Protective Response In Mice Research Articles

DNA immunization in a mouse model of latent tuberculosis: effect of DNA vaccination on reactivation of disease and on reinfection with a secondary challenge.

Individuals who are latently infected with Mycobacterium tuberculosis can develop active disease via either endogenous reactivation of the latent bacilli or exogenous reinfection with a second mycobacterial strain. In this study, we investigated whether immunization with a tuberculosis DNA vaccine cocktail that induces significant protective responses in mice could prevent reactivation of disease in a murine latent-tuberculosis model. In addition, we assessed whether DNA vaccination could retard the growth of a secondary aerogenic infection with M. tuberculosis (exogenous reinfection) in latently infected mice. In the reactivation studies, administration of the DNA vaccine combination did not prevent recrudescence of the latent infection after injection of dexamethasone. Moreover, for the reinfection experiments, only a modest decrease in the growth of a secondary M. tuberculosis challenge in DNA-vaccinated animals, compared to controls, was observed 14 and 28 days after the reinfection of previously exposed mice. Interestingly, although proliferation of the secondary challenge was reduced significantly in a nonvaccinated chronic-infection group relative to the naïve controls, the number of bacilli still increased by 500-fold 1 month after the secondary challenge in mice with active tuberculosis. These results indicate that novel immunotherapeutic approaches will be required to prevent reactivation of infection or reinfection of individuals with latent tuberculosis.

Do Sequence Repeats Play an Equivalent Role in the Choline-binding Module of Pneumococcal LytA Amidase?

Do Sequence Repeats Play an Equivalent Role in the Choline-binding Module of Pneumococcal LytA Amidase?

Immunogenicity of DNA vaccines expressing tuberculosis proteins fused to tissue plasminogen activator signal sequences.

Novel tuberculosis DNA vaccines encoding native ESAT-6, MPT-64, KatG, or HBHA mycobacterial proteins or the same proteins fused to tissue plasminogen activator (TPA) signal sequences were evaluated for their capacity to elicit humoral, cell-mediated, and protective immune responses in vaccinated mice. While all eight plasmids induced specific humoral responses, the constructs expressing the TPA fusions generally evoked higher antibody responses in vaccinated hosts. Although most of the DNA vaccines tested induced a substantial gamma interferon response in the spleen, the antigen-specific lung responses were 2- to 10-fold lower than the splenic responses at the time of challenge. DNA vaccines encoding the ESAT-6, MPT-64, and KatG antigens fused to TPA signal sequences evoked significant protective responses in mice aerogenically challenged with low doses of Mycobacterium tuberculosis Erdman 17 to 21 days after the final immunization. However, the protective response induced by live Mycobacterium bovis BCG vaccine was greater than the response induced by any of the DNA vaccines tested. These results suggest that the tuberculosis DNA vaccines were able to elicit substantial immune responses in suitably vaccinated mice, but further refinements to the constructs or the use of alternative immunization strategies will be needed to improve the efficacy of these vaccine candidates.

Oral or parenteral administration of replication-deficient adenoviruses expressing the measles virus haemagglutinin and fusion proteins: protective immune responses in rodents.

The genes encoding the measles virus (MV) haemagglutinin (H) and fusion (F) proteins were placed under the control of the human cytomegalovirus immediate early promoter in a replication-deficient adenovirus vector. Immunofluorescence and radioimmune precipitation demonstrated the synthesis of each protein and biological activity was confirmed by the detection of haemadsorption and fusion activities in infected cells. Oral as well as parenteral administration of the H-expressing recombinant adenovirus elicited a significant protective response in mice challenged with MV. While the F-expressing adenovirus failed to protect mice, cotton rats immunized with either the H- or F-expressing recombinant showed reduced MV replication in the lungs. Antibodies elicited in mice following immunization with either recombinant had no in vitro neutralizing activity, suggesting a protective mechanism involving a cell-mediated immune response. This study demonstrates the feasibility of using oral administration of adenovirus recombinants to induce protective responses to heterologous proteins.

