猪肺炎支原体高密度发酵工艺的研究

为提高猪肺炎支原体发酵培养的滴度,在500L发酵罐基础上,对猪肺炎支原体J株的发酵工艺进行了一系列优化试验,各项发酵培养条件经优化后,发酵支原体的滴度得到了明显提高,发酵滴度可达到1×109CCU/ml以上,本研究为提供高效、安全和稳定的猪肺炎支原体疫苗产品奠定基础.     

猪肺炎支原体DJ-166株高密度发酵工艺优化

猪肺炎支原体培养存在对营养需求严苛、培养难度大、菌液培养滴度不高等问题.本试验开展了一系列改进猪肺炎支原体高密度发酵培养工艺的研究,采用添加15％猪血清的优化CH液体培养基(初始pH为7.8),按5％接种量接种,在发酵罐中培养、pH降到6.7～6.8时收获菌液,菌液培养滴度可达1×1010 CCU/mL.本试验结果对猪...     

猪肺炎支原体HN0613株高密度发酵工艺研究及效力评价

旨在建立猪肺炎支原体（Mycoplasma hyopneumoniae,Mhp）HN0613株高密度发酵工艺技术。以培养滴度（CCU）作为考查指标,确定Mhp HN0613株的最佳活力代次以及复苏培养时间;通过四因素三水平正交试验筛选最佳基础培养基配方;以通气量、搅拌转速和接种百分比作为单因素考查指标,优化15L发酵工艺参变量,并确定700L高密度发酵放大工艺;按照确定的高密度发酵工艺技术,制备MhpHN0613株灭活疫苗,评价其对兔和仔猪的免疫效力。结果表明,Mhp HN0613株F11代较其他代次CCU水平高,可达1×10^9CCU／mL,为最佳活力代次;其在复苏72h后,pH值降为7．0左右,CCU水平达到最高,为1×10^9CCU／mL,为最佳复苏培养时间;培养体系中,2％的初始葡萄糖添加量、8％接种百分比、初始培养基pH值7．6、15％猪血清添加量为最适培养条件;通气量100L／h、搅拌转速300r／min和8％接种百分比条件最有利于Mhp HN0613株15L发酵培养;首次建立了平衡kLa联动pH值回调的工艺技术路线,并将优化后发酵罐搅拌叶轮结构改为推进式叶轮,降低了叶轮对菌体的剪切作用力,CCU较优化前提高了10倍,且培养周期较优化前缩短了2-3d;700L高密度发酵放大工艺中,Mhp HN0613株培养滴度不低于5×10^9CCU／mL．最高可达1×10^10CCU／mL;免疫兔血清抗体效价均不低于1：40,最高可达1：160;免疫组实验仔猪肺炎减少率均高于80％,临床观察精神及食欲未见异常。该研究为Mhp疫苗规模化发酵生产提供了一定的理论依据。     

绵羊肺炎支原体与猪肺炎支原体交叉抗原研究

为了研究绵羊肺炎支原体标准株Y98与猪肺炎支原体标准株232全菌蛋白免疫原性的异同,试验采用SDS-PAGE和免疫印迹分析技术对其进行了研究,结果表明:绵羊肺炎支原体标准株Y98同猪肺炎支原体标准株232免疫印迹结果基本一致,在70,50,40,25 ku蛋白带处存在共同抗原。     

猪支原体肺炎诊治

随着我国农业的不断发展,目前生猪养殖已成为农户的第二收入来源。而猪支原体肺炎是养殖场较为常见的传染病,对生猪有较大的危害,当混合感染时疾病的诊治难度增加,给猪场和养殖户造成较大的经济损失。猪支原体肺炎又称猪地方性肺炎、猪喘气病,该病病原为猪肺炎支原体,是一种慢性、接触性呼吸道传染病,该病可引起猪只生长迟缓,料肉比上升、日增重减少等,延长了肥猪出栏时间。猪支原体肺炎可对猪造成严重免疫抑制。该病分布较广,是世界养猪业较为关注的重要疫病。     

猪肺炎支原体的研究进展

猪支原体肺炎是一直困扰养猪业发展的重要疾病之一。本文综合国内外对其病原一猪肺炎支原体的最新研究情况，从其对猪生长性能的影响、防治动向、检测、主要抗原的研究、基因工程疫苗研制等五个方面进行讨论。     

猪支原体肺炎防治研究进展

<正>猪支原体肺炎（Mycoplasmal pneumoniaeofswine,MPS）又称猪气喘病,是支原体科猪肺炎支原体（Mycoplasmal hyopneumoniae,Mhp）引起的一种接触性、慢性、呼吸道传染性疾病,其主要表现为猪只咳嗽、生产性能降低。此外,猪鼻支原体（Mycoplasma hyorhinis,Mhr）、絮状支原体（Mycoplasma fl occulare,Mfl oc）及猪滑液支原体（Mycoplasma hyosynoviae,Mhyo）也是猪群中常见的支原体科病原。     

