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مقاله علمی – پژوهشی:‌ سطوح استروئیدهای جنسی در مولدین ماده کوی (Cyprinus carpio carpio) تزریق شده با GnRH نوترکیب Sex steroid levels in female koi (Cyprinus carpio carpio) broodstock injected with recombinant GnRH 1 1 در مطالعه حاضر، اثر تزریق GnRH نوترکیب (rGnRH) با و بدون آنتی دوپامین بر سطوح استروئیدهای جنسی مولدین ماده ماهی کوی (Cyprinus carpio carpio) مورد بررسی قرار گرفت. بدین منظور، تعداد 60 عدد ماهی مولد کوی آماده تکثیر با میانگین وزنی 52/94 ± 25/415 گرم به مدت دو هفته با شرایط نگهداری سازگار شدند. بعد از گذراندن دوره سازگاری و مناسب شدن دمای محیط، مولدین به چهار گروه آزمایشی تقسیم شدند. برای تیمارهای اول و دوم به عنوان گروه‌های شاهد مثبت و منفی به‌ترتیب سرم فیزیولوژی و آنتی دوپامین دامپریدون با دوز 5 میلی‌‌گرم بر کیلوگرم وزن بدن تزریق شد، در تیمار سوم هورمون GnRH نوترکیب با دوز 25 میکروگرم بر کیلوگرم وزن بدن و در تیمار چهارم هورمون GnRH نوترکیب با دوز 25 میکروگرم بر کیلوگرم وزن بدن به همراه آنتی دوپامین دامپریدون با دوز 5 میلی‌‌گرم بر کیلوگرم وزن بدن به صورت یک مرحله ای تزریق شد. از این مولدین برای بررسی استروئیدهای جنسی (17 بتا استرادیول، تستوسترون و پروژسترون) در چهار نوبت (قبل از تزریق، 6، 12 و 24 ساعت بعد از تزریق)، خون‌‌گیری شد. نتایج نشان داد که از نظر تعداد مولدین تخم‌‌ریزی کرده تفاوت معنی‌‌داری بین تیمارها وجود نداشت. سطوح استروئیدهای جنسی تحت تاثیر تیمارهای هورمونی قرار گرفت (05/0>p). سطح هورمون 17 بتا استرادیول در گروه شاهد و تیمار تزریق شده با دامپریدون در مراحل نمونه‌‌برداری بدون تغییر باقی‌‌ماند (05/0p>)، اما سطح این هورمون در تیمارهای دریافت‌کننده هورمون GnRH نوترکیب با و بدون آنتی دوپامین (دامپریدون) 6 ساعت بعد از تزریق افزایش و سپس 12 ساعت بعد از تزریق کاهش یافت (05/0>p). سطح هورمون پروژسترون و تستوسترون در گروه‌‌های شاهد مثبت و منفی بدون تغییر باقی ماند (05/0p>)، اما در تیمارهای دریافت کننده هورمون GnRH نوترکیب با و بدون آنتی دوپامین (دامپریدون) 6 و 12 ساعت بعد از تزریق افزایش و سپس در 24 ساعت بعد از تزریق کاهش یافت (05/0>p). نتایج نشان داد که هورمون GnRH نوترکیب در دوز 25 میکروگرم بر کیلوگرم وزن بدن بدون آنتی دوپامین می‌‌تواند تخم‌‌ریزی را القاء کند. 2 Introduction The control of fish reproduction in a captive setting can be achieved through the manipulation of various environmental elements, including photoperiod, water temperature, and the selection of spawning grounds (Yeganeh et al., 2022). Given the limited understanding of the biological needs of certain species and the impracticability of replicating environmental conditions to achieve natural reproduction, one viable approach is to employ hormones. A problem in final maturity induction in the carp family is the high activity of dopamine as an inhibitor in the final maturity and spawning step. A recombinant GnRH combined with dopamine receptor antagonists was employed to facilitate the final maturation process in both goldfish (Carassius auratus) and koi carp. (Mohammadzadeh et al., 2020b, 2021a; Yeganeh et al., 2022). No study is so far available on the effects of recombinant GnRH (rGnRH) without anti-dopamine antagonist in koi carp. Therefore, this study aims to investigate the possible use of rGnRH without anti-dopamine antagonist in female koi carp. Methodology The investigation was conducted to examine the effects of recombinant gonadotropin-releasing hormone (rGnRH) administered alone or in combination with domperidone on the reproductive function and sex steroid levels in female koi carp (Cyprinus carpio). For the purpose of this study, female koi carps with an average weight of 415.25 ± 94.52 g were divided into four experimental groups. Each group was subjected to a specific injection procedure. The first group received a control injection of 0.9% NaCl. The second group was injected with 5 mg/kg body weight of domperidone (Dop). The third group received an injection of 25 μg/kg body weight of rGnRH (rGn). The fourth group was injected with 25 μg/kg body weight of rGnRH with 5 mg/kg of domperidone (rGnDop). To investigate the levels of sex steroids, samples of blood were obtained prior to administration and at the time points of 6, 12, and 24 hours following the injection. Result All the female koi carp of rGn and rGnDop treatments spawned, but none of the females spawned in the NaCl and Dop treatments. There was notable difference among spawned fish during the latency period, with the rGn treatment yielding the longest latency compared to the shorter latency period in the rGnDop treatment. The hormonal treatments had an impact on the levels of sex steroids, namely 17β estradiol (E2), testosterone (T), and 17α, 20β-dihydroxy-4-pregnen-3-one (DHP). The level of E2 decreased in rGn and rGnDop 6, 12 and 24 h after injection. T and DHP levels increased in rGn and rGnDop treatments at 6 and 12 h after the injection and then decreased at 24 h post-injection. Discussion and conclusion The findings of the final induction of maturity in female koi breeders illustrate that the administration of rGnRH, both with and without anti-dopamine, exerts a natural influence on the physiological process of koi fish reproduction. Furthermore, it is noteworthy that these interventions do not lead to any abnormalities in the reproductive system at different stages of the artificial reproduction of this species. The elevation of DHP levels in breeders treated with rGnRH without domperidone exhibits a comparable pattern to those treated with rGnRH alongside domperidone. Consequently, it is advisable to utilize rGnRH without anti-dopamine for the purpose of inducing the final maturation of koi carp. Conflict of interest The authors declare that they have no conflict of interest. Acknowledgments The authors would like to thank the Sari Agricultural Sciences and Natural Resources University (SANRU). 1 11 2025/06/3 1404/3/13 2026/07/1 1405/4/10 آرش رمضانی Arash Ramzani گروه شیلات، دانشکده علوم دامی و شیلات، دانشگاه علوم کشاورزی و منابع طبیعی ساری، ساری، ایران ramzani.arash@gmail.com سکینه یگانه Sakineh Yeganeh گروه شیلات، دانشکده علوم دامی و شیلات، دانشگاه علوم کشاورزی و منابع طبیعی ساری، ساری، ایران sk.yeganeh@gmail.com صدیقه محمدزاده Sedigheh Mohammadzadeh دانشکده دامپزشکی، دانشگاه تخصصی فناوری‌‌های نوین، آمل، ایران mohammadzadeh.sedigheh@yahoo.com Recombinant GnRH Ovulation synchronization Koi Spawning Anti-dopamine GnRH نوترکیب همزمانی تخم‌‌ریزی ماهی کوی تخم‌‌ریزی آنتی دوپامین Ahmadifar, A., Imanpour, M. R., Amini, K., Zadmajid, V. and Gholampour, T.A., 2016. Different methods using GnRHa on out of season reproductive efficiency in male goldfish (Carassius auratus, Linnaeus 1758). Applied Biology, 6(1): 29-39. 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Advancement and synchronisation of spawning in Salmo gairdneri and S. trutta following administration of LRH-A combined or not with pimozide. Aquaculture, 43(1-3): 57-66. DOI: 10.1016/0044-8486(84)90009-7 ##Brzuska, E., 2021. Reproduction effectiveness of carp (Cyprinus carpio L.) from the Hungarian W breeding line after stimulating ovulation with spawning inducing agents of natural (CPH, hCG, PMSG) and/or synthetic origin (Ovopel, Dagin, Ovaprim, mGnRH-A). Aquaculture, 532: 736023. DOI: 10.1016/j.aquaculture.2020.736023.##Gillet, C., Breton, B. and Mikolajczyk, T., 1996. Effects of GnRHa and pimozide treatments on the timing of ovulation and on egg quality in Arctic charr (Salvelinus alpinus) at 5 and 10 C. Aquatic Living Resources, 9(3): 257-263. DOI: 10.1051/alr:1996029##Kouřil, J., Barth, T., Hamáčková, J. and Flegel, M., 1986. Induced ovulation in tench (Tinca tinca L.) by various LH-RH synthetic analogues: effect of site of administration and temperature. 