Protective responses in mice to vaccination with multiply administered cold-adapted influenza vaccine reassortants and wild-type viruses

Protective responses to influenza vaccine reassortants derived from the cold-adapted ( ca) donor strains A/Leningrad/134/17/57 and B/USSR/60/69 and wild-type epidemic viruses were studied in two strains of mice. Preliminary experiments revealed that, when mixtures of three viruses were inoculated intranasally to mice with 50 μl containing 10 6 EID 50 per 200 μl (10 5.4 EID 50 per mouse), interference between strains did not occur. However, interference with the growth of the influenza reassortant B/60/32/R took place if its concentration in the mixture was reduced to 10 5 (10 4.4 per mouse) or if it was inoculated at 10 6 EID 50 (10 5.4 per mouse) in the presence of the influenza reassortant R 34 and two other influenza A epidemic strains; interference was unrelated to serological responses to infection with B/60/32/R. Despite evidence of interference, mice inoculated with the same mixtures in two identical doses, three weeks apart, were able to clear a challenge from each of seven homotypic and heterotypic influenza A and B strains. Heterotypic clearance of influenza A challenge viruses was greater following mixed infection, indicating that common determinants within the surface antigen glycoproteins contributed to immune responses which were broader than could be expected to be induced by parenteral vaccination.

Protective Activity of Different Immunosera AgainstClostridium chauvoei

The protective activity of several antiserum preparations againstC. chauvoeistrain 5078 challenge is studied. Immunosera were obtained from the following antigenic preparations from 12 h-old cultures ofC. chauvoeiat 37°C in anaerobic conditions: (1) flagellar suspension (F); (2) cellular extract (CE): lyophilised cells (0·5 g) extracted by using veronal buffer, pH = 8·6; (3) formaldehyde-treated cell suspension with 0·5% of formol (FC); and (4) heated cells boiled for 2 h(ØC). Lots of white rabbits were inoculated by i.v. route for each immunogenic preparation. The immunosera agglutination titres were: F:1:476, CE: 1:13186; FC 1:476 and ØC: 1:64. Passive immunisation was carried out by injecting by the i.p. route groups of ten mice with 0·5 mL of pooled sera. Mice were challenged 6 h later with a spore suspension in CaCl22·5% containing 10 LD50. Both immunised and non-immunised animals were observed for 4 days. Anti-CE and anti-FC protected 100% of the animals, while anti-F and anti-ØC protected 75% and 50%, respectively. Results showed that protective antigens were extracted by the veronal buffer, and that heat destroys some of them. Although FC protected 100% of immunised animals, it has been shown elsewhere that CE presented the same protective response in mice when diluted at least 1:500. Results also show that flagella contributes in part to the immunogenicity of the strain.

Construction of a trivalent candidateShigella vaccine strain with host-vector balanced-lethal system

A trivalent liveShigella vaccine candidate FSD01 against S.flexneri 2a, S.sonnei and S.dysenteriae I was constructed. This candidate strain was based on the S.flexneri 2a vaccine T32. By homologous recombination exchange, the chromosomalasd gene of T32 was site-specifically inactivated, resulting in the strain unable to grow normally in LB broth, while anotherasd gene of S.mutans was employed to construct an Asd(+) complementary vector. This combination ofasd (-) host/Asd(+) vector formed a balanced-lethal expression system in T32 strain. By use of this system, two important protective antigen genes coding for S.sonnei Form I antigen and Shiga toxin B subunit were cloned and expressed in T32, which led to the construction of trivalent candidate vaccine FSD01. Experimental results showed that this strain was genetically stable, but its recombinant plasmid was non-resistant. Moreover, it was able to effectively express trivalent antigens in one host and induce protective responses in mice against the challenges of the above threeShigella strains.

Immunogenic and protective response in mice immunized with a purified, inactivated, Dengue-2 virus vaccine prototype made in fetal rhesus lung cells.

The feasibility of a purified, inactivated vaccine (PIV) against dengue type 2 (DEN-2) virus was explored. Dengue-2 virus strain 16681 was used for producing a monotypic PIV. Virus adapted to fetal rhesus lung (FRhL-2) cells was harvested from roller bottle culture supernatant fluids, concentrated, and purified on sucrose gradients. Analysis of purified virus preparations by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blotting showed primarily envelope (E) and premembrane (prM) antigens. These preparations had a purity, estimated from silver-stained gels, of approximately 70%, and a yield, based on recovery of virus and viral antigen, of 10-20%. The purified virus was inactivated with 0.05% formalin at 22 degrees C, or alternatively, with 7 mRads from a 60Co source. Vaccinated mice developed high titers of anti-DEN-2 virus neutralizing antibody and were partially protected from virus challenge. These results warrant further testing and development of PIVs for the other DEN virus serotypes.

