氟苯尼考对六种海洋致病弧菌的体外抗菌活性研究

测定了氟苯尼考等4种抗菌药对6种常见海洋致病弧菌的最小抑菌浓度(MIC)和最小杀菌浓度(MBC)。测试结果显示,氟苯尼考对6株弧菌的MIC值范围为2～4μg／mL,氯霉素为1～32μg／mL,土霉素为4～128μg／mL,恩诺沙星为0．125～1μg／mL;对多数弧菌氟苯尼考与氯霉素的抗菌活性相似。氟苯尼考对6株弧菌的MBC／MIC值范围为2-4,而氯霉素为2～16,表明氟苯尼考的杀菌效果优于氯霉素。对照药物恩诺沙星、土霉素的MBC／MIC值分别为1-2和2-8,恩诺沙星显示出优良的抗菌和杀菌活性。随着接种菌量增加,所测3种抗菌药的MIC随之升高;当接种量达到10^7ufu／mL时．药物对多数弧菌显示出微弱的作用。     

五倍子对3种致病菌的体外抑菌和急性毒性试验

采用试管两倍稀释法测定五倍子水煎煮液对3种鱼类常见致病菌的最小抑菌浓度（MIC）和最小杀菌浓度（MBC）,并进行了对鲤鱼的急性毒性试验.结果表明,五倍子煎煮液对温和气单胞菌、爱德华氏菌、柱状屈挠杆菌的最小抑菌浓度分别为6.250 mg/mL、3.125 mg/mL、0.781 mg/mL;最小杀菌浓度分别为12.500mg/mL、12.50.mg/mL、3.125 mg/mL.五倍子对鲤鱼的48 h半致死浓度为1.1015g/kg,安全浓度为0.1102g/kg.五倍子具有较好的杀菌防病效果.     

中华绒螯蟹的致病菌及抗菌药物的筛选

对河蟹育苗场各期蚤状幼体(Z1～Z5)、大眼幼体至Ⅴ期幼蟹进行了病原菌的分离采样。采用"细菌生化鉴定编码"对育苗期的中华绒螯蟹的致病菌进行鉴定,主要为溶藻弧菌(V.alginolyticus)、霍乱弧菌(V.cholerae)、荧光假单胞菌(P.fluorescens)、豚鼠气单胞菌(A.caviae)和梅氏弧菌(V.metschnikovii)。采用扩散法(K-B法)对上述菌株进行了药物敏感性试验,结果表明:上述各菌株对庆大霉素、氟哌酸、头孢哌酮、丙氟哌酸、氟嗪酸、复方新诺明、环丙沙星具有较高的敏感性,抑菌直径>16mm。同时选择氟哌酸和环丙沙星对荧光假单胞菌、溶藻弧菌进行了最小抑菌浓度(MIC)与最小杀菌浓度(MBC)的测定。氟哌酸对荧光假单胞菌和溶藻弧菌的MIC分别为25μg/mL和0.78μg/mL;环丙沙星对荧光假单胞菌和溶藻弧菌的MIC分别为6.25μg/mL和0.025μg/mL。氟哌酸对荧光假单胞菌和溶藻弧菌的MBC均为6.25μg/mL;环丙沙星对荧光假单胞菌和溶藻弧菌的MBC分别为50μg/mL和0.1μg/mL。     

二氧化氯和三氯异氰脲酸对鱼类3种致病菌的杀灭试验

对肠型点状产气单胞杆菌，嗜水气单胞菌和柱状曲桡杆菌进行复壮．将分离纯化的3种菌用二氧化氯和三氯异氰尿酸进行抑菌、杀菌效果研究。结果表明：二氧化氯对肠型点状气单胞菌、嗜水气单胞菌、柱状曲桡杆菌的最小抑菌浓度分别是0．17、0、17、0．06μg／mL，最小杀菌浓度分别是0．23、0．23、0．17μg／mL；三氯异氰脲酸对上述3种菌的最小抑菌浓度分别是6．25、12．5、6．25μg／mL，最小杀菌浓度分别是12．5、25．0、6．25μg／mL。     

