副猪嗜血杆菌的分离鉴定与药敏试验

对一例临床症状、病理剖检变化疑似副猪嗜血杆菌病的仔猪病料进行实验室诊断,进行了细菌培养特性和生化特性等鉴定,初步怀疑为副猪嗜血杆菌,根据副猪嗜血杆菌的16 S rRNA基因设计特异性引物进行PCR扩增鉴定,结果表明,分离菌为副猪嗜血杆菌.药敏试验显示,该分离菌对阿米卡星、头孢氨苄、新霉素高度敏感.     

副猪嗜血杆菌病的病原分离鉴定与病理学诊断

为了研究副猪嗜血杆菌病的病原学和病理学诊断方法,给该病的临床诊断和防制提供依据,试验采用病理剖检、细菌分离培养、生化鉴定和病理组织学观察等方法对疑似副猪嗜血杆菌感染的病猪进行诊断。结果表明:病猪主要表现为急性败血症和多发性浆膜炎病理变化,并分离到3株副猪嗜血杆菌,分别命名为Hps1、Hps2、Hps3;心脏、肺脏和脾脏等脏器含有大量嗜中性粒细胞,发生化脓性纤维蛋白性浆膜炎,根据病原学和病理学诊断结果确诊为副猪嗜血杆菌病。     

天津地区副猪嗜血杆菌的分离鉴定及敏感药物筛选

为了给天津地区各区县养猪场提供副猪嗜血杆菌病的科学防控及合理的治疗用药方案,本试验对天津地区各区县养猪场疑似感染副猪嗜血杆菌的病猪进行了解剖及病理变化观察,无菌采集36份各猪场病猪组织脏器并进行细菌分离,对分离菌进行染色镜检、卫星试验、接触酶反应试验、微量生化反应和PCR鉴定。结果显示,剖检结果以纤维素性胸膜炎及腹膜炎常见,关节腔切面内有大量黄亮黏稠液体;分离出了9株疑似副猪嗜血杆菌菌株,染色镜检均可见革兰氏阴性多形态菌;分离菌有明显的卫星现象;接触酶反应试验均为阳性;微量生化反应D-核糖、山梨醇、果糖、阿拉伯糖和硫化氢均为阴性,麦芽糖、葡萄糖、蔗糖、甘露醇和脲酶均为阳性,山梨糖和木糖各有1株为阳性,5株菌β-半乳糖为阳性;PCR鉴定结果显示9株疑似副猪嗜血杆菌菌株在822 bp处均与阳性对照处于同一条带上,均判定为副猪嗜血杆菌,分离率达25%。用16种抗菌药物对9株副猪嗜血杆菌进行敏感药物筛选,药敏结果显示头孢曲松钠和氧氟沙星对分离株有良好的抑菌效果。本试验结果可为天津地区副猪嗜血杆菌病的科学防制提供依据。     

规模猪场副猪嗜血杆菌的分离鉴定及药物试验

从数例疑似副猪嗜血杆菌病猪的病料中分离袋3株细菌,通过细菌形态特征、培养特性和生长特性的观察,确诊该病的病原菌是副猪嗜血杆菌.同时,对分离菌进行药敏试验,为今后防控该病提供试验依据.     

副猪嗜血杆菌的分离与鉴定

从福建省龙岩市l例临床症状、病理剖检变化疑似副猪嗜血杆菌病的患病猪的关节液中分离到1株革兰氏染色阴性可疑菌株,经卫星现象、生化鉴定等实验室诊断,进一步以副猪嗜血杆菌的16S rRNA基因设计特异性引物进行PCR鉴定,确定该分离菌为副猪嗜血杆菌。药敏试验结果表明,分离株对环丙沙星、卡那霉素、强力霉素、氨苄西林、阿米卡星、庆大霉素药物敏感;对泰妙菌素、诺氟沙星有抵抗力。     