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مقاله علمی – پژوهشی:‌ اثر جایگزینی آرد ماهی با پودر لارو حشره‌ سرباز سیاه (Hermetia illucens) بدون هیچ‌گونه افزودنی و با غنی‌سازی‌شده با اسپیرولینا بر رشد، شاخص‌های بیوشیمیایی خون در بچه‌ماهی سی‌َبس آسیایی (Lates calcarifer) Effects of replacing fishmeal with black soldier fly larvae (Hermetia illucens) meal, with and without spirulina enrichment, on growth and biochemical plasma indices in Asian sea bass (Lates calcarifer) fry 1 1 پژوهش حاضر، با هدف بررسی تأثیر جایگزینی آرد ماهی با پودر لارو مگس سرباز سیاه (Hermetia illucens) غنی‌شده با جلبک اسپیرولینا بر شاخص‌های عملکرد رشد، ترکیب شیمیایی لاشه و شاخص‌‌های بیوشیمیایی پلاسمای خون در بچه‌ماهیان سی‌بس آسیایی (Lates calcarifer) انجام شد. تعداد ۳۰۰ قطعه ماهی با وزن اولیه 00/0± 64/8 گرم به‌طور تصادفی در ۱۵ مخزن پلی‌اتیلنی با حجم ۳۰۰ لیتر (۲۰ ماهی در هر مخزن) توزیع شده و به ‌مدت ۵۶ روز در شرایط آب دریا با شوری ۲/۴۰ گرم در لیتر پرورش یافتند. جیره‌های آزمایشی در ۵ تیمار شامل شاهد، ۲۵ درصد و ۵۰ درصد پودر حشره معمولی و ۲۵ درصد و ۵۰ درصد پودر حشره غنی‌شده با ۱۵ درصد اسپیرولینا (تیمارهای BSF25%، BSF50%، BSF+SP25% و BSF+SP50%) تنظیم گردیدند. نتایج نشان داد که بالاترین میزان وزن نهایی، درصد افزایش وزن و نرخ رشد ویژه در تیمار (BSF+SP25%) به‌دست آمد. در مقابل، کمترین میزان این شاخص‌های رشدی در سطوح ۵۰ درصد جایگزینی مشاهده گردید. در بررسی ترکیب لاشه، بالاترین درصد چربی در تیمار (BSF+SP50%) ثبت شد. همچنین بیشترین سطح کلسترول و تری‌گلیسرید پلاسما در تیمار (BSF+SP50%) و بالاترین میزان پروتئین کل در تیمارهای (BSF25%) و (BSF50%) مشاهده شد. نتایج تجزیه‌‌وتحلیل بیوشیمیایی شامل گلوکز، کلسترول و تری‌گلیسرید نشان داد که جایگزینی آرد ماهی تا سطح ۵۰ درصد، فاقد اثرات منفی بر سلامت ماهی بود. نتایج پژوهش حاضر، نشان داد که جایگزینی ۲۵ درصد آرد ماهی با پودر لارو غنی‌شده با اسپیرولینا، راهکاری مؤثر و پایدار برای بهبود عملکرد رشد و ارتقاء سلامت این گونه در صنعت آبزی‌پروری است. 2 Introduction Fishmeal (FM) is a cornerstone of aquafeed, but its sustainability and cost challenges necessitate the search for alternative protein sources. Black soldier fly larvae (BSFL, Hermetia illucens) meal has emerged as a promising substitute due to its high protein content and efficiency in converting organic waste into valuable biomass. However, its high fat content, particularly saturated lauric acid, and the presence of chitin can negatively impact nutrient digestibility and growth performance in fish. Recent studies highlight that the nutritional quality of insect larvae, especially their fatty acid profile, can be improved by altering their diet during rearing. Spirulina, a microalga rich in protein, vitamins, minerals, and essential fatty acids, is known to enhance the nutritional value of insect meals for aquaculture. Given that BSFL can bioaccumulate fatty acids from their growth substrate, enriching their diet with spirulina can significantly improve their nutritional value for aquaculture. This study aimed to investigate the effects of replacing fishmeal with conventional BSFL meal and spirulina-enriched BSFL meal on the growth performance, whole-body proximate composition, and plasma biochemical parameters of juvenile Asian seabass (Lates calcarifer). While previous research on various fish species has shown mixed results regarding insect meal inclusion, our study represents the first comprehensive investigation into the combined effects of BSFL meal and spirulina enrichment on Asian seabass. The findings highlight that 25% replacement with spirulina-enriched BSFL (+BSFSP25%) provides optimal growth performance while higher inclusion levels (50%) may have negative effects on growth and biochemical indices. Methodology The 56-day feeding trial was conducted using 300 juvenile Asian seabass with an average initial weight of approximately 8.6–8.7 g. The fish were randomly distributed into five experimental groups with three replicates each, housed in 300-liter tanks. All tanks were supplied with filtered saltwater and continuous aeration. Water quality parameters, including temperature (28–30°C), dissolved oxygen (5–7 mg/L), pH (7.5–8), and salinity (45 g/L), were monitored daily. Fish were fed to apparent satiation twice per day with five experimental diets: a control diet (FM) containing 100% fishmeal, and four diets where fishmeal was replaced by black soldier fly larvae meal (BSF) at 25% (BSF25%) and 50% (BSF50%) levels, and by spirulina-enriched BSFL meal at 25% (+BSFSP25%) and 50% (+BSFSP50%) levels. All diets were formulated to be isoproteic (~40% crude protein) and isolipidic (~9% crude lipid). At the end of the trial, samples were collected to measure growth performance indices, whole-body proximate composition, and plasma biochemical parameters. Growth performance was assessed using Final Body Weight (FBW), Weight Gain (WG), Specific Growth Rate (SGR), Feed Conversion Ratio (FCR), and Survival Rate (SR). Whole-body proximate analysis determined the content of protein, lipid, moisture, and ash. Blood samples from two fish per tank were used for plasma analysis of total protein, glucose, cholesterol, and triglycerides. All data were analyzed using one-way ANOVA followed by Duncan's test at p < 0.05. Results Dietary treatments significantly influenced the growth performance and feed utilization efficiency of the fish (p<0.05). The most pronounced growth outcomes, including final body weight (FBW; 57.4 ± 0.1 g), weight gain (WG; 572.7 ± 0.1%), and specific growth rate (SGR; 3.4 ± 0.0% day⁻¹), were achieved in the +BSFSP25% group. Conversely, the lowest growth metrics were observed in the BSF50% and +BSFSP50% cohorts, both of which exhibited a significant decline compared to the other treatments. While the BSF50% group recorded the highest feed conversion ratio (FCR), this difference remained statistically non-significant. Notably, the survival rate was markedly compromised in the BSF50% treatment compared to all other experimental groups. Proximate composition analysis of the whole body revealed that crude protein levels peaked in the BSF25% and BSF50% treatment groups. In contrast, the highest crude lipid content was recorded in the +BSFSP50% group. A significant reduction in moisture content was observed in the BSF50% and +BSFSP50% cohorts (p<0.05). Furthermore, ash content reached its maximum level in the +BSFSP50% group, whereas the control (FM) group exhibited the lowest ash values. Dietary treatments exerted a statistically significant influence on the plasma biochemical profiles of the fish (p<0.05). Total protein concentrations were markedly elevated in the BSF25% and BSF50% cohorts compared to the other experimental groups. Regarding glucose metabolism, the highest values were recorded in the +BSFSP50% group, while the BSF25% treatment yielded the lowest levels. Furthermore, a clear trend was observed in the lipid profiles, with cholesterol and triglyceride concentrations peaking in the +BSFSP50% and +BSFSP25% groups, respectively. Conversely, the most reduced cholesterol levels were found in the BSF50% group, whereas the BSF25% group exhibited the lowest triglyceride concentrations. Discussion The outcomes of the present study are congruent with prior research, highlighting that the partial substitution of fishmeal with black soldier fly larvae (BSFL) meal, especially when enriched with Spirulina platensis, exerts a substantial influence on the growth performance, carcass composition, and physiological status of Asian seabass. The superior growth metrics observed in the +BSFSP25% cohort underscore the efficacy of this hybrid dietary strategy. This synergistic improvement may be attributed to the well-balanced amino acid and fatty acid profiles of BSFL meal, augmented by the diverse bioactive compounds and potent antioxidative properties of spirulina. Such a nutritional combination likely optimizes metabolic pathways and enhances nutrient assimilation, thereby promoting superior growth. In conclusion, these findings suggest that integrating spirulina-enriched insect meal at a 25% replacement level offers a sustainable and high-performance alternative to conventional fishmeal in Asian seabass aquaculture. Conversely, the growth retardation observed at the 50% substitution level highlights that high dietary inclusion of BSFL meal may exceed the physiological tolerance of Asian seabass. This decline is potentially attributable to the presence of anti-nutritional factors (ANFs), specifically chitin, alongside an imbalanced essential amino acid profile. These findings corroborate that the optimal replacement threshold for fishmeal in this species remains below the 50% mark. Furthermore, the concomitant increase in carcass adiposity and hyperlipidemia observed in the +BSFSP50% group suggests a significant metabolic perturbation, necessitating a more precise calibration of the dietary lipid matrix in high-inclusion formulations. Conclusion In summary, this research offers a viable and sustainable framework for modern aquaculture. The strategic substitution of 25% of fishmeal with spirulina-enriched BSFL meal not only bolsters growth performance and optimizes carcass quality but also enhances the overall physiological resilience of Asian seabass. This dietary intervention represents a significant advancement in the development of eco-friendly and cost-effective aquafeeds, effectively mitigating the industry’s heavy reliance on finite marine-derived proteins. Ultimately, these findings facilitate the transition toward more circular and resource-efficient production systems, promoting the long-term sustainability of the global aquaculture sector. Conflict of interest The authors declare no conflict of interest. Acknowledgment The authors would like to thank the South of Iran Aquaculture Research Centre for providing the experimental facilities and technical support throughout the study. 