An immunoprotective molecule, the major secretory protein of Legionella pneumophila, is not a virulence factor in a guinea pig model of Legionnaires' disease.

ABSTRACT The activities of HMR 3647, HMR 3004, erythromycin, clarithromycin, and levofloxacin for 97 Legionella spp. isolates were determined by microbroth dilution susceptibility testing. Growth inhibition of two Legionella pneumophila strains grown in guinea pig alveolar macrophages was also determined. The concentrations required to inhibit 50% of strains tested were 0.06, 0.02, 0.25, 0.03, and 0.02 μg/ml for HMR 3647, HMR 3004, erythromycin, clarithromycin, and levofloxacin, respectively. BYEα broth did not significantly inhibit the activities of the drugs tested, as judged by the susceptibility of the control Staphylococcus aureus strain; however, when Escherichia coli was used as the test strain, levofloxacin activity tested in BYEα broth was fourfold lower. HMR 3647, HMR 3004, erythromycin, and clarithromycin (0.25 and 1 μg/ml) reduced bacterial counts of two L. pneumophila strains grown in guinea pig alveolar macrophages by 0.5 to 1 log10, but regrowth occurred over a 2-day period. HMR 3647, erythromycin, and clarithromycin appeared to have equivalent intracellular activities which were solely static in nature. HMR 3004 was more active than all drugs tested except levofloxacin. In contrast, levofloxacin (1 μg/ml) was bactericidal against intracellular L. pneumophilaand significantly more active than the other drugs tested. Therapy studies with HMR 3647 and erythromycin were performed in guinea pigs with L. pneumophila pneumonia. When HMR 3647 was given (10 mg/kg of body weight) by the intraperitoneal route to infected guinea pigs, mean peak plasma levels were 1.4 μg/ml at 0.5 h and 1.0 μg/ml at 1 h postinjection. The terminal half-life phase of elimination from plasma was 1.4 h. All 16 L. pneumophila-infected guinea pigs treated with HMR 3647 (10 mg/kg/dose given intraperitoneally once daily) for 5 days survived for 9 days after antimicrobial therapy, as did all 16 guinea pigs treated with the same dose of HMR 3647 given twice daily. Fourteen of 16 erythromycin-treated (30 mg/kg/dose given intraperitoneally twice daily) animals survived, whereas 0 of 12 animals treated with saline survived. HMR 3647 is effective against L. pneumophilain vitro, in infected macrophages, and in a guinea pig model of Legionnaires’ disease. HMR 3647 given once daily should be evaluated as a treatment for Legionnaires’ disease in humans.

Download Full-text

Vaccination with the major secretory protein of Legionella pneumophila induces cell-mediated and protective immunity in a guinea pig model of Legionnaires' disease.

Journal of Experimental Medicine ◽

10.1084/jem.169.3.691 ◽

1989 ◽

Vol 169 (3) ◽

pp. 691-705 ◽

Cited By ~ 51

Author(s):

S J Blander ◽

M A Horwitz

Keyword(s):

Immune Response ◽

Legionella Pneumophila ◽

Guinea Pigs ◽

Protective Immunity ◽

Secretory Protein ◽

Sublethal Dose ◽

Exact Test ◽

Cell Mediated Immune Response ◽

Legionnaires Disease ◽

Guinea Pig Model

We have examined the capacity of the major secretory protein (MSP) of Legionella pneumophila to induce humoral, cell-mediated, and protective immunity in a guinea pig model of Legionnaires' disease. MSP was purified to homogeneity by ammonium sulfate precipitation, molecular sieve chromatography, and ion-exchange chromatography. The purified MSP was nonlethal and nontoxic to guinea pigs upon subcutaneous administration. Guinea pigs immunized with a sublethal dose of aerosolized L. pneumophila or a subcutaneous dose of MSP developed a strong cell-mediated immune response to MSP. Such guinea pigs exhibited marked splenic lymphocyte proliferation and cutaneous delayed-type hypersensitivity to MSP in comparison with control animals. Guinea pigs immunized with MSP also developed a strong humoral immune response to MSP, as assayed by an ELISA. The median reciprocal antibody titer was 362 (range 45 to greater than 2,048) for immunized animals compared with less than 8 for controls. In contrast, guinea pigs immunized with a sublethal dose of L. pneumophila failed to develop anti-MSP antibody. Guinea pigs immunized with MSP and then challenged with a lethal aerosol dose of L. pneumophila exhibited highly significant protective immunity in each of five consecutive experiments. MSP induced protective immunity in dose-dependent fashion (40 greater than 10 greater than 2.5 greater than 0.6 micrograms MSP); vaccination with two doses of as little as 2.5 micrograms MSP induced significant protective immunity (p = 0.01, Fisher's Exact Test, two-tailed). Altogether, 21 (81%) of 26 animals immunized with 40 micrograms MSP survived challenge compared with 0 (0%) of 26 sham-immunized control animals (p = 7 x 10(-10), Fisher's Exact Test, two-tailed). MSP-immunized but not control guinea pigs were able to limit L. pneumophila multiplication in their lungs. This study demonstrates that (a) guinea pigs sublethally infected with L. pneumophila develop a strong cell-mediated immune response to MSP; (b) guinea pigs immunized with MSP develop a strong humoral and cell-mediated immune response to MSP; (c) guinea pigs immunized with MSP develop a very high level of protective immunity to lethal aerosol challenge with L. pneumophila; and (d) MSP-immunized animals are able to limit L. pneumophila multiplication in their lungs. MSP, an extracellular protein of an intracellular pathogen, has potential as a vaccine for the prevention of Legionnaires' disease. Secretory molecules of other intracellular pathogens may also have vaccine potential.

