Clinical Infectious Diseases. 32(10): 1430-1433.Tamada T., Koganemaru H., Mastsumoto K. & Hitomi S. 2009.Urosepsis caused by Edwardsiella tarda Journal of infection and chemotherapy. 15(3): 191-194.Thune R.L., Stanley L.A. & Cooper R.K. 1993. Pathogenesis of gram-negative bacterial infections in warm water fish. Annual Reviem of Fish Diseases. 3: 37-68.Uzal F.A., Plattiner B.L. & Hostetter J.M. 2015. Alimentary system. In: Maxie M.G. (Ed). Jubb, Kennedy, and Palmer’s Pathology of domestic animals.v.2. 6th edn. Saint Louis: Elsevier, pp.167-177. Wang I.K., Kuo H.L., Chen Y.M., Lin C.L., Chang H.Y., Chuang F.R. & Lee M.H. 2005. 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Immune response in the tilapia, Oreochromis mossambicus on exposure to tannery effluent

Protective effects and mechanisms of a probiotic bacterium Lactobacillus rhamnosus against experimental Edwardsiella tarda infection in tilapia ( Oreochromis niloticus)

As a quintessential marine teleost, Paralichthys olivaceus demonstrates vulnerability to a range of pathogens. Long-term infection with Edwardsiella tarda significantly inhibits fish growth and even induces death. Gills, blood, and kidneys, pivotal components of the immune system in teleosts, elicit vital regulatory roles in immune response processes including immune cell differentiation, diseased cell clearance, and other immunity-related mechanisms. This study entailed infecting P. olivaceus with E. tarda for 48 h and examining transcriptome data from the three components at 0, 8, and 48 h post-infection employing weighted gene co-expression network analysis (WGCNA) and protein-protein interaction (PPI) network analysis. Network analyses revealed a series of immune response processes after infection and identified multiple key modules and key, core, and hub genes including xpo1, src, tlr13, stat1, and mefv. By innovatively amalgamating WGCNA and PPI network methodologies, our investigation facilitated an in-depth examination of immune response mechanisms within three significant P. olivaceus components post-E. tarda infection. Our results provided valuable genetic resources for understanding immunity in P. olivaceus immune-related components and assisted us in further exploring the molecular mechanisms of E. tarda infection in teleosts.

Large-scale disease outbreaks that caused by Streptococcus agalactiae (SA) infection have dramatically affected tilapia production worldwide. Patchouli oil (PO) is a traditional Chinese herbal medicine in treating gastrointestinal symptoms and is considered to be a safe and efficient candidate to treat SA infection in tilapia. In this study, we first investigated the changes of intestinal microbiota and host responses in GIFT tilapia after a 28-day dietary supplementation of different concentrations of PO (basic diet, 5 μL PO/g diet, or 15 μL PO/g diet) and followed by SA infection using an integrative analysis of transcriptome and 16 S amplicon profiling. We observed that dietary supplementation of 5 μL PO/g diet improved food intake and growth in tilapia. Histopathological observation confirmed there were increased surface area of intestinal villi and generally decreased ROS as revealed by the MAD, GSH-PX, and T-SOD activities in the tilapia intestines in response to dietary PO supplementation. The intestinal microbial diversities and community structures were significantly affected, and the colonization of probiotics (e.g., Cetobacterium and clostridium_ sensu_stricto_1) in the mucosa was improved in the dietary PO groups. The relative abundance of Streptococcus (0.033%) in the 15 μL PO/g dietary group was much lower than that in the control group (basic diet; 0.86%). A large number of DEGs were enriched in the immune-related pathways, e.g., lysosomal lumen, MHC protein complex, and toll-like receptor 2 signaling pathway. Our findings suggested that dietary supplementation of PO enhanced intestinal physical barriers, increased the colonization of probiotic bacteria and improved intestinal immunity in tilapia. This provides novel knowledge on PO as a dietary supplement to improve intestinal health against SA invasion in tilapia.

Effects on hematological parameters, antioxidant and immune responses, AChE, and stress indicators of olive flounders, Paralichthys olivaceus, raised in bio-floc and seawater challenged by Edwardsiella tarda