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مقاله علمی – پژوهشی:‌ تاثیر کود آمینو اسید تهیه شده از ضایعات کپور معمولی (Cyprinus carpio) بر شاخص‌‌های رشد و ترکیبات گیاه گوجه‌‌فرنگی (Solanum lycopersicum) در شرایط گلخانه‌‌ای Effect of amino acid fertilizer derived from common carp (Cyprinus carpio) waste on growth indices and quality attributes of tomato (Solanum lycopersicum) under greenhouse conditions 1 1 پژوهش حاضر، با هدف استفاده بهینه از ضایعات ماهی کپور معمولی (Cyprinus carpio) و تبدیل آن به محصولی ارزشمند در کشاورزی، به بررسی اثر کود آمینواسید حاصل از هیدرولیز آنزیمی بر شاخص‌های رشد و کیفیت گوجه‌فرنگی (Solanum lycopersicum) در شرایط گلخانه‌ای پرداخته است. ضایعات ماهی پس از فرآیند هیدرولیز آنزیمی با آنزیم‌های آلکالاز و فلاورزایم به پپتیدها و اسیدهای آمینه کوچکتر تبدیل شدند. محصول حاصله از نظر تجزیه‌‌وتحلیل کیفی شامل پپتیدهای محلول، الگوی وزن مولکولی، درجه هیدرولیز، محتوای اسیدهای آمینه کل و آزاد، ترکیبات معدنی اصلی و ریزمغذی‌ها، ویژگی‌های فیزیکی و شاخص‌های میکروبی عمومی و اندیکاتور بررسی شد. سپس کود تولیدی بر پایه هیدرولیز ضایعات کپور معمولی فرموله گردید و پایداری آن طی سه ماه نگهداری مورد ارزیابی قرار گرفت. آزمایش گلخانه‌ای در قالب طرح کاملاً تصادفی با پنج تیمار شامل تیمار شاهد، کود آمینواسید پایه (E)، کود غنی‌شده با پتاسیم و فسفر (EKP)، کود تجاری داخلی (I) و کود وارداتی (Ex)، هر کدام با پنج تکرار انجام شد. شاخص‌های مورفوفیزیولوژیک (ارتفاع بوته، تعداد برگ و وزن خشک اندام‌ها) و شاخص‌های کیفی میوه شامل کلروفیل برگ، ماده جامد محلول (TSS)، لیکوپن و ویتامین C اندازه‌گیری گردید. نتایج نشان داد، کود آمینواسید کپور معمولی دارای محتوای اسیدهای آمینه کل 87/13 درصد و آزاد 67/7 درصد بود و پایداری مناسبی در دوره نگهداری داشت. تیمار EKP بیشترین اثر مثبت را بر رشد گیاه، افزایش وزن و تعداد میوه و بهبود ویژگی‌های کیفی محصول نشان داد و از نظر آماری بر سایر تیمارها برتری داشت. همچنین مقایسه با کودهای تجاری بیانگر کارایی بالاتر کود تولیدی بود. استفاده از کود آمینواسید تولیدی از ضایعات ماهی کپور معمولی، علاوه بر بهبود چشمگیر شاخص‌های رشد و کیفیت گوجه‌فرنگی، با توجه به هزینه تولید پایین و بازگشت سریع سرمایه، می‌تواند به عنوان یک راهکار دانش‌بنیان و پایدار در صنعت کشاورزی معرفی گردد. پژوهش حاضر، گامی مؤثر در عرصه اقتصاد چرخشی و ارزش‌آفرینی از پسماندهای صنایع شیلاتی محسوب می‌شود. 2 Introduction Amino acid fertilizers are recognized as effective bio-stimulants that contain both free and bound amino acids, playing vital roles in enhancing plant growth, development, and stress resilience (Rouphael and Colla, 2020). These compounds serve as fundamental components of proteins, supplying essential nitrogen and carbon, while also boosting metabolic processes and improving crop quality (Garcia and Perez, 2021). Common sources of amino acid fertilizers include plant, microbial, and animal proteins, which undergo hydrolysis to release peptides and amino acids that plants can readily absorb (Chen et al., 2019). Tomato, a major greenhouse crop, has shown significant improvements in chlorophyll content, vegetative growth, yield, and quality when treated with amino acid fertilizers (Gasana and Kim, 2020).  Specific amino acids such as lysine, arginine, and proline are known to help plants withstand environmental stresses like drought, salinity, and temperature variations (Garcia and Perez, 2021). A sustainable and innovative source for these fertilizers is fish processing waste, particularly from common carp, which is rich in proteins. Repurposing these by-products not only reduces environmental impact but also creates valuable agricultural inputs. Previous research confirms that fish waste-derived amino acid fertilizers can enhance plant growth, increase fruit yield, and improve biochemical traits such as protein and chlorophyll levels (Chen et al., 2019). Given the rising demand for sustainable agriculture, this study explores the potential of amino acid fertilizers made from carp waste to support eco-friendly tomato production (Rouphael and Colla, 2020). Methodology The research was carried out at the Caspian Sea Ecology Research Center in Sari, Iran. Common carp waste including skin, viscera, heads, and fins was collected from local markets, frozen, and processed via enzymatic hydrolysis using Alcalase and Flavourzyme (Safari et al., 2016; Safari et al., 2018; Safari et al., 2020). The process included pasteurization, enzyme treatment under controlled pH and temperature, and heat inactivation. The resulting hydrolysate was then centrifuged, filtered, and stored for further analysis. Chemical and physicochemical properties such as total protein, soluble protein, peptide size, degree of hydrolysis, ash, fat, moisture, nitrogen, phosphorus, potassium, pH, salinity, and amino acid composition were evaluated using standard methods (Hesse, 1971; Adler-Nissen and Olsen, 1979; James, 1995; Moore, 2004; AOAC, 2005; ISIRI, 2008a, b, c; APHA, 2017).  Microbiological safety was verified through tests for aerobic bacteria, yeast, mold, coliforms, E. coli, and Salmonella (Sallam, 2007; ISIRI 8923-1, 2007; ISO 21527-1, 2008; Hernández et al., 2009; ISIRI 11166, 2010; ISIRI 10889-3, 2013; ISO 4833-1, 2013; ISIRI 13321-2, 2014). A greenhouse trial using a completely randomized design tested five treatments: control, enzyme hydrolysate, enzyme hydrolysate enriched with phosphorus and potassium, a domestic commercial fertilizer, and an imported commercial fertilizer. Tomato seedlings in pots received weekly foliar applications, and parameters related to growth and quality such as plant height, leaf count, leaf area, fruit yield, chlorophyll, TSS, lycopene, and vitamin C were measured (Abbasi et al., 2003; Boras et al., 2011). Data were analyzed using ANOVA and Duncan’s test. Results The hydrolysis process significantly increased total protein content from 15.17% in raw waste to 76.61% in the final product. Fat and moisture decreased, while ash and organic nitrogen levels rose. The hydrolysate had a hydrolysis degree of 21.11%, soluble protein of 26.87 mg/mL, and contained 13.87% total amino acids and 7.67% free amino acids, including essential types like lysine and leucine. It exhibited desirable physical traits, was water-soluble, and met safety standards with no pathogenic contamination. Storage studies confirmed the product’s stability over three months. In the greenhouse, the enriched hydrolysate treatment led to the best results in plant height, leaf number, leaf area, fruit count, and fruit weight. It also outperformed other treatments in chlorophyll levels, TSS, lycopene, and vitamin C content. Discussion and conclusion Enzymatic hydrolysis successfully transformed carp waste into a plant-friendly fertilizer rich in bioavailable peptides and amino acids (He et al., 2013; Safari et al., 2016; Safari et al., 2018; Safari et al., 2020). The resulting product was safe, stable, and effective as a foliar spray (FAO, 2010; López-Caballero et al., 2010; Ortizo et al., 2020). Key amino acids supported chlorophyll formation and nitrogen metabolism, leading to better growth and fruit quality (Colla et al., 2015; Rouphael et al., 2017; Zhang et al., 2018). The enriched hydrolysate treatment demonstrated synergistic benefits, enhancing both yield and nutritional quality of tomatoes (Colla et al., 2014; Bulgari et al., 2015; Guo et al., 2018; Zhang et al., 2018). This method offers a sustainable way to upcycle fish waste into a high-value agricultural input (Kumar and Singh, 2019; Martinez and Smith, 2020; Ciepiela and Kamińska, 2021). Further optimization and testing on other crops could broaden the application of this biofertilizer (Zhou et al., 2019; He et al., 2020). In summary, common carp waste-derived amino acid fertilizer presents an eco-friendly and efficient alternative to synthetic fertilizers in greenhouse tomato cultivation. Conflict of interest The authors declare no conflict of interest. Acknowledgment The authors extend their gratitude to the Caspian Sea Ecology Research Center for providing the necessary facilities and support. 1 11 2025/10/5 1404/7/13 2026/04/30 1405/2/10 رضا صفری reza safari موسسه تحقیقات علوم شیلاتی کشور، ، سازمان تحقیقات، آموزش و ترویج کشاورزی safari1351@gmail.com زهرا یعقوب زاده zahra yaghoubzadeh موسسه تحقیقات علوم شیلاتی کشور، ، سازمان تحقیقات، آموزش و ترویج کشاورزی za_yaghoub@yahoo.com فائزه ترک پهنابی Faezeh Turk Pahnabi موسسه تحقیقات علوم شیلاتی کشور، ، سازمان تحقیقات، آموزش و ترویج کشاورزی pahnabifaezeh@gmail.com Amino acid fertilizer common carp waste tomato enzymatic hydrolysis کود آمینواسید ضایعات کپور معمولی گوجه‌فرنگی هیدرولیز آنزیمی Abbasi, P.A., Cuppels, D.A., Lazarovits, G., 2003. Effect of foliar applications of neem oil and fish emulsion on bacterial spot and yield of tomatoes and peppers. Canadian Journal of Plant Pathology, 25(1), pp.41–48. https://doi.org/10.1080/07060660309507048.##Adler-Nissen, J., Olsen, H.S., 1979. The influence of peptide chain length on taste and functional properties of enzymatically modified soy protein. Journal of Food Science, 44(2), pp.456–462. https://doi.org/10.1021/bk-1979-0092.ch007.##AOAC, 2005. Official Method of Analysis. 17th ed. Washington DC: Association of Official Analytical Chemists.##APHA, 2017. Standard Methods for the Examination of Water and Wastewater. 17th ed. Washington DC: American Public Health Association.