Emended descriptions of indole negative and indole positive isolates of Brachyspira (Serpulina) hyodysenteriae

Two type/reference strains of Brachyspira (B.) hyodysenteriae, 14 Belgian and German indole negative, and 14 Belgian, German and Swedish indole positive field isolates of strongly β-haemolytic intestinal spirochaetes were compared by pulsed-field gel electrophoresis (PFGE) patterns, biochemical reaction patterns, 16S rDNA sequences and MIC determinations of six antibacterial substances. Three tests for indole production, including a spot indole test, were compared with congruent results. All field isolates were classified as B. hyodysenteriae due to a high genetic and phenotypic similarity with the type strains. The Belgian and German indole negative isolates had identical and unique PFGE patterns for the tested restriction enzymes MluI and SalI, as well as identical 16S rDNA sequences, and they could not be differentiated by any of the methods used. Seven unique PFGE patterns were achieved from the 14 indole positive field isolates. The patterns were identical and unique for epidemiologically related isolates. Type/reference strains and isolates without known relation to other tested isolates showed unique banding patterns. The MICs of tylosin, tiamulin, erythromycin, clindamycin, carbadox and virginiamycin were determined in broth for all isolates. In contrast to Belgian and German isolates, the majority of the Swedish field isolates were susceptible to tylosin, erythromycin and clindamycin. Probable pathways of infection for some of the Swedish isolates were determined. The PFGE patterns of epidemic clones of B. hyodysenteriae remained stable for a period of up to 8 years. In vivo development of resistance to macrolide and lincosamide antibiotics due to use of tylosin was clearly indicated for two epidemic clones.     

不同精油的抑菌效果及协同作用研究

为研究三种精油(肉桂醛、百里香酚、香兰素)的抑菌效果及其协同作用,本试验采用牛津杯法和试管二倍稀释法测定抑菌圈的值径及最小抑菌浓度,结果表明:三种精油对大肠杆菌K88、沙门氏菌、金黄色葡萄球菌都有较强的抑菌效果,在相同浓度下,抑菌效果为肉桂醛>百里香酚>香兰素;当百里香酚与肉桂醛以4∶1复合时,对大肠杆菌K88的抑菌效果最佳;当肉桂醛与香兰素以4∶1复合时,对沙门氏菌和金黄色葡萄球菌的抑菌效果最佳。综上所述,肉桂醛和百里香酚的抑菌效果优于香兰素,百里香酚与香兰素,肉桂醛和香兰素复配时具有协同作用。     

常用中草药对水产动物致病菌体外抑菌效果研究

试验旨在探讨黄柏、诃子、知母、苏木等常用中草药对水产动物致病菌的体外抑菌效果,为今后水产养殖细菌性疾病的防控提供参考。以美人鱼发光杆菌、嗜水气单胞菌、哈维氏弧菌、创伤弧菌临床分离株为试验菌株,以抑菌圈直径、最小抑菌浓度（MIC）、最小杀菌浓度（MBC）及联合抑菌指数（FICI）作为检测指标,研究常用中草药对水产病原菌的抑菌活性。采用水提法提取中草药有效成分,并通过琼脂平板打孔法测定中草药对水产致病菌抑菌圈,微量肉汤稀释法测定中草药MIC和MBC及微量肉汤棋盘稀释法测定联合抑菌指数（FICI）。结果显示,16种中草药对水产致病菌具有不同程度的抑菌作用。药敏试验结果显示,苏木、女贞子、夏枯草、青皮对4种水产致病菌均具有抑菌效果,抑菌圈直径在10.5~29.0 mm之间,而4种病原菌对马齿苋均表现为耐药;MIC检测结果显示,苏木、女贞子、夏枯草、青皮对4种病原菌的MIC值为7.81~250.00 mg/mL;MBC检测结果显示,苏木、女贞子、夏枯草、青皮对4种病原菌的MBC值为7.81~500.00 mg/mL;联合抑菌试验结果显示,苏木与夏枯草、女贞子及青皮联用仅对美人鱼发光杆菌起到协同抑菌作用,对嗜水气单胞菌、创伤弧菌及哈维氏弧菌均无抑菌作用,女贞子和青皮组合仅对哈维氏弧菌抑菌作用为相加作用,该组合在抑菌效果上对其他病原菌表现为无关或颉颃作用,其他药物组合在抑菌效果上对4种病原菌则表现为无关或颉颃作用;联合抑菌圈测定结果表明,中草药联合使用对4种病原菌抑菌作用不一。本试验结果表明常用中草药对水产动物致病菌具有一定程度的抑菌作用。     

