Escherichia coli pre-exposed to a sub-minimal inhibitory concentration (sub-MIC) of several antibiotics elicits an enhanced humoral response which is protective against challenges with untreated homologous and heterologous bacteria. To characterise the specificity of this response we produced murine monoclonal antibodies (MAbs) to aztreonamtreated E. coli 0 6 : K-. This resulted in the identification of MAb MT 1 F, of isotype IgG,, that recognised a 12-kDa protein component of the untreated bacterial cells. After passive transfer, the MAb displayed protective activity in mice infected with lethal doses of live E. coli 0 6 : K-and E. coli 0 1 11 : B4. In ELISA experiments the MAb cross-reacted with structures located on whole cells of E. coli 0 6 : K-, E. coli 0 1 1 1 : B4, E. coli J5 and Salmonella minnesota Re595 and it also exerted a bactericidal activity against live E. coli 0 6 : K-. The modifications induced by antibiotic treatment may unmask bacterial epitopes that may elicit the production of MAbs endowed with protective capacity.

The murine immune response to Escherichia coli O6:K-alone or pre-exposed to 0.1 x MIC of aztreonam was investigated. Relative to mice immunized with untreated bacteria, mice immunized with antibiotic-treated microorganisms presented a significantly enhanced protection towards a challenge of 100 x LD50 of viable E. coli O6:K-. Previous injection of 0.1 mL of serum drawn from mice immunized with treated and untreated bacteria protected non-immunized mice towards a challenge of 10 x LD50 of viable E. coli O6:K--. Serum from mice immunized with treated bacteria also protected non-immunized mice towards a lethal challenge of E. coli O111. The antiserum contained high titre of IgG antibodies that cross-reacted with lipopolysaccharide isolated from smooth and rough Gram-negative bacteria. Immunoblotting showed additional bands of reactivity to the untreated E. coli O6:K-. Immunization with antibiotic-treated bacteria led to the production of type specific and cross reactive antibodies that protected animals against viable homologous and heterologous lethal challenges.

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The capacity of human and murine polyclonal and monoclonal antibodies to inhibit lipopolysaccharide (LPS)-induced tumor necrosis factor-alpha (TNF-alpha) release from human monocytes was investigated. Human pooled immunoglobulin G (IVIG), human IgM monoclonal antibody (HA-1A) directed against the lipid A moiety of LPS, and murine IgG monoclonal antibody (MT-1F) raised in mice against antibiotic-treated Escherichia coli O6:K- were either added simultaneously with LPS to monocytes or preincubated for 1 h at 37 degrees C before being added to monocytes. TNF-alpha content in the monocyte supernatants was then tested. Simultaneous addition of increasing concentrations of IVIG (from 0.3 to 2.5 mg/ml) and 10 micrograms/ml of LPS to monocytes induced an enhanced release of TNF-alpha by monocytes in a dose dependent fashion. Preincubation of IVIG with LPS abolished the additive effect, but did not inhibit LPS-induced TNF-alpha release by monocytes. The simultaneous addition of LPS and HA-1A to monocytes had no additive effect nor did it inhibit TNF-alpha release. On the other hand, inhibition of TNF-alpha release was observed when HA-1A was preincubated with LPS before being added to monocytes. In all instances MT-1F inhibited TNF-alpha release when the monocytes were stimulated with smooth type LPS, but not with LPS isolated from rough mutants.