Protective effects of an oral microencapsulated Mycoplasma hyopneumoniae vaccine against experimental infection in pigs.

Oral microencapsulated Mycoplasma hyopneumoniae vaccines were tested for their ability to prevent mycoplasmal pneumonia in pigs. Eighteen four-week-old specific pathogen free pigs were divided into six groups of three. Each pig of groups 1 to 4 was inoculated intramuscularly with formalin-inactivated M hyopneumoniae in adjuvant and boosted orally 14 days later with four different microencapsulated vaccine microspheres. Group 5 was used as a positive control. All 15 pigs of groups 1 to 5 were challenged at 28 days after the first vaccination by an intratracheal inoculation of pneumonic lung suspension. The three pigs of group 6 were used as a negative control. All four vaccinated groups showed some protection when challenged, but the protection was more solid in pigs boosted with vaccine D (group 4) which contained less porcine serum in the microsphere. The study indicates that oral vaccination with M hyopneumoniae could play a role in prevention and eradication of mycoplasmal pneumonia in the pig.     

Protective efficacy of a live attenuated Mycoplasma hyopneumoniae vaccine with an ISCOM-matrix adjuvant in pigs

An attenuated Mycoplasma hyopneumoniae vaccine that requires intrathoracic administration is commercially available for use against mycoplasmal pneumonia in China. Given the limitations of such a route of administration, this study was undertaken to assess the capacity of an ISCOM-matrix adjuvant to enhance immunogenicity following intramuscular use. Immune responses in pigs following vaccination and subsequent intra-tracheal bacterial inoculation were examined using lymphocyte proliferation, serology and mucosal IgA in both nasal and saliva swabs.Vaccination induced clear lymphocyte proliferation, but only slight serum antibody responses although these were significantly increased following experimental infection. Mucosal IgA was not detected in either nasal or salivary secretions. Following bacterial challenge, animals vaccinated with the adjuvant-containing live vaccine exhibited less severe pulmonary lesions (median score 3.67) than unvaccinated pigs (median score 13.58). The degree of ciliary loss on the respiratory tract surface was reduced in vaccinated pigs compared with experimentally infected controls. The findings indicated that the adjuvant vaccine administered IM provided protection against experimentally induced mycoplasmal pneumonia and could have commercial potential.     

猪支原体肺炎疫苗在生产中的应用效果观察

猪支原体肺炎 (Mycoplasma pneumonia ofswine,MPS)是由猪肺炎支原体引起的一种猪的慢性呼吸道传染病。该病在我国常称为猪喘气病 (或气喘病 ) ,国外常称为猪地方性流行性肺炎 (Swine en-zootic pneumonia)。该病广泛分布于集约化养猪场 ,患猪长期生长不良、继发病原体感染时也会造成严重死亡。 Pointon等 (1 985)报道 ,病猪生长率减少1 2 .7% ,饲料转化率受抑制 1 3 .8% ,体重减少 1 0～2 5kg。 Clark(1 991 )通过调查、研究认为 ,该病每年对美国养猪业造成的损失就达 2～ 1 0亿美元。近年来 ,随着集约化养猪业的不断发展 ,猪支原体肺炎…     

Evaluation of Mycoplasma hyopneumoniae inactivated vaccine in pigs under field conditions

An inactivated vaccine prepared from broth culture supernatant of Mycoplasma hyopneumoniae with an aluminum adjuvant was evaluated in three herds (herd A: specific pathogen-free herd, herd B: high health status herd with no clinical signs of respiratory infection, herd C: low health status herd with serious epidemiological and economical problems). A total of 212 pigs from the three herds were divided into two groups. One group was injected twice with the vaccine at 4-week intervals and the other was a control group. No adverse reactions were noted following the vaccinations either systematically or locally in any of the vaccinated pigs from any of the herds. In herd A, the vaccination provided antibody response within 4 weeks after the second vaccination and antibody responses continued for more than 12 weeks. In herds B and C, the number of pigs with lung lesions, mean percentage of lung lesions, and the numbers of M. hyopneumoniae recovered from pigs at slaughter in the vaccinated group were significantly (P < 0.05) reduced compared to the control group. Furthermore, vaccination resulted in improved average daily weight gain (ADG), improved feed conversion ratio (FCR), and improved days to market weight in herd C, whereas no difference in growth performance was shown in herd B. It is suggested that the inactivated vaccine prepared from broth culture supernatant of M. hyopneumoniae is effective in reducing clinical signs and lung lesions. Also, vaccination resulted in improved growth performance in herds where clinical signs and economic losses were significant.     

