The detection of Vibrio parahaemolyticus and Vibrio alginolyticus using modified culture media

Studies have been completed for the detection of Vibrio(V.) parahaemolyticus and V. alginolyticus in Baltic Sea fish. The author had prepared for that purpose a liquid culturing medium and a solid substrate. The percentage of positive findings is remarkable. The results so far recorded should be a good reason for further studies.     

副溶血弧菌和霍乱弧菌双重荧光定量PCR快速检测方法的建立与应用

建立一种基于TaqMan探针法同步定量检测副溶血弧菌和霍乱弧菌的双重荧光定量PCR方法。根据副溶血弧菌toxR基因和霍乱弧菌ompW基因设计特异性引物与TaqMan探针。toxR和ompW探针5'端分别标记FAM、CY5荧光报告基团,3'端均标记BHQ1荧光淬灭基团。结果显示:该方法的副溶血弧菌和霍乱弧菌最低检测限均达到10 CFU/m L,灵敏度高;特异性试验表明这两种细菌与其他病原菌(大肠杆菌O157、沙门氏菌、拟态弧菌、创伤弧菌)无交叉反应;批间和批内重复性实验表明变异系数均小于1.5%,说明重复性好。采用人工染菌虾肉样品,副溶血弧菌和霍乱弧菌的最低检测限分别为100 CFU/m L和50 CFU/m L,人工染菌贝类样品的最低检测限分别为50 CFU/m L和50 CFU/m L。两种细菌在虾肉中富集2 h后的最低检出限均为1 CFU/m L。结论:本研究所建立的双重荧光PCR检测方法具有灵敏度高、特异性强和重复性好的特点,包括DNA提取的整个检测过程可在1至3 h内完成,是同时快速检测副溶血弧菌和霍乱弧菌的有效手段。     

牛奶中土霉素对青海弧菌和费氏弧菌的毒性作用测定

本文通过应用费氏弧菌和青海弧菌,探究了牛奶中土霉素对这两种发光细菌的毒性抑制作用规律。牛奶中,一定浓度下,土霉素对青海弧菌生物发光的抑制率随作用时间呈对数趋势增加,在0.45mg/L作用下,该关系方程为y=0.3007ln（x）-0.1940,R2=0.9934;作用5min时,土霉素抑制率随着浓度变化的规律呈直线关系,之后呈对数关系,E50值在5min时为0.75mg/L,30min时降低至0.12mg/L;土霉素对费氏弧菌生物发光的抑制率随作用时间呈对数趋势增加,在0.55mg/L作用下,y=0.2742ln（x）-0.0942,R2=0.9961;其抑制率随着浓度变化的规律在5min呈指数关系,之后呈对数关系,在作用10min时,y=0.5524ln（x）＋0.9045,R2=0.9949,E50值在5min时为0.75mg/L,40min时降低至0.09mg/L。     

Vibrio Anguillarum: A Comparative Study of Fish Pathogenic,Environmental, and Reference Strains

An examination of 41 fish pathogenic and 51 environmental strains of Vibrio anguillarum from Danish coastal areas, and of 8 reference strains of Vibrio anguillarum, biotype 1, showed that these strains shared rather broad-spectered activities towards many carbohydrates, amino acids, proteins and lipids.They also exhibited rather uniform growth characteristics, including parameters as salinity (NaCl), temperature, antibiotic resistance, and growth on specific media.A computer analysis of these strains and 3 strains of Vibrio anguillarum, biotype 2 (Vibrio ordalii), showed that the Danish strains all belonged to Vibrio anguillarum, biotype 1, and that no basis existed for setting up new biotypes.Mean values of the G+C content in DNA of strains of the various categories of Vibrio anguillarum, biotype 1, ranged from 45.5 to 46, while the mean value for the Vibrio ordalii strains was 48.9.The biochemically active Vibrio anguillarum seems more suitable for environmental studies than the more fastidious Vibrio ordalii.Key words: Vibrio anguillarum, Vibrio ordalii, biochemical activities, environmental, fish pathogenic, reference strains     