副猪嗜血杆菌病的诊断

从江西省抚州市某规模化养猪场病猪肺脏分离到1株革兰阴性长丝状菌,经细菌生化鉴定、PCR检测鉴定为副猪嗜血杆菌。使用KRG方法对其进行血清分型鉴定,该株副猪嗜血杆菌的血清型为13型。抗生素药物敏感试验结果表明,分离菌株对氨苄西林、头孢类、苯唑西林、氯霉素、阿莫西林表现较高的敏感性。对强力霉素、青霉素G中度敏感,而对磺胺类、链霉素、庆大霉素表现出耐药性;致病性试验结果表明,分离菌株对小白鼠有强致病性,命名为JXFZHP01株。本结果为江西副猪嗜血杆菌病的防治提供了理论依据。     

副猪嗜血杆菌的分离鉴定及药敏试验

黑龙江省近几年频繁发生以多发性浆膜炎、关节炎、脑膜炎以及急性死亡为特征的传染病,为猪场造成严重经济损失,为了确诊是否有副猪嗜血杆菌感染,采集病死猪肝脏、脾脏和肺脏等病料,进行副猪嗜血杆菌的分离;对其理化特性进行鉴定,应用PCR方法对其16S rRNA基因扩增后进行克隆测序,将测序结果在GenBank上进行BLAST分析,把相近的基因序列应用DNAStar软件进行同源性和进化关系分析;用分离菌株进行药物敏感性试验,筛选敏感药物。结果分离出一种具有多形性的NAD依赖性菌株,经鉴定为副猪嗜血杆菌;16S rRNA基因进化分析结果表明,分离菌株与以往报道的副猪嗜血杆菌中国、日本和美国分离株属于同一亚群,而西班牙分离株属于单独的亚群;分离菌株对壮观霉素和头孢拉定高度敏感。     

副猪嗜血杆菌的分离鉴定及中药药敏试验

从数例疑似副猪嗜血杆菌病猪的肺脏和心包积液中分离到3株革兰氏阴性细小杆菌,经涂片镜检、生化试验和PCR鉴定,确定分离的3株菌为副猪嗜血杆菌.中药药敏试验结果表明,副猪嗜血杆菌对金银花及其组成的复方敏感,且复方作用效果优于单方.     

金昌市副猪嗜血杆菌的分离与鉴定

为确诊金昌市金川区某规模养猪场引起猪大量发病死亡的病因,以及时的找到有效的治疗方法,本研究针对该养殖场病死猪进行无菌采集组织病料,送往甘肃农业大学微生物实验室进行细菌学诊断的方法,对采集的病死猪肺脏、关节滑液、胸腔积液和心包积液进行了细菌分离纯化、染色镜检、生化鉴定和药物敏感试验等实验室检测,实验室检测结果显示组织病料抹片和细菌分离后染色镜检发现大量丝状革兰氏阴性杆菌,其生化试验结果与副猪嗜血杆菌生化特性一致,由此可以证实此次引起猪大规模发病死亡的病原微生物为副猪嗜血杆菌,经实验室药物敏感试验结果,可使用先锋霉素Ⅳ、阿米卡星等药物进行临床治疗。     

仔猪副猪嗜血杆菌病的诊治

仔猪副猪嗜血杆菌病是由猪副嗜血杆菌引起的仔猪并能继发其他病原感染的疾病,可引起仔猪发热、呼吸困难、跛行、关节肿胀、皮肤及黏膜发绀、站立困难甚至瘫痪、僵猪或死亡,又称多发性纤维素性浆膜炎和关节炎,也称格拉泽氏病。副猪嗜血杆菌只感染猪,通过空气、飞沫、污染物传播。病猪和带菌猪是该病的传染源,临床多见断奶前后的保育猪和仔猪感染,9周龄后病猪的临床表现并不明显。本文通过患病仔猪的临床症状、剖检变化、实验室诊断等多种方法进行了综合的判断,患病仔猪被确诊为副猪嗜血杆菌病,通过药敏试验分析其对抗生素敏感性,选择敏感的药物进行治疗,取得良好的治疗效果。     