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.     

In vivo and in vitro comparisons of spray-drying and solvent- evaporation preparation of microencapsulated Mycoplasma hyopneumoniae for use as an orally administered vaccine for pigs

OBJECTIVE: To evaluate the efficacy of an orally administered vaccine of Mycoplasma hyopneumoniae that was prepared by spray drying or solvent evaporation. ANIMALS: Thirty 6-week-old, crossbred, specific-pathogen-free (SPF) pigs. PROCEDURE: Pigs were randomly allocated into 5 groups and housed in an SPF facility. Pigs in 2 groups (groups AQ and CAP) were fed M hyopneumoniae enteric-coated vaccine on days 0, 10, and 20. A third group (group IM) received an IM injection of M hyopneumoniae vaccine with aluminium hydroxide as an adjuvant on days 0, 10, and 20. The last 2 groups (non-vaccinated-challenged [NV-C] and nonchallenged [NC]) were fed a sham treatment. All 24 pigs in groups AQ, CAFP IM, and NV-C were challenge exposed with 5 ml of a 10% pneumonic lung suspension administered on day 40 via intubation of the trachea. All pigs were slaughtered and the lungs removed and examined for lesions on day 68. RESULTS: In vitro studies indicated that these 2 microencapsulation techniques formed an effective shell and protected mycoplasmal antigen from gastric acid. Results of inoculation and challenge tests indicated that microencapsulated M hyopneumoniae were sufficiently potent to induce an immune response and provide good protection. CONCLUSIONS AND CLINICAL RELEVANCE: Orally administered microencapsulated M hyopneumoniae vaccines induced an immune response and reduced the severity of lung lesions in challenge-exposed pigs. Results suggest that this novel method can be applied to other antigens, because the spray-drying process yielded an orally administered M hyopneumoniae vaccine that induced a good immune response.     

Comparison of Mycoplasma hyopneumoniae strains by serologic methods

Six field strains of Mycoplasma hyopneumoniae isolated from pneumonic lungs of pigs, reference strains 11 and J of M hyopneumoniae, Ms 42 strain of Mycoplasma flocculare, and BTS 7 strain of Mycoplasma hyorhinis were compared serologically, using hyperimmune antisera produced in rabbits. All strains of M hyopneumoniae were closely related as determined with the disk growth-inhibition test; however, differences in zone sizes indicated that some antigenic heterogeneity existed. Cross-reactions were not detected between M hyopneumoniae, M flocculare, and M hyorhinis with the growth-inhibition test. The metabolic-inhibition test was more useful for detection of intraspecies antigenic difference than was the growth-inhibition test, since antigenic diversity was clearly detected among M hyopneumoniae strains. Slight cross-reactions were observed between M hyopneumoniae and M flocculare. Using 2-dimensional immunoelectrophoresis, antigenic differences were observed among M hyopneumoniae strains, although many common components also were detected in electropherograms. Mycoplasma flocculare possessed a close antigenic relationship to M hyopneumoniae, as determined by two-dimensional immunoelectrophoresis, whereas both organisms were less related to M hyorhinis. Evidence obtained in this study indicated that strains of mycoplasmas tentatively identified as M hyopneumoniae were similar antigenically, but evidence was obtained also of some diversity in antigenic structure among these strains.     