猪致病性大肠杆菌高密度发酵工艺的研究

为了提高猪致病性大肠杆菌(O139血清型)的生产量,缩短生产周期,降低生产成本,首先在10L发酵罐中对影响大肠杆菌发酵的pH值、相对溶氧量、生物量、氨基氮、还原糖、无机磷测定等方面进行了单因子试验优化。研究表明,大肠杆菌在37.5℃,pH值为7.5,4%的接种量,溶氧量为50%,200r/min的转速以及15g/L的葡萄糖添加量,20g/L的氨基氮添加量,100mmol/L的无机盐添加量,培养18h后可以得到理想的培养效果。利用此优化的条件,结合兽医药品GMP原则,在200L发酵罐中进行了发酵试验,结果在发酵结束后菌液的OD600可以达到34.51,相当于菌体浓度为12.94g/L,从而得到了大肠杆菌工业化生产工艺,为大肠杆菌疫苗大规模生产奠定了基础。     

Diagnosis of Mycoplasma hyopneumoniae

Mycoplasma hyopneumoniae, the cause of enzootic pneumonia, remains an important pathogen in the swine industry. This small, complex organism colonizes the ciliated cells of the respiratory tract, resulting in little exposure to the immune system. Confirming the presence of M. hyopneumoniae, as well as identifying its role in respiratory disease and pneumonia, remains challenging to the veterinary profession. While culture of the organism remains the gold standard for identification, the use of serology, the polymerase chain reaction and various assays to detect the presence of M. hyopneumoniae in tissue is common in diagnostic laboratories. Because of the role M. hyopneumoniae plays in increasing the severity of pneumonia associated with concurrent bacterial and viral infections, understanding the pathogenesis and diagnostic assays available is critical for developing effective intervention strategies to control respiratory disease on a herd basis.     

猪肺炎支原体体外抗菌药物敏感性试验研究

为了解猪肺炎支原体对抗菌药物的敏感性,采用宏量肉汤稀释法对4株猪肺炎支原体的最小抑菌浓度（MIC）值进行了测定。结果表明,猪肺炎支原体对泰妙菌素和环丙沙星最为敏感,MIC≤0．03μg／mL,其次分别为四环素类药物（包括四环素和多西环素）、林可霉素和泰乐菌素,而对氟苯尼考的敏感性则较差。本研究可为猪支原体肺炎的防控以及抗菌药物的合理使用提供参考。     

猪肺炎支原体流行病学及防控研究进展

猪肺炎支原体是引起猪地方性肺炎的主要病原,流行于世界各地,给养猪业造成重大经济损失。本文综述了猪肺炎支原体的主要传播途径以及引起的临床症状和肺部病变,总结了其主要致病机理,分析了综合控制该疫病的方法。猪肺炎支原体的主要传播途径是与感染猪密切接触,也可通过空气远距离传播,但无垂直传播相关报道;猪肺炎支原体感染的主要临床特征是间歇性、强度可变的干咳,但在与其他病原继发或混合感染后,临床症状和肺部病变可能会加重;猪肺炎支原体主要通过释放黏附素引起纤毛发生病变、死亡,导致上皮细胞受损或脱落从而影响纤毛功能,也能通过产生剧毒的H2O2等代谢产物引发毒性效应;防治该病首先要通过生物安全措施控制猪肺炎支原体的传入,其次是采用抗菌药物进行治疗,最后是通过接种疫苗降低其传播率。本文为猪养殖场做好猪肺炎支原体病防控提供了参考。     

RAPD and VNTR analyses demonstrate genotypic heterogeneity of Mycoplasma hyopneumoniae isolates from pigs housed in a region with high pig density

Since differences in the virulence of Mycoplasma (M.) hyopneumoniae strains have been described, the isolation of field strains followed by genotypic and phenotypic characterisation has become a major goal in epidemiological studies. The aim of this study was to compare various M. hyopneumoniae isolates from different pig herds and numerous pigs within the same herd. Therefore, pigs of 109 herds located in North-Western Germany were sampled either on-farm or during necropsies. Overall, 52 isolates of M. hyopneumoniae were recovered from 45 pigs originating from 21 herds. The identity of cultures was confirmed by PCR targeting the 16S-23S intergenic spacer region. Typing of isolates was achieved by random amplified polymorphic DNA (RAPD) analysis and multi-locus analysis of variable number of tandem repeats (VNTR) and demonstrated a high degree of heterogeneity of M. hyopneumoniae isolates. Differences among isolates recovered from animals of the same herd or even from the same pig revealed a grouping into different genotypic clusters. This outcome was observed with both methods. It was concluded that more than one strain of M. hyopneumoniae might be present in a pig herd and even in a single pig, suggesting high genetic heterogeneity between isolates of the same epidemiological source. These factors should be considered when applying nucleic amplification techniques for characterising M. hyopneumoniae strains to specify the epidemiology of infection and to evaluate virulence factors triggering the corresponding disease.     