Download Full-text

The Guinea-pig Model of Legionnaires' Disease

Handbook of Animal Models of Infection ◽

10.1016/b978-012775390-4/50173-1 ◽

1999 ◽

pp. 303-314 ◽

Cited By ~ 7

Author(s):

P.H. Edelstein

Keyword(s):

Guinea Pig ◽

Pig Model ◽

Legionnaires Disease ◽

Guinea Pig Model

Download Full-text

Enhancing the Protective Efficacy of Mycobacterium bovis BCG Vaccination against Tuberculosis by Boosting with the Mycobacterium tuberculosis Major Secretory Protein

Infection and Immunity ◽

10.1128/iai.73.8.4676-4683.2005 ◽

2005 ◽

Vol 73 (8) ◽

pp. 4676-4683 ◽

Cited By ~ 35

Author(s):

Marcus A. Horwitz ◽

Günter Harth ◽

Barbara Jane Dillon ◽

Saša Masleša-Galić

Keyword(s):

Mycobacterium Tuberculosis ◽

Pulmonary Tuberculosis ◽

Guinea Pig ◽

Mycobacterium Bovis ◽

Protective Immunity ◽

Protective Efficacy ◽

Secretory Protein ◽

Pig Model ◽

Booster Vaccine ◽

Guinea Pig Model

ABSTRACT Tuberculosis continues to ravage humanity, killing 2 million people yearly. Most cases occur in areas of the world to which the disease is endemic, where almost everyone is vaccinated early in life with Mycobacterium bovis BCG, the currently available vaccine against tuberculosis. Thus, while more-potent vaccines are needed to replace BCG, new vaccines are also needed to boost the immune protection of the 4 billion people already vaccinated with BCG. Until now, no booster vaccine has been shown capable of significantly enhancing the level of protective immunity induced by BCG in the stringent guinea pig model of pulmonary tuberculosis, the “gold standard” for testing tuberculosis vaccines. In this paper, we describe a booster vaccine for BCG comprising the purified recombinant Mycobacterium tuberculosis 30-kDa protein, the major secreted protein of this pathogen. In the guinea pig model of pulmonary tuberculosis, boosting BCG-immunized animals once with the 30-kDa protein greatly increased cell-mediated and humoral immune responses to the protein in three consecutive experiments. Most importantly, boosting BCG-immunized animals once with the 30-kDa protein significantly enhanced protective immunity against aerosol challenge with highly virulent M. tuberculosis, as evidenced by a significantly reduced lung and spleen burden of M. tuberculosis compared with those for nonboosted BCG-immunized animals (mean additional reduction in CFU of 0.4 ± 0.1 log in the lung [P = 0.03] and 0.6 ± 0.1 log in the spleen [P = 0.002]). This study suggests that administering BCG-immunized people a booster vaccine comprising the 30-kDa protein may enhance their level of immunoprotection against tuberculosis.

Download Full-text

A New Vaccine against Tuberculosis Affords Greater Survival after Challenge than the Current Vaccine in the Guinea Pig Model of Pulmonary Tuberculosis

Infection and Immunity ◽

10.1128/iai.71.4.1672-1679.2003 ◽

2003 ◽

Vol 71 (4) ◽

pp. 1672-1679 ◽

Cited By ~ 150

Author(s):

Marcus A. Horwitz ◽

Günter Harth

Keyword(s):

Guinea Pig ◽

Virulent Strain ◽

Secretory Protein ◽

Pig Model ◽

Regional Lymph Nodes ◽

Current Vaccine ◽

Guinea Pig Model ◽

Highly Virulent ◽

Global Health Problem ◽

Recombinant Bcg

ABSTRACT Tuberculosis (TB) remains an enormous global health problem, and a new vaccine against TB more potent than the current inadequate vaccine, Mycobacterium bovis BCG, is urgently needed. We describe a recombinant BCG vaccine (rBCG30) expressing and secreting the 30-kDa major secretory protein of Mycobacterium tuberculosis, the primary causative agent of TB, that affords greater survival after challenge than parental BCG in the highly demanding guinea pig model of pulmonary TB. Animals immunized with rBCG30 and then challenged by aerosol with a highly virulent strain of M. tuberculosis survived significantly longer than animals immunized with conventional BCG. The parental and recombinant vaccine strains are comparably avirulent in guinea pigs, as they display a similar pattern of growth and clearance in the lung, spleen, and regional lymph nodes. The pMTB30 plasmid encoding the 30-kDa protein is neither self-transmissible nor mobilizable to other bacteria, including mycobacteria. The pMTB30 plasmid can be stably maintained in Escherichia coli but is expressed only in mycobacteria. The recombinant and parental strains are sensitive to the same antimycobacterial antibiotics. rBCG30, the first vaccine against TB more potent than nearly century-old BCG, is being readied for human clinical trials.

Download Full-text