中药单味药及复方对嗜水气单胞菌的体外抑菌作用

本试验用煎煮法提取中药有效成分,采用琼脂平板扩散法探讨了黄连、五倍子、大黄、诃子等中药单味药及中药复方对嗜水气单胞菌(Aeromonas hydrophila)的体外抑菌作用,测定了抑菌圈大小.采用双倍试管稀释法测得了最小抑菌浓度(MIC)和最小杀菌浓度(MBC).试验结果表明,16种中药提取物对嗜水气单胞菌有不同程度的抑制作用,其中五倍子、黄连、诃子、大黄的水提物的抑菌效果最好,MIC值分别为7.81 mg/mL、15,63 mg/mL、31.25 mg/mL、62.50 mg/mL.五倍子作为主药,分别与诃子、黄连、大黄接一定比例组成的复方的MIC分别为3.906 mg/mL、7.813 mg/mL、15.625 mg/mL.通过对筛选的复方与市场上销售的3种成品复方的体外抑菌活性的比较,发现本实验室筛选的复方对嗜水气单胞菌的体外抑菌活性更强.     

Rosemary oil,clove oil,and a mix of thyme-orange essential oils inhibit Salmonella and Campylobacter in vitro

The demand for foods that are free of pathogens and chemical residues has increased interest in the use of plant-based products as natural antimicrobials. Essential oils (EO) from plants are natural compounds that have been shown to have antimicrobial properties against food-borne pathogens. The objective of the current study was to determine the ability of various concentrations of 4 selected EO to inhibit Salmonella enterica (3 different serovars and a cocktail of all 3) and Campylobacter (2 strains of Campylobacter jejuni, one strain of Campylobacter coli, and a cocktail of all 3). The disc diffusion method was used to screen the oils of thyme, orange, rosemary, and clove oil. The minimum inhibitory concentration or minimum bactericidal concentration of the EO was determined using a 2-fold broth dilution method at concentrations ranging from 0.0008 to 1.000% (vol/vol). Two independent experiments were performed. Zones of inhibition (ZI) were expressed in millimeters and concentrations were expressed in percentages. All the oils demonstrated antibacterial activity against the strains tested. However, thyme oil demonstrated the strongest inhibitory activity than other oils against Salmonella (ZI of 18.5 mm). In general, Campylobacter was more susceptible to the antibacterial activity of EO, with plates containing thyme or clove oil showing no growth. Orange oil was also highly effective on Campylobacter, with a mean ZI of 17.5 mm. The least expensive treatment effective against both Salmonella and Campylobacter was a combination of 100% concentrations of thyme and orange oil combined on a 50:50 proportion. Tested on the same strains of bacteria, the thyme-orange combination (TOC) had a mean ZI of 20.5 mm for Salmonella and 21.3 mm for Campylobacter. Thyme-orange combination demonstrated a synergetic effect against Salmonella, but no such effect was noticed for Campylobacter. On average, 0.14% TOC was required to inhibit both pathogens. Hence, TOC can be considered as a potential antimicrobial for future studies on food systems.     

不同中药与盐酸恩诺沙星联用对大肠杆菌耐药菌株体外抑菌作用研究

本试验旨在研究不同中药与盐酸恩诺沙星联用对大肠杆菌耐药菌株的体外抑菌作用。采用微量肉汤法测定临床上一种常用药物盐酸恩诺沙星和不同中药对大肠杆菌耐药菌株及其联合使用的最小抑菌浓度(minimum inhibitory concentration,MIC)。结果发现,单味中药水提物中侧柏叶、金樱子和五味子的抑菌效果较好,其MIC值分别为3.91、3.91和7.81 mg/mL;单味中药的醇提物中,侧柏叶、马齿苋抑菌效果较好,其MIC值分别为7.81和3.91 mg/mL。盐酸恩诺沙星分别与五味子和金樱子水提物联合作用后,对大肠杆菌耐药菌株的MIC值分别降低到0.0078和0.0039 mg/mL;与马齿苋、赤芍、侧柏叶和金樱子的醇提物联合后,对大肠杆菌耐药菌株MIC值均明显降低。盐酸恩诺沙星分别与鱼腥草水提物、赤芍水提物、椿皮水提物、马齿苋醇提物、苍术醇提物、三颗针醇提物、赤芍醇提物、薤白醇提物、拳参醇提物、侧柏叶醇提物、金樱子醇提物和败酱草醇提物联合作用后,对大肠耐药菌株呈现相加或协同抑菌作用,其FIC值分别为0.75、0.62、0.62、0.27、0.75、0.75、0.31、0.62、0.50、0.53、0.28和0.62。初步认为不同中药中含有能有效降低大肠杆菌耐药性的中药,也含有与盐酸恩诺沙星组合,具有协同或相加作用的中药,为寻找有效防治大肠杆菌病的药物组合来对抗多重耐药菌提供了可能。     