##Araújo, E.F., dos Santos, V.L., de Almeida, M.V., 2015. Microbiological safety of fish protein hydrolysates produced in Brazil. Food Control, 50, pp.832–838. https://doi.org/10.1016/j.foodcont.2014.09.023.##Boran, M., Karaçal, S., 2021. Fish protein hydrolysates as a source of bioactive peptides for food and agriculture. Food Chemistry, 357, 129805. https://doi.org/10.1016/j.foodchem.2021.129805.##Boras, M., Zidan, R., Halloum, W., 2011. Effect of amino acids on growth, production and quality of tomato in plastic greenhouse. 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Frontiers in Plant Science, 5, p.448. https://doi.org/10.3389/fpls.2014.00448.##FAO, 2010. Guidelines for the safe use of wastewater, excreta and greywater. FAO Water Reports 35. Available at: http://www.fao.org/3/i1954e/i1954e.pdf.##Garcia, M. and Perez, L., 2021. The role of fish waste-derived amino acids in mitigating abiotic stress in crops. Environmental Agriculture, 7(4), pp.200–208.##Gasana, Q.G. and Kim, D.H., 2020. Effect of foliar spraying mixed with fish amino acids (FAA) and oriental herbal nutrient (OHN) extract on growth, yield and quality of watermelon (Citrullus lanatus). Rwanda Journal of Agricultural Sciences, 2(1), pp.127–136.##Guo, W., Zhang, H., Sun, C. & Xie, C., 2018. Impact of fish protein hydrolysates on plant growth and antioxidant activity in tomato. Journal of Plant Nutrition, 41(3), pp.346–355. https://doi.org/10.1080/01904167.2017.1357894.##He, S., Zhang, X. & Xu, X., 2013. Degree of hydrolysis and molecular weight distribution of fish protein hydrolysate and its effects on plant growth. Journal of Agricultural and Food Chemistry, 61(18), pp.4531–4538.##He, S., Zhang, Y., Liu, H., Wang, L. & Liu, Z., 2020. Optimization of enzymatic hydrolysis of fish processing by-products and characterization of protein hydrolysates. Food Chemistry, 312, 126033. https://doi.org/10.1016/j.foodchem.2019.126033. ##Hernández, F., Sgorbati, S. & Chiesa, L.M., 2019. Effect of fish hydrolysate and amino acid fertilizer on tomato yield and quality. Scientia Horticulturae, 246, pp.99–105.##Hesse, P. R 1971. A Text Book of Soil Chemical Analysis. John Nurray Williams Clowes and sons Ltd. London 324pp.##James, C.S. 1995. Analytical Chemistry of Foods, Springer, New York, doi:10.1007/978-1-4615-2165-5. https://doi.org/10.1007/978-1-4615-2165-5. ##Kim, S.-K. and Mendis, E., 2006. Bioactive compounds from marine processing byproducts – A review. Food Research International, 39(4), pp.383–393. https://doi.org/10.1016/j.foodres.2005.10.010.##Kumar, A. and Singh, P., 2019. Use of fish protein hydrolysate in organic tomato cultivation. Journal of Organic Agriculture, 9(1), pp.15–22.##López-Caballero, M.E., Gómez-Guillén, M.C. and Montero, P., 2010. Antioxidant activity of fish protein hydrolysates: Application to tomato growth. Food Chemistry, 123(1), pp.33–40.##Martinez, L., and Smith, D. (2020). Sustainable Agriculture Using Fish Waste Amino Acids. Journal of Organic Agriculture, 12(2), 98-105.##Moore, J. 2004. Amino acid analysis of hydrolysates (feed, fxal, etc), Michign State University, Department of Animal Sciences, Nathalie Trottiers Laboratory.##Rouphael, Y. and Colla, G., 2020. Toward a sustainable agriculture through plant biostimulants: From experimental data to practical applications. Agriculture, 10(10), p.146. https://doi.org/10.3390/agriculture1010146.##Rouphael, Y., Colla, G., Giordano, M., El-Nakhel, C., Kyriacou, M.C. and De Pascale, S., 2017. Foliar applications of a legume-derived protein hydrolysate elicit dose-dependent increases of growth, leaf mineral composition, yield and fruit quality in two greenhouse tomato cultivars. Scientia horticulturae, 226, pp.353-360. https://doi.org/10.1016/j.scienta.2017.09.007.##Safari, S., Ghorbani, R., Motamedzadegan, A. and Hosseini, S.V., 2016. Optimization of enzymatic hydrolysis conditions for rainbow trout waste. Journal of Aquatic Food Product Technology, 25(5), pp.667–678.##Safari, S., Motamedzadegan, A. and Hosseini, S.V., 2020. Shelf-life and nutrient stability of fish-based amino acid fertilizers. Journal of Food Processing and Preservation, 44(3), e14321.##Safari, S., Motamedzadegan, A., Ghorbani, R. and Hosseini, S.V., 2018. 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مقاله علمی – پژوهشی:‌ افزودن نانوکپسول‌‌های حامل و فرم آزاد پپتیدهای زیست‌‌فعال حاصل از هیدرولیز آنزیمی فانوس‌ماهی (Benthosema pterotum) به فرمولاسیون دوغ و ارزیابی خصوصیات میکروبی، شیمیایی و حسی محصول Adding carrier nanocapsules and free form of bioactive peptides derived from enzymatic hydrolysis of lanternfish (Benthosema pterotum) to doogh formulation and evaluating the microbial, chemical and sensory properties of the product 1 1 عوارض خطرناک نگهدارنده‌‌های شیمیایی برای سلامت مصرف‌‌کنندگان، حدود دو دهه است که توجه محققین را به سمت استخراج و تولید ترکیبات طبیعی قابل جایگزین با نگهدارنده‌‌های شیمیایی معطوف کرده است. از مهم‌‌ترین ترکیبات با منشاء طبیعی که فعالیت ضد باکتریایی و آنتی‌‌اکسیدانی آنها در تحقیقات مختلف تائید شده است، می‌‌توان به پروتئین‌‌های هیدرولیزشده یا پپتیدهای زیست‌‌فعال اشاره کرد. در تحقیق حاضر، پپتیدهایی با وزن مولکولی کمتر از 3 کیلودالتون از فانوس ماهی (Benthosema pterotum) به روش آنزیمی جداسازی و سپس با کمک تکنیک خشک‌‌کردن انجمادی و پوشش ترکیبی مالتودکسترین-صمغ عربی (با نسبت برابر) نانوریزپوشانی شدند. سپس نانوکپسول‌‌های حامل (200 و 400 میکروگرم بر میلی‌‌لیتر) و پپتیدهای آزاد (400 و 500 میکروگرم بر میلی‌‌لیتر) به فرمولاسیون دوغ اضافه و خصوصیات کیفی محصول طی 60 روز نگهداری در دمای یخچال ارزیابی گردید. نتایج نشان داد که تیمارهای حاوی نانوکپسول‌‌های حامل نسبت به شاهد و تیمارهای حاوی پپتیدهای آزاد، در زمینه مهار رشد Staphylococcus aureus  و Escherichia coli و حذف رادیکال آزاد ABTS موفق‌‌تر عمل کردند (05/0p<). ضمن این‌که pH، اسیدیته و فعالیت حذف رادیکال این تیمارها برخلاف شاهد و تیمارهای حاوی پپتیدهای آزاد، طی دوره نگهداری فاقد تغییر معنی‌‌داری بودند (05/0<p). با افزایش غلظت پپتیدهای آزاد و نانوکپسول‌‌های حامل، شمارش باکتری‌‌ها و قدرت مهار رادیکال ABTS در تیمارها به‌ترتیب کاهش و افزایش یافتند (05/0p<). اما pH و اسیدیته تحت تأثیر قرار نگرفتند. ارزیابی رنگ، بو و مزه تیمارها نشان داد که فقط امتیازات تیمار حاوی 400 میکروگرم بر میلی‌‌لیتر نانوکپسول‌‌های حامل در روز اول و 60 برابر بود (05/0<p) اما در سایر تیمارها، امتیازات شاخص‌‌های حسی در طول دوره نگهداری به صورت معنی‌‌داری کاهش یافتند (05/0p<). یافته‌‌های این پژوهش نشان داد که نانوکپسولاسیون پپتیدهای زیست‌‌فعال با استفاده از روش خشک‌‌کردن انجمادی و پوشش ترکیبی مالتودکسترین-صمغ عربی، تکنیک موثری به منظور ارتقاء و پایداری خواص نگهدارندگی آنها برای استفاده در صنعت لبنیات (به‌ویژه دوغ) است. 2 Introduction The chemical types of antioxidants and antimicrobial compounds used in food industries as preservatives causes some diseases in consumers due to their harmful effects (Reyhani et al., 2024). For this reason, over the last two decades, various researches have been conducted to produce and extract preservative compounds of natural origin, and have yielded acceptable results. Hydrolyzed proteins or bioactive peptides that one of the sources of their production are aquatic animals (with low marketability) and the wastes from their processing, are antioxidant (Ng and Wong, 2024) and antimicrobial (Rivera-Pérez et al., 2023) compounds of natural origin. The use of bioactive peptides in food formulations as preservatives always requires the use of a suitable protection system in harsh environmental conditions. Because the structure of peptides may be destroyed in thermal processes during food production. One of the appropriate protection systems for active compounds is encapsulation, which is usually done by spray drying and freeze drying methods (Sarabandi et al., 2020). During this process, a microcapsule or nanocapsule is created around the bioactive peptides, which protects these compounds from possible changes in adverse conditions. Meanwhile, the capsules formed around the peptide in this technique release their contents at a controlled rate and under special conditions (Rao et al., 2016). Doogh is one of the favorite drinks of Iranians, which is produced from the lactic fermentation of milk. Due to its low pH and rich in nutrients, this product suffers from bacterial contamination, especially at ambient temperature, which ultimately causes spoilage and changes in its aroma during storage (Karimi et al., 2022). This is an important challenge in the milk industry. On the other hand, due to the disadvantages of chemical preservatives, it is very important to use natural preservatives that can solve this problem. The aim of the present research in the first stage is the nanoencapsulation of bioactive peptides obtained from lanternfish (Benthosema pterotum) by freeze drying method and maltodextrin-gum Arabic combined coating. In the next step, carrier nanocapsules and free peptides are used in doogh formulation and the microbial, chemical and sensory properties of the product are evaluated during 60 days of storage at refrigerator temperature. Methodology Lanternfish (B. pterotum) were hydrolyzed using Protamex enzyme under optimal conditions (50ºC, Amount of enzyme: 30 Anson units, 90 min) and then by ultrafiltration, peptides with a molecular weight of less than 3 kilodaltons were isolated. Then these peptides were encapsulated using the freeze drying method and combined