13 23 2025/06/32025/09/21 1404/6/30 2026/07/12026/07/1 1405/4/10 کیانوش آبگون Kianoush Abgoun گروه شیلات، دانشکده منابع طبیعی دریا، دانشگاه علوم و فنون دریایی خرمشهر، خرمشهر، ایران kabgoon@yahoo.com ابراهیم رجب زاده قطرمی Ebrahim Rajabzadeh Ghatrami گروه شیلات، دانشکده منابع طبیعی دریا، دانشگاه علوم و فنون دریایی خرمشهر، خرمشهر، ایران rajabzadeh48@gmail.com منصور طرفی موزان زاده Mansour Torfi Mozanzadeh پژوهشکده آبزی‌پروری جنوب کشور، مؤسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، اهواز، ایران mansour.torfi@gmail.com ایمان سوری نژاد Iman Sourinezhad گروه شیلات، دانشکده علوم و فنون دریایی، دانشگاه هرمزگان، بندرعباس، ایران i_sourinezhad@yahoo.com Asian seabass (Lates calcarifer) Black soldier fly (Hermetia illucens) Fishmeal replacement Growth performance Sustainable aquaculture سی‌بس آسیایی حشره‌ سرباز سیاه جایگزینی آرد ماهی عملکرد رشد آبزی‌پروری پایدار Abdel-Tawwab, M., Khalil, R.H., Metwally, A.A., Shakweer, M.S., Khallaf, M.A. and Abdel-Latif, H.M., 2020. 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مقاله علمی – پژوهشی:‌ اثر زیست‌‌توده سیانوباکتری Anabaena sp. حاوی سم Microcystin-LR، رشد یافته در غلظت‌‌های مختلف فسفر بر زئوپلانکتون Daphnia magna Effect of biomass of Anabaena sp. containing the microcystin-LR, grown at different phosphorus concentrations on the zooplankton, Daphnia magna 1 1 سیانوباکتری‌های تولیدکننده سم به‌عنوان یکی از عوامل تهدیدکننده زنجیره‌های غذایی آبزیان شناخته می‌شوند. هدف از پژوهش حاضر، بررسی اثر حاد و مزمن زیست‌توده سیانوباکتریAnabaena sp.  حاوی سم Microcystin-LR، پرورش‌یافته در غلظت‌های مختلف فسفر، بر زئوپلانکتونDaphnia magna  بود. بدین منظور، سیانوباکتریAnabaena sp.  به مدت ۱۰ روز در معرض غلظت‌های مختلف فسفر شامل غلظت 1/7 میلی‌‌گرم بر لیتر به‌‌عنوان تیمار شاهد و غلظت‌‌های 74/7، 38/8 و 66/9 میلی‌‌گرم بر لیتر فسفر در دمای 28 درجه سانتی‌‌گراد و نور فلورسنت با شدت 35 میکرومول فوتون بر ثانیه کشت داده شد. نتایج نشان داد که میزان تولید سم Microcystin-LR به‌ طور معنی‌داری تحت تأثیر غلظت فسفر قرار دارد (05/0>p) به‌طوری‌که بیشترین مقدار سم در تیمار 38/8 میلی‌‌گرم بر لیتر فسفر با 5/300 پیکوگرم بر میلی‌لیتر و کمترین مقدار آن در تیمار شاهد با 41/33 پیکوگرم بر میلی‌لیتر اندازه‌گیری شد. ارزیابی سمیت حاد زیست‌توده سیانوباکتری بر زئوپلانکتونD. magna  نشان داد که کمترین مقدار LC₅₀ ۹۶ ساعته (بیشترین سمیت) در تیمار 38/8 میلی گرم بر لیتر فسفر مشاهده شد که 12/271 میلی‌گرم وزن خشک زیست‌توده بر لیتر بود و با سایر تیمارها اختلاف معنی‌‌دار داشت (05/0>p). در آزمایش سمیت مزمن نیز بیشترین میزان مرگ‌ومیر در تیمار با 38/8 میلی‌‌گرم در لیتر فسفر با میزان زیست‌‌توده خشک 100 میلی‌‌گرم بر لیتر مشاهده شد که 60 درصد بود. هیچ مرگ‌ومیری در تیمار شاهد مشاهده نشد. بیشترین تعداد نوزادان در تیمار شاهد و کمترین تعداد نوزاد متولد شده نیز مربوط به تیمار 38/8 میلی‌‌گرم بر لیتر فسفر بود که به‌‌ترتیب 21 و 10 عدد شمارش شدند. اولین زمان تولیدمثل برای تیمار شاهد روز 8 و برای سایر تیمارها روز 9 ثبت گردید. نتایج حاصل بیانگر آن است که افزایش غلظت فسفر محیط می‌تواند از طریق افزایش تولید Microcystin-LR، سمیت زیست‌توده سیانوباکتریAnabaena sp.  را تشدید کرده و تهدیدی جدی برای زئوپلانکتون‌ها و پایداری اکوسیستم‌های آبی ایجاد کند. 2 Introduction Eutrophication of freshwater ecosystems has led to a worldwide increase in the frequency of harmful cyanobacterial blooms (Paerl and Otten, 2013; Preece et al., 2017). Many cyanobacteria produce secondary metabolites known as cyanotoxins, among which microcystins are the most widespread and ecologically significant (Spoof and Catherine, 2017). Microcystin-LR (MC-LR) is recognized as the most toxic and prevalent microcystin variant in freshwater systems (Li et al., 2017). Species within the genus Anabaena are globally distributed and capable of producing microcystins under favorable environmental conditions (Welker and Döhren, 2006). Zooplankton—particularly cladocerans such as Daphnia magna—play a pivotal role in aquatic food webs and are widely regarded as sensitive indicators of cyanobacterial toxicity (Ferrao-Filho et al., 2008; Ger et al., 2016). Cyanotoxins are thought to act as chemical defenses against grazing, thereby constraining energy transfer from primary producers to higher trophic levels (Ferrao-Filho and Kozlowsky-Suzuki, 2011). In addition to their acute lethal effects, a growing body of evidence indicates that microcystins can induce a broad range of sublethal and chronic responses in zooplankton, including reduced feeding activity, impaired growth, delayed maturation, and diminished reproductive performance (Ferrao-Filho et al., 2000; Vilar et al., 2014). These chronic effects are ecologically significant because they may alter the population dynamics of primary consumers even at toxin concentrations below those causing immediate mortality (Ferrao-Filho and Kozlowsky-Suzuki, 2011). Moreover, prolonged exposure to cyanobacterial biomass can promote the accumulation of microcystins in zooplankton tissues, thereby increasing the potential for trophic transfer and biomagnification within aquatic food webs (Ferrao-Filho et al., 2014; Pham and Utsumi, 2018). Despite extensive research on cyanobacterial toxicity, relatively little is known about how phosphorus availability influences microcystin production and the resulting acute and chronic effects on zooplankton. Methodology Anabaena sp. was cultured under four phosphorus concentrations, including a control treatment corresponding to the basal BG-11 medium phosphorus (7.1 mg/L P) and three elevated phosphorus levels (7.74, 8.38 and 9.66 mg/L P). Cyanobacterial biomass was harvested, freeze-dried, and used for toxin quantification and toxicity experiments. Microcystin-LR concentrations were measured using an ELISA method following standard protocols (Fan et al., 2022). Acute toxicity tests were conducted using neonates (<24 h old) of D. magna exposed to different concentrations of freeze-dried cyanobacterial biomass (0–1000 mg dry weight L⁻¹) for 96 hours, and mortality was recorded periodically (Ferrao-Filho et al., 2014). Median lethal concentration (LC50) values were calculated using Probit analysis. Chronic toxicity tests were performed over a 15-day exposure period at lower biomass concentrations (12.5, 25, and 50 mg DW/L). Survival, time to first reproduction, and total offspring production were recorded according to established methods (Smutna et al., 2014; Herrera et al., 2015). At the end of the exposure period, microcystin-LR accumulation in D. magna tissues was quantified using ELISA. Results Phosphorus concentration significantly influenced microcystin-LR (MC-LR) production in Anabaena sp. cultured under different experimental treatments. ELISA analysis showed that MC-LR concentrations in freeze-dried cyanobacterial biomass ranged from 33.41 to 300.5 pg /mL. The highest MC-LR content was recorded in the treatment with 8.38 mg/L P, followed by 9.66 mg/L P, while the lowest toxin concentration was observed in the control treatment (7.1 mg/L P). Statistical analysis revealed significant differences among treatments (p < 0.05), indicating that phosphorus availability markedly affected toxin production. Acute toxicity assays demonstrated clear dose-dependent effects of cyanobacterial biomass on the survival of D. magna. Mortality increased with increasing biomass concentration in all treatments. Biomass derived from the 8.38 mg/L P treatment caused the highest mortality, reaching 100% at 1000 mg DW/L within 96 hours. At intermediate concentrations (500 and 250 mg DW/L), mortality rates of approximately 70% and 50%, respectively, were observed for the 8.38 mg/L P treatment. In contrast, the control treatment exhibited the lowest toxicity, with mortality rates of 40%, 20%, and 10% at biomass concentrations of 1000, 500, and 250 mg DW/L, respectively. The lowest 96-h LC50 value (271.12 mg DW/L) was observed for the 8.38 mg/L P treatment, whereas higher LC50 values were observed for the remaining treatments, indicating lower toxicity. Chronic toxicity experiments revealed