In vitro and in vivo screening of herbal extracts against Streptococcus agalactiae in Nile tilapia (Oreochromis niloticus)
In vitro and in vivo screening of herbal extracts against Streptococcus agalactiae in Nile tilapia (Oreochromis niloticus)
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针对尼罗罗非鱼（Oreochromis niloticus）无乳链球菌的草药提取物的体外和体内筛选
DOI：
10.1016/j.aquaculture.2019.01.024
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发表时间：
2019-03-30
期刊：
影响因子：
4.5
通讯作者：
Zhou, Yong-Can
中科院分区：
文献类型：
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作者：
Guo, Wei-Liang;Deng, Heng-Wei;Zhou, Yong-Can
本研究以115种中草药为材料，采用4种提取方法制备了354种中草药提取物。采用微孔板法和琼脂扩散法对提取物对无乳链球菌的体外抗菌活性进行了研究。然后将体外抗菌活性高的草药提取物以100 mg/g（干草药/干饲料，w：w）的比例混合到饲料中，并饲喂具有S.用于体内评价它们的治疗功效。随后，根据拟合方程计算具有最高治疗功效的三种草药提取物的EC 50值（导致相对存活百分比（RPS）为50%的浓度）。最后，使用超高效液相色谱-Q Exactive Orbitrap-MS（UPLC-QE-MS）对具有最高治疗功效的草药进行植物化学分析。结果表明：采用微孔板法检测，354种候选中药提取物中有62种具有抑菌活性，抑菌率在5%以上;采用琼脂孔扩散法检测，62种候选中药提取物中有29种抑菌圈清晰，33种抑菌圈半清晰。在50 ℃下浸渍3小时制备的马兜铃（果实）、日本野牡丹（花）、鸡血藤（茎）和槟榔（果实）提取物的RPS分别为100.0%、55.6%、75.0%和33.3%;人参（叶）、大黄（叶）和槟榔（叶）的RPS分别为100.0%、55.6%、75.0%和33.3%。在室温下浸泡72 h制备的高良姜（根）和高良姜（种子）提取物的RPS分别为77.8%、25.0%和12.5%;在室温下超声提取30 min制备的板蓝根（根）提取物的RPS为62.5%。A. debilis、人参P.三种提取物的EC（50）分别为44.03、41.19、39.54 mg/g。A.的植物化学分析debilis、人参P.使用UPLC-QE-MS对腹直肌提取物进行分析得到以下结果：阴性模型分别检测到270、623和326个峰，并鉴定出54、70和52种化合物;阳性模型分别检测到446、1401和400个峰，并鉴定出61、107和44种化合物。这些结果表明，A. debilis、人参P.下直肌可用于治疗S.无乳症感染的尼罗罗非鱼，并建议适当的生物活性化合物，从这三种草药提取物可开发作为替代抗生素。
This study investigated 354 herbal extracts prepared from 115 herbs via 4 extraction methods. The extracts' antibacterial activities against Streptococcus agalactiae were sequentially evaluated in vitro by two methods: (1) microplate assay and (2) agar well diffusion. Then the herbal extracts with high antibacterial activity in vitro were mixed into feed at 100 mg/g (dry herb/dry feed, w: w) and fed to fishes with S. agalactiae infection for in vivo evaluation of their therapeutic efficacy. Subsequently, the EC50 values (the concentrations result in the relative percent survival (RPS) of 50%) of the three herbal extracts with the highest therapeutic efficacies were calculated according to fit equations. Finally, phytochemical analyses of the herbs with the highest therapeutic efficacies were conducted using ultra performance liquid chromatography-Q Exactive Orbitrap-Mass spectrometry (UPLC-QE-MS). The results showed the following: in detections from the microplate assay method, 62 of the 354 candidate herbal extracts had antibacterial activity with inhibition ratios above 5%; in detections from the agar well diffusion method, 29 of the 62 the candidate herbal extracts showed clear inhibition zones and 33 showed semi-clear inhibition zones. Aristolochia debilis (fruit), Lonicera japonica (flower), Spatholobus suberectus (stem), and Arecae Semen (fruit) extracts prepared by 3 h of maceration at 50 degrees C had RPSs of 100.0%, 55.6%, 75.0%, and 33.3%, respectively; Panax ginseng (leaf), Rheum palmatum L. (root), and Semen Alpiniae (seed) extracts prepared by 72 h of maceration at room temperature had RPSs of 77.8%, 25.0%, and 12.5%, respectively; and Radix Isatidis (root) extract prepared by 30 min of ultrasonic-extraction at room temperature had an RPS of 62.5%. A. debilis, P. ginseng, and S. suberectus extracts had EC(50)s of 44.03, 41.19, 39.54 mg/g, respectively. Phytochemical analyses of A. debilis, P. ginseng, and S. suberectus extracts using UPLC-QE-MS produced the following results: the negative model detected 270, 623, and 326 peaks and identified 54, 70, and 52 compounds, respectively; the positive model detected 446, 1401, and 400 peaks and identified 61, 107, and 44 compounds, respectively. Taken together, these results indicate that the extracts of A. debilis, P. ginseng, and S. suberectus could prove useful for the treatment of S. agalactiae infection in Nile tilapia and suggest that appropriate bioactive compounds from these three herbal extracts may be developed as alternates to antibiotics.