maltodextrin-gum Arabic coating. In the next step, carrier nanocapsules (200 and 400 µg/ml) and free peptides (400 and 500 µg/ml) were added to the doogh formulation. Finally, Microbial (Staphylococcus aureus and Escherichia coli count), antioxidant (ABTS radical scavenging activity), chemical (pH and acidity) and sensory (color, aroma and taste) properties of the product were evaluated during 60 days of storage at refrigerator temperature (4±1°C). The data were analyzed by one-way analysis of variance (One-Way ANOVA in SPSS22 software) and the difference between the means was evaluated by Duncan's test at 95% confidence level (p<0.05). Result The results showed that the treatments containing nanocapsules carrying bioactive peptides (200 and 400 µg/ml) were more successful in inhibiting the growth and proliferation of S. aureus and E. coli bacteria as well as removing the ABTS free radical during the storage period compared to the control and the treatments containing free peptides (400 and 500 µg/ml) (p<0.05). The number of S. aureus bacteria in treatments containing 400 and 500 µg/ml of bioactive peptides as well as treatments containing 200 and 400 µg/ml of carrier nanocapsules on the first day of storage was 3.26 ± 0.08, 2.25±0.12, 2.24±0.09 and 1.11±0.06 Log CFU/mL, respectively. These numbers ​​for Escherichia coli were recorded as 4.42±0.09, 3.21±0.17, 3.23±0.11, and 1.94±0.08 Log CFU/mL, respectively. The ABTS free radical scavenging rate in these treatments was measured on the same day as 29.73±1.11, 35.52±1.72, 39.65±2.12, and 47.14±1.18 Unit/ml, respectively. By increasing the concentration of free peptides (from 400 to 500 µg/ml) and carrier nanocapsules (from 200 to 400 µg/ml), as well as the storage time (1, 20, 40 and 60 days), the number of bacteria under study in the treatments decreased significantly (p<0.05). pH, acidity and ABTS radical scavenging power of treatments containing 200 and 400 µg/ml carrier nanocapsules did not change significantly during the storage period (p>0.05) and had a constant trend. While in treatments containing free peptides and control, antioxidant activity and pH decreased (p<0.05), but acidity increased by increasing storage time (p<0.05). The pH values ​​for the control, treatments containing 400 and 500 μg/mL of bioactive peptides, as well as 200 and 400 μg/mL of carrier nanocapsules on day 60 of storage were measured as 3.03±0.02, 3.55±0.01, 3.56±0.05, 4.74±0.04, and 4.75±0.09, respectively. These values ​​for acidity were recorded as 0.85±0.03, 0.67±0.02, 0.68±0.01, 0.36±0.07, and 0.37±0.03, respectively. By increasing the concentration of free peptides (from 400 to 500 µg/ml) and carrier nanocapsules (from 200 to 400 µg/ml) in doogh, the antioxidant activity (ABTS free radical scavenging power) of the treatments increased significantly (p<0.05), but the pH and acidity values ​​of them did not change (p>0.05). The evaluation of sensory indicators (color, smell and taste) of doogh samples showed that only the treatment containing 400 µg/ml of carrier nanocapsules on the first and 60 day earned equal points (p>0.05), but in other treatments, the sensory indicators during the storage period were significantly have decreased (p<0.05). Of course, the treatment containing 200µg/ml also had equal points in two color and smell indices on the first day and 60, but the score of the taste index in this treatment decreased on the 60th day compared to the first day. The result of examining the sensory indicators of different treatments compared to the control on the first day showed that adding bioactive peptides and carrier nanocapsules to the doogh formulation had no significant effect (positive or negative) on the color, smell and taste of the product. Discussion and conclusion The reduction in the number of S. aureus and E. coli bacteria in treatments with the addition of bioactive peptides and carrier nanocapsules indicates the ability of the aforementioned compounds to combat the growth and proliferation of the studied bacteria That in this regard and also ABTS free radica scavenging, nanocapsules carrying bioactive peptides were more successful and stronger. This finding has been recorded due to the higher active surface area of ​​the nanocapsules, as well as the capsule's (maltodextrin-gum Arabic) ability to protect the core (peptides) structure in adverse environmental conditions and its gradual release. Two mechanisms of peptides antibacterial activity are: A- Peptides are attached to the membrane of bacterial cells and create irreversible pores on the surface of the membrane, as a result of which the cytoplasm flows out of the cell and the cell is completely destroyed. Bioactive Peptides pass through the cell membrane and bind to intracellular organelles such as ribosomes and genomes and disrupt the normal growth process of the cell (Amissah, 2012). In the present research prove that the sensitivity of S. aureus, which is a Gram-positive bacteria, to free peptides and carrier nanocapsules is higher than E. coli. This higher sensitivity is related to the absence of the lipopolysaccharide layer in the cell wall of gram-positive bacteria (Reyhani Poul and Yeganeh, 2023). The results of the evaluation of pH, acidity and sensory properties of treatments during 60 days of storage showed that only in two treatments containing carrier nanocapsules, this indexes remained constant. But in the control and two treatments containing free peptides, mentioned indexes changed in a negative direction (decrease in the quality of dough). This finding indicates that in regard of controlling chemical and microbial reactions that cause food spoilage, bioactive peptides carrier nanocapsules are more powerful than free peptides. According to the obtained results, bioactive peptides from lanternfish with a molecular weight of less than 3 kilodaltons can be used as a natural preservative in the formulation of doogh. Also, if these peptides are nanoencapsulated using a combined maltodextrin-gum Arabic coating and freeze-drying technique, the resulting nanocapsules more actively combat oxidative and bacterial spoilage in doogh during storage at refrigerated temperatures. Conflict of interest The authors declare no conflicts of interest. Acknowledgment The Authors would like to thank the Sari Agricultural Sciences and Natural Resources University (SANRU), for financial support of this research under contract number, 03-1404-03. 13 31 2025/10/52025/08/26 1404/6/4 2026/04/302026/04/30 1405/2/10 سکینه یگانه Sakineh Yeganeh گروه شیلات، دانشکده علوم دامی و شیلات، دانشگاه علوم کشاورزی و منابع طبیعی ساری، ساری، ایران skyeganeh@gmail.com سهیل ریحانی پول Soheyl Reyhani Poul گروه فرآوری محصولات شیلاتی، دانشکده شیلات و محیط زیست، دانشگاه علوم کشاورزی و منابع طبیعی گرگان، گرگان، ایران soheylreyhani@gmail.com فائزه ترک پهنابی Faezeh Tork pahnabi گروه شیلات، دانشکده علوم دامی و شیلات، دانشگاه علوم کشاورزی و منابع طبیعی ساری، ساری، ایران pahnabifaezeh@gmail.com فرزانه شاکریان Farzaneh Shakerian گروه شیلات، دانشکده علوم دامی و شیلات، دانشگاه علوم کشاورزی و منابع طبیعی ساری، ساری، ایران farzaneshakerian64@gmail.com Bioactive peptides Doogh Antioxidant and antimicrobial activity پپتیدهای زیست‌‌فعال صمغ عربی دوغ فعالیت آنتی‌‌اکسیدانی و ضد میکروبی Abd Rashid, N.Y., Manan, M.A., Pa'ee, K.F., Saari, N. and Wong, F.W.F., 2022. 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مقاله علمی – پژوهشی:‌ بررسی کیفیت آب و وضعیت تروفیک (تغذیه ای) با تکیه بر شاخص‌های زیستی (پریفیتون و زئوپلانکتون) در رودخانه خرسان، استان چهارمحال و بختیاری Assessment of water quality and trophic status using biological indices (periphyton and zooplankton) in the Khersan River, Charmahal and Bakhtiari Province 1 1 پژوهش حاضر با هدف بررسی کیفیت آب و وضعیت تروفیک(تغذیه ای) رودخانه خرسان در شهرستان لردگان (استان چهارمحال و بختیاری) انجام شد. برای ارزیابی شرایط زیستی، شاخص‌های Shannon (تنوع گونه)، Evenness (یکنواختی گونه)، Saprobic (آلودگی آلی) و TDI (شاخص تروفی دیاتومه) بر اساس جوامع پریفیتون و زئوپلانکتون مورد استفاده قرار گرفت. نمونه‌برداری به‌ صورت فصلی از پاییز 1402 لغایت تابستان 1403 در پنج ایستگاه انجام شد. نتایج نشان داد که در جوامع پریفیتونی پنج شاخه شامل 30 جنس و 38 گونه و در جوامع زئوپلانکتونی چهار شاخه شامل یک خانواده، 4 جنس و 13 گونه شناسایی شد. شاخه Bacillariophyta با 74 درصد و شاخه Rotifera با 56 درصد دارای بیشترین سهم گونه‌ای بودند. شاخص تنوع Shannon در بیشتر ایستگاه‌ها برای هر دو گروه زیستی در محدوده 1 الی 3 قرار گرفت که بیانگر شرایط آلودگی متوسط بود. شاخص یکنواختی گونه برای پریفیتون‌ها 97/0 -67/0و برای زئوپلانکتون‌ها 98/0 -44/0 محاسبه شد. شاخص Saproby بر اساس پریفیتون در محدوده 01/2-78/1 و بر اساس زئوپلانکتون در محدوده 01/2-51/1 قرار داشت. این نتایج نشان‌دهنده شرایط آلودگی آلی متوسط[1] در همه ایستگاه‌ها و فصول بود. شاخص دیاتومه نیز در محدوده 74/3-17/2 قرار گرفت که شرایط مزوتروفی تا یوتروفی را در بهار و تابستان و شرایط یوتروفی را در پاییز و زمستان نشان داد. این وضعیت بیانگر بارگذاری متوسط تا زیاد مواد مغذی در رودخانه است. بررسی‌ها نشان داد که با وجود غناء فصلی مواد مغذی به‌ویژه فسفر، محدودیت‌های فیزیکی (جریان شدید آب و مواد معلق)، مانع افزایش زی‌توده شدند. شاخهBacillariophyta  نه با تراکم بیشتر بلکه با مقاومت و رقابت مؤثر، غلبه داشتند. کنترل بارگذاری مواد مغذی برای حفظ تعادل توزیع گونه‌ها در آینده ضروری است. همچنین زئوپلانکتون‌های غالب در همه ایستگاه‌ها شاخصی از آلودگی آلی متوسط بودند که با نتایج پریفیتون‌ها هم‌خوانی داشت.   [1] Beta-mesosaprobic 2 Introduction Hydrological processes and the physicochemical characteristics of rivers have been altered by climate change, particularly due to anthropogenic influences, resulting in significant consequences for riverine habitats and aquatic ecosystems (Gilvear et al., 2002).The combined effects of severe pollutant loading and disrupted hydrodynamics associated with dam construction further diminish the ecological capacity of river systems (Zhang et al., 2019). Advances in our understanding of the structural and functional dynamics of aquatic communities now provide opportunities for more effective management of environmental instability and pollution (Ekpo, 2013). Aquatic organisms, through their continuous interactions with one another and with the surrounding water environment, play a critical role in sustaining ecosystem integrity (Caroni et al., 2013). In flowing waters, where hydrological changes are rapid and difficult to estimate, physicochemical measurements due to their instantaneous nature cannot reflect the integration of multiple environmental factors or the long-term stability of river ecosystems. Consequently, biological monitoring, employing living organisms as bioindicators, is indispensable for assessing ecological conditions and detecting change (Oertel and Salanki, 2003). This study, conducted for the first time in the Khersan River basin, focuses on the Khersan 3 Dam, a double-arch concrete dam currently under construction in Lordegan County, Chaharmahal and Bakhtiari Province. In this study, biological Indices based on periphyton and zooplankton communities were used to assess the water quality status of the Marbareh and Bashar tributaries entering the Khersan River, in order to better understanding and implement necessary conservation strategies. Methodology Five stations were selected for investigation: St1: Marbareh River (under the entrance bridge of Ab-Malakh village); St2: Marbareh River (under the bridge of Narmeh village);St3: Upstream of the dam site (at the beginning of the reservoir); St4: Confluence of the Bashar and Marbareh rivers (before the village of Dowrah Ghamashkeh); St5: Location of the Khersan-3 Dam reservoir (the dam is under construction). Seasonal sampling was conducted throughout 2023 and 2024. Laboratory analyses of periphyton and zooplankton followed the protocols of APHA (2017), Wetzel and Linkens (1991), and Postel et al. (2000). Taxonomic identification was guided by references including Tiffany and Britton (1971), Habit and Pankow (1976), Hartley et al. (1996), Boltovskov (2000), Kuticova (1970), and Manolova (1964). Indices of species diversity and evenness were calculated according to Mason (1982). The saprobic index was computed following Sladechek (1973), Papadimitriou et al. (2010), and Li et al. (2017), while the Diatom Trophic Index was determined using the method of Kelly and Whitton (1995). Physicochemical parameters were measured in accordance with APHA (2017) standards. Statistical analyses were performed using SPSS version 22. Data were tested for normality, and non-normal distributions were normalized through logarithmic transformation. Temporal and spatial variations were examined using one-way ANOVA and Duncan’s post hoc test, and Pearson correlation was applied to explore relationships between selected physicochemical factors and biological indices. Results The annual qualitative and quantitative data analysis of microalgae in periphyton at sampling stations revealed the presence of 68 species. Periphyton organisms were identified across five phyla:Bacillariophyta,Cyanobacteria,Chlorophyta, Euglenozoa, and Xanthophyta. Bacillariophyta accounted for the highest percentage of species (74%). Dominant species were introduced at each station.Variance analysis comparing the mean total density of periphyton showed no significant differences between stations (p≥05/0).The lower densities occurred in warmer seasons and higher densities in colder seasons.The annual analysis of zooplankton at sampling stations revealed 13 species, 4 genera, and 1 family. Zooplankton were identified across four phyla: Rotifera, Ciliophora, Cladocera, and Copepoda. Rotifera accounted for the highest percentage of species (56%). Dominant species were introduced at each station. Duncan’s test in the comparative analysis of mean total density across stations identified Station 4 as significantly different (p≤05/0). zooplankton, also showed lower densities in warmer seasons and higher densities in colder seasons. Abiotic factors were measured to support biological data analysis and index values. Most stations, based on Shannon diversity index for both biological groups, fell within the range of 1 to 3, indicating moderate pollution. Species evenness index ranged: For periphyton: 0.67 to 0.97, For zooplankton: 0.44 to 0.98. Saprobic index based on periphyton ranged from 1.78 to 2.01, and for zooplankton from 1.51 to 2.01. The organic pollution index across all stations and seasons for both groups indicated β-mesosaprobic conditions, or moderate organic pollution. Diatom index values ranged from 2.17 to 3.74, indicating mesotrophic to eutrophic conditions in spring and summer, and eutrophic conditions in colder seasons suggesting moderate to high nutrient loading. Discussion and conclusion The biological data from various stations along Khersan river during the 2023–2024 indicated  that the ecological conditions of these rivers generally fall within the range of moderate organic pollution. The Shannon species diversity index for both biological groups ranged between 1 and 3, corresponding to semi-stable conditions and moderate pollution levels.The saprobic index, calculated based on periphyton (1.78–2.01) and zooplankton (1.51–2.01), also placed all stations within the β-mesosaprobic range, indicating moderate organic pollution. The species evenness index showed relative balance, especially among periphyton, and species indicative of severe pollution were neither abundant nor dominant. The widespread presence of dominant diatoms such as Diatoma vulgaris and Navicula sp. both indicators of moderate pollution aligns with the saprobic index values. Similarly, the dominant zooplankton species across all stations were indicators of moderate organic pollution (Kulas et al., 2021). Station 4 exhibited the highest density of both periphyton and zooplankton communities, as well as the highest species diversity index for both groups. However, the lowest populations of these organisms were recorded in spring, likely due to flood conditions and high water discharge (50–66 m³/s) across all stations. The lowest Shannon diversity index values were also calculated for spring. The Shannon index for periphyton showed a strong inverse correlation with water flow (r = -0.67) and total suspended solids (TSS) (r=-0.59). The latter maybe affects light penetration, sediment abrasion, and substrate conditions (Yuli Herawati et al., 2024). The Shannon index for zooplankton showed a strong direct correlation with dissolved oxygen concentration (r = 0.5) and nitrate concentration (r = 0.53), and a strong inverse correlation with total phosphorus (r = -0.59). This may be influenced by the positive correlation (r=0.55) between phytoplankton and zooplankton diversity indices in the basin. In spring, floodwaters and wastewater inflows increased total phosphorus concentrations across all stations compared to colder seasons, while total nitrogen concentrations decreased in warmer seasons. This imbalance in the nitrogen-to-phosphorus ratio with high discharge and suspended solids, likely reduced light penetration resulting in lower diversity and density of periphyton, phytoplankton, and subsequently zooplankton (Liu et al., 2023). The saprobic index for periphyton showed a moderate direct correlation with BOD₅ (r = 0.45). Higher BOD₅ levels in winter, along with elevated saprobic index values, suggest increased organic pollution, altered nutrient cycles, and higher trophic levels (Sulastri et al., 2021). The Diatom Trophic Index (TDI), indicated eutrophic conditions with clear nutrient loading in autumn and winter, and mesotrophic to eutrophic conditions with moderate to critical nutrient loading in spring and summer.The dominance and species richness of Bacillariophyta (diatoms) in most stations (2–5) suggest favorable conditions for their high density compared to other microalgae like Cyanobacteria and Chlorophyta. Diatoms thrive in cold temperatures and turbid waters with high suspended solids, and their ability to attach to substrates and form biofilms helps them resist being washed away in high-flow environments (Yoo et al., 2025). The Khersan River basin represents a system with moderate to high nutrient loading. However, due to physical constraints such as strong water flow, turbidity, and high suspended solids, this nutrient richness does not translate into high biomass. Bacillariophyta dominate not by sheer density but through resilience and competitive advantage under harsh and variable conditions. Upstream activities, flow changes due to dam construction, and gradual inflow of runoff and organic pollutants may soon disrupt the relative balance of biological species distribution. This study is the first of its kind in this watershed, and future monitoring at shorter intervals is essential to assess nutrient loading and trophic status. Conflict of interest The authors declare that they have no conflict of interest. Acknowledgment We sincerely express our gratitude to the Iranian Fisheries Science Research Institute for its support, which provided the scientific and laboratory framework for this study. This research was derived from the project entitled “Investigation of the abundance and biodiversity of plankton and periphyton communities in the sub‑basin rivers of Khersan 3 Dam” with the approved code14-76-12-114-02049-021030. 