恶喹酸与诺氟沙星对异育银鲫体内嗜水气单胞菌的抑菌效果比较

在水温(25±2)℃下,给体质量150~200g的异育银鲫分别口灌3种剂量(20、30、40mg/kg)的诺氟沙星和恶喹酸,结合这两种药物对嗜水气单胞菌AH10的体外药效学研究和对异育银鲫单次口灌不同剂量的诺氟沙星、恶喹酸的体内药代动力学,研究诺氟沙星、恶喹酸对异育银鲫体内嗜水气单胞菌AH10的抑菌效果。试验结果表明,恶喹酸和诺氟沙星对嗜水气单胞菌AH10的最小抑菌质量浓度分别为1μg/mL和0.5μg/mL。口灌上述3种剂量恶喹酸、诺氟沙星后,异育银鲫血浆中恶喹酸的最大药物质量浓度分别为4.1μg/mL、6.0μg/mL和8.89μg/mL;诺氟沙星的最大药物质量浓度分别为11.5μg/mL、15.1μg/mL和18.9μg/mL;恶喹酸的最大药物质量浓度/最小抑菌质量浓度分别为4.1、6.0和8.89;诺氟沙星的最大药物质量浓度/最小抑菌质量浓度分别为23、30.2和37.8;恶喹酸0~24h内药—时曲线下面积/最小抑菌质量浓度分别为21.6214、33.1449、39.1846;诺氟沙星0~24h内药—时曲线下面积/最小抑菌质量浓度分别为274.75、451.55、578.35。综合0~24h内药—时曲线下面积/最小抑菌质量浓度、最大药物质量浓度/最小抑菌质量浓度这两个指标可知,诺氟沙星对异育银鲫体内的嗜水气单胞菌AH10抑制效果强于恶喹酸。     

22味中草药对创伤弧菌和灿烂弧菌体外抑菌作用筛选

研究了22味中草药对创伤弧菌(Vibrio vulnificus)和灿烂弧菌(V.splendidus)的体外抑菌作用。结果显示,22味中草药对其均有不同程度的抑制作用;其中,黄连和连翘对创伤弧菌的抑菌和杀菌效果最好,最小抑菌浓度分别为0.49mg/mL和1.95mg/mL,最小杀菌浓度与最小抑菌浓度相同;黄连、黄芩、连翘和黄柏对灿烂弧菌的抑菌和杀菌效果较好,最小抑菌浓度分别为0.98、1.95、1.95和3.90mg/mL,最小杀菌浓度分别为0.98、1.95、1.95和7.81mg/mL。     

22味中草药对创伤弧菌和灿烂弧菌体外抑菌作用筛选

研究了22 味中草药对创伤弧菌(Vibrio vulnificus)和灿烂弧菌(V. splendidus)的体外抑菌作用.结果显示,22味中草药对其均有不同程度的抑制作用;其中,黄连和连翘对创伤弧菌的抑菌和杀菌效果最好,最小抑菌浓度分别为0.49 mg/mL和 1.95 mg/mL,最小杀菌浓度与最小抑菌浓度相同;黄连、黄芩、连翘和黄柏对灿烂弧菌的抑菌和杀菌效果较好,最小抑菌浓度分别为0.98、1.95、1.95和3.90 mg/mL,最小杀菌浓度分别为0.98、1.95、1.95和7.81 mg/mL.     

单过硫酸氢钾粉对5种水产常见致病菌的体内外抑菌作用

研究了单过硫酸氢钾粉对5种水产常见致病菌的体内、外抑菌作用,测定了体外最小抑菌浓度(MIC)、最小杀菌浓度(MBC)及以保护率为指标的体内抑菌效果。结果表明,体外抑菌实验中,单过硫酸氢钾粉对爱德华氏菌Edwardsiella、温和气单胞菌Aeromonas sobria、嗜水气单胞菌Aeromonas hydrophila、迟钝爱德华氏菌Edwardsiella tard、荧光假单胞菌Psudomonas fluorescens均有明显的抑制作用,其对5种菌的MIC分别为1、0.25、0.125、0.25、0.5μg/ml;MBC与MIC相同。而体内抑菌实验中,随着单过硫酸氢钾粉浓度的增加,由5种致病菌引起鲫鱼的死亡率逐步降低。统计分析发现,使用0.25μg/ml单过硫酸氢钾粉药浴鲫鱼,即可对由5种致病菌引起的鲫鱼病害有治疗的作用。     

41种中草药对3种鲟源病原菌的体外抑菌效果

采用平板打孔法在观察了弗氏柠檬酸杆菌(Citrobacter freundii)、鲁氏耶尔森氏菌(Yersinia ruckeri)、嗜水气单胞菌(Aeromonas hydrophila)等3种鲟源病原菌对41种中草药敏感性的基础上,进一步采用二倍稀释法测定了具有良好抑菌活性的中草药对3种病原菌的最小抑菌浓度和最小杀菌浓度。实验结果表明,3种病原菌对乌梅、石榴皮、地榆、杞子等均高度敏感,对杜仲、柴胡、丹参、熟地黄等均中度敏感,对麦冬、当归、胖大海、玉竹均不敏感。此外,乌梅、石榴皮、地榆、杞子对3种病原菌的最小抑菌浓度和最小杀菌浓度分别为7.81 mg/mL和15.625 mg/mL、15.625 mg/mL和31.25 mg/mL、15.625 mg/mL和31.25 mg/mL以及15.625 mg/mL和31.25 mg/mL,而杜仲、柴胡、丹参、熟地黄等中度敏感中草药对3种病原菌的最小抑菌浓度和最小杀菌浓度均分别高于31.25 mg/mL和62.5 mg/mL。     