Immunological analysis of the protective responses to the chimeric synthetic peptide representing T- and B-cell epitopes from the fusion protein of measles virus

The role of different adjuvant formulations and routes of immunization on the antibody responses and protection induced in mice was determined by a synthetic peptide representing T- and B- cell epitopes from the measles virus (MV) fusion (F) protein (TTB peptide) which has previously been shown to induce protective responses against MV encephalitis in mice. When the peptide TTB was administered in Freud's complete adjuvant (FCA), Freud's imcomplete adjuvant (FIA), alum or with IL-2, anti-peptide antibody responses and anti-MV antibody responses were detected. Interestingly, immunization with TTB without adjuvant resulted in the induction of anti-peptide antibody responses which did not cross react with the MV. The use of FIA as an adjuvant led to a significantly higher IgG2a antibody response compared with FCA and alum, whereas alum led to a significantly lower IgG3 response. Immunization with TTB in FCA, FIA and alum led to the generation of high affinity antibodies (with alum generating the highest affinity), whereas immunization of peptide with IL-2 or in PBS resulted in the induction of antibodies of lower affinity. Only the FCA, FIA and alum formulations led to the induction of protective responses in mice against MV-induced encephalitis. When the subcutaneous route (s.c.) of immunization was compared with the intraperitoneal route (i.p.), s.c. immunization with the TTB peptide led to higher anti-peptide and anti-MV antibody responses and higher affinity antibodies compared to those induced following i.p. immunization. Mice receiving the TTB peptide via the s.c. route had a higher percentage survival after MV challenge than those immunized via the i.p. route. These results show that the nature of the adjuvant used as well as the route of immunization play an important role in the induction of protective anti-TTB peptide antibody responses against MV-induced encephalitis in mice.

An antigenic recombinant fusion protein from Trichinella spiralis induces a protective response in BALB/c mice.

A previously reported recombinant lambda Ts39 fusion protein (FP) derived from Trichinella spiralis larvae was purified by affinity chromatography using an anti-beta-galactosidase antibody conjugated column and the effect of FP on induction of a protective response in mice was studied. BALB/c mice were injected three times at weekly intervals with FP emulsified in an equal volume of Freund's complete adjuvant and one week after the last injection mice were each challenged with a lethal dose of 1200 L1 larvae of Trichinella spiralis. As a result, the FP induced significant protection. Mice were also infected orally with 250 L1 larvae each after injection of the FP. On the 35th day of infection, immunized mice with the FP harboured 78% fewer muscle larvae than saline controls. Also, the numbers of adult worms in the small intestine were smaller in FP injected mice than saline controls on day 6 and 9 of the infection. These results suggest that the recombinant lambda Ts39 FP is a potentially valuable antigen for vaccine development.

Induction of antibody and protective responses in mice by Brucella O-polysaccharide-BSA conjugate

A Brucella conjugate vaccine was synthesized by covalently coupling the O-polysaccharide (PS) obtained from Brucella melitensis 16M, to bovine serum albumin (BSA). The conjugate (PS-BSA) was non-toxic for mice. When injected at two doses (10 or 1 micrograms) in mice, it induced the production of antibodies and the boost was shown to be efficient if administered after 10 weeks. Immunization of mice with 10 micrograms of conjugate conferred significant protection against virulent challenge with B. melitensis H38, similar to that obtained with the reference vaccine H38.

Increased local complement levels upon intraperitoneal injection of mice with Listeria monocytogenes and regulation by polyanions

Alternative complement pathway activity in mouse peritoneal fluid was determined by a sensitive microtiter assay with rabbit erythrocytes as target cells and cobra venom factor as the inducer of bystander hemolysis. The intra- and interstrain variations in five mouse strains were 30% and, maximally, 64%, respectively. No activity was detected in genetically C5-deficient mice. In C5-sufficient mice, intraperitoneal injection of heat-inactivated Listeria monocytogenes resulted in a 250% increase in local complement activity within 30 min. This effect was reversed by simultaneous injection of dextran sulfate but not heparin. These results are discussed in relation to the ability of listeriae to activate the mouse alternative complement pathway in vitro and the effectiveness of dextran sulfate and the failure of heparin in functioning as an adjuvant in the induction of a protective response in mice by a heat-inactivated listeria vaccine.