33 48 2025/10/52025/08/262025/10/10 1404/7/18 2026/04/302026/04/302026/04/30 1405/2/10 رحیمه رحمتی Rahimeh Rahmati پژوهشکده اکولوژی دریای خزر، موسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، ساری، ایران rahmati764@gmail.com آسیه مخلوق Asieh Makhlough پژوهشکده اکولوژی دریای خزر، موسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، ساری، ایران asieh_makhlough@yahoo.com نرگس عالیشاه Narges Alishah پژوهشکده اکولوژی دریای خزر، موسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، ساری، ایران n.alishah69@gmail.com حسن نصراله زاده ساروی Hasan Nasrolahzadeh Saravi پژوهشکده اکولوژی دریای خزر، موسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، ساری، ایران hnsaravi@gmail.com حسن فضلی Hasan Fazli پژوهشکده اکولوژی دریای خزر، موسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، ساری، ایران fazlihasan@gmail.com بهزاد رهنما Behzad Rahnama پژوهشکده اکولوژی دریای خزر، موسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، ساری، ایران rahnama.behzad@gmail.com احد احمدنژاد چهره Ahad Ahmadnejad Chehreh پژوهشکده اکولوژی دریای خزر، موسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، ساری، ایران Ahmadi.ahad@yahoo.com محمد کاردر رستمی Mohammad Kardar Rostami پژوهشکده اکولوژی دریای خزر، موسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، ساری، ایران MohammadKardarrostami@gmail.com Khersan River water quality Periphyton Zooplankton Biological Indecies رودخانه خرسان کیفیت آب پریفیتون زئوپلانکتون شاخص زیستی APHA (American Public Health Association)., 2017. 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مقاله علمی – پژوهشی:‌ اثر مکمل نمک‌‌های صفراوی بر عملکرد رشد، تغذیه و برخی شاخص‌های بیوشیمیایی پلاسما در ماهی باس دریایی آسیایی جوان (Lates calcarifer) Effect of bile salt supplementation on growth performance, feed efficiency and some plasma biochemical indices in Asian sea bass (Lates calcarifer) juveniles 1 1 مطالعه‌ای به مدت 120 روز جهت ارزیابی اثرات مکمل نمک‌های صفراوی بر عملکرد رشد، ضریب تبدیل غذایی و برخی از شاخص‌‌های فیزیولوژی ماهی باس دریایی (Lates calcarifer) با متوسط وزن اولیه 5/0±182 گرم انجام شد. به جیره پایه (پروتئین 44 درصد و چربی 15 درصد) غلظت‌های صفر (کنترل)، (BA400)400، (BA800 )800، ( BA1200) 1200و (BA1600) 1600 میلی‌‌گرم بر کیلوگرم مکمل نمک‌های صفراوی (رونئون، 30 درصد) اضافه شد. 255 عدد ماهی در 15 عدد مخزن پلی اتیلنی 1000 لیتری استوانه ای ذخیره سازی شدند و به مدت 120 روز با جیره های آزمایشی تا حد سیری (2 درصد وزن بدن)، تغذیه شدند. ماهیان در تیمار BA800 دارای وزن نهایی بیشتری نسبت به گروه های کنترل و BA400 بودند (05/0p<). آنزیم‌های سرمی آلانین و آسپارتات آمینو ترانسفراز در تیمار BA800 در مقایسه با سایر تیمار، به طور معنی‌داری کمتر بودند (05/0p<). سطح گلوکز سرم در تیمار BA400 و BA1600 به‌ترتیب دارای کمترین (1/84 میلی‌گرم در میلی‌لیتر) و بیشترین (1/243 میلی‌گرم در میلی‌لیتر) سطوح بودند. میزان لایزوزیم سرم در تیمار BA800 بیشتر از گروه‌‌های کنترل، BA1200 و BA1600 بود (05/0p<). بر اساس نتایج حاصله در مطالعه حاضر، افزودن 800 میلی‌گرم نمک‌های صفراوی بر کیلوگرم جیره غذایی نه‌تنها سبب افزایش رشد و بهبود ضریب تبدیل غذایی بلکه سبب بهبود شاخص‌های فیزیولوژی در ماهی باس دریایی آسیایی گردید. 2 Introduction The Asian sea bass, Lates calcarifer, is a carnivorous and marketable fish, with economic value in sea cages in the Persian Gulf and the Makran Sea. The culture yield of this species in the country by 2022 was about 6000 tons, and its production rate in the world is more than 120,000 tons (FAO, 2024; Mozanzadeh et al., 2022). The best levels of protein and fat in the diet for feeding the fattening stage of this species are 45-48 and 12-15 percent, respectively (Williams et al., 2003). Currently, one of the strategies for preventing and treating fatty liver in fish is mainly based on feed additives, including bile acids (Romano et al., 2020). Bile acids are defined as sterol compounds with hydrophilic and hydrophobic properties (Romano et al., 2020), playing an important role in the digestion and absorption of dietary fats by helping to suspend fats in the digestive tract and increasing the activity of bile salt-activated lipase (Romano et al., 2020). Accordingly, a feeding trial was designed to investigate the potential of using bile acids (supplemented with Runeon 1) in formulated diets of juvenile Asian sea bass and the effect on growth performance and feed efficiency. Methodology A 120-day study was conducted to investigate the effects of bile salt supplementation on growth performance, feed conversion ratio and some health indices of L. calcarifer with an average initial weight of 182±0.5 g. The basal diet (44% protein and 15% fat) was supplemented with bile salt supplements (Ronion 1, 30% purity) at concentrations of zero (control), 400 (BA400), 800 (BA800), 1200 (BA1200) and 1600 (BA1600) mg/kg. Two hundred and twenty-five fish were stored in 15 1000-liter cylindrical polyethylene tanks (15 fish per tank) and fed the experimental diets to satiety for 120 days. The average water temperature during the period was 30.5±1.2°C and the water salinity was 46±0.2 g/L. Growth parameters, including final weight, weight gain, specific growth rate and feed conversion ratio were determined. In addition, serum biochemical and immunological factors also were evaluated. Results Fish in the BA800 treatment had higher final weight, body weight gain percentage and specific growth rate than the control and BA400 groups (p<0.05, Table 1), but there was no significant difference with the BA1200 and BA1600 treatments. Feed conversion ratio in the BA800 and BA1200 treatments were lower than the other treatments. Liver enzymes alanine aminotransferase and aspartate aminotransferase were significantly lower in the BA800 treatment compared to the other treatments (p<0.05). Serum glucose levels in the BA400 and BA1600 treatments had the lowest and highest levels, respectively, and the other groups had intermediate numbers. Serum lysozyme levels in the BA800 treatment were higher than those in the control, BA1200, and BA1600 groups, but were not significantly different from those in the BA400 treatment (p<0.05). There was no significant difference in serum complement activity levels among the experimental treatments. Antiprotease activity levels in the bile acid-fed treatments were significantly higher than those in the control group. Discussion and conclusion The use of bile acid supplements in formulated diets has improved the growth of many farmed aquatic species (Wang et al., 2023). The results of the present study showed that adding 800 mg/kg of bile salts to the diet of Asian sea bass increased growth by improving feed conversion ratio in this species. Appropriate levels of bile acids increase the digestibility and absorption of fats by increasing the efficiency of nutrient absorption and activating digestive enzymes, especially lipases in the intestine, and by improving the efficiency of the diet, they lead to increased growth in aquatic animals (Romano et al., 2020). However, excessive and inappropriate levels of bile acids may have toxic effects on cells and lead to reduced growth and feed conversion ratio, and the level of this level varies depending on the type of farmed species (Romano et al., 2020). This supplement also has positive effects on liver health by reducing liver enzyme levels and improving non-specific immunity by increasing serum lysozyme and antitrypsin activity. However, adding high levels of bile acids (1600 mg/kg) to the diet will reduce growth, feed conversion ratio, and increase liver enzymes and serum glucose. Based on the results obtained in this study, adding 800 mg of bile salts per kilogram of diet increased growth, decreased serum liver enzymes, and increased serum lysozyme and antitrypsin enzyme activity levels. Conflict of interest We wish to confirm that there are no known conflicts of interest associated with this study. Acknowledgements The authors of this article would like to thank and appreciate the efforts of Mr. Moghaddis Zadeh and Mr. Pazhand from the Bandar Imam Khomeini Marine Fish Research Station for maintaining the fish. 49 59 2025/10/52025/08/262025/10/102025/11/22 1404/9/1 2026/04/302026/04/302026/04/302026/04/30 1405/2/10 منصور طرفی موزان زاده Mansour Torfi Mozanzadeh پژوهشکده آبزی‌‌پروری جنوب کشور، مؤسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش وترویج کشاورزی، اهواز، ایران mansour.torfi@gmail.com سمیرا ناظم رعایا Samira Nazemroaya پژوهشکده آبزی‌‌پروری جنوب کشور، مؤسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش وترویج کشاورزی، اهواز، ایران samira.nazemroaya@gmail.com فاطمه