Effect of normal and immune sera on Haemophilus ducreyi 35000HP and its isogenic MOMP and LOS mutants

A bactericidal assay was developed in order to test the effect of hyperimmu ne rabbit sera on the viability of serum-resistant Haemophilus ducreyi 3500 0HP. Testing of several lots of rabbit complement and time course experimen ts showed that the serum-sensitive H. ducreyi CIPA77 was killed efficiently by 25% complement at 35 degrees C in 3 h. We hypothesized that incubation of 35000HP under these conditions with the appropriate bactericidal antibod y would kill this strain. A panel of high titre rabbit antisera was develop ed and tested against 35000HP. The panel included antisera raised to whole cells, total membranes, Sarkosyl-insoluble outer membrane proteins, the H. ducreyi lipoprotein, and the peptidoglycan-associated lipoprotein. None of the antisera convincingly showed bactericidal activity. The bactericidal as say was also used to determine the effect of normal human serum (NHS) on is ogenic mutants of 35000HP. 35000HP-RSM2, an Omega kan insertion mutant that expresses a truncated lipooligosaccharide, was as resistant to NHS as its parent. A mutant deficient in expression of the major outer membrane protei n (35000.60) was sensitive to NHS. We conclude that 35000HP is relatively r esistant to normal and hyperimmune sera, and that the major outer membrane protein contributes to this resistance. (C) 1999 Academic Press.