Salmonella enteritidis temperature-sensitive mutants protect mice against challenge with virulent Salmonella strains of different serotypes

The protection conferred by temperature-sensitive mutants of Salmonella enteritidis against different wild-type Salmonella serotypes was investigated. Oral immunization with the single temperature-sensitive mutant E/1/3 or with a temperature-sensitive thymine-requiring double mutant (E/1/3T) conferred: (i) significant protection against the homologous wild-type Salmonella strains; (ii) significant cross-protection toward high challenge doses of S. typhimurium. Significant antibody levels against homologous lipopolysaccharide and against homologous and heterologous protein antigens were detected in sera from immunized mice. Moreover, a wide range of protein antigens from different Salmonella O serotypes were recognized by sera from immunized animals. Besides, primed lymphocytes from E/1/3 immunized mice recognized Salmonella antigens from different serotypes. Taken together, these results indicate that temperature-sensitive mutants of S. enteritidis are good candidates for the construction of live vaccines against Salmonella.     

Human C-reactive protein does not protect against acute lipopolysaccharide challenge in mice

The physiological and pathophysiological functions of C-reactive protein (CRP), the classical acute-phase protein, are not well established, despite many reports of biological effects of CRP in vitro and in model systems in vivo. Limited, small scale experiments have suggested that rabbit and human CRP may both protect mice against lethal toxicity of Gram-negative bacterial LPS. However, in substantial well-controlled studies in C57BL/6 mice challenged with Escherichia coli O111:B4 LPS, we show in this work that significant protection against lethality was conferred neither by an autologous acute-phase response to sterile inflammatory stimuli given to wild-type mice 24 h before LPS challenge, nor by human CRP, whether passively administered or expressed transgenically. Male mice transgenic for human CRP, which mount a major acute-phase response of human CRP after LPS injection, were also not protected against the lethality of LPS from either E. coli O55:B5 or Salmonella typhimurium. Even when the acute-phase human CRP response was actively stimulated in transgenic mice before LPS challenge, no protection against LPS toxicity was observed. Indeed, male mice transgenic for human CRP that were pretreated with casein to stimulate an acute-phase response 24 h before LPS challenge suffered significantly greater mortality than unstimulated human CRP transgenic controls. Rather than being protective in this situation, human CRP may thus have pathogenic proinflammatory effects in vivo.     

Broadly neutralizing antibodies protect against hepatitis C virus quasispecies challenge

A major problem in hepatitis C virus (HCV) immunotherapy or vaccine design is the extreme variability of the virus. We identified human monoclonal antibodies (mAbs) that neutralize genetically diverse HCV isolates and protect against heterologous HCV quasispecies challenge in a human liver-chimeric mouse model. The results provide evidence that broadly neutralizing antibodies to HCV protect against heterologous viral infection and suggest that a prophylactic vaccine against HCV may be achievable.     

鼻腔接种伤寒杆菌Fe-SOD对鼠伤寒杆菌攻击小鼠的免疫保护作用

为探讨鼻腔接种伤寒杆菌Fe-SOD对鼠伤寒杆菌攻击小鼠的交叉免疫保护作用,用IL－1作为佐剂,将伤寒杆菌Fe-SOD经鼻腔接种小鼠,再以不同剂鼠伤寒杆菌攻击,观察小鼠的存活情况,当用IL－1作为佐剂时,经鼻腔接种伤寒杆菌Fe-SOD的小鼠在2LD50鼠伤寒杆菌攻击后,3天和7天的存活率均明显高于对照组（P＜0．01或P＜0．05）,当以5LD50鼠直菌攻击时,小鼠7天的存活率明显高于对照组（P＜0．01）。结果说明伤寒杆菌Fe-SOD经鼻腔接种后对鼠伤寒杆菌的攻击可产生一定的免疫保护作用,也进一步说明了Fe-SOD是沙门氏菌的共同保护性抗原。     

Monoclonal antibodies against defined Leishmania antigens protect mice against infection by different species of Leishmania

Mice monoclonal antibodies (IgG) have been raised against Leishmania infantum promastigotes by fusing SP 2/0 myeloma cells and immunized mice splenic cells. The monoclonal antibodies have been detected by indirect immunofluorescence. In vivo tests showed that some of them could inhibit the life cycle of several Leishmania species from the Old and the New World. Studies of these protective monoclonals by the western blot technique showed the presence of three constant antigens (40 kD, 70 kD and 113 kD) amongst the Leishmania species studied.     