猪肺炎支原体及其生物学研究进展

猪肺炎支原体是引起猪支气管肺炎的主要病原 ,严重危害着养猪事业的发展。虽然猪肺炎支原体的结构简单 ,但由于它对营养要求较高 ,故培养难度大 ,也是人们不能对其深入研究的一个主要原因。近年来利用克隆技术 ,对猪肺炎支原体从基因水平上进行了多种研究 ,已经取得了一些可喜成果。文章从它的生物学特性、荚膜结构、膜蛋白研究以及基因水平的研究等方面进行了综合性论述 ,并简明地指出了目前研究中存在的问题以及未来展望     

Immunological characterization of Mycoplasma hyopneumoniae recombinant proteins

Mycoplasma hyopneumoniae, the primary pathogen of enzootic pneumonia, is highly prevalent worldwide and causes major economic losses to the pig industry. Commercial vaccines are widely used in the control of this disease, however, they provide only partial protection. The aim of this study was to evaluate 34 recombinant proteins of M. hyopneumoniae expressed in Escherichia coli. Antigenic and immunogenic properties of these proteins were analyzed. For this, the proteins were tested against hyperimmune and convalescent pig sera through ELISA and Western blot. Immunogenicity of the recombinant proteins was evaluated in BALB/c mice following intramuscular inoculation. Most antigens were able to induce a strong immune response and sera from inoculated mice were able to recognize native proteins by cell ELISA and Western blot. Several recombinant proteins were specifically recognized by convalescent pig sera, indicating they are expressed during infection. These data may help to develop more efficacious vaccines against M. hyopneumoniae.     

Control of Mycoplasma hyopneumoniae infections in pigs

Mycoplasma hyopneumoniae, the primary pathogen of enzootic pneumonia, occurs worldwide and causes major economic losses to the pig industry. The organism adheres to and damages the ciliated epithelium of the respiratory tract. Affected pigs show chronic coughing, are more susceptible to other respiratory infections and have a reduced performance. Control of the disease can be accomplished in a number of ways. First, management practices and housing conditions in the herd should be optimized. These include all-in/all-out production, limiting factors that may destabilize herd immunity, maintaining optimal stocking densities, prevention of other respiratory diseases, and optimal housing and climatic conditions. Strategic medication with antimicrobials active against M. hyopneumoniae and, preferably, also against major secondary bacteria may be useful during periods when the pigs are at risk for respiratory disease. Finally, commercial bacterins are widely used to control M. hyopneumoniae infections. The main effects of vaccination include less clinical symptoms, lung lesions and medication use, and improved performance. However, bacterins provide only partial protection and do not prevent colonization of the organism. Different vaccination strategies (timing of vaccination, vaccination of sows, vaccination combined with antimicrobial medication) can be used, depending on the type of herd, the production system and management practices, the infection pattern and the preferences of the pig producer. Research on new vaccines is actively occurring, including aerosol and feed-based vaccines as well as subunit and DNA vaccines. Eradication of the infection at herd level based on age-segregation and medication is possible, but there is a permanent risk for re-infections.     

猪支原体肺炎活疫苗（168株）肺内免疫机制研究

为研究猪支原体肺炎活疫苗(168株)的免疫机制,通过肺内接种免疫5 ～ 10日龄仔猪,并于免疫后不同时间点检测血清中IgG抗体效价、全血中淋巴细胞转化效率、呼吸道局部的IFN-γ浓度和特异性SIgA滴度,于免疫后28 d剖杀采集呼吸道上皮组织,通过扫描电镜法与原位杂交检测法观察疫苗株在呼吸道的存留以及对纤毛的影响情况.结果发现,免疫后猪血液中淋巴细胞转化增强1.52～2.01倍,支气管表面IFN-γ浓度和特异性SIgA滴度持续增加,但血清抗体一直未检测到.扫描电镜与原位杂交检测结果发现疫苗株能有效地黏附在支气管纤毛上皮细胞上,但对纤毛的影响较小.由此表明,猪支原体肺炎活疫苗(168株)通过肺内免疫可有效激活全身细胞免疫及呼吸道局部的黏膜免疫与细胞免疫反应,而且还可以通过黏附支气管纤毛上皮细胞产生占位效应而对上皮组织不产生损伤.     

猪肺炎支原体 MY-99株的致病性

将猪肺炎支原体MY-99株人工培养F100株经鼻腔接种于健康成华×大约克杂交猪,2周后60%的猪出现气喘,个别猪伴有轻微咳嗽;病理切片可见到支气管周围多量淋巴样细胞浸润、肺泡壁增厚等典型的间质性肺炎病变.试验猪平均日增重比对照猪显著降低,接种后0～14 d期间降低了31.05%(P<0.01),15～28 d期间降低了42.01%(P<0.01),29～42 d期间降低了0.1%(P>0.05).试验猪血清中的猪肺炎支原体抗体水平在2周后最高(1∶24.2),4周后降至1∶23.9,6周后最低(1∶22.5),显示猪肺炎支原体MY-99株诱发了急性猪地方流行性肺炎,同时从试验猪肺脏中分离出了猪肺炎支原体.超微切片显示,其菌体大小为0.33～0.48 μm,并缺乏细胞壁;革兰氏染色为阴性类球形菌体,在压力下可以通过0.45 μm的细菌滤膜,菌落呈现煎蛋状.