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。     

云南鸡源副禽嗜血杆菌的分离鉴定与耐药性分析

无菌棉签深入病死鸡鼻窦腔深部无菌采集病料进行细菌分离,对分离菌进行形态学观察、生理生化试验、革兰氏染色、血凝活性测定、16S rDNA序列测定和药敏试验。结果在富含V因子血琼脂平板上分离到一株灰白色、圆形、边缘整齐的光滑型小菌落,菌落在血琼脂平板、麦康凯等培养基上无明显菌落生长;镜检观察为革兰氏阴性菌,具有血凝活性,符合副禽嗜血杆菌生化培养特性;细菌16S rDNA核酸序列Blast比对结果显示为副禽嗜血杆菌。分离菌株对妥布霉素、阿奇霉素、庆大霉素、阿米卡星具有敏感性。     

小熊猫肺炎克雷伯菌的分离与鉴定

为探究1例10余岁小熊猫死亡的病因,无菌采集死亡小熊猫的肝、脾组织进行细菌分离培养。实验结果显示,在培养基上均有细菌生长,且菌落生长形态相似;分离菌革兰染色镜检、生化鉴定结果表明,各分离菌株特征一致,为肺炎克雷伯菌。小白鼠致病性试验结果显示,分离菌株具有较高致病性。     

环境因子对东北林蛙胚胎期和变态期影响的研究进展

东北林蛙因其具有药用价值与经济价值,日益受到人们的重视。近年来,我国学者开展了关于东北林蛙养殖和生态学的研究。文章着重介绍温度对东北林蛙生长发育、性别分化的影响和营养因子对东北林蛙变态期的影响,同时就环境因子对东北林蛙不同发育阶段影响的研究现状进行了综述,并分析了在东北林蛙养殖中应注意的与环境相关的问题。     

东北林蛙烂趾病病原菌的分离鉴定及药敏试验

对东北林蛙烂趾病病原菌进行分离鉴定,并对其药物敏感性进行研究。结果表明:从东北林蛙患病趾处分离到1株溶血性致病菌,经16S rDNA序列分析法鉴定为嗜水气单胞菌。通过对23种抗生素的药敏试验表明,此菌对氟喹诺酮类(如环丙沙星、培氟沙星、诺氟沙星),部分头孢类抗生素(如头孢呋辛、头孢噻肟、头孢吡肟),β-内酰胺类抗生素(如亚胺培南、氨曲南)高度敏感;对部分头孢类抗生素(如头孢噻吩、头孢西丁、头孢他啶),氨基糖苷类抗生素(如卡那霉素、庆大霉素、链霉素)中度敏感;对青霉素类如(青霉素、苯唑西林、氨苄西林、阿莫西林)有耐药性。说明可以考虑用氟喹诺酮类、头孢类或β-内酰胺类抗生素防治东北林蛙烂趾病。     

低温环境对东北林蛙心率的影响

为了分析低温环境下东北林蛙(Rana dybowskii)心率的变化,为其养殖、运输或暂养提供基础数据,试验通过采集低温环境下东北林蛙在体和离体心脏的心电图(ECG),分析了低温环境对东北林蛙心率的影响,探究了低温环境下或冬眠过程中两栖类动物心率的变化规律。结果表明:15℃时东北林蛙在体心率和离体心率分别约39次/min和38次/min,而由10℃下降至0℃时在体心率迅速下降为15～10次/min、由10℃下降至2℃时离体心率迅速下降为13～5次/min,-2℃时东北林蛙在体心率可短时间维持在9次/min以下,-3℃时东北林蛙在体心率为0;东北林蛙在体心率和离体心率均随温度下降而下降、或温度回升而升高;相同低温环境下东北林蛙在体心率显著高于离体心率。说明进入冬眠时东北林蛙心率迅速下降并维持在较低水平;在可耐受的极限低温下(-2℃)东北林蛙仍维持较低心率;低温环境下东北林蛙心率与温度呈正相关,其机体可通过神经或体液正向调节心脏活动,进而满足低温环境下机体新陈代谢对血流量的需求。     

几株桑天牛成虫肠道优势细菌的分离与鉴定

为了研究桑天牛成虫肠道微生态环境,探索破坏桑天牛营养利用等功能的生物防治新途径,对健康桑天牛成虫肠道的优势细菌进行分离鉴定。选择平皿上出现3个以上的菌落作为肠道优势细菌进行分离纯化,共获得6个菌株。6个菌株均呈杆状,革兰染色和氧化酶反应均呈阴性,在25～35℃、pH 5.5～9.5的环境中生长良好,但各菌株的菌落形态及多种生化性状有差异。通过菌体形态观察、染色反应、生理生化特征分析及16S rDNA碱基序列测定与同源性分析,鉴定1、2、3号菌株分别为产酸克雷伯氏菌(Klebsiella oxytoca)、大肠埃希氏菌(Escherichia coli)、荧光假单胞菌(Pseudomonas fluorescens),4、5、6号菌株分别属于不动杆菌属(Acinetobacter)、沙雷氏菌属(Serratia)、肠杆菌属(Enterobacter)。     