Antimicrobial action of nisin against Salmonella typhimurium lipopolysaccharide mutants.

The antimicrobial activity of nisin against outer membrane lipopolysaccharide mutants of Salmonella typhimurium LT2 was investigated. Nisin sensitivity was associated with the extent of saccharide deletions from the outer membrane core oligosaccharide. The results indicated that the core oligosaccharide in lipopolysaccharide plays a role in nisin sensitivity.     

Antimicrobial action of nisin against Salmonella typhimurium lipopolysaccharide mutants.

The antimicrobial activity of nisin against outer membrane lipopolysaccharide mutants of Salmonella typhimurium LT2 was investigated. Nisin sensitivity was associated with the extent of saccharide deletions from the outer membrane core oligosaccharide. The results indicated that the core oligosaccharide in lipopolysaccharide plays a role in nisin sensitivity.     

Protective Capacity of L-Form Salmonella typhimurium against Murine Typhoid in C3H/HeJ Mice

L forms of Salmonella typhimurium LT2 conferred strong protection to a lethal challenge with its parental bacterium on innately hypersusceptible C3H/HeJ mice, and its minimal protective dose was approximately 150 L-forming units. Although L-form S. typhimurium was avirulent for C3H/HeJ mice, it multiplied slowly in both the liver and spleen with the maximal growth 2–3 weeks after immunization and thereafter it persisted in the liver until 24 weeks. Protective immunity began to work between 4 and 6 weeks after immunization, and it remained active as long as the L forms colonized the liver (until 24 weeks after immunization). Vaccination with the L form induced a population of T cells responding to L-form whole-cell lysate (WCL), while delayed-type hypersensitivity (DTH) to the extract of S. typhimurium was induced after the establishment of solid immunity. Moreover, neither T-cell responses nor DTH to heat-killed S. typhimurium was generated. In addition, antibody responses were elicited to WCL but not to heat-killed S. typhimurium. These results indicate that protection conferred by the L forms is attributable to the persistent colonization of the L forms rather than the presence of DTH, and also that Salmonella cytoplasmic antigens are involved in induction of immunological responses by vaccination with the L forms.     

Immunization with SsEno fails to protect mice against challenge with Streptococcus suis serotype 2

In our ongoing efforts to develop a vaccine against Streptococcus suis infection, we tested the potential of S. suis enolase (SsEno), a recently described S. suis adhesin with fibronectin-binding activity, as a vaccine candidate in a mouse model of S. suis -induced septicemia and meningitis. Here, we show that SsEno is highly recognized by sera from convalescent pigs and is highly immunogenic in mice. Subcutaneous immunization of mice with SsEno elicited strong immunoglobulin G (IgG) antibody responses. All four IgG subclasses were induced, with IgG1, IgG2a and IgG2b representing the highest titers followed by IgG3. However, SsEno-vaccinated and nonvaccinated control groups showed similar mortality rates after challenge infection with the highly virulent S. suis strain 166'. Similar results were obtained upon passive immunization of mice with hyperimmunized rabbit IgG anti-SsEno. We also showed that anti-SsEno antibodies did not increase the ability of mouse phagocytes to kill S. suis in vitro . In conclusion, these data demonstrate that although recombinant SsEno formulated with Quil A triggers a strong antibody response, it does not confer effective protection against infection with S. suis serotype 2 in mice.     

Interaction of the globular domain of human C1q with Salmonella typhimurium lipopolysaccharide

Gram-negative bacteria can bind complement protein C1q in an antibody-independent manner and activate classical pathway via their lipopolysaccharides (LPS). Earlier studies have implicated the collagen-like region of human C1q in binding LPS. In recent years, a number of C1q target molecules, previously considered to interact with collagen-like region of C1q, have been shown to bind via the globular domain (gC1q). Here we report, using recombinant forms of the globular head regions of C1q A, B and C chains, that LPS derived from Salmonella typhimurium interact specifically with the B-chain of the gC1q domain in a calcium-dependent manner. LPS and IgG-binding sites on the gC1q domain appear to be overlapping and this interaction can be inhibited by a synthetic C1q inhibitor, suggesting common interacting mechanisms.     