Bacteriostatic Test in vitro of Traditional Chinese Medicine Radix Dichroa Powder on Common Pathogens

The purpose of this study was to indicate the antibacterial effect of traditional Chinese medicine Radix dichroa powder (RDP) in vitro. Minimal inhibitory concentration (MIC) of RDP for 12 pathogens were determined by two fold dilution method and agar dilution method, and the dose-effect relationship of antibiotic effect was studied, and the antimicrobial effect was also observed by Oxford-cup method. The results from two fold dilution method indicated that RDP had stronger inhibited effect to Streptococcus and Bacillus subtilis (MIC were from 15.6 to 62.5 mg/mL) than that of S. aureus (MIC was about 250 mg/mL), Enterobacteria (MIC was about 500 mg/mL) and fungi (MIC > 500 mg/mL); The results of agar dilution method showed that RDP had some antibacterial activities on Proteusbacillus vulgaris, Candida albicans And aspergillus niger, MIC was about 500 mg/mL, while it had weak antibacterial effect on other pathogens (MIC > 500 mg/mL). The results of Oxford-cup method showed that RDP (500 mg/mL) had some degree antimicrobial effect on pathogens, and the bacteria-inhibiting ring diameter of RDP not exceeded 10 mm, while drug-action ring was observed obviously around the Oxford-cup, and the number of bacteria was significantly decreased within the ring. In conclusion, RDP had some degree antibacterial effect on common pathogens, but the effects were not strong because of charicteristics of RDP itself and lower effective content.     

中药常山散的体外抑菌作用研究

试验旨在研究中药常山散的体外抑菌活性。分别采用试管二倍稀释法联合琼脂平板稀释法及营养琼脂稀释法对选用的12种致病菌进行抑菌试验,测定最低抑菌浓度（MIC）,进行抗菌作用量效关系研究,并通过牛津杯法观察药物抑菌效果。试管二倍稀释法联合琼脂平板法结果表明,常山散对链球菌属和芽孢杆菌的抑菌效果较强,MIC在15.6~62.5 mg/mL之间;对金黄色葡萄球菌和肠杆菌有较弱的抑菌作用,MIC在250~500 mg/mL之间;对真菌的抑菌作用最弱,MIC值 > 500 mg/mL,其抑菌强度大小依次为：链球菌属、芽孢杆菌、肠杆菌、真菌。营养琼脂稀释法结果表明,常山散对变形杆菌、白色念珠菌及黑曲霉具有一定的抑菌作用,MIC约为500 mg/mL,而对其他链球菌属、肠杆菌属及芽孢杆菌属细菌的抑菌效果较弱,MIC值均 > 500 mg/mL。牛津杯法抑菌活性研究结果显示,常山散药液（500 mg/mL）对12种菌均有一定的抑菌效果,但抑菌效果较弱,绝大多数抑菌环直径≤10 mm。牛津杯周围可见明显的药物作用圈,作用圈内细菌数量较其他部位明显减少。综合以上试验结果,中药常山散对常见致病菌均具有一定抑菌效果,但由于药物本身的特性及有效组分含量较低,其抑菌作用效果较弱。     

阿散酸-TMP的制备与体外抗菌活性研究

以阿散酸和甲氧苄啶(TMP)为原料,制备阿散酸-TMP,采用IR、熔点对其进行了表征;正交试验确定了最优反应条件为:反应温度80 ℃,反应时间3 h,反应物料比1.1∶1(阿散酸∶TMP)。通过体外抑菌试验来测试其抑菌效果,结果表明,阿散酸-TMP对5种供试菌的抑菌圈直径都在20 mm以上,显著大于阿散酸与TMP的混合物(P＜0.05);阿散酸-TMP对埃希氏大肠杆菌、金黄色葡萄球菌、鸡白痢沙门氏菌、福氏志贺氏菌、藤黄微球菌的MIC分别为32、64、256、32、128 μg/mL,均小于阿散酸与TMP的混合物。阿散酸-TMP对供试菌的抑菌效果好于阿散酸、TMP及阿散酸与TMP的混合物。     