حکمت پور Fatemeh Hekmatpour پژوهشکده آبزی‌‌پروری جنوب کشور، مؤسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش وترویج کشاورزی، اهواز، ایران hekmatpourf@gmail.com علیرضا قائدی Alireza Ghaedi بخش تحقیقات علوم دامی و شیلات، مرکز تحقیقات و آموزش کشاورزی و منابع طبیعی استان چهارمحال و بختیـاری، سـازمان تحقیقـات، آمـوزش و تـرو یج،کشـاورزی، شهرکرد، ایران aliangler@gmail.com رضا کاظم پور Reza Kazempour دانشکده دامپزشکی، واحد علوم و تحقیقات، دانشگاه آزاد اسلامی، تهران، ایران kazempour@gmail.com لفته محسنی نژاد Lefteh Mohseninejad پژوهشکده آبزی‌‌پروری جنوب کشور، مؤسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش وترویج کشاورزی، اهواز، ایران lmohseni1@yahoo.com حسین هوشمند Hossein Houshmand پژوهشکده آبزی‌‌پروری جنوب کشور، مؤسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش وترویج کشاورزی، اهواز، ایران houshmand.hossein@gmail.com مجتبی ذبایح نجف آبادی Mojtaba Zabayeh Najafabadi پژوهشکده آبزی‌‌پروری جنوب کشور، مؤسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش وترویج کشاورزی، اهواز، ایران zabayeh2005@gmail.com حمید سقاوی Hamid Saghavi پژوهشکده آبزی‌‌پروری جنوب کشور، مؤسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش وترویج کشاورزی، اهواز، ایران Saghavih@gmail.com محمد یونس زاده فشالمی Mohammad Youneszadeh Feshalami پژوهشکده آبزی‌‌پروری جنوب کشور، مؤسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش وترویج کشاورزی، اهواز، ایران m_yooneszadeh@yahoo.com مینا آهنگر زاده Mina Ahangarzadeh پژوهشکده آبزی‌‌پروری جنوب کشور، مؤسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش وترویج کشاورزی، اهواز، ایران m.ahangarzadeh@yahoo.com عبدالرحیم اصولی Abdolrahim Oosooli پژوهشکده آبزی‌‌پروری جنوب کشور، مؤسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش وترویج کشاورزی، اهواز، ایران rahimoosooli@gmail.com راضیه عناناتی Razieh Ananati پژوهشکده آبزی‌‌پروری جنوب کشور، مؤسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش وترویج کشاورزی، اهواز، ایران Razieh-Ananati@gmail.com مصطفی صادقی پناه Mostafa Sadeghi Panah پژوهشکده آبزی‌‌پروری جنوب کشور، مؤسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش وترویج کشاورزی، اهواز، ایران Sadeghipanah@gmail.com شاپور مهرجویان Shapour Mehrjooyan پژوهشکده آبزی‌‌پروری جنوب کشور، مؤسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش وترویج کشاورزی، اهواز، ایران Mehrjoo.Shapour@gmail.com Bile acids growth performance feed efficiency health indicators Asian sea bass اسیدصفراوی کارایی رشد بازده تغذیه شاخص های سلامت باس دریایی آسیایی Abdel‑Tawwab, M., Abdel‑Latif, H.M.R., El Basuini, M.F., El‑Nokrashy, A.M., Khaled, A.A., Kord, M., Soliman, A.A., Zaki, M., Nour, A.-E., Labib, E.-M. 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مقاله علمی – پژوهشی:‌ اثر پروبیوتیک تک‌سل پلاس بر فراوانی باکتری‌های جنس Vibrioدر استخرهای پرورش مولدین میگو سفید غربی (Litopenaeus vannamei) Effect of Tek Cell Plus probiotic on the abundance of Vibrio genus bacteria in western white shrimp (Litopenaeus vannamei) broodstock rearing ponds 1 1 در حال حاضر، بیماری‌‌های فراوانی پرورش میگوی سراسر جهان را تحت تاثیر قرار داده‌‌اند. عوامل عفونی باکتریایی مهم‌‌ترین معضل بهداشتی میگوی پرورشی هستند. یک جایگزین مناسب برای عوامل ضد میکروبی شیمیایی برای جلوگیری از بیماری در آبزی پروری میگو، استفاده از پروبیوتیک‌‌هاست. به طور کلی، بیماری‌‌های ناشی ازVibrio spp. ، علت اصلی مرگ‌ومیر در هچری‌‌های میگو به‌ویژه در مراحل اولیه لاروی است. در بررسی حاضر، نمونه‌ آب مورد مطالعه از 3 استخر مزارع مولدسازی در شهرستان کنارک از عمق 30 سانتی‌متری از سطح آب به ‌صورت تصادفی برداشت شد. اطلاعات مدیریتی مزرعه پرورش میگو (نحوه مدیریت آب و غذادهی) و برخی شاخص‌های کیفی آب (شوری، pH و درجه حرارت آب)، اندازه‌گیری و ثبت ‎گردید. در آزمایشگاه نمونه‌ها همگن شده و کشت باکتریایی در محیط کشت‌های MYP، TSA و TCBS در سه تکرار به ‌منظور جداسازی اولیه Vibrio  غالب از آب استخر انجام شد. نتایج شمارش کل باکتری‌‌های جنس Vibrio نشان داد که پس از افزودن پروبیوتیک به آب استخرها، رشد باکتری‌‌های Vibrio شناسایی شده، نسبت به کنترل مثبت (تعداد باکتری‌‌های Vibrio قبل از افزودن پروبیوتیک)، به طور معنی‌داری کاهش یافت. تعداد کل باکتری جنس Vibrio و تعداد کل باکتری هتروترف در سه استخر شماره 1، 2 و 3 قبل از افزودن پروبیوتیک به‌ترتیب اعداد 102×2/3، 102×3/4 و 108×5 واحد کلنی بر میلی‌لیتر و بعد از افزودن پروبیوتیک به‌ترتیب اعداد  102×3 ، 102×3/1 و 102×4 واحد کلنی بر میلی‌لیتر نشان داد. نتایج حاصل از تعیین توانایی بازدارندگی رشد به روش انتشار از طریق چاهک بیشترین و کمترین اثر بازدارندگی محصول تجاری پروبیوتک و استفاده از سوسپانسیون آماده شده (C1) به‌ترتیب در A1-MYP (5/7 میلی‌متر) و V.p-TSA (3/6 میلی‌متر) مشاهده شد و در باکتری خالص شده از محصول تجاری پروبیوتیک در محیط کشت اختصاصی (C2) بیشترین و کمترین اثر بازدارندگی مربوط به A1-MYP (5/8 میلی‌متر) و B2-MYP (6 میلی‌متر) مشاهده گردید. بیشترین فعالیت مهار کنندگی پروبیوتیک در تست چاهک گذاری علیه A1 و V.P مشاهده شد. نتایج نشان داد پروبیوتیک تک سل پلاس دارای خاصیت آنتی‌‌باکتریال در برابر Vibrio غالب جداسازی شده از استخر و Vibrio parahaemolyticus بود. 2 Introduction Aquaculture, as one of the most important sources of animal protein, plays an increasingly significant role in global food security, and its importance has grown further due to the depletion of natural stocks and the rising demand for seafood (Madhana et al., 2021). In this context, shrimp farming has expanded rapidly, particularly in tropical countries, and has become a key sector of aquaculture production (HAU and Haryana, 2022). The Pacific white shrimp Litopenaeus vannamei is considered one of the most important cultured species due to its fast growth, high survival rate, relative resistance to diseases, and tolerance to a wide range of salinities, making it especially suitable for the environmental conditions of southern Iran (Chong‑Robles et al., 2014; Laramore et al., 2001). Despite advances in management practices, bacterial diseases remain among the main challenges faced by shrimp farms, with vibriosis caused by Vibrio spp. being a major contributor to mortality and economic losses (Muthu et al., 2024). Vibrios are Gram‑negative pathogenic bacteria that not only cause diseases in aquatic animals but can also lead to foodborne infections in humans (Bintsis, 2017). The excessive use of antibiotics to control these diseases has resulted in the emergence of antibiotic‑resistant bacteria and disruption of the microbial balance in aquaculture environments (Kesarcodi‑Watson et al., 2008). In this regard, environmentally friendly and preventive approaches, such as the use of probiotics as suitable alternatives to antibiotics, have gained increasing attention (Defoirdt et al., 2011; Proespraiwong et al., 2023). Probiotics contribute to disease reduction and improved survival by enhancing water quality, strengthening host immunity, and competing with pathogenic bacteria, particularly Vibrio spp. (Lahay et al., 2023; Verschuere et al., 2000). The efficacy of these compounds is commonly evaluated through in vitro antagonism assays (Kesarcodi‑Watson et al., 2008).The native probiotic “Tek Cell Plus,” which was isolated from shrimp farms in Bushehr Province and has reached industrial-scale production following global registration and extensive testing (Ghaednia et al., 2024; Mahjoub et al., 2019), was evaluated in the present study for its ability to reduce the bacterial load of vibrios in the pond water of Pacific white shrimp broodstock culture systems, as well as for its inhibitory effects on vibrio growth under laboratory conditions. Methodology Aqueous samples were procured from three discrete broodstock production ponds in the Konarak district utilizing stratified random sampling techniques at a standardized depth of 30 cm below the water surface interface. Comprehensive aquaculture management parameters were documented, including water quality management protocols, nutritional regimes, and physicochemical variables (salinity, pH, and thermal gradient). Laboratory analyses incorporated sample homogenization followed by selective and differential microbiological cultivation on MYP, TSA, and TCBS media, with triplicate inoculations to ensure statistical validity. The antagonistic properties of the commercial probiotic against indigenous Vibrio isolates were evaluated through quantitative inhibition zone assays using the agar well diffusion methodology. Positive controls consisted of pre-intervention Vibrio population densities, allowing for comparative efficacy assessment. Result Microbiological enumeration demonstrated a statistically significant reduction (p<0.05) in Vibrio spp. population densities following probiotic application across all experimental units. Specifically, Total Vibrio Count (TVC) and Total Heterotrophic Bacteria (THB) in ponds 1, 2, and 3 exhibited pre-intervention values of 3.2×10², 4.3×10², and 5×10⁸ CFU/mL, respectively. Post-application analyses revealed substantial reductions to 3×10², 1.3×10², and 4×10² CFU/mL, respectively, indicating a logarithmic decrease in bacterial load, particularly in pond 3. In vitro antagonism assays utilizing the well diffusion technique revealed differential inhibitory capacities. The commercial probiotic suspension (C1) demonstrated maximum and minimum inhibition growth zones against A1-MYP (7.5±0.2 mm) and V.p-TSA (6.3±0.1 mm), respectively. When utilizing purified bacterial isolates from the commercial probiotic (C2), enhanced inhibitory efficacy was observed, with maximum and minimum