Monoclonal, but not polyclonal, antibodies protect against Plasmodium yoelii sporozoites

One of the primary strategies for malaria vaccine development has been to design subunit vaccines that induce protective levels of antibodies against the circumsporozoite (CS) protein of malaria sporozoites. In the Plasmodium yoelii mouse model system such vaccines have been uniformly unsuccessful in protecting against sporozoite-induced malaria. To demonstrate that antibodies to P. yoelii CS protein could provide protection we established a passive transfer model. Passive transfer of Navy yoelii sporozoite 1 (NYS1), an IgG3 mAb against the P. yoelii CS protein, protected 100% of mice against challenge with 5000 P. yoelii sporozoites. Binding of NYS1 to sporozoites was inhibited by incubation with (QGPGAP)2, a synthetic peptide derived from the repeat region of the P. yoelii CS protein, indicating that the epitope on sporozoites recognized by this mAb was included within this peptide. The levels of antibodies to (QGPGAP)2 by ELISA, and to sporozoites by indirect fluorescent antibody test and CS precipitation reaction were similar in sera from mice that received NYS1 in passive transfer and were protected against challenge with 5000 sporozoites, and from mice that had been immunized with subunit vaccines containing (QGPGAP)2 but were not protected against challenge with 40-200 sporozoites. To determine if antibody avidity, not absolute concentration could explain the striking differences in protection, we established a thiocyanate elution assay. The results suggest that NYS1, the protective mAb, has a lower avidity for (QGPGAP)2 and for sporozoites than do the vaccine-induced antibodies. Although the results of the conventional antibody assays did not correlate with protection, sera from the protected animals inhibited sporozoite development in mouse hepatocyte cultures significantly more than did the sera from the unprotected, subunit vaccine-immunized animals, correlating with protection. The data clearly demonstrate that antibodies to the CS protein can protect against intense sporozoite infection. Improved understanding of the differences between protective mAb and nonprotective polyclonal antibodies will be important in the further development of malaria vaccines.     

Macrophage-mediated mitogenic suppression induced in mice of the C3H lineage by a vaccine strain of Salmonella typhimurium

Salmonella typhimurium, strain SL3235, an avirulent organism, has been used as a live vaccine in mice of the C3H lineage and has been found to confer high levels of protection. In the present study, it was found that intraperitoneal injection of approximately 5 X 10(5) live SL3235 induced potent suppression of spleen cell mitogenic responses to a panel of B- and T-cell mitogens in the Salmonella-hypersusceptible C3H/HeJ and C3HeB/FeJ, and the inherently resistant C3H/HeNCrlBR mice. Maximal suppression (greater than 99%) was seen at 1 week, and was still significant but waning (50%) at 3 weeks postimmunization. In contrast, cells of mice receiving acetone-killed cells were not suppressed. Removal of macrophages, but not T or B cells, restored responsiveness, indicating that suppression was macrophage mediated. Prostaglandins were not the major mediator of suppression, as in vitro administration of indomethacin failed to abrogate suppression. As mitogenic suppression occurred in mice with high levels of Salmonella immunity, the suppression is interpreted as a marker of a powerful immunomodulatory process induced by live cells, rather than as an indication of poor immune status of the host.     

Complex of recombinant heat shock protein with lipopolysaccharide induces rapid protection of mice against Salmonella typhimurium and avirulent for humans avian influenza virus H5N2

Protective effect of immunization with heat shock protein (HSP) against bacterial and viral infections in mice was studied. Recombinant HSP 70 kDa of Mycobacterium tuberculosis contaminated with lypopolysaccharide (0.185 mcg/ml) was used for experiments. One intraperitoneal injection of 100 or 400 mcg of HSP induced rapid protection against intraperitoneal challen e with 125 LD50 of Salmonella typhimurium (on 3rd-6th day) and against intranasal challenge with 10 LD50 of avirulent for humans avian influenza virus H5N2 (A/ mallard/Pennsylvania/10218/84) (on 5th-8th day). Three daily injections with 10 mcg of HSP induced rapid, significant and long-term protection against S. typhimurium. Immunization with HSP protected 100% of mice during 3 days after the challenge, 50% of immunized animals survived during 21 days (duration of the study). All nonimmunized mice died on 6th day.     

Cytotoxic antibodies against tumor cells in the blood of intact C3H mice]

Cytotoxic antibodies against mouse mammary tumour cells, L-cells and hepatoma 22a cells have been found in the serum of C3H/f and C3H/He mice over 8 months of age. Analogues antibodies were found in the serum of young and old BALB/c mice, but not in C57BL/6 mice. The cytotoxic activity of antimammary tumour cell serum has been completely abolished by its depletion by renal tissue of syngeneic and allogeneic animals.     

