HISTOPLASMA-CAPSULATUM MODULATES THE ACIDIFICATION OF PHAGOLYSOSOMES

The phagolysosome is perhaps the most effective antimicrobial site wit hin macrophages due both to its acidity and to its variety of hydrolyt ic enzymes. Few species of pathogens survive and multiply in these ves icles. However, one strategy for microbial survival would be to induce a higher pH within these organelles, thus interfering with the activi ty of many lysosomal enzymes. Altering the intravesicular milieu might also profoundly influence antigen processing, antimicrobial drug deli very, and drug activity. Here we report the first example of an organi sm proliferating within phagolysosomes that maintain a relatively neut ral pH for a sustained period of time. We inoculated P388D1 macrophage s with fluorescein isothiocyanate (FITC)-labeled Histoplasma capsulatu m or zymosan. Using the ratio of fluorescence excitations at 495 and 4 50 nm, we determined that vesicles containing either virulent or aviru lent FITC-labeled H. capsulatum yeasts had a pH one to two units highe r than vesicles containing either zymosan or methanol-killed H. capsul atum. The difference in pH remained stable for at least 5.5 h postinoc ulation. Longer-term studies using cells preincubated with acridine or ange indicated that phagolysosomes containing live Histoplasma continu ed to maintain a relatively neutral pH for at least 30 h. Many agents raise the pH of multiple vesicles within the same cell. In contrast, H . capsulatum affects only the phagolysosome in which it is located; du ring coinoculation of cells with unlabeled Histoplasma and labeled zym osan, organelles containing zymosan still acidified normally. Similarl y, unlabeled zymosan had no influence on the elevated pH of vesicles h ousing labeled Histoplasma. Thus, zymosan and Histoplasma were segrega ted into separate phagolysosomes that responded independently to their phagocytized contents. This localized effect might reflect an intrins ic difference between phagosomes housing the two particle types, activ e buffering by the microbe, or altered ion transport across the phagol ysosomal membrane such that acidification is inhibited.