蛋源摩氏摩根菌分离、鉴定及遗传进化分析

本研究利用传统细菌鉴定方法、革兰氏阴性细菌生化鉴定系统及16S rDNA 序列分析对分离自市售新鲜鸡蛋中的菌株进行形态学、培养特性及生化特性等生物学特性鉴定及遗传进化分析,结果表明分离株为摩氏摩根菌。药物敏感性试验结果表明,该菌株对痢特灵、利福平、四环素、磷霉素、多黏菌素B和阿奇霉素有耐药性,属于多重耐药菌株。序列分析结果显示,该分离菌株与目前已分离的其他食源性摩氏摩根菌菌株高度同源。     

鲈鱼摩氏摩根菌的鉴定及药敏试验

从患病鲈鱼体内分离到1株优势菌LY2014,对其进行形态观察、生理生化试验、16SrDNA基因序列分析,同时采用Kriby-Bauer纸片扩散法进行耐药性分析。结果该菌为发酵型、具有动力的革兰阴性杆菌,根据生化试验结果初步确定该菌为摩氏摩根菌。以该菌的基因组为模板PCR扩增获得的16SrDNA序列长度为1 407bp(GenBank登入号为KJ830813)。该序列与GenBank数据库中摩氏摩根菌的16SrDNA基因序列相似性高达99%,系统进化树显示其与摩氏摩根菌亲缘关系最近,从而确定菌株LY2014为摩氏摩根菌。该菌株对磷霉素、菌必治和多黏霉素B高度敏感,对痢特灵、红霉素和诺氟沙星等5种药物中度敏感,对氨苄西林和氟苯尼考等11种药物耐药。     

患病大鲵摩氏摩根菌的分离与鉴定

从患病大鲵肝脏中分离到1株优势菌SX01,通过Biolog微生物自动鉴定系统、16S rDNA序列分析及构建系统发育树的方法对纯培养的细菌进行鉴定,并进行人工感染试验及药敏试验。结果表明,该分离菌为摩氏摩根菌(Morganella morganii),对大鲵具有致病性。药物敏感性试验表明,该菌株对头孢曲松、头孢吡肟、氨曲南等10种药物高度敏感。     

1株猪源摩氏摩根菌的分离及鉴定

从猪小肠内容物分离得到1株疑似致病菌进行鉴定。革兰染色显示,该菌为革兰阴性短杆菌;16S rDNA及同源性结果,确定其为摩氏摩根菌,命名QX01。回归试验表明,该菌能够引起昆明系小鼠肝脏和肺脏不同程度充血和出血,具有较强的致病力。19种药物的敏感性试验结果表明,该菌对头孢噻肟、左氧氟沙星和环丙沙星等10种药物敏感,对恩诺沙星、磺胺二甲嘧啶、红霉素等5种药物耐受。本试验首次从猪小肠内容物分离的摩氏摩根菌具有较强致病力,其潜在危害应引起重视。     

重庆黑山羊摩氏摩根菌的分离鉴定与药敏试验

从重庆健康黑山羊上呼吸道分离到1株革兰阴性短杆菌,通过形态学观察、16S r DNA及系统发育树分析,确定该菌株为摩氏摩根菌,命名为RC01。致病性试验显示,菌株RC01对小鼠致病性较强。药物敏感性试验显示,RC01对卡那霉素、萘啶酸、头孢噻肟、左氧氟沙星较为敏感;对青霉素、氨苄西林、大观霉素、克拉霉素耐药。本研究首次从健康山羊上呼吸道分离到对小鼠具有较强致病性的摩氏摩根菌病原,其潜在危害应引起人们的重视。     