significant lethal and sublethal effects of Anabaena sp. biomass on D. magna during the 15-day exposure period. Survival rates declined with increasing biomass concentration, particularly in treatments associated with higher phosphorus levels. The highest mortality rates were observed in the 8.38 and 9.66 mg/L P treatments at biomass concentrations of 50 and 100 mg DW/L. In contrast, no mortality was recorded in the control group without cyanobacterial biomass. Reproductive performance of D. magna was markedly affected by chronic exposure to cyanobacterial biomass. The total number of offspring produced over the experimental period decreased significantly with increasing biomass concentration in all treatments. The lowest cumulative number of neonates was recorded in the 8.38 mg/L P treatment, particularly at 100 mg DW/L, where reproduction was strongly suppressed. In contrast, the control group showed the highest reproductive output. Additionally, the time to first reproduction was delayed in all exposed groups compared to the control, with reproduction occurring one day later in treatments containing cyanobacterial biomass. Analysis of microcystin-LR accumulation in D. magna tissues indicated relatively low toxin concentrations at the end of the chronic exposure period. No detectable MC-LR was found in zooplankton exposed to 25 mg DW/L across all treatments. However, measurable toxin levels were detected at higher biomass concentrations, particularly in the 8.38 and 9.66 mg/L P treatments. The highest accumulation was observed at 100 mg DW/L, although concentrations remained low across all treatments. Discussion and conclusion The enhanced toxicity observed under phosphorus-enriched treatments can be attributed to increased microcystin-LR production and shifts in the biochemical composition of cyanobacterial biomass. Previous studies have shown that phosphorus can indirectly regulate microcystin production by promoting biomass accumulation and increasing cellular energy availability (Halstvedt et al., 2007; Wang et al., 2010). Phosphorus also plays a central role in ATP synthesis, providing the energy required for microcystin biosynthesis (Li et al., 2023). Chronic-exposure experiments revealed significant lethal and sublethal effects in D. magna, including reduced survival, delayed reproduction, and decreased offspring production. These responses are characteristic of chronic cyanobacterial toxicity and have been widely documented in cladocerans exposed to microcystins (Ferrao-Filho and Kozlowsky-Suzuki, 2011; Vilar et al., 2014). Although microcystin accumulation in D. magna tissues was relatively low overall, accumulation increased with prolonged exposure and under phosphorus-enriched conditions, consistent with previous reports of microcystin bioaccumulation and trophic transfer (Ferrao-Filho et al., 2014; Pham and Utsumi, 2018). Overall, these findings suggest that nutrient-driven cyanobacterial blooms may have long-term impacts on zooplankton populations through both direct toxicity and disruption of key life-history traits. In conclusion, cyanobacterial toxicity to D. magna appears to depend strongly on biomass dose and phosphorus-mediated toxin production, and chronic exposure may pose substantial ecological risks even at sublethal concentrations. Ultimately, these effects may compromise the population stability of primary consumers and reduce energy transfer in eutrophic freshwater ecosystems. Conflict of interest The authors declare no conflict of interest. Acknowledgement The authors gratefully acknowledge the Faculty of Natural Resources, Isfahan University of Technology, for providing laboratory facilities and technical support for this research. They also thank the faculty members and laboratory staff who assisted with the experimental work. 25 37 2025/06/32025/09/212026/02/12 1404/11/23 2026/07/12026/07/12026/07/1 1405/4/10 فاطمه رستمی Fatemeh Rostami گروه شیلات، دانشکده منابع طبیعی، دانشگاه صنعتی اصفهان، اصفهان، ایران rostami.1370@yahoo.com امیدوار فرهادیان Omidvar Farhadian گروه شیلات، دانشکده منابع طبیعی، دانشگاه صنعتی اصفهان، اصفهان، ایران omfarhad@iut.ac.ir نصراله محبوبی صوفیانی Nasrollah Mahboobi Soofiani گروه شیلات، دانشکده منابع طبیعی، دانشگاه صنعتی اصفهان، اصفهان، ایران soofiani@iut.ac.ir Anabaena sp. Daphnia magna Microcystin-LR Phosphorus Toxin Anabaena sp. Microcystin-LR سمیت Daphnia magna فسفر Apel, K. and Hirt, H., 2004. Reactive oxygen species: metabolism, oxidative stress, and signal transduction. 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مقاله علمی – پژوهشی:‌ تاثیر سطوح مختلف آستاگزانتین جیره بر شاخص‌های رسیدگی جنسی مولدین فیل‌ماهی (Huso huso) Different dietary levels of astaxanthin on growth indices, sex hormones and sexual maturity indices in pre-broodstock beluga (Huso huso) 1 1 اثر مکمل آستاگزانتین جیره بر شاخص‌های رشد، گناد و شاخص‌های تولید‌مثلی در فیل‌ماهی ماده پرورشی مورد بررسی قرار گرفت. تعداد 15 عدد فیل‌ماهی ماده در سن 32 -28 ماه در مرحله رسیدگی جنسی II، با متوسط وزن 16/0±16/16 کیلوگرم و طول 2/13±65/ 140سانتی‌متر با جیره‌های حاوی 50 درصد پروتئین، 14 درصد چربی و 18 مگاژول بر کیلوگرم انرژی: AST (فاقد آستاگزانتین و 5/8 درصد روغن ماهی)‌ AST 50، AST 100، AST 150 ، AST 200 (50، 100، 150 و 200 میلی‌گرم در کیلوگرم آستاگزانتین) به مدت 615 روز تغذیه شدند. در پایان دوره تغذیه فیل‌ماهیان تغذیه‌شده با جیره AST200 دارای بیشترین وزن نهایی (10/2±60/45 کیلوگرم)، درصد افزایش وزن (6/32±46/183 درصد)، رشد روزانه (04/5±49/45 گرم) و ضریب رشد ویژه (01/0±16/0 درصد/ روز) بودند (05/0p<). نمونه‌برداری برای وضعیت گناد و تغییرات هورمون‌های جنسی در چهار مرحله انجام شد. در آخرین مرحله نمونه‌برداری میزان ترشح هورمون استرادیول در مولدین تغذیه‌شده با جیره‌های حاوی آستاگزانتین 50، 100 و 200 میلی‌‌گرم در کیلوگرم به طور معنی‌داری افزایش یافت و میزان تستوسترون سرم مولدین تغذیه‌شده با جیره دارای 200 میلی‌‌گرم در کیلوگرم آستاگزانتین در مقایسه با مولدین تیمار شاهد، بیشتر بود، اما میزان آلفا هیدروکسی پروژسترون در مولدین تغذیه‌شده با جیره 200 میلی‌گرم در کیلوگرم آستاگزانتین به ‌طور معنی‌داری کمتر از مولدین تیمار شاهد بود (05/0p<). نتایج به‌دست آمده از مشاهدات لاپاراسکوپی از گناد ماهیان نشان دادکه مولدین تغذیه‌شده با جیره‌های حاوی 200 میلی‌گرم آستاگزانتین در مرحله ورود به مرحله IV بوده و اووسیت‌ها و تخم‌های نارس در بافت تخمدان قابل مشاهده بود. نتایج این آزمایش نشان داد که افزودن مکمل آستاگزانتین به میزان 200 میلی‌گرم در کیلوگرم موجب افزایش ترشح هورمون‌های رسیدگی جنسی، رشد گناد و افزایش ترشح هورمون‌های جنسی در گونه فیل‌ماهی می‌شود. در شرایط عادی، تغییر از مرحله رسیدگی جنسی II بهIV  یا 6-5 سال به‌طول می‌انجامد درحالی‌که در این آزمایش این مرحله به 33 ماه کاهش یافت. 2 Introduction Prolonged sexual maturation is a significant challenge in Huso huso aquaculture for both caviar and fry production. The most extended period of sexual maturation occurs during the II-III and III-IV stages (7–8 years), which are associated with substantial stagnation in capital investment. The decrease of the sexual maturation period and quality of gametes is greatly influenced by vitellogenesis quality that vitellogenesis is depended on time secretion of hormone 17 beta-estradiol. Under normal conditions, after temperature and photoperiod changes, hypothalamus secretes hypothalamic releasing hormone (GnRH), which releases gonadotropin hormones GTH1 and GTH2. GTH1 in the ovary triggers follicle cell receptors to produce 17-beta-estradiol, (the main sex steroid). 17-beta-estradiol diffuses across the membrane of liver cells, attaches to estrogen receptors, activates vitellogenin transcription and translation, and allows that fish to enter stage IV sexual development. Astaxanthin, a small molecule containing hydroxyl and unsaturated ketone groups, is a 17β-estradiol secretion stimulator, steroidogenesis accelerator, estrogen and progesterone secretion enhancer that lead to a sexual maturity period reduction in fish. Few studies have been undertaken to determine the effect of astaxanthin on gonadal growth and sexual maturation indicators in sturgeon. This study conducted on growth indices, hormone secretion, and sexual maturity in sturgeon prebroodstocks at sexual maturation stage II fed by dietary astaxanthin. Methodology Astaxanthin was derived from Carophyll® pink 10% (DSM-Bright Science, BrighterLiving™, Netherlands; CAS No.: 7542-45-2) provided by Abzico Company, Yasuj, Kohlakoyeh and Boyer-Ahmad Province, Iran. The experimental diets were formulated based on the nutrition requirements of Siberian sturgeon broodstock and available references (protein was 50%, lipid 1 15%, and energy was 18 mj/ kg dry matter in Excel Solver program. AST 50, AST 100, AST 150, and AST 200 diets were created by supplementing 50, 100, 150, and 200 mg/kg astaxanthin, respectively.  