Investigation of the effects of some phytochemicals on Yersinia ruckeri and antimicrobial resistance

Abstract In this study, it is aimed to investigate the effects of Moringa oleifera and Sorbus domestica plant extracts on bacterial disease agents Yersinia ruckeri in aquaculture. Morphological and biochemical properties of 2 different Y. ruckeri isolates were determined. Then, Real-Time PCR analysis and gene sequencing of the isolates were identified. Phytochemicals (M. oleifera and S. domestica) and antibiotics (Oxytetracycline (OX) and Enrofloxacin (ENR)) were used together in the antibiogram test of antibiotics compared to the effect status of antibiotics. Also, the effects of phytochemicals on Y. ruckeri growth was examined comparatively by spectrophotometrically measuring at 600 nm wavelength every 2 hours according to bacterial growth densities with 10 different groups formed on TSB medium. As a result of the study, it was observed that the isolates formed Gram negative, catalase positive, oxidase negative, mobile and typical Y. ruckeri colonies. After the biochemical tests performed with Microgen ID panel, 99.85% similarity was determined. The isolates overlap with the 16S rRNA gene region after sequence analysis, and 99% of the isolates were similar in phylogenetic analysis. After the antibiogram test, Oxytetracycline and Enrofloxacin antibiotics were resistant to Y. ruckeri but the effects of phytochemicals were less on solid medium (MHA). As a result of the measurements carried out in liquid medium (TSB), it was observed that phytochemicals such as M. oliefera and S. domestica inhibit the growth of bacteria by 40-50%. As the importance of antibiotic resistance is increasing day by day, we believe that these phytochemicals will give positive results in treatment instead of using antibiotics.

摘要 本研究旨在探究辣木（Moringa oleifera）与花楸（Sorbus domestica）植物提取物对水产养殖中细菌性致病菌鲁氏耶尔森菌（Yersinia ruckeri）的作用效果。研究首先明确了两株鲁氏耶尔森菌分离株的形态学与生化特性，随后对分离株开展实时荧光定量PCR（Real-Time PCR）分析与基因测序。在药敏试验中，将植物提取物与土霉素（Oxytetracycline, OX）、恩诺沙星（Enrofloxacin, ENR）两类抗生素联合使用，对比其与单独使用抗生素的抑菌效果差异。此外，本研究在胰蛋白胨大豆肉汤（TSB）培养基上设置10个实验组，每2小时以分光光度法检测600nm波长处的吸光度以反映细菌生长密度，以此对比分析植物提取物对鲁氏耶尔森菌生长的抑制作用。研究结果显示，分离株呈现革兰氏阴性、过氧化氢酶阳性、氧化酶阴性、具有运动性的典型鲁氏耶尔森菌菌落形态。通过Microgen细菌鉴定试剂盒（Microgen ID panel）完成生化鉴定后，两株分离株的鉴定相似性达99.85%。序列分析表明，分离株的16S核糖体RNA（16S rRNA）基因区域具有同源性，系统发育分析显示99%的分离株具有较高的序列相似性。药敏试验结果表明，鲁氏耶尔森菌对土霉素与恩诺沙星表现出耐药性，而植物提取物在固体培养基穆勒希顿琼脂（MHA）上的抑菌效果较弱。液体培养基（TSB）中的生长检测结果显示，辣木与花楸提取物可抑制细菌生长达40%~50%。鉴于抗生素耐药性问题日益严峻，本研究认为此类植物提取物有望替代抗生素，为水产养殖细菌性病害防治提供潜在解决方案。