Immunodetection of lipopolysaccharide in macrophages during the processing of non invasive Shigella dysenteriae

The location of lipopolysaccharide (LPS) was studied by immunofluorescence and immunoelectron microscopy in macrophages infected with a non-invasive Shigella dysenteriae 1 strain. Bacterial degradation began only 3 h after the end of infection. The first visible sign of degradation was detected by immunogold labelling at the level of LPS which detached from the bacterial surface and was transferred to the perinuclear lysosomes. After a few hours, it was found in small vesicles spread over the whole macrophage cytoplasm in which it remained visible for 72 h. These vesicles seemed to belong to a compartment in which slowly or non-degradable compounds are stored. LPS separation from the bacterial surface was immediately followed by the degradation of the intrabacterial constituents. The long lag period observed before initiation of bacterial degradation was not due to a lack of phagosome acidification, since DAMP, a lysosomotropic drug was found in all phagosomes at the end of the ingestion period. The frequency of phagosome-lysosome fusion was 30% for S dysenteriae and 72% for B subtilis used as a reference of high fusion frequency. The low frequency of fusion of S dysenteriae may play an important role in the survival of the virulent strains in macrophage by providing bacteria enough time to lyse the phagosome membrane before lysosome fusion occurs.