猪肺炎支原体和猪鼻支原体双重PCR检测方法的建立与应用

根据猪肺炎支原体(Mhp)和猪鼻支原体(Mhr)的16S rRNA基因设计3条引物, 建立Mhp和Mhr的双重PCR检测方法,并对该方法进行了特异性和敏感性试验,并使用建立的方法检测了临床样品和疫苗样品。结果显示该方法具有良好的特异性,最低可检测到0.66ng 的Mhp基因组DNA和0.58 ng Mhr基因组DNA,临床样品和疫苗样品检测结果与普通PCR检测结果一致。该双重PCR方法,可用于Mhp与Mhr的鉴别、诊断以及疫苗纯粹性检查,快速而准确。     

Mycoplasma hyopneumoniae colonization of pigs sired by different boars

Differences in Mycoplasma hyopneumoniae colonization were evaluated in experimentally inoculated pigs sired by 3 different boars of the same genetic line. Forty-six pigs were used, including a treatment group and positive and negative control groups. The pigs were intratracheally inoculated with an M. hyopneumoniae suspension or with Friis media as a placebo. To evaluate differences in the magnitude of colonization during a 35-day period, nasal and tracheal swabs were collected weekly and tested by nested polymerase chain reaction (N-PCR). Temperature, weight and circulating antibodies were measured for 35 days. At 11 and 35 d postinoculation the pigs were necropsied and macroscopic and microscopic lesions were determined. A section of bronchus was tested by the indirect immunofluorescence test (IFAT), scanning electron microscopy (SEM) and N-PCR. The N-PCR results from the nasal and tracheal swabs showed that the pigs sired by one boar (B3) had a distinctive colonization pattern, different from that of the pigs from the other 2 boars and from the positive controls. SEM studies demonstrated that at 35 d postinoculation a higher proportion of B3 pigs had lower numbers of mycoplasmas attached to the cilia compared with B1 and B2 offspring. No significant differences were observed in temperature and weight gain among groups by ANOVA; however, with use of a 2 × 2 table, temperature differences were observed between pigs sired by boars B1 and B2 at 4 d postinoculation. No pigs seroconverted, showed gross or microscopic lesions, or had positive IFAT results. These results provide evidence of differences in patterns of colonization between pigs sired by different boars, suggesting a possible genetic effect.     

Studies of the field efficacy and safety of a single-dose Mycoplasma hyopneumoniae vaccine for pigs

The field efficacy and safety of a single-dose Mycoplasma hyopneumoniae vaccine were evaluated in three-to five-week-old pigs. Two field efficacy studies were conducted, one in England with 673 pigs, and one in Germany with 719 pigs. The pigs were injected intramuscularly with either the vaccine or saline (control) at a ratio of 2:1 and reared under commercial conditions to slaughter weight. The efficacy of the vaccine was evaluated by comparing the lung lesions associated with infection with M. hyopneumoniae in the control and vaccinated animals postmortem. In both countries the vaccinated pigs had a significantly lower percentage of lung lesion scores, in England 5.7 v 10.2 per cent (P = 0.0022) and in Germany 3.9 v 7.7 per cent (P = 0.0056). In Germany the average daily weight gain (ADG) of the vaccinated pigs was significantly higher (639 g v 616 g) (P = 0.0205). In both countries and in both the treated and control animals there was a significant negative correlation between the ADG and the lung lesion score (P = 0.0001). Two safety trials were conducted, one in England and one in Germany, each with 75 pigs, and in each case 50 pigs were given the maximum batch release antigen titre of the vaccine and 25 were given saline. The safety of the vaccine was evaluated by observation for local and systemic reactions and any increases in rectal temperature. No abnormal reactions were observed in the vaccinated pigs and there was no significant difference between the mean peak rectal temperatures of the vaccinated and control pigs in either trial.     