台湾泥鳅死亡致病菌的分离鉴定及药敏试验

为确定捕捞转场后造成台湾泥鳅（Paramisgurnus dabryanus ssp.Taiwan）死亡的病原,本试验对台湾泥鳅进行病理解剖、病原分离。对分离菌进行形态学观察、生理生化特性测定及16S rDNA基因测序鉴定,并进行人工感染试验、药敏试验。结果显示,从泥鳅肝脏、脾脏、肾脏及肠道组织中分离到形态一致的优势菌株,经纯化保存,命名为XJC。菌株XJC葡萄糖发酵、D-葡萄糖、鸟氨酸、苯丙氨酸及尿素利用阳性,其他所测生化指标均为阴性,与摩氏摩根菌（Morganella morganii）一致;16S rDNA基因序列与摩氏摩根菌相似性达99.6%,被鉴定为摩氏摩根菌。感染试验结果表明,菌株XJC对台湾泥鳅具有致病性,被感染的泥鳅出现了肠道充血典型病理变化。从发病泥鳅体内分离到与菌株XJC一致的菌株,可以判定菌株XJC是泥鳅病原菌。药敏试验结果表明,菌株XJC对氨基糖苷类、喹诺酮类及第三代头孢类药物表现出敏感,对四环素、青霉素、红霉素、多肽类、呋喃唑酮、洁霉素及第一、第二代头孢类药物均等表现耐药,生产中可选用氨基糖苷类、喹诺酮类药物进行摩氏摩根菌疾病的防治。本试验结果可为台湾泥鳅摩氏摩根菌病诊断和防治提供参考。     

1例摩氏摩根菌引起BALB/c小鼠感染的诊治

本试验对某实验动物养殖场送检的一批BALB/c小鼠进行剖检,观察病理变化,对病变组织进行细菌的分离纯化鉴定。利用16S rRNA基因对分离细菌进行PCR扩增、测序及系统进化树分析。分离菌株进行动物回归试验和药敏试验。经形态学及分子生物学鉴定,分离菌株为摩氏摩根菌,对小鼠有很强的致病性。药敏试验表明,该菌耐药性高,仅对新霉素、链霉素、左氧氟沙星、氟苯尼考敏感,选用敏感药物新霉素和氟苯尼考进行联合用药,疫情得到有效控制。     

青海湖裸鲤体表白点病病原菌的分离与药敏试验

为确定养殖青海湖裸鲤(Gymnocypris przewaiskii)体表白点病的病原,分别对患病鱼白色点状囊泡样物进行了细菌分离、生理生化特性鉴定、16SrRNA基因分析和药敏感试验,分离到了以弗氏柠檬酸杆菌(Citrobacter freundii)、维氏气单胞菌(Aeromonas veronii,AV)和摩氏摩根菌(Morganella fulton)为代表的菌落形态一致、革兰阴性细菌;且分离到的3种菌都对头孢吡肟、丁胺卡那霉素、庆大霉素等药物敏感。     

节尾狐猴摩氏摩根菌的分离鉴定与药敏试验

为探究1例2月龄雄性节尾狐猴幼崽死亡的病因,取其肝脏、肾脏等组织样本进行细菌培养,分离出的菌株通过菌落形态观察、革兰染色镜检、生理生化特性鉴定,确定为摩氏摩根菌。并对分离菌株进行了药物敏感试验,显示该菌对临床多种常用抗生素的耐药性较高,应引起足够的重视。     

兔源肺炎克雷伯氏菌的生物学特性研究

为探究兔源肺炎克雷伯氏菌的生物学特性,本研究对从病兔中采集的病料进行细菌的分离培养、细菌特征性基因(Khe基因)的PCR检测、生化鉴定、药敏试验、致病性试验、毒力基因的扩增及序列分析对细菌分离株进行分析。结果显示麦康凯平板上生长有奶油状菌落且镜检为革兰氏阴性菌;Khe基因扩增为阳性;生化鉴定结果表明分离菌为肺炎克雷伯氏菌;药敏结果显示该菌对诺氟沙星、氧氟沙星、头孢曲松等9种药物高敏,对链霉素、卡那霉素、头孢呋辛等5种药物中敏,对先锋V、庆大霉素等7种药物不敏感甚至耐药;致病性结果显示,分离株能使小白鼠在24h内死亡;分离株携带有wabG、uge和fimH三种毒力基因。本研究结果表明该肺炎克雷伯氏菌具有很强的致病性,是导致兔死亡的主要病原菌。     