zones of inhibition registered against A1-MYP (8.5±0.2 mm) and B2-MYP (6.0±0.1 mm), respectively. The most pronounced antimicrobial activity was consistently observed against the A1 isolate and Vibrio parahaemolyticus, suggesting strain-specific antagonistic mechanisms. Discussion and conclusions The present findings demonstrate that the native probiotic Tek Cell Plus, identified as Bacillus vallismortis with the GenBank accession number JQ085958.1, significantly reduced the bacterial load of Vibrio, the dominant bacterial genus in shrimp pond water, relative to the total bacterial population. Similar probiotic effects have been reported for Bacillus cereus and Paenibacillus spp., which were evaluated at concentrations of 10⁴ and 10⁵ CFU mL⁻¹ in post‑larval shrimp ponds. In vivo experiments revealed that these bacteria exerted strong probiotic activity against Vibrio species, resulting in a significant reduction in shrimp larval mortality. The antagonistic activity of B. cereus and Paenibacillus spp. against vibrios has been attributed to the production of bioactive compounds such as zwittermicin‑A and kanosamine (Ravi et al., 2007). These mechanisms are consistent with the outcomes of the present study, wherein the application of B. vallismortis led to a marked reduction in Vibrio abundance in pond water. Supporting evidence is provided by Temario et al. (2022), who demonstrated that dietary supplementation with Bacillus subtilis BF12 significantly increased survival, improved growth performance, and reduced the intestinal abundance of Vibrio parahaemolyticus in Penaeus monodon. Additional studies have emphasized the efficacy of Bacillus-based probiotics in L. vannamei culture systems. In a 45‑day in vivo trial, B. subtilis IPA‑S.51 and Shewanella algae IPA‑S.111 inhibited V. parahaemolyticus growth while also enhancing shrimp growth performance. Agar‑well diffusion assays further confirmed that probiotics, given sufficient contact time, can produce clear inhibition zones against pathogenic vibrios. Ramesh et al. (2014) isolated twelve Bacillus spp. strains from the gut of P. monodon, several of which showed strong inhibitory activity against V. harveyi VSH5, with inhibition zones reaching up to 19.0 ± 0.1 mm. These effects were attributed to pH modification, production of inhibitory metabolites, nutrient competition, and lactonase enzyme activity. Probiotic efficacy has also been demonstrated using other bacterial groups. Studies by Ravi et al. (2007) and Ferreira et al. (2015) confirmed that Bacillus spp. significantly reduced Vibrio abundance in shrimp ponds, while Thompson et al. (2022) showed that lactic acid bacteria, including Lactobacillus curvatus subsp. curvatus, L. plantarum, and Pediococcus acidolactici, inhibited several pathogenic vibrios through the release of antimicrobial compounds. Acute hepatopancreatic necrosis disease (AHPND/EMS), primarily caused by toxigenic strains of V. parahaemolyticus, represents one of the most severe bacterial diseases in shrimp aquaculture. A meta‑analysis by Ghaednia et al. (2024) concluded that direct addition of single‑strain Bacillus-based probiotics to pond water is the most effective application method for AHPND control. The present study aligns with these findings, as water application of B. vallismortis yielded clear suppressive effects on pathogenic vibrios. Finally, bioinformatic analysis identified two Vibrio isolates closely related to Vibrio sagamiensis and Vibrio fortis, both belonging to the class Gammaproteobacteria. Previous studies have reported V. sagamiensis in tropical marine environments (Yoshizawa et al., 2010) and V. fortis as an emerging pathogen in aquatic organisms (Thompson et al., 2003; Wang et al., 2016), suggesting potential pathogenic risks in shrimp farming systems. Overall, the findings confirm the effectiveness of Bacillus vallismortis as a robust probiotic candidate for sustainable control of Vibrio-associated diseases in shrimp aquaculture. Conflict of interest All authors declare that they have no competing interests. Acknowledgment The authors would like to express their sincere appreciation to all those who contributed to this research through their guidance and scientific support. 61 73 2025/10/52025/08/262025/10/102025/11/222023/11/22 1402/9/1 2026/04/302026/04/302026/04/302026/04/302026/04/30 1405/2/10 سولماز بازدار Solmaz Bazdar گروه شیلات، دانشکده علوم دریایی، دانشگاه دریانوردی و علوم دریایی چابهار، چابهار، ایران bazdarsolmaz4@gmail.com مصطفی غفاری Mostafa Ghaffari گروه شیلات، دانشکده علوم دریایی، دانشگاه دریانوردی و علوم دریایی چابهار، چابهار، ایران mgmostafaghaffari@gmail.com اشکان اژدری Ashkan Ajdari مرکز تحقیقات شیلاتی آبهای دور، موسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، چابهار، ایران a_arzhan@yahoo.com زینب کرد zeinab kord گروه شیلات، دانشکده علوم دریایی، دانشگاه دریانوردی و علوم دریایی چابهار، چابهار، ایران kord.zeynab97@gmail.com Key words: single cell plus bacteria probiotic Viprio Spp Vibrio parahaemolyticus breeder shrimp breeding Litopenaeus vannamei breeding pool پروبیوتیک تک سل پلاس Vibrio Vibrio parahaemolyticus مولد میگو Litopenaeus vannamei Basim, Y., Mohebali, G., Jorfi, S., Nabizadeh, R., Ghadiri, A., Moghadam, M. 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مقاله علمی – پژوهشی:‌ تأثیر افزودن پروبیوتیک تک‌سل (Bacillus subtilis) بر شاخص‌های رشد، بقاء و کیفیت آب میگوی, Boone, 1931 Litopenaeus vannamei پرورش‌یافته در سیستم بیوفلاک Effect of supplementing single cell probiotic (Bacillus subtilis) on growth performance, survival, and water quality of Litopenaeus vannamei, Boone, 1931 cultured in a biofloc system 1 1 پرورش میگوی پا سفید غربی (Litopenaeus vannamei, Boone, 1931) نیازمند راهکارهایی برای حفظ کیفیت آب و بهبود شاخص‌های رشد است. یکی از رویکردهای نوین در این زمینه، استفاده هم‌زمان از فناوری بیوفلاک و مکمل زیستی مانند پروبیوتیک است. پژوهش حاضر، با هدف ارزیابی تأثیر کاربرد بیوفلاک و پروبیوتیک تک‌سل بر کیفیت آب و عملکرد رشد میگوی پا سفید غربی در مزارع پرورش میگو واقع در سایت پرورش میگوی گمیشان، انجام شد. پست لاروهای مرحله 12 (PL12) با میانگین وزنی 05/0±52/0 گرم با تراکم 200 هزار قطعه در هکتار در مزارع پرورش میگو ذخیره‌‌سازی شدند. در طول دوره پرورش، شاخص‌‌های فیزیکوشیمیایی (شوری، هدایت الکتریکی، دما، (pH به‌ صورت منظم پایش گردید. تغذیه میگوها با استفاده از خوراک تجاری کنسانتره، به میزان 4-2 وعده در روز و در محدوده 10-5/2 درصد زیست توده کل، از خرداد لغایت شهریور 1402 انجام شد. طرح آزمایشی به‌ صورت کاملاً تصادفی با یک گروه شاهد و سه تیمار آزمایشی، هر کدام با سه تکرار، اجرا گردید: گروه شاهد: (بدون افزودنی)، تیمار 1: (افزودن بیوفلاک و پروبیوتیک تک‌‌سل با غلظت 300 گرم در هکتار)؛ تیمار 2 (افزودن بیوفلاک و پروبیوتیک تک‌‌سل با غلظت 400 گرم در هکتار)؛  تیمار 3 (افزودن بیوفلاک و پروبیوتیک تک‌‌سل با غلظت 500 گرم در هکتار). نتایج حاصل از تجزیه‌وتحلیل آماری نشان داد که تفاوت معنی‌‌داری در درصد بازماندگی بین تیمارها وجود نداشت. با این‌حال‌، تیمار حاوی 300 گرم پروبیوتیک در هکتار، بالاترین میانگین وزن نهایی، بیشترین نرخ رشد روزانه و کمترین ضریب تبدیل غذایی را نسبت به سایر تیمارها به‌ویژه تیمار شاهد و تیمار 500 گرم در هکتار (غلظت پیشنهادی کارخانه سازنده)، نشان داد. بر این اساس، به‌نظر می‌‌رسد که استفاده از غلظت 300 گرم پروبیوتیک تک‌سل در هکتار، ضمن بهبود عملکرد رشد، از نظر اقتصادی نیز مقرون به‌صرفه‌تر است و می‌تواند به‌ عنوان دوز بهینه در سیستم‌های پرورش میگوی پا سفید غربی، پیشنهاد شود. 2 Introduction Aquaculture now represents a primary sector in global food production, having surpassed capture fisheries in aquatic animal production as of 2022 (Abdel-Rahim et al., 2023; Caputo et al., 2023). Among the farmed species, Litopenaeus vannamei (Boone, 1931) has become the dominant species due to its rapid growth, omnivorous diet, high metabolic rate, and adaptability to tropical marine environments (De Silva et al., 2021; Huang et al., 2025; Fadel et al., 2025). Although this species is native to the eastern Pacific, more than 85% of its global aquaculture production now occurs in Asian countries (FAO, 2020; Amiin et al., 2023). Specifically, in Iran, more than 180,000 hectares are suitable for shrimp farming, including 4,000 hectares in Golestan Province. Nevertheless, intensive shrimp farming is confronted with ecological challenges, particularly the accumulation of nutrients and deterioration of water quality due to animal feed (Chaikaew et al., 2019). Maintaining optimal water parameters including temperature, dissolved oxygen, salinity, pH, and conductivity is critical for preventing stress and disease in shrimp culture (Kautsky et al., 2000; Kuncha et al., 2025). Biofloc technology (BFT), which stimulates the formation of beneficial microbial flocs through the addition of carbon sources, has been shown to improve productivity and promote environmental sustainability (Kumar et al., 2014; Mansour et al., 2022; Iber et al., 2025). The incorporation of probiotics into recirculating aquaculture systems further enhances water quality, gut health, and immune function (Menaga et al., 2023; Kaya, 2025). Probiotics exert their effects by competitively excluding pathogens, producing antimicrobial substances, and stimulating host immune responses (Angahar, 2016; Hoseinifar et al., 2018); certain strains also facilitate the reduction of nitrogenous waste (Dalmin et al., 2001; Mang et al., 