Twenty-seven pre-broodstock fish with average weight and length of 16.16±0.16 and 140.65±13.2 cm at sexual maturity II were selected. The fish were fed 0.5% of their body weight for 645 days. the fish Gonadal development was examined with a monitor model (LG, 19M38HB, Korae) by making a longitudinal incision in the abdomen. sexual maturation hormones levels (testosterone, 17 beta estradiol and 17 alpha hydroxyprogesterone) were determined by blood sampling collected from caudal vein. Dry matter was measured by drying the sample at 105°C until constant weight was reached; crude protein was measured using the Kjeldahl method in three stages of digestion, distillation, titration, and multiplying the nitrogen obtained from each gram of dry matter by 6.25; ash was measured by burning the sample in an electric furnace (Muffle Furances, RHF 16/3/3216 P1 Model, England, Plymouth) at 550°C.Crude fat were measured by Soxhlet lipid extraction using ether solvent reaching a boiling point of 50-60°C for 4-6 hours in a Soxhlet extractor (Gerhart Soxthoterm SOX Hamburg, Hamburg, German), and total energy were measured using a bomb calorimeter (Calorimeteradiabatic C-400 IKA, Heiterbeini, GMBIL, Brussels, Belgium) (AOAC, 1995).17 alpha hydroxyprogesterone was determined in ng/gr using the Monobind ELISA kit, 17 beta estradiol and testosterone levels were determined using kits (Immunotech, France) and the II25 detector via radioimmunoassay (RIA) using a GammaCounter (LKB, France) in ng/ml using the modified method of Cattaldi et al. (1998). Results Fish fed the C200 diet exhibited the highest final weight, body weight gain percentage, and specific growth rate, along with the lowest feed conversion ratio (p<0.05). There were no significant variations in estradiol, testosterone, and alpha-hydroxyprogesterone levels in fish during the first biometric period following the introduction of pre-broodstock to the pond (sexual maturation II) (p>0.05). In second biometry, there was no significant difference in estradiol and testosterone hormones secretion after 10 months of feeding. Fish fed the AST0 had an estradiol level of around 0.3±0.05 ng/ml. However, fish fed the AST 50, AST 100, and AST 150 had levels of 0.27±0.01, 0.25±0.03, and 0.25±0.01 ng/ml, respectively (p>0.05). Fish fed AST 200 had higher levels (0.35±0.04 ng/ml). The testosterone level in fish fed AST 0 was 0.27±0.04 ng/ml. Testosterone levels decreased in fish fed AST 50, AST 100, and AST 150 (0.17±0.00, 0.18±0.006, 0.18±0.01 ng/ml), whereas increased in fish fed AST 200 compared to AST 0 (p>0.05). There was no significant difference in the amount of 17 alpha-hydroxyprogesterone between treatments, however the highest amount was seen in the blood serum of fish fed a AST 200. In third biometry, Broodstock fed by AST 200 had the highest testosterone levels (0.71±0.19 ng/mg), significantly higher than other treatments (p<0.05). Fish fed with AST150 and AST200 showed the highest amounts of testosterone (0.78 ± 0.4 and 0.71 ± 0.19 ng/mg, respectively) (p<0.05). also, Broodstock fed AST100 and AST200 showed highest 17-alpha-ydroxyprogesterone (42±0.08 and 43±0.08 ng/ml, respectively) compared to fish fed AST0 dietary (p<0.05). But, Estradiol hormone secretion in fish fed with AST 50, AST 100, and AST 200 had a significant increase at last sampling stage and reached to 0.76±0.092 ng/ml (p<0.05). The testosterone level in fish fed a C0 was 1.63±0.24 ng/ml. No significant difference in testosterone secretion was seen in broodstock fed with C0 compared to AST 50, C100, and AST 150. However, broodstock serum testosterone fed with C200 increased and reached 7.5±1.1 ng/mL. Fish fed with AST 50, AST 100, and AST 150 had significantly lower levels of alpha-ydroxyprogesterone compare to AST 0. Fish fed with C200 also had lower levels of alpha-hydroxyprogesterone in their serum compared to fish fed AST 0 diet (p<0.05). On the other hand, The results obtained from fish laparoscopic observations showed that the broodstock fed a diet containing C200 were entering stage IV and immature oocytes and eggs were visible in the ovarian tissue, while the broodstock fed a diet lacking astaxanthin as well as the broodstock of AST 50, AST 100 and AST 150 treatments were stopped at stage III sexual maturation. Conclusion and discussion  In agreement with our result, Xie et al. (2020) showed that astaxanthin supplementation at levels of 75 and 100 mg/kg improved growth parameters in largemouth bass (Micropterus salmoides). Similarly, astaxanthin supplementation in the diets of white leg shrimp (Litopenaeus vannamei), pacu (Piaractus mesopotamicus) (Bacchetta et al., 2019), and tiger shrimp (Penaeus monodon) (Niu et al., 2015) increased growth and decreased feed conversion ratio. According to Zhang et al. (2013b), carotenoid pigments improve nutritional digestion, absorption, and feed utilization by enhancing digestive enzyme activity and increasing feed intake. Carotenoids have sexual maturation-stimulating properties that can induce vitellogenesisy the of sex steroids synthesis increasing (17β-estradiol) and quicker oocyte maturation accelerate in rainbow trout (Örn et al., 2003). Fish that were provided the C200 diet showed higher levels of estradiol and 17α-hydroxyprogesterone compared to those fed the AST0 diet after 16 months of feeding. Laparoscopic observations confirmed that the fish fed AST200 had passed the III stage of sexual maturation and vitellogenesis had occurred. Similar results were reported in the bull tongue shoe (Cynoglossus semilaevis) (Xu et al., 2017), the large freshwater shrimp (Macrobrachium rosenbergii) (Tao et al., 2025). In this study, broodstock fed with AST200 showed a rapid decrease in 17α-hydroxyprogesterone levels as they near approach of stage IV. During oocyte maturation, HSDH-20β converts 17α-hydroxyprogesterone in granulosa cells to 17α-20β-hydroxy progesterone, it seems be, 17α-hydroxyprogesterone decreasing in C200 treated broodstock may be due to oocyte maturation. At this stage, follicles are poised to produce steroids in order to stimulate ultimate maturation. Conflict of interest There is no conflict of interest between authors Acknowledgment This article is derived from the project "Investigation of the effect of dietary astaxanthin and HUFA fatty acids supplementation on gonads and reproductive indices in farmed female Huso huso" with code 990585-98001-007-12-32-12. The authors express their gratitude to all individuals who provided scientific and practical supports. 39 60 2025/06/32025/09/212026/02/122025/10/1 1404/7/9 2026/07/12026/07/12026/07/12026/07/1 1405/4/10 میرحامد سید حسنی Mir Hamed SDayed Hassani انستیتو تحقیقات بین المللی تاسماهیان دریای خزر، موسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، رشت، ایران mirhamedhassani@yahoo.com ایوب یوسفی جوردهی Ayoub Yousefi Jourdehi انستیتو تحقیقات بین المللی تاسماهیان دریای خزر، موسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، رشت، ایران Ayoub2222002@yahoo.com علی حلاجیان Ali Hallajian انستیتو تحقیقات بین المللی تاسماهیان دریای خزر، موسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، رشت، ایران alihallajian@gmail.com محمود محسنی Mahmoud Mohseni انستیتو تحقیقات بین المللی تاسماهیان دریای خزر، موسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، رشت، ایران mahmoudmohseni73@gmail.com تورج سهرابی Tooraj Sohrabi انستیتو تحقیقات بین المللی تاسماهیان دریای خزر، موسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، رشت، ایران tsohrabi@gmail.com هوشنگ یگانه Hooshang Yeganeh انستیتو تحقیقات بین المللی تاسماهیان دریای خزر، موسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، رشت، ایران سجاد قاسمیان Sajjad Ghasemian انستیتو تحقیقات بین المللی تاسماهیان دریای خزر، موسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، رشت، ایران فاطمه فداکار ماسوله Fatemeh Fadakar Masouleh انستیتو تحقیقات بین المللی تاسماهیان دریای خزر، موسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، رشت، ایران Key word: Huso huso Astaxanthin Sexual maturation فیل‌ماهی آستاگزانتین شاخص‌های رشد هورمون‌های جنسی رسیدگی جنسی Abdel-Ghani, M. 