陕南白山羊伪结核棒状杆菌分离鉴定及灭活铝胶疫苗研制

羊干酪性淋巴结炎又称羊伪结核病,是由伪结核棒状杆菌感染引起的一种人畜共患慢性接触性传染病,对山羊危害最为严重,目前尚无有效治疗措施。陕西省石泉县陕南白山羊饲养量较大,羊群也存在伪结核棒状杆菌的感染,造成较大的经济损失。为筛选对当地伪结核棒状杆菌敏感的药物和研制灭活疫苗,本研究从疑似伪结核病患羊采取多份新鲜的脓液,进行细菌分离鉴定;对获得的1株分离菌进行药物敏感性试验并用其研制灭活疫苗。结果显示,分离菌在固体培养基上形成白色、干燥、扁平、不透明、边缘整齐的中等大小菌落,革兰染色阳性,经16SrRNA检测和序列分析,证实分离到的菌株为羊伪结核棒状杆菌。药物敏感性试验结果显示,该分离菌对环丙沙星、头孢曲松、红霉素、卡那霉素、头孢噻肟、妥布霉素、强力霉素、庆大霉素、氯霉素高度敏感;对四环素、利福平、链霉素中度敏感。以该分离菌为种子菌制备的灭活铝胶疫苗无菌检验合格,试验接种山羊无不良反应,目前已经在采样羊场进行临床应用,进一步评价免疫效果。     

一起仔猪水肿病的诊断与治疗

通过对湖南省新化县某养猪场的40 kg以下的发病仔猪进行临床症状观察、病理剖检和实验室诊断,确诊为仔猪水肿病.通过采用肌肉注射强力水肿灵进行治疗,并对易感猪群肌注仔猪水肿病三价灭活疫苗进行紧接免疫接种,有效控制了病情.     

A:L1型鸭源多杀性巴氏杆菌灭活疫苗制备及免疫效力研究

【目的】制备鸭多杀性巴氏杆菌灭活疫苗,有效控制贵州省鸭多杀性巴氏杆菌发生和流行。【方法】以实验室分离保存的A：L1型多杀性巴氏杆菌地方流行株为菌种,通过涂板法测定该菌株生长曲线,并采用改良寇氏法计算该菌株对鸭的半数致死量（median lethal dose,LD₅₀）,将该菌培养至终浓度为1×10¹⁰ CFU/mL后分别制备白油佐剂灭活疫苗和卡波姆佐剂灭活疫苗,经疫苗质量检验后进行免疫雏鸭试验;通过攻毒保护试验评价比较两种疫苗的保护率,并对攻毒试验鸭肝脏、肺脏、脾脏进行病理组织学观察。【结果】A：L1型多杀性巴氏杆菌疫苗株培养至18 h到达峰值,活菌数可达1.0×10¹⁰ CFU/mL;LD₅₀为5 CFU;制备的两种疫苗安全性良好;攻毒保护试验结果显示,一免后卡波姆佐剂灭活疫苗免疫保护率可达62.5%,高于白油佐剂灭活疫苗（50.0%）及商品灭活疫苗（50.0%）。二免后卡波姆佐剂灭活疫苗优势更为明显,免疫保护率可达87.5%,高于白油佐剂灭活疫苗（75.0%）及商品灭活疫苗（62.5%）。病理组织学结果显示,一免后卡波姆佐剂灭活疫苗能对肺脏提供较好的保护效果,二免后在肝脏、肺脏和脾脏的保护效果上,卡波姆佐剂灭活疫苗组优势明显。【结论】利用贵州地区流行的A：L1型菌株所制备的卡波姆佐剂灭活疫苗免疫效果明显,能对贵州地区多杀性巴氏杆菌病的防控起到重要作用。     

Vaccines against Aujeszky's disease: Evaluation of their efficacy under standardized laboratory conditions

Summary

A standardized test was developed to compare the efficacy of Aujeszky's disease virus (ADV) vaccines under laboratory conditions. Per test 3 groups of 6 to 8 sero‐negative pigs were used. The first vaccination was done at 10 weeks of age. One group was vaccinated once, another was vaccinated twice and the 3rd served as control. Pigs were challenge exposed to the virulent NIA‐3 strain of ADV 12 weeks after the first vaccination. Apart from mortality, average periods of growth arrest, fever and virus shedding after challenge were used as parameters to evaluate vaccine efficacy.