Study on the Effect of the Rosemarinic Acid Combined with Antibacterial Agents on the Bacterial with Gene fosA3

This research was aimed to investigate the effect of rosemarinic acid combined with antibacterial agents against bacterial carrying gene fosA3 in vitro.Resistance gene types were determined by identification of bacteria isolated in clinics.Antibacterial activity and fungicidal activity of rosemarinic acid were tested by oxford cup method and spread-plate method.The minimal inhibiton concertration (MIC) of rosemarinic acid and antibacterial agents were tested through twice micro-dilution method,fractional inhibitory concentration index (FICI) of rosemarinic acid combined with antibacterial agents were determined by microdilution checkerboard techniques.Escheriohia coli carrying multidrug resistance gene fosA3 was isolated.The MIC of rosemarinic acid was 640 μg/mL,when application of rosemarinic acid combined with ciprofloxacin,levofloxacin,coly-mycin,gatifloxacin,amikacin,ceftazidime,ceftiofur sodium,FICI≤0.5,showed an additive effect;With ceftriaxone sodium,norfloxacin,mequindox,0.5fosA3 Escheriohia coli.Antibacterial activities of antibacterial agents against the fosfomycin resistance gene fosA3 Escherichia coli were enhanced significantly by rosemarinic acid.     

迷迭香酸与抗菌药联合对含fosA3基因细菌抑菌效果的研究

试验旨在探讨迷迭香酸与抗菌药联合体外抗含fosA3耐药基因大肠杆菌的效果。本试验对临床分离的细菌进行鉴定,确定细菌所含耐药基因种类,用牛津杯法测定迷迭香酸的抑菌活性,平板涂布法测定迷迭香酸的杀菌活性,采用96孔反应板二倍微量稀释法分别检测迷迭香酸与抗菌药的最小抑菌浓度(MIC),再用微量棋盘稀释法测定迷迭香酸与抗菌药联合作用后的分级抑菌浓度指数(FICI)。结果显示,分离到含fosA3耐药基因大肠杆菌;迷迭香酸的MIC为640 μg/mL,迷迭香酸与环丙沙星、左旋氧氟沙星、黏杆菌素、加替沙星、阿米卡星、头孢他啶、头孢噻呋钠联合应用后其FICI≤0.5,有协同作用;与头孢曲松钠、诺氟沙星、痢菌净联合应用后0.5fosA3基因大肠杆菌有一定的抑菌活性和杀菌活性,其与部分抗菌药联合应用后,抗菌药体外抗菌活性显著增强。     

中药水提物及其复方对大肠杆菌的体外抑菌效果

为了评价中药及其复方对大肠杆菌的体外抑菌效果,取乌梅、五倍子、山茱萸、黄芩等30种中药,通过试管二倍稀释法测定中药水提物对大肠杆菌的最小抑菌浓度（MIC）与最小杀菌浓度（MBC）,并筛选出抗菌作用较强的中药水提物与其他中药水提物组成复方,测定中药复方对大肠杆菌的体外抑菌效果。结果表明,五倍子、乌梅、金银花、大青叶、黄芩、薄荷、五味子、青蒿、秦皮对大肠杆菌有明显的抑制作用,其中五倍子对大肠杆菌的抑菌活性最强（MIC为6.25 mg/mL）,而五味子和乌梅对大肠杆菌杀菌活性最强（MBC为50 mg/mL）。选取9种中药水提物作为主药与其他中药水提物分别组成复方后,大部分复方对大肠杆菌的抑制作用明显增强。     

4味中药及其与抗菌药的复方制剂的MIC测定

用改进的试管两倍稀释法测定了黄连、鱼腥草、大青叶和苦参4味中药及其相互配伍（1：1）对标准大肠杆菌的最小抑菌浓度和4味中药与抗菌药配伍（1000：1）对临床分离鸡大肠杆菌的最小抑菌浓度。结果表明，4味中药及其相互配伍对大肠杆菌标准株有一定的抑制作用，其MIC值介于3．91—62．50mg／mL之间，其中大青叶和鱼腥草以1：l配伍对大肠杆菌抑菌效果最好，MIC为3．91mg／mL；对临床分离株，单方药中黄连抑菌效果最好，MIC为31．25mg／mL，24种复方药中，黄连和盐酸多西环素或加替沙星以1000：1配伍的抑菌效果最好，其MIC为3．91mg／mL。     

