Induction of beta-defensin resistance in the oral anaerobe Porphyromonas gingivalis

Induction of resistance of oral anaerobes to the effects of human beta-defensin 1 (hbetaD-1) to hbetaD-4 was investigated by pretreating cells with either sublethal levels of defensins or environmental factors, followed by a challenge with lethal levels of defensins. Cultures of Porphyromonas gingivalis were (i) pretreated with defensins at 1 ng/ml, (ii) heated to 42degreesC (heat stress), (iii) exposed to normal atmosphere (oxidative stress), or (iv) exposed to 1 mM hydrogen peroxide (peroxide stress). Samples (10 mul) were distributed among the wells of sterile 384-well plates containing hbetaD-1 to -4 (100 mug/ml). Plates were incubated at 37degreesC for 36 h in an anaerobe chamber. Growth inhibition was determined by a system that measures the total nucleic acid of a sample with a DNA binding dye. The MICs of the four defensins for P. gingivalis were 3 to 12 mug/ml. We found that sublethal levels of the defensins and beat and peroxide stress, but not oxidative stress, induced resistance to 100 mug of defensin per ml in P. gingivalis. Resistance induced by sublethal levels of hbetaD-2 lasted 90 min, and the resistance induced by each defensin was effective against the other three. Multiple strains exposed to hbetaD-2 all evidenced resistance induction. Defensin resistance is vital to the pathogenic potential of several human pathogens. This is the first report describing the induction of defensin resistance in the oral periodontal pathogen P. gingivalis. Such resistance may have an effect on the ability of oral pathogens to persist in the mouth and to withstand innate human immunity.