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مقاله علمی – پژوهشی:‌ تأثیر جایگزینی بخشی از آرد ماهی با آرد حشرات (کرم سوسک زرد و شفیره کرم ابریشم) در جیره غذایی بر شاخص‌های رشد و بیان ژن‌های مرتبط با رشد تاس‌ماهی سیبری جوان Effects of dietary partial substitute of insect meal (meal worm larvae and silk worm pupa) for fish meal on growth and related-gene expression of Siberian sturgeon 1 1 مطالعه حاضر با هدف ارزیابی تأثیر جایگزینی بخشی از آرد ماهی با آرد کرم سوسک زرد (Tenebrio moliter)، آرد شفیره کرم ابریشم (Bombyx mori) و ترکیب آرد این دو حشره در جیره غذایی تاس‌ماهیان سیبری (Acipenser baerii) بر عملکرد رشد و بیان ژن‌های مرتبط با رشد و متابولیسم انجام شد. آزمایش تغذیه‌ای با تهیه 1 جیره شاهد و 3 تیمار با جیره آزمایشی شامل آرد ماهی بدون افزودن پودر حشره (جیره غذایی شاهد) و سه جیره غذایی با جایگزینی ۱۵ درصد آرد ماهی با آرد کرم سوسک زرد (تیمار 1)، 15 درصد آرد شفیره کرم ابریشم (تیمار 2) یا ترکیبی از 5/7 درصد کرم سوسک زرد و 5/7 درصد شفیره کرم ابریشم (تیمار 3)، انجام شد. نتایج تحقیق نشان داد که جیره‌‌های دارای منبع پروتئین آرد حشرات به‌ویژه جیره غذایی ترکیبی (تیمار 3)، سبب افزایش معنی‌دار در وزن نهایی، میزان افزایش وزن، نرخ رشد ویژه و بهره‌وری پروتئین شدند و ضریب تبدیل غذایی را نسبت به جیره غذایی مبتنی بر آرد ماهی بهبود دادند. تحلیل بیان ژن نشان داد که تیمارهای حاوی حشرات موجب افزایش قابل‌توجه بیان Ghrelin (تنظیم اشتها و تعادل انرژی)، TOR  (تنظیم سنتز پروتئین و رشد سلولی) و IGF1 (رشد سیستمیک) شدند. همچنین افزایش بیان ژن ApoE  در تیمارهای 2 و 4 بیانگر بهبود در انتقال و متابولیسم لیپیدها بود. به ‌طور کلی، نتایج این پژوهش نشان داد که آرد حشرات می‌تواند همزمان رشد و متابولیسم را بهبود دهد و جایگزینی مؤثر و پایدار برای آرد ماهی در تغذیه تاس‌‌ماهی سیبری باشد. بنابراین، استفاده از منابع پروتئینی حشرات می‌تواند راهکاری کاربردی برای توسعه پایدار صنعت پرورش تاس‌ماهی سیبری باشد. 2 Introduction Sustainable aquaculture requires reliable, high-quality protein sources. Fishmeal, widely used in sturgeon diets, faces challenges such as limited supply, high cost, and environmental impact (Alfiko et al., 2022; Majluf et al., 2024). Insects, including yellow mealworm larvae (Tenebrio molitor) and silkworm pupae (Bombyx mori), are rich in protein (42–63%), essential amino acids, lipids, and micronutrients, and can be reared on organic waste (Alfiko et al., 2022; p). In Iran, sturgeon aquaculture has developed significantly, and finding suitable alternatives to fishmeal is of particular importance (Hosseinzadeh Sahafi, 2024, In Persian). This study aimed to evaluate the effects of partial replacement (15%) of fishmeal with insect meals, alone or in combination, on growth performance and the expression of genes related to growth, metabolism, and immunity in Siberian sturgeon (Acipenser baerii). Methodology Preparation of experimental diets and rearing conditions: Yellow mealworm larvae and silkworm pupae were ground and used to prepare three isonitrogenous and isolipidic experimental diets. Diet formulation was based on the nutritional requirements of Siberian sturgeon (Caimi et al., 2020). Treatments included control (fishmeal only), Treatment 1 (15% mealworm larvae), Treatment 2 (15% silkworm pupae), and Treatment 3 (7.5% mealworm larvae + 7.5% silkworm pupae). Five-month-old Siberian sturgeon (initial weight: 250.46 ± 76.59 g) were acclimated for two weeks and randomly distributed into 400 L fiberglass tanks. The feeding trial lasted 90 days at 24 ± 2.1°C with a natural photoperiod. Fish were fed by hand to apparent satiation three times daily (08:00, 14:00, 20:00), six days per week. Uneaten feed was collected daily, dried, and weighed (Caimi et al., 2020; Sheikh Veisi et al., 2022). Fish were fasted for 24 hours and weighed every two weeks (Bagheri et al., 2025). Growth performance: Growth indices including weight gain (WG), specific growth rate (SGR), feed conversion ratio (FCR), condition factor (CF), and protein efficiency ratio (PER) were calculated using standard formulas (Caimi et al., 2020). Gene expression: Tissue samples from brain, liver, and intestine were collected and stored at -80°C. Gene expression was evaluated for GH, IGF1, ghrelin (growth and appetite), TOR (protein metabolism), and ApoE (lipid metabolism) using RT-qPCR. NCBI accession numbers for all primers are presented in Table 2. Statistical analysis: Data were analyzed using SPSS software (version 9.5.1.733). One-way ANOVA followed by Tukey's test was used for group comparisons. Results are presented as mean ± SD (n=4). Results Growth performance: Treatment 1 (15% mealworm larvae) achieved significantly higher final weight (967.25 g) and weight gain (636.25 g) compared to other treatments, while the control group showed the lowest values (828 g and 498 g, respectively) (p≤0.05). PER in Treatment 1 (1.6) was significantly higher than other treatments. FCR in Treatments 1, 2, and 3 was similar (1.1), while the control group showed a higher FCR (1.21). Experimental diets had no significant effect on condition factor (p>0.05). Gene expression: GH expression in brain increased in Treatments 1 and 2, while Treatment 3 showed the most significant increase (p≤0.05). IGF1 expression in liver significantly increased in Treatment 2, followed by Treatment 3 (p≤0.05). Intestinal ghrelin expression significantly increased in Treatment 3 (p≤0.05). ApoE expression in liver increased in Treatment 1, followed by Treatment 3 (p≤0.05). TOR expression significantly increased in all insect-based treatments, with the highest level in Treatment 3 (p≤0.05). The lowest expression levels for all genes belonged to the control group. Discussion and conclusion The results demonstrated that partial replacement of fishmeal with insect meals improves growth performance and activates molecular pathways related to growth, metabolism, and immunity in Siberian sturgeon. The significant increase in growth indices and FCR improvement in insect-based diets indicated that insect proteins enhance feed efficiency and nutrient conversion. These findings are consistent with previous studies showing that 15–30% replacement of fishmeal with insect meal improves growth without negative effects on sturgeon health (Zhu et al., 2011; Caimi et al., 2020; Rawski et al., 2020). The lack of significant differences in CF among treatments suggests that insect proteins do not alter body morphology (Jackson et al., 2002). The marked increase in ghrelin expression in Treatment 3 indicates a strong effect of the combined insect diet on appetite regulation and energy balance. Ghrelin plays a key role in stimulating feed intake and improving metabolic efficiency (Zarantoniello et al., 2021). The significant increase in TOR expression in all insect-based treatments indicates activation of anabolic pathways and increased protein synthesis capacity, consistent with reports that insect meal can stimulate the TOR pathway in fish (Lanes et al., 2021). Increased IGF1 expression in Treatments 2 and 3 indicates stimulation of the GH-IGF growth axis. Increased ApoE expression in Treatments 1 and 3 indicates improved lipid transport and energy metabolism, consistent with the presence of bioactive fatty acids in insect meals (Weththasinghe et al., 2022). The combination of mealworm larvae and silkworm pupae created a synergistic effect by providing complementary nutritional profiles. The results demonstrate that 15% replacement of fishmeal with insect protein sources, particularly the combination of yellow mealworm larvae and silkworm pupae, enhances growth, lipid metabolism, and protein metabolism pathways in Siberian sturgeon. Activation of the ghrelin-TOR-IGF1 pathway and increased ApoE expression provide clear evidence of improved anabolism and nutritional efficiency. Insect protein can serve as a sustainable alternative to fishmeal while improving growth performance and molecular health in sturgeon. Therefore, insect meal is a suitable option for optimizing commercial feed formulation in sturgeon aquaculture. Conflict of interest The authors declare no conflict of interest. Acknowledgements This study was supported by the Iran National Science Foundation (INSF) under Grant No. 4027280. The authors thank the Iranian Fisheries Science Research Institute and the Inland Water Aquatic Resources Research Center (Gorgan) for their support. 