Immunization of mice with recombinant S-adenosyl-L-homocysteine hydrolase protein confers protection against Brucella melitensis infection

Due to drawbacks of live attenuated vaccines, much attention has been focused on screening Brucella-protective antigens as subunit vaccine candidates. Here, an immunoproteomic assay was used to identify the immunogenic soluble proteins of Brucella melitensis 16M. In the present study, 27 unique immunogenic proteins were identified from the two-dimensional electrophoresis immunoblot profiles by liquid chromatography tandem MS (LC-MS/MS). From this set, the gene encoding one immunodominant protein of interest, S-adenosyl-l-homocysteine hydrolase (AdoHcyase), was expressed in Escherichia coli. The recombinant AdoHcyase induced a strong antibody response in BALB/c mice, and the polyclonal antibody could recognize a band of approximately 52 kDa in the immunoblots of soluble protein extracts from five Brucella strains. rAdoHcyase significantly stimulated the production of interferon-γ and interleukin-2, and induced a high level of protection against B. melitensis 16M challenge at 4 weeks postchallenge. Our results indicated that rAdoHcyase could be a useful candidate for the development of subunit vaccines against B. melitensis.     

Reversion of a streptomycin-dependent strain of Escherichia coli

Summary A streptomycin dependent, spectinomycin resistant mutant ofEscherichia coli was used to select spontaneous phenotypic revertants to non-dependence on streptomycin. The ribosomes from one such revertant, which is inhibited by both streptomycin and spectinomycin, were analyzedin vitro. The altered protein responsible for the suppression of the streptomycin dependent phenotype was identified; this protein is 30S-10. The genetic locus for this mutation is a newly identified locus and it has been positioned close to thestr locus. The identification of the altered component responsible for the suppression of the spectinomycin resistant phenotype may be the same as that for the streptomycin dependent phenotype, but this is unproven.     

The candidate vaccine strain Brucella ovis ∆abcBA is protective against Brucella melitensis infection in mice

Brucellosis is a major zoonotic disease, and Brucella melitensis is the species most often associated with human infection. Vaccination is the most efficient tool for controlling animal brucellosis, with a consequent decrease of incidence of human infections. Commercially available live attenuated vaccines provide some degree of protection, but retain residual pathogenicity to human and animals. In this study, Brucella ovis ∆abcBA (Bo∆abcBA), a live attenuated candidate vaccine strain, was tested in two formulations (encapsulated with alginate and alginate plus vitelline protein B [VpB]) to immunize mice against experimental challenge with B. melitensis strain 16M. One week after infection, livers and spleens of immunized mice had reduced numbers of the challenge strain B. melitensis 16M when compared with those of nonimmunized mice, with a reduction of approximately 1-log₁₀ of B. melitensis 16M count in the spleens from immunized mice. Moreover, splenocytes stimulated with B. melitensis antigens in vitro secreted IFN-γ when mice had been immunized with Bo∆abcBA encapsulated with alginate plus VpB, but not with alginate alone. Body and liver weights were similar among groups, although spleens from mice immunized with Bo∆abcBA encapsulated with alginate were larger than those immunized with Bo∆abcBA encapsulated with alginate plus VpB or nonimmunized mice. This study demonstrated that two vaccine formulations containing Bo∆abcBA protected mice against experimental challenge with B. melitensis.     

Immunization with a pentameric L1 fusion protein protects against papillomavirus infection

The prophylactic papillomavirus vaccines currently in clinical trials are composed of viral L1 capsid protein that is synthesized in eukaryotic expression systems and purified in the form of virus-like particles (VLPs). To evaluate whether VLPs are necessary for effective vaccination, we expressed the L1 protein as a glutathione S-transferase (GST) fusion protein in Escherichia coli and assayed its immunogenic activity in an established canine oral papillomavirus (COPV) model that previously validated the efficacy of VLP vaccines. The GST-COPV L1 fusion protein formed pentamers, but these capsomere-like structures did not assemble into VLPs. Despite the lack of VLP formation, the GST-COPV L1 protein retained its native conformation as determined by reactivity with conformation-specific anti-COPV antibodies. Most importantly, the GST-COPV L1 pentamers completely protected dogs from high-dose viral infection of their oral mucosa. L1 fusion proteins expressed in bacteria represent an economical alternative to VLPs as a human papillomavirus vaccine.     

带有氯霉素抗性基因标记的重组布氏杆菌S2的构建及其生物学特性

无法区分免疫接种和自然感染是限制布氏杆菌疫苗应用的主要原因。本研究通过基因同源重组技术,用氯霉素抗性基因(Cmr)替代布氏杆菌弱毒S2株的WbkC基因,筛选获得重组布氏杆菌rS2-WbkC株。试验发现,重组菌rS2-WbkC由原先的光滑型转变成粗糙型。rS2-WbkC在TSA培养基上传25代后仍能稳定表达氯霉素抗性蛋白。小鼠动物试验模型表明,rS2-WbkC与S2有相似的保护性,但rS2比S2具有更高的安全性。rS2-WbkC免疫小鼠后,其抗血清可通过平板凝集试验与S2免疫的血清相区分。本研究为布氏杆菌标记疫苗的研制提供了技术平台。     

Immunization of sheep against infection with larvae of the blowfly Lucilia cuprina

Sheep were repeatedly exposed to a second stage excretory-secretory antigen preparation of Lucilia cuprina by intradermal injection (S group) or intranasal aerosol (I group) in an attempt to induce immunity to the larvae. Hypersensitivity responses to the injections were monitored and correlated with larval numbers at subsequent challenge. Intradermal injections showed that the immediate or IgE-mediated and the intermediate or Arthus response were the major skin hypersensitivity reactions to the larval antigens. At challenge there was no significant reduction in larval numbers between the S and the control group, however the Arthus reaction did show some correlation with larval recoveries in the S group. There was a significant reduction in larval numbers (P < 0.005 Mann-Whitney U Test) between the I and the control group. In addition, the animals which showed respiratory hypersensitivity to the aerosol during the immunization period had the lowest larval recoveries. The results of a second challenge in the I group did not show continued protection. It is suggested that the protective response was suppressed by exposure to antigens at the first challenge infection. Exudate samples recovered during the infection were analysed for protein amount and the results were correlated with larval survival. They suggested that two separate mechanisms of resistance are operating in this experiment. The first occurs early in the infection, is probably associated with immediate hypersensitivity, and may control the initiation of protein leakage. The second occurs later in the infection and may result in the leakage of proteins able to control larval survival.