Serum antibody response to Pseudomonas aeruginosa antigens during corneal infection

Abstract

Previous studies in our laboratory have indicated that naturally resistant, inbred DBA/2J mice mount a greater serum antibody response to Pseudomonas aeruginosa 19660 than susceptible C57BL/6J mice. However, the specificity of the antibody produced was not known. The present study examines the specificity and kinetics of the humoral response of these mouse strains to potential virulence factors produced by the organism during both a primary and a secondary corneal infection administered 4 weeks after the primary infection. Serum antibody levels specific for lipopolysaccharide (LPS), exotoxin A, phospholipase C (PLC), alkaline protease, elastase, and flagella were measured by enzyme-linked immunosorbent assay. Little or no antibody to either alkaline protease or elastase was detected during either primary or secondary infection. Immunoglobulin G (IgG) antibodies specific to exotoxin A, PLC, and flagella were detected 2 weeks after primary infection, and a rapid response to these antigens was measured 1 week after secondary infection. During primary infection, detectable LPS-specific antibody was only IgM, while IgG appeared only after secondary infection. The kinetics of the humoral response in susceptible C57BL/6J mice-were similar to those in resistant DBA/2J mice, although the magnitude of the response varied according to the antigen tested. These results indicate that LPS, exotoxin A, PLC, and flagella are present or produced in amounts that are immunogenic during corneal infection by P. aeruginosa 19660 in the mouse strains tested.Pseudomonas aeruginosa is an opportunistic pathogen which causes severe corneal infections in humans and experimental animals (6,8,14,37,38). The infection is usually preceded by corneal trauma. Thereafter, the infection rapidly spreads and may result in permanent ocular damage. Previous experimental studies from our laboratory have indicated that DBA/2J mice can spontaneously restore corneal clarity within a few weeks after infection and are therefore classified as naturally resistant (4, 18). On the other hand, C57BL/6J mice initially exhibit the same severity of ocular infection at 24 to 72 h postinfection but are unable to restore corneal clarity within a period of 4 weeks and are classified as susceptible (4, 18). Consequently, corneal infection in these mice usually leads to corneal perforation, phthisis bulbi (shrinkage), or both. However, when the mice were reinfected in the contralateral control eye 2 months after the primary corneal infection, most of the mice restored corneal clarity within 3 to 6 days after the secondary infection (7). Recent studies suggest that genetic factors of the host regulate the corneal response of the two mouse strains and that one to two resistance and susceptibility genes may be involved (2, 5). Extending these studies further, we have recently described a correlation between the dissimilar corneal response of the two mouse strains with the ability to mount a specific, rapid, and protective humoral response to P. aeruginosa prior to the develo...

Rapid and sensitive method for evaluating Pseudomonas aeruginosa virulence factors during corneal infections in mice

“…Immune response to infection. Previous works have established the pattern of immune responses to P. aeruginosa antigens during corneal infection with strain 19660 (20,22,23). In this study, we sought to determine whether P. aeruginosaspecific serum IgG could be elicited after infection with lowlevel challenge doses.…”

“…In this model, scratches are made on the corneas of mice anesthetized with inhalants such as ether, and the challenge strain of P. aeruginosa is applied in a small volume (5 l). However, only a few strains of P. aeruginosa have been reported to be pathogenic under these conditions, and inocula of 10 7 to 10 8 CFU are needed to initiate infection (1,2,11,15,20,22,23). The limited number of virulent strains and the need for high-challenge inocula has restricted the use of this model in the evaluation of microbial and host factors involved in the pathogenesis of P. aeruginosa corneal infections.…”

Kinetics of serum, tear, and corneal antibody responses in resistant and susceptible mice intracorneally infected with Pseudomonas aeruginosa

“…Recent substrate specificity studies that examined the serum antibody response to various * Corresponding author. 885 exoenzymes and surface antigens of P. aenrginosa during corneal infection showed that susceptible C57BL/6 mice not only were capable of responding to flagella and the exoenzymes exotoxin A and phospholipase C during both primary and secondary infection but in some cases exceeded the response mounted by DBA/2 mice (29). Neither mouse strain produced antibody to elastase or alkaline protease after either primary or secondary infection (29).…”

“…885 exoenzymes and surface antigens of P. aenrginosa during corneal infection showed that susceptible C57BL/6 mice not only were capable of responding to flagella and the exoenzymes exotoxin A and phospholipase C during both primary and secondary infection but in some cases exceeded the response mounted by DBA/2 mice (29). Neither mouse strain produced antibody to elastase or alkaline protease after either primary or secondary infection (29). These data suggested that the hyporesponsiveness of C57BL/6 mice is restricted to as-yet-unidentified antigens present on the surfaces of P. aeruginosa cells.…”

Pseudomonas aeruginosa antigens as potential vaccines

“…Antigenic properties of exoproducts P. aeruginosa produces a number of secreted virulence factors including exotoxin A, exoenzyme S, proteases I (neutral protease) and III (alkaline phosphatase), elastase, hemolysins (thermolabile phosphatase C and thermostable acid glycolipid), enter- Table 1 Probable role of extracellular products of P. aeruginosa in virulence and immunogenicity Extracellular antigen Role in virulence Antibody or protective response Reference Proteases : (i) Elastase Induces hemorrhagic lesions and necrosis, Abs in patients with chronic and/or active [18^20] tissue invasion, destruction of host defenses, P. aeruginosa infections; provide protections against e.g. complements and opsonins severe lung lesions in animal models (ii) Alkaline protease Destruction of non-speci¢c host defenses, Abs detected in burn wound patients [21] tissue invasion Phospholipase C Destruction of pulmonary surfactant Rise in Ab titer in CF patients colonized with [22,23] Generation of in£ammatory mediators P. aeruginosa, and in animal model infections Rhamnolipids…”