61 73 2025/06/32025/09/212026/02/122025/10/12025/12/30 1404/10/9 2026/07/12026/07/12026/07/12026/07/12026/04/12 1405/1/23 طاهره باقری Tahereh Bagheri مرکز تحقیقات ذخایر آبزیان آبهای داخلی، موسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، ترویج و آموزش کشاورزی، گرگان، ایران Bagheri1360@gmail.com محمود حافظیه Mahmoud Hafezieh موسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، تهران، ایران jhafezieh@yahoo.com عیسی شریف پور Issa Sharifpour موسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، تهران، ایران isharifpour@yahoo.com همایون حسین زاده صحافی Homayoun Hossein Zadeh Sahafi موسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، تهران، ایران H_Hosseinzadeh@yahoo.com محمود محسنی Mahmoud Mohseni انستیتو تحقیقات بین المللی ماهیان خاویاری، موسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، رشت، ایران mahmoudmohseni73@gmail.com منصور شریفیان Mansour Sharifian موسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، تهران، ایران sharif_23m@yahoo.com شهره مسائلی Shohre Masaeli موسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، تهران، ایران Shohre.masaeli@yahoo.com رامین عبدلی Ramin Abdoli مرکز تحقیقات ابریشم کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، رشت، ایران ramin.abdoli.ramin.abdoli@gmail.com اسمعیل پقه Esmaeil Pagheh مرکز تحقیقات ذخایر آبزیان آبهای داخلی، موسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، ترویج و آموزش کشاورزی، گرگان، ایران esmaeilpaghe@gmail.com Siberian sturgeon Fish meal Insect meal Replacement Gene expression Growth performance تاس ماهی سیبری آرد ماهی آرد حشرات جایگزینی بیان ژن رشد Alfiko, Y., Xie, D., Astuti, R.T., Wong, J. and Wang, L., 2022. 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مقاله علمی – پژوهشی:‌ اثرات پوشش ترکیبی آلژینات و صمغ دانه شاهی (Lepidium sativum) همراه با پروتئین هیدرولیز شده دانه چیا (Salvia hispanica) بر ویژگی‌های کیفی، شیمیایی و حسی ناگت ماهی کپور نقره‌ای (Hypophthalmichthys molitrix) Effect of a combined coating of alginate and watercress seed gum with hydrolyzed chia seed protein on the qualitative, chemical and sensory properties of silver carp (Hypophthalmichthys molitrix) nuggets 1 1 در مطالعه حاضر، اثرات پوشش ترکیبی آلژینات، صمغ دانه شاهی(Lepidium sativum) به همراه پپتیدهای زیست فعال حاصل هیدرولیز آنزیمی از دانه چیا (Salvia hispanica) بر ویژگی‌های کیفی و فساد اکسیداسیونی ناگت ماهی کپور نقره‌ای (Hypophthalmichthys) مورد بررسی قرار گرفت . بدین منظور، خواص آنتی اکسیدانی پروتئین هیدرولیز شده دانه چیا با استفاده از آنزیم‌های پروتئاز میکرو Alcalase و Flavourzyme تعیین شد. سپس تاثیر غلظت‌های مختلف پروتئین هیدرولیز شده (0/5 و1 درصد) به همراه پوشش ترکیبی آلژینات، صمغ دانه شاهی بر ویژگی‌های فیزیکوشیمیایی و حسی ناگت ماهی در ابتدای دوره نگهداری و ویژگی‌های شیمیایی شامل پراکسید (PV)، مقادیر کلی بازهای نیتروژنی فرار (TVB-N) و تیوباربیتوریک اسید ((TBA طی دوره نگهداری 16روزه در یخچال مورد بررسی قرار گرفت.نتایج مربوط به ویژگی‌های پروتئین هیدرولیز شده نشان داد که آنزیم Alcalase می‌تواند پروتئین هیدرولیزی با درجه هیدرولیز(19/22 درصد)، محتوای پروتئینی(70/93 درصد) و بازیافت پروتئینی و خاصیت آنتی‌اکسیدانی بالاتری(02/82 درصد) تولید کند و افزایش زمان هیدرولیز تاثیر مثبتی بر ویژگی‌های مذکور داشت (05/0>p). نتایج مربوط به ویژگی‌های کیفی ناگت نشان داد که افزودن آلژینات، صمغ دانه شاهی و پروتئین هیدرولیز شده سبب کاهش جذب روغن (15/12 درصد) و افزایش رطوبت، درصد پوشش دهی و راندمان سرخ کردن و نرمی بافت (41/7 درصد) ناگت ماهی سرخ‌شده، شد. همچنین پوشش آلژینات، صمغ دانه شاهی و پروتئین هیدرولیز شده روند فساد اکسیداسیونی را در ناگت به طور معنی‌داری به تعویق انداخت و عمر ماندگاری ناگت را افزایش داد (05/0>p) و با افزایش غلظت پروتئین هیدرولیز شده، نتایج بهتری مشاهده شد (05/0>p).به طورکلی، می‌توان نتیجه گرفت که استفاده از پوشش ترکیبی آلژینات (2 درصد) ، صمغ دانه شاهی‌(‌1 درصد) به همراه پروتئین هیدرولیز شده دانه چیا در سطح 1 درصد (هیدرولیز شده با آنزیم آلکالاز در زمان 30 دقیقه)، نقش به‌سزایی در بهبود ویژگی‌های کیفی ناگت سرخ‌شده و افزایش ماندگاری آن طی دوره نگهداری دارد. 2 Introduction Silver carp (Hypophthalmichthys molitrix), owing to its easy accessibility and lower cost compared to marine fish, presents economic opportunities for developing by-products (Ojagh et al., 2018). The introduction of fish nuggets, with their desirable flavor and taste, can stimulate consumer demand for aquatic products. Ready-to-eat and semi-prepared foods hold a significant market share, valued for their crispy texture, attractive appearance, color, and enhanced flavor. Deep-frying is a common food processing technique where food is cooked in hot oil at temperatures exceeding the boiling point of water (Chen et al., 2009). This method aims for rapid food processing to extend shelf life while simultaneously creating unique textural, color, aroma, and flavor attributes. Strategies to minimize oil absorption during deep-frying include coating with hydrocolloids (such as edible coatings and gums) and the utilization of bioactive peptides from plant sources. Chia seeds (Salvia hispanica) are recognized for their nutritional richness, including high protein content (15–23%), antioxidants, dietary fiber, vitamins, and minerals. Notably, their oil is abundant in alpha-linolenic acid (Omega-3). Their unique rheological properties and ease of extraction have led to their application as thickening and gelling agents in the food industry, with products formulated using chia seed gum achieving positive consumer acceptance. The text also references Lepidium sativum (garden cress) concerning its oil content and thickening capabilities; however, clarification on the specific scientific name employed in the study may be beneficial. Enzymatic protein hydrolysis, employing either endogenous enzymes (autolysis process) or commercial enzymes, is utilized to improve protein characteristics and enhance bioactivity (Karazhiyan et al., 2009). This study specifically utilized Alcalase (a protease from Bacillus licheniformis with an activity of 4.2 AU/mL) and Flavourzyme (an exopeptidase with an activity of 1.5 AU/mL) (Oveysipour et al., 2012). The primary objective of this research was to assess the antioxidant properties of chia seed protein hydrolysates generated by these enzymes. Furthermore, the study aimed to investigate the impact of a composite coating, comprising alginate and garden cress gum, on the physicochemical and sensory properties of fish nuggets. This research endeavors to leverage these components to enhance the quality and appeal of fish nugget products. Methodology Preparation of chia seeds and (Alcalase enzyme - Flowerzyme) extracted from Novazyme, Denmark. The protein content of chia seeds was measured by the Kjeldahl method (Villanueva-Lazo et al., 2022). Preparation of fat-free flour and protein isolate from chia seeds was carried out using the method of Zhou et al. (2006). To measure the hydrolysis of chia seed protein isolate in the present study, 6 treatments were selected. Treatment 1: Flowerzyme + 10 minutes of hydrolysis time. Treatment 2: Flowerzyme + 20 minutes of hydrolysis time. Treatment 3: Flowerzyme + 30 minutes of hydrolysis time. Treatment 4: Alcalase + 10 minutes of hydrolysis time. Treatment 5: Alcalase + 20 minutes of hydrolysis time. Treatment 6: Alcalase + 30 minutes of hydrolysis time. The degree of hydrolysis was calculated using the method of (Villanueva-Lazo et al., 2022). The approximate length of the peptides resulting from hydrolysis was measured (Chatterjee et al., 2015). The protein recovery rate was determined by the Biuret method and the antioxidant activity was determined by the DPPH method (He et al., 2013). Extraction of (Lepidium sativum) seed gum was performed using the method of Karazhiyan et al. (2011). Preparation of wheat flour coating with different percentages by the method (Chen et al., 2009). Fish nuggets (Hypophthalmichthys molitrix) were prepared and physical, chemical, textural and sensory tests were performed (Asadi Farsani, 2018). The moisture content and the amount of absorbed oil were measured using the method (Avak and Glaser, 2005). The texture measurement test was calculated using the Instron texture measurement device according to the method of (Das et al., 2008). The coating percentage and frying efficiency of fish nuggets were calculated according to the formula (Darai Garmkhanei et al., 2009). Peroxide value was measured using the method of (Ronald and Ronald, 1991). (TBA) was measured using a colorimetric method (Hedayatifard and Miri, 2017). The (TVB-N) index was determined by the Kjeldahl method (Avak and Glaser, 2005). Sensory evaluation was investigated using the hedonic test method (Asadi Farsani, 2018). Results The initial protein measurements of chia seeds and its protein isolate were (23.91±1.00) and (40.82±1.24), respectively. The