en fish disease fish disease پژوهشي Orginal research papers <span lang="EN" style="font-size:11.0pt"><span style="font-family:&quot;Calibri&quot;,sans-serif"><span style="color:black">This study aimed to produce and evaluate the antibacterial activity of recombinant rainbow trout pro-hepcidin (rpro-RtHep). T</span></span></span><span lang="TR" style="font-size:11.0pt"><span style="font-family:&quot;Calibri&quot;,sans-serif"><span style="color:black">he gene sequence encoding&nbsp;</span></span></span><span lang="EN" style="font-size:11.0pt"><span style="font-family:&quot;Calibri&quot;,sans-serif"><span style="color:black">rainbow trout pro-hepcidin was synthesized and cloned into the pET28a+ expression vector. The recombinant vector pET28a+/pro-RtHep was transformed into <i>Escherichia coli</i> BL21 (DE3) Rosetta for protein expression. The recombinant bacteria were cultured at 37&deg;C and induced with 0.5 mM Isopropyl &beta;-D-1-thiogalactopyranoside (IPTG). </span></span></span><span lang="TR" style="font-size:11.0pt"><span style="font-family:&quot;Calibri&quot;,sans-serif"><span style="color:black">The presence of a 10 kDa band on SDS-PAGE confirmed successful peptide expression; subsequently, purification was carried out using affinity chromatography, and the single 10 kDa band was verified by SDS-PAGE. The biological activity of purified&nbsp;rpro-RtHep&nbsp;was evaluated by examining its inhibitory effects on the growth of selected <i>Aeromonas hydrophila, Streptococcus iniae</i> and<i> Yersinia</i><b> </b><i>ruckeri</i>. The results showed that&nbsp;rpro-RtHep&nbsp;effectively inhibited the growth of these bacterial species. </span></span></span><span lang="EN" style="font-size:11.0pt"><span style="font-family:&quot;Calibri&quot;,sans-serif"><span style="color:black">Additionally, the survival rate of zebrafish (<i>Danio</i> <i>rerio</i>) and with the rpro-RtHep (25 &micro;g g^-1) was assessed and the results revealed that </span></span></span><span lang="TR" style="font-size:11.0pt"><span style="font-family:&quot;Calibri&quot;,sans-serif"><span style="color:black">survival rate</span></span></span><span lang="EN" style="font-size:11.0pt"><span style="font-family:&quot;Calibri&quot;,sans-serif"><span style="color:black"> was significantly higher compared to that of the control group exposed to <i>A.</i> <i>hydrophila</i>. </span></span></span><span lang="TR" style="font-size:11.0pt"><span style="font-family:&quot;Calibri&quot;,sans-serif"><span style="color:black">Immune factors, including lysozyme levels, ACH50, total protein, and total immunoglobulin demonstrated significant increases in fish injected with rpro-RtHep compared to the control group (<i>p</i><0.05</span></span></span><span lang="EN" style="font-size:11.0pt"><span style="font-family:&quot;Calibri&quot;,sans-serif"><span style="color:black">). The study demonstrated that rpro-RtHep </span></span></span><span lang="TR" style="font-size:11.0pt"><span style="font-family:&quot;Calibri&quot;,sans-serif"><span style="color:black">protects zebrafish<i> </i>against <i>A</i>. <i>hydrophila </i>infection via regulating of immune factors.</span></span></span> Bacterial resistance, Antibiotic alternative, Hepcidin, Immune factors 663 674 http://jifro.ir/browse.php?a_code=A-10-4999-1&slc_lang=en&sid=1 S. Mohammadzadeh mohammadzadeh.sedigheh@yahoo.com 100319475328460059224 100319475328460059224 No Faculty of Veterinary Medicine, Amol University of Special Modern Technologies, Amol, Iran R. Khoshbakht r.khoshbakt@gmail.com 100319475328460059225 100319475328460059225 Yes Department of Pathobiology, Faculty of Veterinary Medicine, Amol University of Special Modern Technologies, Amol, Iran P. Shohreh p.shohreh@yahoo.com 100319475328460059226 100319475328460059226 No Department of Clinical Science, Faculty of Veterinary Medicine, Amol University of Special Modern Technologies, Amol, Iran M. Ranjbar mojtaba.ranjbar@gmail.com 100319475328460059228 100319475328460059228 No Department of Microbial Biotechnology, Faculty of Biotechnology, Amol University of Special Modern Technologies, Amol, Iran Fatemeh Moradian f_moradi@yahoo.com 100319475328460059227 100319475328460059227 No Department of Basic Sciences, Sari Agricultural Sciences and Natural Resources University, Sari, Iran Ehsan Ahmadifar Ehsan.ahmadifar@gmail.com 100319475328460059229 100319475328460059229 No Department of Fisheries, Faculty of Natural Resources, University of Zabol, Zabol, Iran