Two inactivated and 4 attenuated vaccines were tested. Two attenuated vaccine viruses were excreted after vaccination. Despite maximal standardization, a considerable variation still existed between the experiments in mortality and growth arrest periods of control pigs after challenge. However, the controls were always more severely affected than the vaccinated pigs. All vaccines except one were effective in preventing death after challenge, but none conferred complete protection. Most vaccinated pigs still lost weight, developed fever and shed virus after challenge. Revaccination after 3 or 4 weeks had little effect, particularly with the attenuated vaccines. The results of the present study indicate that 2 of the attenuated vaccines conferred the best protection, I attenuated vaccine appeared to be as effective as the 2 inactivated ones, and the 4th attenuated vaccine was least effective.     

Vaccines against Aujeszky's disease: evaluation of their efficacy under standardized laboratory conditions

A standardized test was developed to compare the efficacy of Aujeszky's disease virus (ADV) vaccines under laboratory conditions. Per test 3 groups of 6 to 8 sero-negative pigs were used. The first vaccination was done at 10 weeks of age. One group was vaccinated once, another was vaccinated twice and the 3rd served as control. Pigs were challenge exposed to the virulent NIA-3 strain of ADV 12 weeks after the first vaccination. Apart from mortality, average periods of growth arrest, fever and virus shedding after challenge were used as parameters to evaluate vaccine efficacy. Two inactivated and 4 attenuated vaccines were tested. Two attenuated vaccine viruses were excreted after vaccination. Despite maximal standardization, a considerable variation still existed between the experiments in mortality and growth arrest periods of control pigs after challenge. However, the controls were always more severely affected than the vaccinated pigs. All vaccines except one were effective in preventing death after challenge, but none conferred complete protection. Most vaccinated pigs still lost weight, developed fever and shed virus after challenge. Revaccination after 3 or 4 weeks had little effect, particularly with the attenuated vaccines. The results of the present study indicate that 2 of the attenuated vaccines conferred the best protection, 1 attenuated vaccine appeared to be as effective as the 2 inactivated ones, and the 4th attenuated vaccine was least effective.     

4型禽腺病毒HLJ1701株灭活疫苗的研制

利用4型禽腺病毒HLJ1701株进行灭活疫苗的研制,并对疫苗的免疫效果进行评价,为家禽4型禽腺病毒的防控提供数据及参考。将HLJ1701株用灭菌生理盐水作10~4倍稀释后,接种9日龄SPF鸡胚,37℃孵育72 h后收获感染鸡胚尿囊液,经甲醛灭活后,加白油佐剂乳化制成油乳剂灭活疫苗,对制备疫苗的性状、安全性、免疫效力等进行检验。结果显示,制备的3批4型禽腺病毒灭活疫苗(HLJ1701株)均为油包水型,黏度均在50 cP以内,对3批疫苗取样,样品经3000 r/min离心15 min,管底无水相析出。安全性试验结果显示,将疫苗按1 mL/只超剂量接种3周龄SPF鸡,试验鸡在观察期内全部健活,未出现局部或全身不良反应,表明疫苗对SPF鸡具有良好的安全性;免疫效力及攻毒保护试验结果显示,用疫苗按0.2 mL/只的剂量免疫接种3周龄SPF鸡1次,免疫接种后21d试验鸡血清中HLJ1701株的抗体平均效价可达2~8以上,使用4型禽腺病毒(HLJ1701株)接种0.2 mL/只(100 LD_(50))对免疫鸡进行攻毒,疫苗对免疫鸡的保护率均为100%。研究表明,实验室条件下研制的4型禽腺病毒(HLJ1701株)灭活疫苗的各项指标均符合标准。     

水貂肠炎细小病毒灭活疫苗抗体消长规律的研究

应用血凝抑制试验对水貂肠炎细小病毒灭活疫苗免疫水貂进行抗体水平动态监测,结果表明,疫苗接种14 d抗体达到保护,21~30 d抗体水平达到高峰;免疫180 d抗体仍在保护值以上。攻毒试验证实,免疫180 d后90%免疫水貂获得保护,确定水貂细小病毒性肠炎灭活疫苗免疫保护期可持续6个月。     

Study on Synergistic Effect of VA5 Immunopotentiator on Pigeon NDV Inactivated Vaccine