皂树皂苷QS21增强猪支原体肺炎活疫苗滴鼻免疫小鼠效果评价

为提高猪支原体肺炎活疫苗的滴鼻免疫效果,以猪支原体肺炎(168株)活疫苗为抗原,以QS21为免疫佐剂混合后滴鼻免疫小鼠,ELISA 法检测免疫效果,包括血清和支气管肺泡灌洗液中抗原特异性抗体水平,并测定了Th1型细胞因子IFN-γ、Th2型细胞因子IL-4和Th17型细胞因子IL-17的分泌情况。结果表明,QS21能激活黏膜和全身性的体液免疫,显著升高血清和支气管肺泡灌洗液中IgG,IgG1、IgG2a抗体含量,并能提高支气管肺泡灌洗液中黏膜免疫抗体sIgA的含量。QS21还能显著增加支气管肺泡灌洗液中IL-4、IFN-γ和IL-17的分泌,激活Th1、Th2和Th17型细胞免疫应答。以上结果表明QS21能全面激活免疫系统,具有开发为猪支原体肺炎活疫苗黏膜免疫佐剂的应用前景。     

Oral immunization using formalin-inactivated Actinobacillus pleuropneumoniae antigens entrapped in microspheres with aqueous dispersion polymers prepared using a co-spray drying process

Oral-vaccine microspheres based on formalin-inactivated Actinobacillus pleuropneumoniae serotype 1 (AP-1) antigens and enteric-coated polymers were prepared using a co-spray drying process. We evaluated using this for a peroral vaccine. We measured specific-antibody titers and protection from challenge in mouse and pig models. In mice (24 per group), a subcutaneous aluminum-adjuvant vaccine or oral vaccination with three doses of AQ6-AP microspheres provided similar protection against intranasal challenge with 5 x 10(8) colony-formation units (cfu) of AP-1 bacterial culture broth. Two weeks after four oral vaccinations with 600 mg of AQ6-AP microsphere acetate solution (containing formalin-inactivated AP-1 antigens of 1.0 x 10(10) cfu bacterial broth), pigs (9 per group) were challenged intranasally with 1 ml of AP-1 bacterial culture broth (5 x 10(9) cfu). The clinical signs, percentage of pig survival ratio, lung lesion areas, and microscopic examinations indicated that the oral AQ6-AP vaccine provided more protection than vaccinating pigs intramuscularly with AP-1 aluminum vaccine.     

A field trial to evaluate a Mycoplasma hyopneumoniae vaccine: effects on lung lesions and growth rates in swine.

A killed Mycoplasma hyopneumoniae vaccine was evaluated in a single swine herd in which the farrowing barn and weaner rooms were on one Mycoplasma-free farm, while the growing and finishing barn was on a separate farm on which Mycoplasma was present. The study was carried out in a cohort of pigs born in a 12-week period. Pigs born in 6 of the 12 wk were vaccinated and the rest were left as controls. The vaccine was administered twice at approximately 3 and 6 wk of age. Carcass characteristics, lung lesions, and growth rates were recorded on 893, 390, and 220 pigs, respectively. The vaccine reduced the prevalence of pneumonic lesions in slaughter hogs from 69% to 36% (P < 0.001). It also appeared to reduce the prevalence of pleuritis from 20% to 13%, but the difference was only statistically significant at P = 0.07. The vaccine had no effects on carcass characteristics except that carcasses of vaccinated pigs were, on average, 1 kg heavier than those of nonvaccinated pigs, and a smaller percentage of vaccinated pigs were shipped "light" (carcass weight < 70 kg). Two methods were used to estimate the effect of the vaccine on growth rates (as measured by days to 80 kg carcass weight) resulting in estimates of 11 and 2 d reduction attributable to vaccination, respectively. The latter estimate was probably an underestimate for reasons discussed in the paper.     

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.     

猪支原体肺炎疫苗研究进展

猪肺炎支原体是引起猪支原体肺炎的病原,该病呈世界性流行,死亡率虽然不高,但由于长期性和消耗性的流行,使饲料转化率降低,并引起猪的多种并发病,是造成养猪业经济损失最重要的疾病之一.目前疫苗免疫是预防猪支原体肺炎和减少经济损失最有效的手段之一,猪支原体肺炎疫苗主要有灭活疫苗和减毒活疫苗.这些疫苗已得到广泛的应用,并在临床上...     

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.     

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.