金黄色葡萄球菌gyrA、gyrB、grlA、grlB和norA基因突变与环丙沙星耐药性的关系

采用药敏纸片法测定了临床分离的160株金黄色葡萄球菌对14种抗菌药物的耐药性，同时进行了MRSA的检测，并采用琼脂稀释法测定纸片法筛选出的45株金黄色葡萄球菌对环丙沙星的最低抑菌浓度（MIC）。根据金黄色葡萄球菌gyrA、gyrB、grlA、grlB和norA基因的核苷酸序列设计引物、以筛选出的45株金黄色葡萄球菌染色体DNA为模板．对gyrA、gyrB、grlA、grlB和norA基因片段进行PCR扩增，回收PCR产物．并与pMD18-T载体连接．转化DH5a感受态大肠杆菌．提取质粒并测序。进一步分析考查金黄色葡萄球菌gyrA、gyrB、grlA、grlB和norA基因及其启动子区突变与环丙沙星耐药性的关系。结果表明．grlA中的2个突变（位于80和84密码子）及gyrA中的2个突变（位于84和88密码子）与环丙沙星耐药性明显相关。     

中草药提取物对多重耐药猪胸膜肺炎放线杆菌体外抑菌试验及抑菌效果研究

为筛选对猪接触传染性胸膜肺炎放线杆菌多重耐药菌株有较好抑菌作用的中草药。挑选了穿心莲、黄芩、苦参、白头翁、芒果叶和百里香6种中草药,采用水提取和65%乙醇提取相结合的方法制备单剂中草药的提取液,然后浓缩至含原生药物1g/mL。采用二倍稀释法分别测定这6种中草药对传染性胸膜肺炎放线杆菌多重耐药菌株的最小抑菌浓度(minimal inhibition concentration, MIC)和最小杀菌浓度(minimal bacteriocidal concentration, MBC)。以筛选出体外抑菌效果较好的中草药。结果表明,胸膜肺炎放线杆菌多重耐药菌株抑菌圈最大的是百里香和芒果叶,其抑菌圈直径分别为40 mm和35 mm.,其次为白头翁、黄芩、穿心莲和苦参,其抑菌圈直径在8~16 mm。百里香、芒果叶、白头翁、黄芩、穿心莲和苦参的最小抑菌浓度分别为0.198 mg/mL、0.198 mg/mL、0.396 mg/mL、0.396 mg/mL、25 mg/mL和25 mg/mL。百里香、芒果叶、白头翁、黄芩、穿心莲和苦参的最小杀菌浓度分别为0.396 mg/mL、0.396 mg/mL、0.792 mg/mL、0.792 mg/mL、50 mg/mL和50 mg/mL。说明除穿心莲和苦参外,其余4种中草药对胸膜肺炎放线杆菌多重耐药菌株有很多好的抑菌和杀菌效果,其中百里香和芒果叶效果最佳。本结果为由传染性胸膜肺炎放线杆菌多重耐药菌引起的猪接触传染性胸膜肺炎的防控和治疗奠定了基础,为开发中草药相关无抗产品以及防治由多重耐药菌引起的猪接触传染性胸膜肺炎提供了参考。     

32种中药及其复方对嗜水气单胞菌体外抑菌效果

为评价中药对嗜水气单胞菌体外抑菌效果,取五倍子、秦皮、乌梅等32种中药,采用水提法制备药液．通过微量二倍稀释法测定各药液对体外培养的嗜水气单胞菌最小抑菌浓度（MIC）及最小杀菌浓度（MBC）,筛选出抗菌作用较强的中药作为主药,结合中兽医理论,与牡丹皮、龙胆草、蒲公英等18种中药组成复方,测定中药复方对嗜水气单胞菌的MIC和MBC。结果表明,五倍子、诃子、栀子等8种中药对嗜水气单胞茵有较强抑菌作用．8种主药与清热凉血、清热燥湿、清热解毒药分别组成复方,其中五倍子、诃子、乌梅与其他多种中药组成的复方抗菌作用增强,同时,主药与主药组成复方后抗菌效果明显提高。     

苍术油对四种细菌最小抑菌浓度的研究

本研究通过试管2倍稀释法测定了苍术油对大肠杆菌(44102)、金黄色葡萄球菌(26003)、枯草芽孢杆菌(63501)和沙门氏菌的最小抑菌浓度(MIC)。结果:苍术油对大肠杆菌、金黄色葡萄球菌和枯草芽孢杆菌的最小抑菌浓度分别为:117.2μg/ml、625μg/ml和312.5μg/ml,而对沙门氏菌没有任何抑菌作用。研究证明苍术油对部分细菌具有良好的抑制作用。