results showed that the degree of hydrolysis of the alkalase enzyme was higher than that of flowerzyme (p<0.05). The measurement of protein recovery values showed that the alkalase enzyme was higher than that of flowerzyme (p<0.05). The results of determining the peptide chain length by alkalase were lower than that of flowerzyme (p<0.05). The results of determining the protein levels in different treatments showed that the alkalase enzyme was higher than that of flowerzyme (p<0.05). The results of the DPPH free radical scavenging activity in different treatments showed that the highest activity was related to the alkalase enzyme (82.02%). The results showed that the lowest oil absorption rate was observed in the alginate + gum + 1% hydrolyzed protein (12.15%) treatment (p<0.05). The highest moisture absorption was observed in the treatments of alginate + gum + hydrolyzed protein 0.5 and 1% (56.84 and 57.45 percent, respectively) (p<0.05). The lowest tissue stiffness was observed in the treatment of alginate + gum + 1% hydrolyzed protein (41.7%) (p<0.05). The highest coverage was observed in the treatments of alginate + gum + hydrolyzed protein 0.5 and 1% (36.05 and 36.54%, respectively) (p<0.05). The highest frying efficiency values were observed in the treatments of alginate + gum + hydrolyzed protein with concentrations of 0.5 and 1% (78.93 and 79.24 percent, respectively) (p<0.05). The results of peroxide value under the influence of time and treatment showed that the highest values were observed in the control treatment and the lowest values were observed in the alginate + gum + 1% hydrolyzed protein treatment (p<0.05). The results of statistical analysis of thiobarbituric acid values during the storage period showed that the highest values were observed in the control treatment and the lowest values were observed in the alginate + gum + 1% hydrolyzed protein treatment (p<0.05). The lowest values of volatile nitrogen bases were observed in the alginate + gum + hydrolyzed protein treatment (0.5 and 1 percent) (p<0.05). The results of sensory evaluations showed that the sensory score of the alginate + gum + hydrolyzed protein treatment was 0.5 and 1 percent lower than the other treatments, respectively (p<0.05). In general, the results showed that the alginate and watercress seed gum coating has antioxidant properties and hydrolyzed chia seed protein increased its antioxidant properties, so that the alginate, watercress seed gum and hydrolyzed protein coating significantly delayed the oxidative spoilage process in the nugget and increased the shelf life of the nugget, and in all tests it had a better effect than the synthetic BHA preservative. Therefore, the combination of alginate (2%), watercress seed gum (1%) and hydrolyzed chia seed protein at the level (1% hydrolyzed) with the alcalase enzyme for (30 minutes) can play a significant role in improving the quality characteristics of fried nuggets and increasing their shelf life during storage, and also meet the consumer need for better and safer seafood products. Conclusions The protein recovery rate is affected by the increase in the degree of hydrolysis (Ovisipour et al., 2013). The reduction in the peptide chain length of the alcalase enzyme is inversely proportional to the degree of hydrolysis (Namati et al., 2012). DPPH is related to factors such as process time, degree of hydrolysis, and the performance of each enzyme in producing active peptides and releasing lipophilic antioxidant amino acids (He et al., 2013). The reason for the reduction in oil absorption in the treatment is strongly influenced by the solubility and hydrophobicity of the protein surface (Mazloumi-Kiapi et al., 2019). The decrease and increase in moisture in the treatments is due to the gel formation characteristic during heating and the high degree of coating and the creation of a relatively thick layer (Bahrami and Khadami, 2020). Hydrocolloids with their water retention capacity soften the texture of the nugget and reduce the stiffness of the texture (Altunakar et al., 2006). The high coating rate in the two treatments of 0.5 and 1% is due to the amount of adhesive that adheres to the fish nuggets (DaraiGarmkhanei et al., 2014). The frying process causes cell collapse, less oil absorption and increases frying efficiency. (Mokhtariyan and Tavakolipour, 2014). The amount of thiobarbituric acid, due to the coating formed on the surface of the nugget, reduces the amount of product contact with oxygen, the rate of initial oxidation of fats and the subsequent formation of hydroperoxides (Mohan et al., 2007). The amount of nitrogenous bases, by adding a coating, delays the presence of bacteria and the process of autolysis and protein decomposition (Darocha et al., 2018). The overall acceptance index in the 0.5 and 1% treatments was approved by the evaluator (Bahrami and Khademi, 2020 In conclusion, the present study suggests several promising avenues for future research. These include exploring the functional and antioxidant properties of hydrolyzed chia seed proteins through enzymatic hydrolysis using proteases such as bromelain and ficin. Additionally, it is proposed to assess the potential of hydrolyzed chia seed proteins as a sustainable nitrogen source in bacterial culture media, with implications for both food science and biotechnology. Furthermore, investigating the synergistic effect of hydrolyzed chia seed proteins and vacuum packaging could offer a novel approach to enhancing the shelf life of marine products, thereby contributing to more effective food preservation techniques. Conflict of Interest  The authors declare that they have no conflict of interest Acknowledgment  The authors would like to express their sincere gratitude to the esteemed Professor Dr. Hedayati Far for his valuable guidance throughout all stages of preparing this article, and to the Caspian Sea Ecology Research Institute for providing the necessary facilities and support for conducting this research 75 90 2025/06/32025/09/212026/02/122025/10/12025/12/302025/11/22 1404/9/1 2026/07/12026/07/12026/07/12026/07/12026/04/122026/07/1 1405/4/10 مسیب ناصری Mosayeb Naseri گروه شیلات و غذاهای دریایی، دانشکده منابع طبیعی، واحد قائم‌شهر، دانشگاه آزاد اسلامی، قائم شهر، ایران Naseri.m1992@gmail.com مسعود هدایتی فر Masuod Hedayatifr گروه شیلات و غذاهای دریایی، دانشکده منابع طبیعی، واحد قائم‌شهر، دانشگاه آزاد اسلامی، قائم شهر، ایران persiafush@gmail.com روح الله جوادیان Ruhollah Javadiyan گروه شیلات و غذاهای دریایی، دانشکده منابع طبیعی، واحد قائم‌شهر، دانشگاه آزاد اسلامی، قائم شهر، ایران 2142081029@iau.ir سمیه بهرام Somayeh Bahram گروه شیلات و غذاهای دریایی، دانشکده منابع طبیعی، واحد قائم‌شهر، دانشگاه آزاد اسلامی، قائم شهر، ایران 2141507839@iau.ir وحید رنجبر Vahid Ranjbar پژوهشکده اکولوژی دریای خزر، مؤسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، ساری، ایران vahid.ranjbar.k123@gmail.com فرشیده حبیبی Farshideh Habibi پژوهشکده اکولوژی دریای خزر، مؤسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، ساری، ایران farshidehhabibi@gmail.com ایوب داودی Ayoub Davoudi پژوهشکده اکولوژی دریای خزر، مؤسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، ساری، ایران ayubdavudi@gmail.com غلامرضا رازقیان Gholamreza Razeghiyan پژوهشکده اکولوژی دریای خزر، مؤسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، ساری، ایران rostamireza@yahoo.com مجتبی رکابی Mojtaba Rekabi پژوهشکده اکولوژی دریای خزر، مؤسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، ساری، ایران zenapurge@gmail.com جواد تقوی Javad Taghavi پژوهشکده اکولوژی دریای خزر، مؤسسه تحقیقات علوم شیلاتی کشور، سازمان تحقیقات، آموزش و ترویج کشاورزی، ساری، ایران j_taghavi2001@gmail.com Alginate watercress seed gum lipid oxidation oil absorption chia seeds(Salvia hispanica) آلژینات صمغ دانه شاهی اکسیداسیون لیپید جذب روغن دانه چیا Asadi Farsani, O., Kordjazi, M., Shabanpour, B., Ojagh, S.M. and Jamshidi, A. 2018##Avak, R. and Glaser, R., 2005##Altunakar., B, Sahin., S. and Sumnu., G. 2006##Bahrami,S. and Khademi, D. 2020##Chen, S.D., Chen H.H., Chao Y.C. and Lin, R.S. 2009##Chatterjee, R., Day, T.K., Ghosh, M. and Dhar, P. 2015##Chen, S.D., Chen, H.H., Chao, Y.C. and Lin, R.S. 2009##Das, A.K., Anjaneyulu, A.S.R., Gadekar, Y.P., Singh, R.P. and Pragati, H. 2008##Daraei Garmehkhani, A., Mirzaei, H.A., Maghsoudlou, Y. and Kashaninezhad, M., 2009##Daraei Garmakhany, A., Mirzaei, H. 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