Feeding truncated heat shock protein 70s protect Artemia franciscana against virulent Vibrio campbellii challenge

The 70 kDa heat shock proteins (Hsp70s) are highly conserved in evolution, leading to striking similarities in structure and composition between eukaryotic Hsp70s and their homologs in prokaryotes. The eukaryotic Hsp70 like the DnaK (Escherichia coli equivalent Hsp70) protein, consist of three functionally distinct domains: an N-terminal 44-kDa ATPase portion, an 18-kDa peptide-binding domain and a C-terminal 10-kDa fragment. Previously, the amino acid sequence of eukaryotic (the brine shrimp Artemia franciscana) Hsp70 and DnaK proteins were shown to share a high degree of homology, particularly in the peptide-binding domain (59.6%, the putative innate immunity-activating portion) compared to the N-terminal ATPase (48.8%) and the C-terminal lid domains (19.4%). Next to this remarkable conservation, these proteins have been shown to generate protective immunity in Artemia against pathogenic Vibrio campbellii. This study, aimed to unravel the Vibrio-protective domain of Hsp70s in vivo, demonstrated that gnotobiotically cultured Artemia fed with recombinant C-terminal fragment (containing the conserved peptide binding domain) of Artemia Hsp70 or DnaK protein were well protected against subsequent Vibrio challenge. In addition, the prophenoloxidase (proPO) system, at both mRNA and protein activity levels, was also markedly induced by these truncated proteins, suggesting epitope(s) responsible for priming the proPO system and presumably other immune-related genes, consequently boosting Artemia survival upon challenge with V. campbellii, might be located within this conserved region of the peptide binding domain.     

Monoclonal antibodies against distinct determinants of histone H5 bind to chromatin

A series of monoclonal antibodies specific for distinguishable epitopes in chromosomal protein histone H5 were obtained from mice immunized with either free H5 or H5 . RNA complexes. The antibodies elicited by H5 could be distinguished from those elicited by H5 . RNA by their binding to native or acid-denatured H5, by their interaction with the globular region of H5, and by their cross-reactivity with H1o. The specificity of the antibodies was assessed by enzyme-linked immunosorbent assay (ELISA) and immunoblotting experiments. The antibodies could distinguish between H5 and the closely related histones H1 and H1o. The binding of some of the antibodies to the antigens was dependent on the type of assay used, suggesting nonrandom binding of the antigen to the solid supports used in ELISA and immunoblotting. Competitive ELISA experiments indicate that 8 of the 11 antibodies characterized bind to distinct epitopes. Three monoclonal antibodies bind to epitopes which are in close spatial proximity, causing mutual steric hindrance. The monoclonal antibodies bind to nuclei of fixed cells and to isolated chromatin, indicating that the epitopes are present both in the purified protein and in chromatin-complexed H5. These monoclonal antibodies can be used to study the organization of distinct regions of histones H5 and H1o in chromatin and chromosomes.     

Immunisation with Salmonella typhimurium-delivered glyceraldehyde-3-phosphate dehydrogenase protects mice against challenge infection with Echinococcus multilocularis eggs

Recombinant glyceraldehyde-3-phosphate dehydrogenase of the cestode parasite Echinococcus multilocularis was expressed in Escherichia coli and in Salmonella typhimurium. The potential of different forms of the recombinant antigen to protect BALB/c mice against oral challenge infections with E. multilocularis eggs was evaluated. Oral or intraperitoneal immunisation with live attenuated S. typhimurium as a carrier for recombinant glyceraldehyde-3-phosphate dehydrogenase of the E. multilocularis resulted in significant protection, reducing the number of developing metacestodes up to 79.8%. The sera of protected animals did not contain detectable amounts of antibody against glyceraldehyde-3-phosphate dehydrogenase of E. multilocularis. By contrast, although anti-glyceraldehyde-3-phosphate dehydrogenase of E. multilocularis antibodies were detectable in the sera, immunisation with E. coli-expressed recombinant glutathione-S-transferase-fusion protein or with glyceraldehyde-3-phosphate dehydrogenase of E. multilocularis fused to a 6HIS-tag failed to protect the animals against oral challenge infections. These data emphasise that antigen delivery systems play a critical role in vaccination and the induction of protective immunity against helminth parasites.