培氟沙星对淡水鱼类细菌出血性败血病药效学研究

为研究培氟沙星对淡水鱼类细菌出血性败血病的防治效果,从而指导临床合理用药,有效控制细菌出血性败血病,进行了体外和体内抗菌药效试验,并通过生产防治试验进行了药效验证。体外抗菌试验采用试管稀释法对致病菌嗜水气单胞菌和温和气单胞菌进行了最小抑菌浓度试验,结果表明,培氟沙星对两种致病菌的最小抑菌浓度和最小杀菌浓度均为0.01μg·mL-1和0.02μg·mL-1,而对照药物氟苯尼考、左氧氟沙星、庆大霉素、恩诺沙星对嗜水气单胞菌的最小抑菌浓度分别为10、1.25、2.5、0.039μg·mL-1,最小杀菌浓度分别为20、5、10、0.078μg·mL-1;对温和气单胞菌最小抑菌浓度为10、5、5、0.005μg·mL-1,最小杀菌浓度分别为40、10、20、0.02μg·mL-1;除恩诺沙星对温和气单胞菌的抑菌效果好于培氟沙星外,培氟沙星对两种致病菌的抑菌效果均好于其他药物。培氟沙星对人工感染嗜水气单胞菌的鲤鱼体内预防和治疗试验以及生产防治试验的结果,治疗组的成活率达到90%,预防组的成活率为87%;生产防治试验面积1500亩(15亩=1 hm2,下同),有效率达95.46%。     

豹纹鳃棘鲈类结节症病原的分离鉴定

为查明养殖豹纹鳃棘鲈类结节症的病因,自患病豹纹鳃棘鲈内脏中分离到1株优势菌株061101,人工感染试验证实,该菌具有强毒力,能够引起被感染鱼内脏出现白色结节并死亡,为引起此次豹纹鳃棘鲈类结节症的病原菌。采用形态学、生化特性测定对菌株061101进行鉴定,同时采用细菌16S rRNA通用引物进行PCR扩增得到该菌的16S rRNA基因序列,测序后比对,构建系统发育树。试验结果表明,菌株061101与发光杆菌属中的美人鱼发光杆菌杀鱼亚种基本特征相符;基于16S rRNA基因序列构建的系统发育树将菌株061101与美人鱼发光杆菌杀鱼亚种聚为一支,置信度达100%。据此确定,此次豹纹鳃棘鲈类结节症的病原为美人鱼发光杆菌杀鱼亚种。药敏结果显示,该菌株仅对链霉素(300μg/片)、庆大霉素(120μg/片)敏感,对多黏菌素B、阿洛西林等中度敏感,对氟苯尼考、恩诺沙星、强力霉素等均耐药。     

Resistance of Aeromonas salmonicida to amoxicillin

Abstract. Isolates of Aeromonas salmonicida were obtained from farmed Atlantic salmon, salmo salar L., with amoxicillin minimum inhibitory concentrations (MICs) of 2·5–10 μg ml^-1 Several antibiotic sensitivity patterns were found among the isolates including resistance to the four UK-licensed antibacterial drugs. Combination of clavulanic acid with amoxicillin was not effective, but all isolates were sensitive to florfenicol and to a cephalosporin, the latter having very low MICs.     

肌注和口灌氟苯尼考在吉富罗非鱼体内的药代动力学

通过肌内注射、口灌两种给药方式,研究氟苯尼考在罗非鱼体内的药物代谢动力学特征。把吉富罗非鱼(GIFT Oreochromis niloticus)随机分成2组,控制水温在30℃,以15 mg/kg分别单剂量肌内注射、口灌给药。经高效液相色谱法(HPLC)测定血浆中氟苯尼考浓度,用Win Nonlin药动学软件分析药动学参数。结果表明:肌内注射氟苯尼考后,药物吸收较慢,消除较快,达峰时间(T_(max))=4 h,峰浓度(C_(max))=4.64μg/mL,消除半衰期(T_(1/2λ)z L)=10.45 h,药-时曲线下面积(AUC)=91.06μg·h/mL。口灌氟苯尼考后,药物吸收较快,消除较慢,T_(max)=1 h,C_(max)=5.92μg/mL,T_(1/2λ)z L=13.13 h,AUC=61.96μg·h/mL。肌内注射、口灌氟苯尼考后,二者的药动学参数差异显著,这一差异表明肌内注射给药吸收相对较慢,但更为完全(肌内注射氟苯尼考的AUC明显较大),消除相对较快。     