2024). Moreover, probiotics are recognized as cost-effective, readily isolatable, and ecologically sustainable interventions (Gullian et al., 2004; Vidhya and Thomas, 2023). In larval stages, probiotics enhance digestion through enzyme secretion (Bairagi et al., 2002; Lara-Flores, 2011; Qiu et al., 2023; Vulla et al., 2024). When combined with biofloc technology (BFT), they improve growth performance, feed efficiency, disease resistance, and water quality (Dash et al., 2018; Pratiwi et al., 2020; Qiu et al., 2023). This study aims to evaluate the synergistic effects of probiotics and BFT on L. vannamei with a focus on water quality improvement, enhanced survival, optimized growth, and increased pathogen resistance. The integrated approach offers a sustainable framework for efficient and environmentally friendly shrimp aquaculture. Methodology This study was conducted at the shrimp aquaculture site in Gomishan County, located in northern Golestan Province, Iran. Field experiments were carried out in 12 shrimp ponds to evaluate the effects of biofloc and the probiotic Tak-Cell (containing Bacillus subtilis) on the growth performance and survival of Pacific white shrimp (L. vannamei). A total of 30 kg of the probiotic was applied over a 120-day period across nine ponds, with an additional 500 g administered during the pond preparation phase. Biofloc was prepared using carbon sources such as sugarcane molasses, rice flour, and wheat flour. These substrates were incubated in warm water (40°C) and subsequently diluted at a specific ratio before being introduced into the culture system. The carbon and nitrogen ratio (C: N) was optimized based on the assumption that 50% of nitrogen from feed is excreted into the aquatic environment. Post-larvae with an initial mean weight of 0.52±0.12 g were stocked at a density of 200,000 individuals per hectare in aerated ponds. Throughout the culture period (June to September), water quality parameters including salinity, temperature, pH, ammonia, nitrate, and phosphate were regularly monitored. Shrimp were fed commercial formulated diets 2–4 times daily at feeding rates ranging from 2.5% to 10% of biomass. The experimental design followed a completely randomized structure with four treatments: (1) biofloc without probiotic (control), and (2–4) biofloc combined with probiotic at concentrations of 300, 400, and 500 g/ha, respectively. Each treatment was replicated three times. Growth parameters including mean body weight, average daily gain, specific growth rate, feed conversion ratio, and survival rate were calculated using standard formulas. Data normality was assessed via the Shapiro–Wilk test. Upon confirmation of normal distribution, one-way ANOVA and LSD post hoc tests were conducted at a 5% significance level using SPSS version 23. Result During the period of shrimp farming in the test ponds, considerable fluctuations in air temperature were observed between the morning and evening hours, occasionally reaching up to 15°C. However, the water temperature, salinity and pH remained relatively stable throughout the sampling period and were consistently within the optimum range for shrimp farming. Statistical analysis revealed no significant differences in survival rates between the different treatments (p>0.05). Nevertheless, the 500 g/ha single-cell probiotic treatment showed the least variation in survival rate, indicating more stable culture conditions. In terms of growth performance, the highest mean final weight was observed in the 300 g/ha of probiotic treatment. Although this difference was not statistically significant compared to the 400 g/ha treatment, it was significantly higher than that of 500 g/ha and control groups. The highest specific growth rate (SGR) was recorded in the 400 g/ha treatment, which was not significantly different from other treatments, including the control. The highest average daily growth rate (ADGR) was also recorded in the 300 g/ha group, but no statistically significant differences were observed between treatments. For live biomass, the 300 g/ha treatment yielded the highest values and showed a statistically significant difference compared to the 500 g/ha group. Feed consumption was highest in the control group and was significantly higher than in the other treatments. The lowest feed conversion ratio (FCR) was found in the 300 g/ha treatment, which differed significantly only from the control group. Discussion and conclusion Probiotics play a crucial role in improving growth performance in aquaculture by optimising metabolic processes and creating favourable ecological conditions (Liu et al., 2009; Todorov et al., 2024; Tao et al., 2025). Their enzymatic activity improves digestion and nutrient uptake, leading to better growth results (Ghosh et al., 2023; Calcagnile et al., 2025). In the present study, water quality parameters such as temperature, salinity, and pH remained within the optimal range, which is consistent with the findings of Naik and Srinivasulu Reddy (2020b) and Van Wyk and Scarpa (1999), who emphasized the importance of a stable environment for shrimp health. Survival rates showed no significant differences between treatments, which is consistent with previous Biofloc studies (Naik and Srinivasulu Reddy, 2020b), although the 500 g/ha probiotic treatment showed greater stability. In particular, the 300 g/ha treatment gave better results for final weight, biomass, and feed conversion ratio (FCR), outperforming the manufacturer‑recommended dose. This result contrasts with Llario et al. (2020), who reported limited growth benefits from Bacillus amyloliquefaciens alone, but supports studies emphasizing the improved efficacy of probiotics when used in combination (Souza et al., 2012; Qiu et al., 2023; Preena et al., 2025). The addition of carbon sources such as molasses and rice bran contributed to improved water quality and plankton density, which is consistent with the results of Naik and Srinivasulu Reddy (2020a), Menaga et al. (2023), and Aparna et al. (2024). These improvements probably supported the observed growth and feed efficiency. The synergistic effects of probiotics and biofloces reduction of microbial load, immune stimulation and supplementary nutrition; are well documented (Serra et al., 2015; Santos et al., 2024; Marimuthu et al., 2024) and were reflected in the result of the present study, with FCR values around 2 and final weights between 13.21 and 20.80g. Overall, the use of 300 g/ha of single-cell probiotic proved to be more effective than the recommended dose and provided both biological and economic benefits. This supports the view that optimised probiotic dosing in conjunction with carbon supplementation and biofloc management can significantly improve the sustainability and productivity of shrimp aquaculture (Khanjani et al., 2024; Ziaei-Nejad et al., 2006; Megahed et al., 2019; Kaya et al., 2022). Based on the results of this study, it is recommended that future research focus on the synergistic application of multi-strain probiotic formulations in combination with optimized carbon sources. This approach can further improve growth performance, feed efficiency, and environmental stability in intensive shrimp aquaculture systems. In addition, adjusting probiotic dosing based on empirical evidence rather than manufacturer guidelines may improve both biological efficacy and economic sustainability. Conflict of interest The authors declare that there are no known financial conflicts of interest or personal relationships that could have appeared to influence the work presented in this article. Acknowledgment The authors sincerely express their appreciation to all colleagues and experts who provided assistance at various stages of this research. This study was financially supported by Gorgan University of Agricultural Sciences and Natural Resources, under the research project entitled “Improving the growth performance and survival of Pacific white shrimp (L. vannamei) through the addition of probiotics and biofloc to pond water” (Project ID: 21‑484‑02). 75 91 2025/10/52025/08/262025/10/102025/11/222023/11/222025/08/2 1404/5/11 2026/04/302026/04/302026/04/302026/04/302026/04/302026/02/8 1404/11/19 رسول قربانی Rasoul Ghorbani دانشگاه علوم کشاورزی و منابع طبیعی گرگان، استان گلستان، گرگان، ایران rasoulghorbani@gmail.com سید عباس حسینی Seyed Abbas Hosseini دانشگاه علوم کشاورزی و منابع طبیعی گرگان، استان گلستان، گرگان، ایران seyedabbas_hosseini@yahoo.com فاطمه عباسی Fatemeh Abbasi دانشگاه علوم کشاورزی و منابع طبیعی گرگان، استان گلستان، گرگان، ایران f.abbasi59@yahoo.com میثم سبزه Meysam Sabzeh دانشگاه علوم کشاورزی و منابع طبیعی گرگان، استان گلستان، گرگان، ایران meysam.sabzeh2002@gmail.com حدیثه کشیری Hadiseh Kashiri دانشگاه علوم کشاورزی و منابع طبیعی گرگان، استان گلستان، گرگان، ایران hadiskashiri@gmail.com عبدالعظیم فاضل Abdol Azim Fazel مرکز تحقیقات ذخایر آبزیان آبهای داخلی، استان گلستان، گرگان، ایران f.fazel58@gmail.com Biofloc Single cell probiotic Pacific white shrimp Shrimp culture Growth performance بیوفلاک پروبیوتیک تک‌سل میگوی پاسفید غربی پرورش میگو شاخص رشد Abdel-Rahim, M.M., Elhetawy, A.I., Mansour, A.T., Mohamed, R.A., Lotfy, A.M., Sallam, A.E. and Shahin, S.A., 2024. 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