A B S T R A C T Serum antibodies to exotoxin A and type-specific lipopolysaccharide were measured by passive hemagglutination in 52 patients with Pseudomonas aeruginosa septicemia. Their comparative protective activities were evaluated by relating the titers of each at the onset of bacteremia to subsequent outcome. High acute serum antitoxin and antilipopolysaccharide titers (log2 reciprocal mean titers >5) were associated with survival (76% of 17 with high vs. 46% of 24 with low antitoxin titers, P = 0.05; 85% of 13 with high vs. 48% of29 with low antilipopolysaccharide titers, P = 0.03). In contrast, neither antibody titer was significantly associated (P c 0.05) with patients' age or sex, severity ofunderlying disease, presence ofleukopenia, steroid or immunosuppressive therapy. Despite a correlation between acute titers of the two antibodies (r = 0.33, P = 0.06), they appeared to protect independently and additively. Whereas 75% of 8 patients with high antitoxin titers and only 38% of 16 with low titers survived with low antilipopolysaccharide titers (P = 0.10), 100% (6/6), 73% (8/11), and 38% (6/16) survived, respectively, when both, one, or neither antibody was present in high titer (P = 0.01). Furthermore, the association between high acute serum antitoxin titers and survival was more pronounced in patients with rapidly fatal underlying disease (P = 0.06) and leukopenia (P = 0.12) than in more favorable prognostic and immune categories. These data indicate that serum antibodies to exotoxin A and lipopolysaccharide are found in most patients with P. aeruginosa septicemiaThe opinions and assertions contained herein are the private ones of the writers and are not to be construed as official or reflecting the views of the Navy Department or the naval service at large.

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100 years after the discovery of a bacterial 'endotoxin', 50 years after the introduction of antibiotics and 25 years after the routine use of intensive care units to support septic shock patients, Gram-negative infections continue to account for significant morbidity and mortality. In the coming decade, basic research on the structure/function of LPS, the cytokine cascade, and receptor-mediated intracellular signalling responses to LPS and cytokines will provide a greater understanding of the molecular, cellular and systemic responses to endotoxin and infection. New therapeutic agents now emerging from research, and better designed clinical trials to assess those agents will contribute to the next significant decline in sepsis- and shock-related morbidity and mortality. This article summarizes the findings of a workshop convened at the National Institutes of Health (NIH) to examine current research on endotoxin and Gram-negative septic shock.

The ability of six antibiotics from different classes to release radiolabeled lipopolysaccharide (LPS) from a phenotypically smooth galE mutant of Escherichia coli O111:B4 was examined. Antibiotic concentrations were 0.0625-512 micrograms/mL. LPS release increased as a function of the antibiotic concentration, reaching a limit at or near the concentration that killed the majority of bacteria. The maximum amount of LPS released by polymyxin B was 40.6% +/- 0.9%, by gentamicin 58.2% +/- 2.5%, by ciprofloxacin 65.8% +/- 2.5%, by ceftazidime 73.1% +/- 0.9%, by tetracycline 75.3% +/- 10.0%, and by imipenem 79.7% +/- 2.3%. In timed experiments, ceftazidime released 61.9% +/- 1.2%, imipenem 51.1% +/- 8.8%, and tetracycline 39.7% +/- 4.4% of the LPS within the first hour of incubation, whereas polymyxin B released 13.5% +/- 1.9%, gentamicin 9.8% +/- 3.6%, and ciprofloxacin 12.7% +/- 2.6% of the LPS (P < .05). Fluoro-radiography and immunoblot analyses revealed similar migration patterns for antibiotic-released and cell-bound LPS on SDS-PAGE gels, suggesting similar O-polysaccharide content in the two LPS fractions. The amount and rate of LPS release from an E. coli strain was dependent upon antibiotic class and concentration.

The protective effect of intravenously administred rabbit antitoxin serum was studied in lethal Pseudomonas aeruginosa bum infections in mice. Survival after infection with 2 median lethal doses of a toxigenic, low-protease-producing strain (PA103) was enhanced in antitoxin-treated mice, as compared with controls that had received anti-bovine serum albumin serum (P = 0.0004). Survival time was prolonged in other antitoxin-treated mice infected with toxigenic, highprotease-producing strains (PA86 and PA220, P = 0.0003 and P = 0.01, respectively). In contrast, antitoxin had no protective effect in mice challenged with a nontoxigenic strain (WR 5, P = 0.57). There were fewer viable bacteria in blood and liver of antitoxin-treated mice than in those of anti-bovine serum albumintreated controls after infection with toxigenic organisms, whereas there were no significant differences between the two groups after challenge with the nontoxigenic strain. These data suggest that P. aeruginosa exotoxin A contributes to lethality in this burn infection model, and this effect is diminished by passive immunization with antitoxin.

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Twenty-nine murine monoclonal antibodies (MAbs) were prepared against antigenic determinants in the core and lipid A regions of Escherichia coli and Salmonella minnesota lipopolysaccharide (LPS). At least eight distinct MAb specificities were identified. Epitopes recognized by MAbs bearing these specificities were localized in the hexose, heptose, and 2-keto-3-deoxy-D-manno-octulosonic acid regions of the core oligosaccharide and on lipid A. Two groups of MAbs exhibited multispecificity for similar but distinct core- and lipid A-related epitopes. Some core-reactive MAbs cross-reacted with corresponding E. coli and Salmonella rough mutant chemotypes; others were specific for E. coli J5 LPS. Lipid A-specific MAbs reacted with free lipid A from diverse sources. Few MAbs reacted with smooth LPS. Antibody cross-reactivity was restricted by inter- and intraspecies differences in covalent core structure and by epitope concealment by overlying O-side chain and core sugars. The putative cross-reactive and antiendotoxic properties of MAbs specific for the core-lipid A complex may be limited by the inability of such MAbs to recognize determinants on "native" LPS.