Modified resazurin microtiter assay for in vitro and in vivo assessment of sulfamonomethoxine activity against the fish pathogen <Emphasis Type="Italic">Nocardia seriolae</Emphasis>

Resazurin microtiter assay (REMA) was carried out using four sulfonamides, three culture media, and four inoculum sizes as a first screening step to establish an easy-to-interpret sulfonamides susceptibility testing method for Nocardia seriolae. The in vitro activity of sulfamonomethoxine (SMM) against 190 clinical N. seriolae isolates was then examined, and in vivo experimental treatment was performed. When the culture medium and the inoculum size were considered in tandem, a 0.5× the original concentration of cation-adjusted Mueller–Hinton broth and an inoculum size of 10² CFU/well showed the clearest endpoint reading for all tested drugs, and the REMA-generated data were in excellent agreement with those generated by the reference Etest method. SMM activity showed minimum inhibitory concentration (MIC) values of 4–32 μg/ml against all tested N. seriolae isolates. Treatment of amberjack groups experimentally infected with N. seriolae isolates having SMM MICs of 4 and 32 μg/ml, resulted in survival rates of 100% and 87.5% in the two groups, respectively. In this study, we developed a simple visual method to test SMM activity against N. seriolae.     

不同浓度的枯草芽孢杆菌对罗非鱼鱼苗的养殖水体水质及其抗病力的影响

罗非鱼鱼苗的养殖水体中引入不同浓度的枯草芽孢杆菌,检测水体的水质指标、鱼苗体内与免疫相关酶的酶活力、鱼苗的生长率和成活率。实验结束时,引入1.0×10^4 cfu/mL枯草芽孢杆菌实验组,氨氮和亚硝酸盐氮含量分别为2.72 mg/L、0.15 mg/L,显著低于对照组（P〈0.05）;AKP活力、抗菌活力分别达249.9 U/g prot、0.59μg/mL,显著高于对照组（P〈0.05）;鱼苗成活率也显著高于对照组（P〈0.05）,比对照组提高了11.0%。结果显示：适合浓度的枯草芽孢杆菌能有效地改善鱼苗养殖水体的水质,提高机体免疫力和成活率。     

双氟沙星与异育银鲫血浆蛋白的结合率

为了研究双氟沙星(difloxacin,DIF)与异育银鲫(Carassais auratus gibebio)血浆蛋白的结合性质,采用平衡透析法测定了DIF与异育银鲫血浆蛋白的体外结合率,并研究了温度、血浆浓度及药物浓度对其的影响。结果显示:DIF血浆蛋白结合率在4℃、15℃、20℃、25℃、30℃、37℃条件下变化差异不显著(P>0.05);药物血浆蛋白结合率显著高于二倍稀释血浆的结合率,也显著高于四倍稀释血浆的结合率(P<0.05)。当DIF浓度在0.5～5μg/mL范围之间时,药物与血浆蛋白结合率在32.4%～19.6%之间,并随着药物浓度的升高而降低,最大结合能力(β)和解离常数(Kdp)分别为3.5×10-7 mol/g和4.04×10-6 mol/L;当DIF质量浓度为5～40μg/mL范围内时,血浆蛋白结合率的变化差异不显著(P>0.05)。此外,游离药物与总药物浓度呈线性关系,关系式为y=0.77x-0.18。结果表明,DIF与异育银鲫血浆蛋白结合率在本实验条件下不受温度影响,且具有浓度依赖性;在0.5～5μg/mL范围内结合率为非线性,而在5～40μg/mL范围内为线性。本研究旨在为DIF药物代谢动力学理论研究及其临床用药提供理论参考。     

The in vitro susceptibility of Aeromonas salmonicida and other fish-pathogenic bacteria to 29 antimicrobial agents