Plasma biochemical reference intervals for koi

OBJECTIVE: To assess reproducibility of an in-house tabletop biochemical analyzer for measurement of plasma biochemical analytes and establish reference intervals in adult koi. DESIGN: Prospective study. ANIMALS: 71 healthy adult koi. PROCEDURES: Plasma was analyzed for concentrations or activities of albumin, alkaline phosphatase, alanine aminotransferase, aspartate aminotransferase, bile acids, BUN, calcium, cholesterol, creatine kinase, gamma-glutamyltransferase, globulin, glucose, K, Na, P, total bilirubin, total protein, and uric acid. Duplicate samples were evaluated by use of the intraclass correlation coefficient to determine reproducibility. To assess the magnitude of differences between replicate samples, the absolute mean difference, SD, and minimum and maximum values were calculated for each analyte. Median values and reference intervals were calculated. RESULTS: Intraclass correlation coefficient values were excellent for all analytes except alanine aminotransferase (good), Na (poor), gamma-glutamyltransferase (poor), and P (poor). Reference intervals were established. CONCLUSIONS AND CLINICAL RELEVANCE: The in-house tabletop biochemical analyzer had good precision for measuring most plasma biochemical analytes. Further research and comparison with other reference procedures are needed before reference intervals and precision can be established for globulin, Na, P, K, and albumin. Aquatic veterinarians may be able to use the reference intervals for adult koi as an important diagnostic tool or as part of a fish wellness program, as commonly done in other domestic species.     

The delimitation of intestine segments of koi carp (Cyprinus carpio var. koi) based on histological features

In this study, the delimitation of intestine segments of koi carp (Cyprinus carpio var. koi) was conducted using a histological approach with the measurements of height of mucosa folds (HF), width of mucosa folds (WF), thickness of muscularis (TM) and cross-sectional area (CSA). According to the change trends for these four parameters, the intestine of the koi carp was divided into anterior intestine, middle intestine and posterior intestine. The locations of the three intestine segments were defined, and their ratios along the entire intestine were accounted for 23.84 ± 1.18%, 46.77 ± 2.29% and 29.39 ± 1.65%, respectively. The anterior intestine had a significantly higher HF, compared with the middle (p < .01) or posterior intestines (p < .01). The muscularis became thin gradually from the anterior intestine to posterior part. TM was significantly different among the anterior, middle and posterior intestines (p < .01). The anterior intestine had a significantly higher CSA, compared with the middle (p < .01) and posterior intestines (p < .01), and the latter two segments had similar CSA values (p > .05). The procedure of the delimitation of the koi carp intestine segments can offer useful information for future studies on other fish species. The presented results are meaningful for studies on differential functions of the different intestine segments in fish.     

Surveillance of herpesviruses in koi carp Cyprinus carpio koi and goldfish Carassius auratus cultured in West Bengal,India

Background: Viral diseases create one of the greatest challenges for the rapidly expanding ornamental fish culture sector. In recent years, the host-specific herpesviruses are receiving much attention due to the intensification in aquaculture globally. Cyprinid herpesvirus-2 (CyHV-2, goldfish hematopoietic necrosis virus), and Cyprinid herpesvirus-3 (CyHV-3, koi herpesvirus) are the lethal pathogens of koi carp Cyprinus carpio koi and goldfish Carassius auratus, respectively. In India, West Bengal is the leading state in ornamental fish production and export. Methods: The surveillance of CyHV-2 and CyHV-3 in koi carp and goldfish cultured in West Bengal was carried out between November 2014 and January 2017 as per Office International des Epizooties guidelines. Results: Both fish species were negative for CyHV-3. The CyHV-2 infection was detected in both gill and kidney tissues of apparently healthy and diseased C. auratus from December 2014 to March 2015. Severe necrosis was observed in the gills of infected C. auratus. Coinfection with members of the bacterial genera Aeromonas and Flavobacterium was also observed in the kidney of infected goldfish. The histopathological observations in the kidney of CyHV-2 infected C. auratus demonstrated the pathogenic potential of CyHV-2 and tissue tropism. Conclusions and Clinical relevance: Environmental stressors like low water temperature and the use of wastewater for culture played a vital role in the onset of CyHV-2 infection in the stressed goldfish. The spread of CyHV-2 can likely pose a certain degree of threat to aquaculture especially the unrestricted movement of goldfish. The results of the present study emphasize the necessity of an organized risk assessment to plan for the management strategies on the control and spread of CyHV-2 in Indian aquaculture.     