7种鸭源细菌的分离与16S rDNA测序鉴定

从病死鸭的脏器中无菌分离细菌.根据培养特性、菌落形态、革兰染色和生化特性,鉴定出7种细菌分离株.以分离株的基因组DNA为模板,用16S rDNA试剂盒扩增其DNA片段,测序后NCBI网站进行BLAST搜索比对,鉴定出细菌种类,分别为鼠伤寒沙门菌、大肠杆菌、鸭疫里默氏杆菌、铜绿假单胞杆菌、施氏假单胞菌、浅绿气球菌和麦氏棒杆菌.前3种为鸭的常见分离菌,其余的较为少见.因此在鸭病控制过程中应注意可能存在一些不常见细菌的混合感染.     

东北林蛙消化系统SOD和POD同工酶电泳分析

为了对东北林蛙的人工养殖和深入研究提供基础的生理生化指标依据,试验采用聚丙烯酰胺凝胶垂直板电泳方法,对东北林蛙春季消化系统中食道、胃、肠道、肝脏4种器官组织的超氧化物歧化酶(SOD)和过氧化物酶(POD)同工酶进行分离、分析。结果表明:东北林蛙消化系统中共分离出SOD1~SOD_(16)、电泳迁移率为0.649~0.034的16条谱带;共分离出POD_1~POD_(13)、电泳迁移率为0.814~0.093的13条谱带。4种器官组织中SOD和POD活性顺序相同,均为肝脏﹥食道﹥胃﹥肠道。     

黑龙江省两种林蛙耐寒能力比较研究——黑龙江林蛙较东北林蛙耐寒的新证据

为了比较分布在黑龙江省的东北林蛙(Rana dybowskii)和黑龙江林蛙(Rana amurensis)的耐寒能力,试验测定了低温及低温持续时间对两种林蛙存活率的影响,低温下两种林蛙的过冷却点及血糖水平,比较并分析了两种林蛙的耐寒能力。结果表明:在潮湿环境中,东北林蛙和黑龙江林蛙能较长时间(≥10 d)耐受的最低温度分别为-1℃和-2℃;在低温胁迫下,黑龙江林蛙与东北林蛙均可通过升高机体血糖水平维持较低的过冷却点免受冰冻,但黑龙江林蛙可通过保持更高水平的血糖维持更低的过冷却点。说明黑龙江林蛙比东北林蛙具有更强的耐寒能力,因而其地理分布的最北纬度界限也明显较高。     

林蛙养殖场地土壤菌群结构及潜在病原菌分析

试验旨在了解林蛙养殖场地土壤菌群组成和结构特征、喷洒益生菌后的土壤菌群变化和潜在致病菌的种类与数量。试验设置3个组：养殖场地组、益生菌喷洒组和半人工养殖组,用高通量测序技术进行检测。3个试验组共获得191 588条有效序列,1 369个操作分类单元（operational taxonomic units,OTU）,经分类学鉴定分属于438个属,29个门。3组样本菌群数量较大,多样性水平也相对较高。优势菌门为变形菌门（Proteobacteria）、拟杆菌门（Bacteroidetes）、蓝细菌门（Cyanobacteria）、芽单胞菌门（Gemmatimonadetes）、绿弯菌门（Chloroflexi）、放线菌门（Actinobacteria）、酸杆菌门（Acidobacteria）、厚壁菌门（Firmicutes）;优势菌属为黄杆菌属（Flavobacterium）、热单胞菌属（Thermomonas）、Sphingopyxis、紫杆菌属（Porphyrobacter）、蓝细菌纲中未定种类（norank_c__Cyanobacteria）。各分组之间菌群多样性差异不显著,各组菌群结构相似度高。土壤中潜在的红腿病致病菌在属的水平有气单胞菌属（Aeromonas）、柠檬酸杆菌属（Citrobacter）、金黄杆菌属（Chryseobacterium）、假单胞菌属（Pseudomonas）。喷洒益生菌后土壤菌群变化不明显。本研究为全面了解东北林蛙养殖场地土壤微生物组成及变化、发掘有益微生物种类和东北林蛙疾病的预防和治疗提供理论依据。     

