Chlorella vulgaris algae ameliorates chlorpyrifos toxicity in Nile tilapia with special reference to antioxidant enzymes and Streptococcus agalactiae infection

Molecular Biology Reports (2024) 51:616 https://doi.org/10.1007/s11033-024-09535-0 ORIGINAL ARTICLE Chlorella vulgaris algae ameliorates chlorpyrifos toxicity in Nile tilapia with special reference to antioxidant enzymes and Streptococcus agalactiae infection Walaa S. Tawfeek1 · Amina S. Kassab1 · Eman T. Al-Sokary2 · Mona E. Abass2 · Ahmed H. Sherif1 Received: 9 February 2024 / Accepted: 8 April 2024 © The Author(s) 2024 Abstract Background Chlorpyrifos (CPF) is a widely used pesticide in the production of plant crops. Despite rapid CPF biodegradation, fish were exposed to wastewater containing detectable residues. Recently, medicinal plants and algae were intensively used in aquaculture to replace antibiotics and ameliorate stress impacts. Methods and results An indoor experiment was conducted to evaluate the deleterious impacts of CPF pollution on Nile tilapia health and the potential mitigation role of Chlorella vulgaris algae. Firstly, the median lethal concentration LC50 − 72 h of CPF was determined to be 85.8 µg /L in Nile tilapia (35.6 ± 0.5 g body weight) at a water temperature of 27.5 °C. Secondly, fish were exposed to 10% of LC50 − 72 h for six weeks, and tissue samples were collected and examined every two weeks. Also, Nile tilapia were experimentally infected with Streptococcus agalactiae. Exposed fish were immunosuppressed expressed with a decrease in gene expressions of interleukin (IL) 1β, IL-10, and tumor necrosis factor (TNF)-α. Also, a decline was recorded in glutathione peroxidase (GPx), superoxide dismutase (SOD), and catalase (CAT) gene expression in the head kidney tissue. A high mortality rate (MR) of 100% was recorded in fish exposed to CPF for six weeks and challenged with S. agalactiae. Fish that received dietary C. vulgaris could restore gene expression cytokines and antioxidants compared to the control. After six weeks of CPF exposure, fish suffered from anemia as red blood cell count (RBCs), hemoglobin (Hb), and packed cell volume (PCV) significantly declined along with downregulation of serum total protein (TP), globulin (GLO), and albumin (ALB). Liver enzymes were significantly upregulated in fish exposed to CPF pollution, alanine aminotransferase (ALT) (42.5, 53.3, and 61.7 IU/L) and aspartate aminotransferase (AST) (30.1, 31.2, and 22.8) after 2, 4, and 6 weeks, respectively. On S. agalactiae challenge, high MR was recorded in Nile tilapia exposed to CPF (G3) 60%, 60%, and 100% in week 2, week 4, and week 6, and C. vulgaris provided a relative protection level (RPL) of 0, 14.29, and 20%, respectively. Conclusions It was concluded that CPF pollution induces immunosuppressed status, oxidative stress, and anemic signs in Nile tilapia. In contrast, C. vulgaris at a 50 g/kg fish feed dose could partially ameliorate such withdrawals, restoring normal physiological parameters. Keywords Antioxidant enzymes · Chlorpyrifos · Chlorella vulgaris · Cytokines · Oreochromis niloticus · Streptococcus agalactiae Introduction Ahmed H. Sherif 1 Fish Disease Department, Animal Health Research Institute AHRI, Agriculture Research Center ARC, Dokki, Kafrelsheikh 12619, Egypt 2 Biochemistry, Nutritional Deficiency Diseases and Toxicology Unit, Animal Health Research Institute AHRI, Agriculture Research Center ARC, Kafrelsheikh 12619, Egypt According to the latest FAO report, Nile tilapia (Oreochromis niloticus) is one of the highest-farmed fish species globally; Egypt became among the highest producers, ranking 11th [1]. Pesticides are widely used in plant crop production, and they can reach water streams and aquatic environments [2], allowing increasing bioaccumulation in the tissues of different aquatic animals [3]. Pesticides enter aquatic 13 616 Page 2 of 12 Molecular Biology Reports environments through their extensive agricultural and domestic use, and approximately 64% of agricultural water sheds worldwide are at risk of pesticide pollution [4]. Chlorpyrifos (CPF) [O, O-diethyl-O-(3,5,6-trichloro2-pyridyl) phosphorothioate] is considered one of the widely used organophosphate pesticides owing to the broad-spectrum of eradicating plant pests and eliminating mosquitoes, the released CPF contaminate the aquatic environment and remain for 8–53 days till decomposition [5]. CPF has highly absorbable properties via the gills, skin, and digestive system of aquatic animals; it bioaccumulates in their tissues (liver and kidney), and its residues have been discovered in farmed and wild fishes, hindering their normal metabolic functions and threatening their life, in addition to causing genotoxicity [6, 7]. Pesticide exposure suppresses fish immunity, adversely impacting cytokine gene expressions such as interleukin (IL)-1β, IL-8, and tumor necrosis factor (TNF)-α [8], making them vulnerable to infectious diseases [9]. Also, Pesticide bioaccumulation could generate reactive oxygen species (ROS), injuring different fish tissues, oxidative stress has recently been hypothesized to be the main mode of CPF toxicity. Antioxidant enzymes are released to detoxify generated ROS, such as glutathione-S transferase (GST), superoxide dismutase (SOD), catalase (CAT), and glutathione (GSH) to counteract oxidative damage [10]. The exposure of fish to pesticides is unavoidable, despite legal restrictions on the use of pesticides that can effectively reduce environmental contamination. Also, improving the biological and physiological status of the fish can ameliorate the toxic withdrawals of pesticides [11]. Pollution, antibiotics, and chemotherapy mainly affect the antioxidant status of aquatic animals. In the case of bacterial infection, the mortality rate, clinical signs, and treatment efficacy are impacted by the antioxidant-immune status of diseased fish. So, pollution and bacterial infection could act synergistically to increase the mortality rate and provoke more prominent clinical signs. Some natural products could mitigate the impacts of several toxicants by increasing antioxidant capacity protecting fish tissues [12–19]. Supplementation with dietary lycopene, chlorella, or citric acid could fully or partially mitigate the impacts of environmental toxicants, which could Table 1 List of fish feed ingredients Ingredient % Ingredient Corn 24 MCP Soya (44%) 33 Salt Fish meal 21 Methionine DDGs 4.5 Choline chloride Corn gluten 15 Minerals premix Soya oil 1 Vitamins premix Note DDGs = Dried Distilled Grains 13 % 1 0.15 0.05 0.05 0.1 0.1 enhance the antioxidant capacity in African sharp-tooth catfish (Clarias gariepinus) [20]. Chlorella vulgaris is a freshwater green algae that contains different components: 60% protein and 18 amino acids, fiber, vitamins, and minerals, in addition to bioactive substances such as antioxidants and chlorophylls. Recently, C. vulgaris became one of the most frequently used microalgae in aquatic animal diet formulation. Many studies have assessed the ability of C. vulgaris to improve growth, immune responses, and stress ameliorati (...truncated)