This study was designed to evaluate the synergistic effect of VA5 immunopotentiator on pigeon ND4416 inactivated vaccine.160 healthy pigeons were randomly divided into four groups,including ND4416 strain inactivated vaccine group (NDV group),ND4416 strain inactivated vaccine and immunopotentiator (VA5) mixed group (NDV+VA5 group),La Sota inactivated vaccine group (La Sota group) and normal saline as the control group (C group),to assess their vaccine efficacy against virulent pigeon NDV by serological analysis and animal testing.Pigeons sera were collected at different time points after immunization and measured the HI antibody titer of each group.The results showed that VA5 immunopotentiator significantly improved the serum antibody level of pigeons immunized with pigeon Newcastle disease inactivated vaccine (P<0.05).In addition,comparative test of spleen lymphocyte transformation was conducted at various time points after immunization.The results indicated that VA5 effectively stimulated the lymphocyte transformation of immune pigeon.Pigeons in each groups were challenged with ND4416 strain at the 30th,90th and 180th d after immunization.The results presented that the NDV+VA5 group had 100% protection rate and higher than La Sota group.The duration of immunization test showed that the antibody titer of NDV+VA5 group reached peak 11.20log₂ at the 21st d,remained 7.50log₂ at 180th d,and the protection rate remained 100% at 180th d.It indicated that VA5 immunopotentiator sustained the immune duration of pigeon NDV vaccine up to 180 d.Moreover,the in vitro detoxification test results suggested that VA5 immunopotentiator reduced the in vitro detoxification cycle of pigeons after challenge.Overall,this study suggested VA5 immunopotentiator could significantly improve the immune efficacy of pigeon Newcastle disease inactivated vaccine,which provided a basis for further enhancing the efficacy of Newcastle disease vaccine,as well increased experimental data for the application of immunopotentiators.     

VA5免疫增强剂对鸽新城疫病毒灭活疫苗的增效作用研究

试验旨在探究VA5免疫增强剂对鸽新城疫病毒（NDV）灭活疫苗（ND4416株）的免疫增效作用。选取160只健康鸽随机分为4组：ND4416株灭活疫苗组（NDV组）、ND4416株灭活疫苗与免疫增强剂（VA5）混合组（NDV+VA5组）、La Sota灭活疫苗组（La Sota组）及生理盐水空白对照组（C组）,进行免疫效力及免疫持续期试验。采用血凝抑制试验检测各组鸽免疫后不同时间点的血清抗体效价,结果显示,VA5免疫增强剂能够显著提高鸽NDV灭活疫苗血清抗体水平（P<0.05）。脾脏淋巴细胞转化试验结果显示,VA5能够有效刺激免疫鸽的淋巴细胞转化。在免疫后第30、90、180天,各组鸽子采用ND4416株进行攻毒试验,结果显示,NDV+VA5组在3个时间点的攻毒保护率均为100%,均高于La Sota组。免疫持续期试验结果显示,NDV+VA5组血清抗体水平在免疫后第21天达峰值11.20log₂,第180天仍能达到7.50log₂,攻毒保护率达100%,表明VA5免疫增强剂可延长鸽新城疫疫苗的免疫效力持续期至180 d。攻毒后体外排毒检测结果表明,VA5免疫增强剂能够缩短攻毒后鸽的体外排毒周期,减少体外排毒。综上,VA5免疫增强剂能够显著提高鸽NDV灭活疫苗的免疫效果,为研制效果更好的鸽NDV疫苗提供了基础依据,同时也为免疫增强剂的应用增加了试验数据。     

牛用口蹄疫AsiaI—O型双价灭活苗与猪用口蹄疫O型灭活苗对猪的免疫效果比较试验

使用牛用口蹄疫AsiaI-O型双价灭活苗与猪用口蹄疫O型灭活苗分别接种50d商品猪和怀孕90d种猪,免疫前和免疫后的3w及7w进行抗体水平检测。O型口蹄疫采用正向间接血凝试验、AsiaI型口蹄疫采用液相阻断ELISA检测。同时对接种猪进行免疫应激观察。结果显示：猪使用牛用口蹄疫AsiaI—O型双价灭活苗3w后口蹄疫AsiaI的抗体水平十分低,最高只有5％,口蹄疫O型抗体合格率在35％以上;而注射猪用口蹄疫O型灭活苗的O型抗体水平合格率只有35％。而牛用口蹄疫AsiaI-O型双价灭活苗两次接种后,AsiaI和O型抗体水平均达到大于70％的要求。