东北林蛙烂趾病病原菌的分离鉴定及药敏试验

对东北林蛙烂趾病病原菌进行分离鉴定,并对其药物敏感性进行研究。结果表明：从东北林蛙患病趾处分离到1株溶血性致病茵,经16SrDNA序列分析法鉴定为嗜水气单胞菌。通过对23种抗生素的药敏试验表明,此菌对氟喹诺酮类（如环丙沙星、培氟沙星、诺氟沙星）,部分头孢类抗生素（如头孢呋辛、头孢噻肟、头孢吡肟）,β-内酰胺类抗生素（如亚胺培南、氨曲南）高度敏感;对部分头孢类抗生素（如头孢噻吩、头孢西丁、头孢他啶）,氨基糖苷类抗生素（如卡那霉素、庆大霉素、链霉素）中度敏感;对青霉素类如（青霉素、苯唑西林、氨苄西林、阿莫西林）有耐药性。说明可以考虑用氟喹诺酮类、头孢类或β-内酰胺类抗生素防治东北林蛙烂趾病。     

Comparison of three preoperative skin preparation techniques in ponies

The purpose of this study was to evaluate the efficacy of 3 preoperative skin preparations (povidone‐iodine [PI] removed with 70% isopropanol, and 4% chlorhexidine gluconate [CG] removed with either 70% isopropanol [CA] or sterile saline solution [CS]) in ponies. Eighteen ponies were randomly assigned to one of the 3 preoperative skin preparation groups. The skin of ponies was aseptically prepared with PI removed with alcohol, or 4% CG removed with either alcohol or sterile saline solution. The antimicrobial efficacy of each skin preparation technique was assessed quantitatively by culturing for surface bacteria with replicating organism detection and counting plates. The percentage of negative cultures, the percentage of cultures with >5 colony forming units (CFU) and the percentage bacterial reduction after the cleansing scrub, the sterile scrub, and the surgical procedures were compared. There was a significant difference between CG and PI in percentage bacterial reduction after the cleansing scrub, but no significant difference at any other time (sterile scrub, post operative skin sampling). In a comparison of the number of negative microbial cultures at each sampling point, there were no significant differences among the 3 skin preparation techniques. There were no significant differences among the treatment groups comparing the number of cultures with a high colony count (>5 CFU) after the cleansing scrub. Skin preparation with CS and PI resulted in significantly fewer cultures with >5 CFU after the sterile scrub than CA. Post operatively, CA had a higher number of samples with >5 CFU than CS and PI. PI removed with alcohol and 4% CS are equally effective in the reduction of skin bacteria after a sterile skin scrub in the operating room; however, CG was more effective in reducing bacterial numbers after the cleansing scrub. The number of cultures with high bacterial counts was greater post operatively, when alcohol was used as a rinse with chlorhexidine. In cases where a sterile scrub is not performed following a cleansing scrub, CG may be a better choice than PI. CA should not be used as a presurgical skin preparation method in ponies.     

人工养殖东北林蛙生长曲线的拟合与分析

为了解东北林蛙的生长规律,运用Gompertz、Logistic和Von Bertalanffy3种非线性模型分别对东北林蛙一龄和二龄生长期间体重数据进行了曲线拟合和分析。结果表明:3种模型都能很好地模拟东北林蛙的生长曲线,但Logistic曲线模型的拟合效果最好。结果显示,运用3种模型对东北林蛙进行生长曲线的拟合和分析是可行的。     

低温胁迫对东北林蛙雄性成体GP、PEPCK比活性的影响

采用葡萄糖氧化酶-过氧化物酶法、蒽酮比色定糖法分别检测低温胁迫下东北林蛙血糖和肝糖原、肌糖原的含量,并通过6-磷酸葡萄糖脱氢酶偶联比色法、乳酸脱氢酶偶联比色法分别检测低温胁迫下东北林蛙肝脏、肌肉糖原磷酸化酶(Glycogen phosphorylase,GP)和肝脏、肾脏磷酸烯醇式丙酮酸羧激酶(PEPCK)的比活性,以探讨低温对东北林蛙GP、PEPCK比活性的影响。结果表明:随着环境温度降低,东北林蛙血糖水平逐步升高,糖原含量逐步降低,与对照组相比,各低温组血糖升高极显著(P<0.01),而肝糖原和肌糖原含量降低极显著(P<0.01);与对照组相比,0℃组、-1℃组肝脏、肌肉组织GP比活性升高极显著(P<0.01),且肝脏、肾脏PEPCK比活性升高极显著(P<0.01)。说明低温胁迫促使东北林蛙糖原含量降低,糖原分解和糖异生限速酶比活性升高,进而血糖含量升高,提示血糖作为抗冻保护剂在东北林蛙冬眠中发挥重要作用。     