副溶血弧菌和溶藻弧菌双重PCR检测方法的建立与初步应用

为建立同时快速检测海产品中的副溶血弧菌(VP)和溶藻弧菌(VA)的双重PCR方法,本研究根据VP和VA的toxR基因序列设计针对这两种细菌的两对特异性引物,建立能够快速同时检测这两种细菌的双重PCR方法,并对该反应体系的特异性和灵敏度进行检测。结果显示纯培养细菌VP和VA的检测灵敏度分别为2.32×103cfu/mL和2.56×103cfu/mL,临床病料检测灵敏度分别为2 cfu和3 cfu;与枸橼酸杆菌、沙门氏菌、美人鱼弧菌、霍乱弧菌、麦氏弧菌无交叉反应。研究表明本实验方法操作简便快速、特异性强、灵敏度高、稳定性好,并且经济实惠,值得推广应用。     

Antibiotic Resistance of Vibrio anguillarum,in Relation to Serovar and Plasmid Contents

A total of 520 Vibrio anguillarum strains, isolated from fish and the environment, were tested for their sensitivity to 20 different antibiotics. Most isolates were of European origin. The results were compared with data on the O-serogroup and plasmid contents. All strains were sensitive to neomycin, spectinomycin, nitrofurantoin, flumequine and oxolinic acid, while most strains were sensitive to streptomycin, Oxytetracycline, chloramphenicol, sulphonamides, trimethoprim, sulphonamides with trimethoprim, nalidixan, rifampicin, novobiocin and O/129. A major part of the strains were resistant to the macrolides, spiramycin and lincomycin. For ampicillin, cephalothin, and Colistin marked differences were recorded with respect to O-serogroup. Most O1 strains were resistant to Colistin and sensitive to ampicillin and cephalothin, while most O2 strains were sensitive to Colistin but resistant to ampicillin and cephalothin. Some antibiotic resistant strains carried plasmids but no conjugation experiments were carried out to detect possible R factors.     

圆眼燕鱼哈维弧菌的分离鉴定及药敏试验

从患病圆眼燕鱼(Platax orbicularis)体内分离到一批形态一致的优势菌株,通过细菌鉴定并检测其药物敏感性,以期为圆眼燕鱼的疾病防治提供参考。对分离菌株进行形态观察、生理生化试验以及16S rRNA基因序列分析,结果显示该菌株与弧菌表型特征一致,16SrRNA与GenBank中哈维弧菌(Vibrio harveyi)的同源性高达99%,在系统进化树中与哈维弧菌(KU245727)聚为一分支,结合该菌的生理生化特征和基因序列分析,鉴定该优势菌株为哈维弧菌。药敏试验结果显示,该分离菌对头孢噻肟、强力霉素、恩诺沙星、萘啶酸、卡那霉素、妥布霉素、氯霉素、复方新诺明等14种药物敏感,对阿莫西林、青霉素G、乙酰螺旋霉素、杆菌肽、多黏菌素B、利福平、红霉素和磺胺异恶唑等8种药物表现为耐药。采用PCR方法检测该菌株携带TEM、Sul2、ANT(3′′)-1耐药基因。研究结果可为哈维弧菌的鉴定及圆眼燕鱼的疾病诊治提供参考。     

Pathogenicity of Vibrio Coli for Swine II. Experimental Infection of Conventional Pigs with Vibrio Coli

The pathogenicity of V. coli for conventional swine was studied by inoculating pigs with cultures of V. coli and V. coli infected gut of gnotobiotic pigs. Thus, six conventional pigs were inoculated with strains of V. coli freshly isolated from infected gnotobiotic pigs. The cultures were grown in simulated sows milk, and added to the feed. Two other groups, of three pigs each, were infected by administration of minced intestine from two gnotobiotic pigs, heavily infected with the organism. Vibrio was isolated from all pigs, including five of the six controls, but larger numbers were isolated from the inoculated groups, especially from those fed macerated gut. Clinical signs of disease were not observed.     

Pathogenicity of Vibrio Coli for Swine I. Experimental Infection of gnotobiotic Pigs with Vibrio Coli

Fifteen gnotobiotic pigs varying in age from three to eight weeks were exposed to 23 strains of V. coli isolated from swine with clinical and/or pathological signs of swine dysentery and also from clinically healthy pigs. Clinical or pathological signs of swine dysentery were not produced, although the organism was readly established in the sedimentary tract. Culture of feces, alimentary tract and environment revealed V. coli in large numbers, but no bacterial growth was obtained from other organs. The histopathology and serology are discussed.     

副溶血性弧菌分离鉴定

利用TCBS琼脂培养基和科玛嘉弧菌显色培养基对细菌进行分离,采用糖分解试验、氧化酶试验、IMVi试验等生化试验鉴定海产品中副溶血性弧菌,为防治疾病提供依据。