Abstract. The minimum inhibitory concentrations (MICs) of 29 antimicrobial agents were determined for some representative Aeromonas salmonidda, Yersinia ruckeri and Vibrio isolates. Eleven of them, including three widely used in clinical practice, were further tested against 28–36 recent field isolates of A. salmonidda. The fluoroquinolones were inhibitory at very low concentrations with 50% of A. salmonidda isolates having a MIC of enro-floxacin ≤0·02μg ml-1. The distribution of MICs to the fluoroquinolones and to oxolinic acid were similar indicating possible cross resistance. Amoxycillin, florfenicol, minocycline, rifampicin and the potentiated sulphonamides had a single peak MIC distribution with values of ≤1·25μg ml-1. Minimum inhibitory concentrations of the fluoroquinolones were four or more times greater at 10°C compared with 22°C; this was not so with amoxycillin, florfenicol or the sulphachlorpyridazine-trimethoprim (S-TP) combination. A. salmonicida was rapidly killed at 22°C by the fluoroquinolones and S-TP whereas oxolinic acid and amoxycillin killed within 24h. Florfenicol was bacteriostatic. The therapeutic potential of some of these antibiotics is considered.     

氟苯尼考及氟苯尼考胺在鲤体内的残留

在水温(18±1)℃下,采用高效液相色谱串联质谱法研究了氟苯尼考及其主要代谢物氟苯尼考胺在体质量(100±10)g鲤(Cyprinus carpio)体内的代谢及残留规律,以制定休药期。实验鱼间隔24h按15 mg.kg-1体质量的剂量口灌给药,连续给药3次,在给药后0.5h、1 h、2h、4h、8h、12h、24h、2d、3d、5d、7d、10d、15d、20d检测肌肉、皮肤内氟苯尼考和氟苯尼考胺的含量。结果表明:氟苯尼考在鲤体内的代谢速度快,2~4 h出现峰值;肌肉中氟苯尼考和氟苯尼考胺的残留浓度高于皮肤,肌肉和皮肤中氟苯尼考的含量均高于氟苯尼考胺,皮肤中的代谢速度较快。按欧盟标准相应的休药期不少于2d,按日本标准则不少于10 d。     

2种水温条件下罗非鱼体内氟苯尼考的药物动力学比较

采用药饵给药,药物剂量为10mg·kg^-1,比较研究了22和28℃水温条件下奥尼罗非鱼（Oreochromis niloticus×O．aureus）体内氟苯尼考的药物动力学。结果得出,22℃组和28℃组罗非鱼血浆的峰药浓度（Cmax）分别为4．46和3．90μg·mL^-1,达峰时间（Tmax）均为12h,消除半衰期（T1/2β）分别为10．03和8．12h,药-时曲线下面积（AUC）分别为86．68和72．44h·μg·mL^-1。相应条件下的肌肉Cmax分别为6．88和4．59μg·g^-1,Tmax均为12h,T1／2β分别为10．97和8．03h,AUC分别为112．71和73．66h·μg·g^-1。低温组罗非鱼血浆和肌肉中药物的T1／2β均长于高温组,前者分别比后者长1．91和2．96h,表明低温组罗非鱼体内药物的消除速度慢于高温组。虽然2个水温组血浆和肌肉中药物的Tmax相同,但低温组血浆、肌肉的Cmax和AUC均明显高于高温组,表明低温组罗非鱼吸收利用药物程度高于高温组。     

Depletion study and estimation of withdrawal periods for florfenicol and florfenicol amine in pacu (Piaractus mesopotamicus)

The intensive production of farmed fish is at a global all‐time high, and the control of bacteria proliferation in fish farms requires the frequent use of antimicrobials. This practice raises important environmental concerns related to the emergence of antimicrobial‐resistant bacteria strains. Only a few antimicrobial drugs have been approved for use in aquaculture, one of which is florfenicol. This work studies the depletion and withdrawal period of florfenicol and its main metabolite, florfenicol amine, in pacu (Piaractus mesopotamicus), a neotropical characin widely farmed in the southern hemisphere. Juvenile pacu (average weight of 724 g) were stocked in a closed‐loop laboratory system with controlled water temperature (25.8°C), and were fed for 10 consecutive days with a diet containing an intended dose of 10 mg/florfenicol per kg bw. Muscle and skin tissues were collected at 1, 3, 6, 8, 10, 12 and 16 days post‐treatment, and florfenicol and florfenicol amine were quantified using a validated ultra‐high performance liquid chromatography‐tandem mass spectrometry (UPLC‐MS/MS) method. The limits of quantitation for florfenicol and florfenicol amine were 10 ng/g in muscle and 50 ng/g in skin. Considering a maximum residue limit of 1000 ng/g for the sum of florfenicol and florfenicol amine in muscle with skin in natural proportions a withdrawal period of 5 days (water temperature 25.8°C) or 129 degree days was calculated on the basis of the upper limit of the one‐sided 95% confidence interval for the 99^th percentile derived from the residue depletion study.