Bacterial flora on Cascades frogs in the Klamath mountains of California

Amphibians are experiencing global declines due in part to the infectious disease chytridiomycosis. Some symbiotic bacteria residents on frog skin have been shown to inhibit the growth of Batrachochytrium dendrobatitis (Bd) but few studies have attempted to fully describe the resident bacterial flora of frog skin. We cultured and sequenced 130 bacterial isolates from frogs collected from the California Klamath Range, recovering predominantly Gram-negative bacteria from 20 higher order taxa and 31 genera. There were also a large number of unclassifiable isolates. Forty-three isolates were assessed for their ability to inhibit the growth of Bd in vitro; of these, two had strong and three had slight anti-Bd activity. We suggest that many bacterial species may play a secondary role in Bd resistance, acting synergistically with inhibitory species. Future research is required in order to characterize these interactions. Understanding the relationships between bacterial strains may be important in predicting and managing the effects of future anti-Bd treatments such as antimicrobial compounds or probiotic bacteria.     

Microbiology of equine wounds and evidence of bacterial biofilms

Horse wounds have a high risk of becoming infected due to their environment. Infected wounds harbour diverse populations of microorganisms, however in some cases these microorganisms can be difficult to identify and fail to respond to antibiotic treatment, resulting in chronic non-healing wounds. In human wounds this has been attributed to the ability of bacteria to survive in a biofilm phenotypic state. Biofilms are known to delay wound healing, principally due to their recalcitrance towards antimicrobial therapies and components of the innate immune response. This study describes the presence of bacterial biofilms within equine wounds. Thirteen 8-mm diameter tissue samples were collected from (n=18) chronic wounds. Following histological staining, samples were observed for evidence of biofilms. Fifty one wounds and control skin sites were sampled using sterile swabs. Control skin sites were on the uninjured side of the horse at the same anatomical location as the wound. The isolated bacteria were cultured aerobically and anaerobically. The biofilm forming potential of all the isolated bacteria was determined using a standard crystal violet microtitre plate assay. Stained tissue samples provided evidence of biofilms within 61.5% (8 out of 13) equine wounds. In total 340 bacterial isolates were identified from all the equine wound and skin samples. Pseudomonas aeruginosa and Enterococcus faecium were the most predominantly isolated bacterial species from equine wound and skin samples respectively. Staphylococcus was the most commonly isolated genus in both environments. Bacteria cultured from chronic and acute wounds showed significantly (P<0.05) higher biofilm forming potential than bacteria isolated from skin. This paper highlights preliminary evidence supporting the presence of biofilms and a high microbial diversity in equine chronic wounds. The presence of biofilms in equine wounds partly explains the reluctance of many lower limb wounds to heal. Non-healing limb wounds in horses are a well documented welfare and economic concern. This knowledge can be used to shape future treatments in order to increase the healing rate and decrease the costs and suffering associate with equine wounds.     

大肠杆菌胁迫下东北林蛙皮肤抗菌肽分泌特性研究

以东北林蛙(Rana dybowskii)为试验对象,采用大肠杆菌(Escherichia coli)作为环境胁迫因子,通过RP-HPLC分析、抑菌活性和蛋白浓度比较,观察其皮肤分泌物的特异性分泌,以期揭示两栖类动物抵御环境微生物时皮肤分泌抗菌肽的变化规律。研究发现,用E.coli胁迫1~18d后皮肤分泌物粗提物洗脱峰数量明显减少了61.54%~84.62%,对4个持续出现的8、9、11和14号洗脱峰的抑菌活性试验显示,8、9号峰对G-和G+菌均表现较强的活性抑制,而11、14号峰仅对G+菌有较强的抑菌活性。此外,E.coli胁迫后第7天组皮肤分泌物的洗脱峰数量、持续出现峰的峰面积值及蛋白浓度与对照组比均明显增加,第23天组均恢复至较高水平。研究结果表明:两栖类皮肤抗菌肽可能在参与机体先天免疫系统防御和启动后天免疫应答的关键环节中发挥重要的桥梁作用。     

东北林蛙捕食行为研究

2010年7～9月,利用焦点动物取样法并结合全事件取样法对东北林蛙的捕食行为研究表明：对运动敏感的视觉特性对东北林蛙的捕食行为有重要影响,其所能捕获的对象均为移动物体;东北林蛙整个捕食行为存在多种模式和环节,分别由相应条件触发;猎物的形状、运动方向和活动性的强弱均影响东北林蛙的捕食行为;相对于移动的目标,振动和闪动的目标更容易引起东北林蛙的捕食;东北林蛙对猎物运动的敏感度较高,但处理食物能力较弱。     

东北林蛙皮肤及其腺体组织形态学观察

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