锦鲤疱疹病毒GY1506株的分离鉴定

对江苏省某养殖场患病鲤鱼进行病毒分离及鉴定。现场采样发现,发病鱼体表黏液增多,眼球凹陷,背鳍和鳃部溃烂。剖检后见肝脏、肾脏、胆囊肿大,肠充血。取病鱼组织匀浆上清接种CCB细胞,14 d后可观察到细胞变圆、脱落、空泡化等典型的细胞病变。采用世界动物卫生组织(OIE)推荐的锦鲤疱疹病毒(KHV)检测引物进行PCR扩增,鳃和肠均能扩增出预期大小的特异性产物。序列分析显示,扩增序列与KHV-J毒株胸苷激酶(TK)基因核苷酸序列同源性为100%。根据TK基因序列建立系统进化树,证实该毒株为KHV亚洲型毒株,命名为KHVGY1506株。     

利用PCR方法检测锦鲤疱疹病毒3个主要靶基因

为建立锦鲤疱疹病毒(KHV)多靶基因PCR检测方法,本实验将KHV接种鲤鱼鳍条细胞,收获病变细胞悬液,提取DNA,根据GenBank中登录的KHV基因序列及出入境检验检疫行业标准推荐的基因(ORF7),设计合成3对特异性引物,针对胸苷激酶基因(TK)、聚合酶基因(Sph)和ORF7基因进行PCR检测.通过优化后的反应体系进行特异性、敏感性试验和样品检测.结果表明:3对引物能够分别特异性扩增出409bp、292 bp和484bp片段;敏感性试验表明对TK基因检测的敏感性高于Sph和ORF7基因,其最低检测量为1.9×106copies/μL;采用优化的3个PCR方法对8个有临床症状的样品进行检测,其中3份样品的3个基因PCR扩增结果均为阳性.因此,本研究选取的3个基因均可用于KHV的检测及确证实验.     

Reproductive medicine in koi (Cyprinus carpio).

Each year more veterinarians are seeing ornamental fish patients and our knowledge of how to manage the pet fish patient is increasing at a rapid pace. A large percentage of these patients are nishikigoi (Cyprinus carpio) which are taxonomically an ornamental carp. In this country and in many parts of the world we refer to these fish as koi. Reproductive medicine problems are well documented in koi and a thorough knowledge of this subject will aid the fish practitioner in diagnosis and treatment.     

红白锦鲤的人工催产及胚胎发育研究

通过对红白锦鲤的胚胎发育形态特点及发育特点的研究,红白锦鲤成熟卵为黏性卵,卵径1.8 mm。发育期间水温为18~23℃。30 min受精后,受精卵吸水膨胀到最大,胚盘隆起,55 min后进入卵裂期,6 h后进入囊胚期,11 h后进入原肠胚期,15 h 22 min胚孔封闭,30 h 29 min后肌节收缩,31 h 21 min后心跳开始,43 h54 min出膜。初孵仔鱼长7 mm,出膜后2~3 d,胸鳍、鳃、口、体内血管等器官和色素相继发育完全,99 h后鳔充气,鱼苗开始平游和进食。红白锦鲤胚胎整个发育期,卵裂期、原肠期、尾芽期发育较慢,而在尾芽出现后发育较快。通过试验发现红白锦鲤自然产卵产生的卵质量优于人工催产。胚胎发育最适温度在20~23℃,出苗数目1~1.3万条/m3,与尼罗罗非鱼、盘丽鱼比较,在神经胚期、眼基形成期及出膜仔鱼体色上有所差异。     

Herpesvirus-associated papillomas in koi carp (Cyprinus carpio).

From January through November 1994, 32% (7/22) of koi carp (Cyprinus carpio) maintained in indoor aquariums developed proliferative cutaneous lesions that consisted of single to multiple 2-10-mm whitish to pink fleshy masses usually associated with fin rays. Although scaleless koi were more commonly affected (3/6) than were normally scaled koi (4/16), the difference in incidence rates was not significant (chi2 text, P > 0.05). Lesions typically resolved spontaneously in 1-3 wk, occasionally persisted for >3 mo, and recurred in several fish after 2-5 mo. Fish were otherwise asymptomatic. Wet mount preparations from lesions were densely cellular and consisted of hyperplastic epidermal cells of normal morphology without parasites or inflammatory cells. Histologically, biopsies were consistent with papillomas and were characterized by a marked benign epidermal hyperplasia without inclusion bodies or inflammatory infiltrate. Transmission electron microscopic examination revealed intranuclear and intracytoplasmic herpesvirus virions. Virus isolation attempts were unsuccessful.