Construction of chimeric phagosomes that shelter Mycobacterium avium and Coxiella burnetii (phase II) in doubly infected mouse macrophages: an ultrastructural study

Dual infection of cells may divert pathogens to intracellular compartments different from those occupied in mono-infected cells. In the present studie s, mouse bone marrow in vitro-derived macrophages were first infected with virulent Mycobacterium avium, which are normally singly lodged within tight phagosomes. These phagosomes do not mature; they undergo homotypic fusion with early endosomes and do not fuse with lysosomes. Seven days later, the cultures were superinfected with phase II (non-virulent) Coxiella burnetii, organisms sheltered in lysosome- (or prelysosome)-like, multi-occupancy ph agosomes. The latter can attain large size and engage in efficient homo- an d heterotypic fusion with other phagosomes. Cultures were fixed for transmi ssion electron microscopy 6, 12, 24, and 48 h later. Other M. avium-infecte d cultures were superinfected with amastigotes of the trypanosomatid flagel late Leishmania amazonensis, which are also sheltered in lysosome- (or prel ysosome)-like multi occupancy vacuoles, and fixed at the same time periods. Chimeric phagosomes containing both M. avium and C. burnetii, were found a lready at 6 h and the proportion of M. avium that colocalized with C. burne tii in the same phagosomes reached over 90 % after 48 h. In such phagosomes , both organisms were ultrastructurally well preserved. In contrast, coloca lization of M. avium and L. amazonensis was rarely found. Speculative scena rios that could underlie the formation of chimeric phagosomes could involve delayed maturation of C. burnetii-containing phagosomes in presence of M. avium, which would allow for fusion of C. burnetii- and M. avium-containing phagosomes; the production, by C. burnetii, of molecules that upregulate t he fusion of M. avium-containing phagosomes with those that contain C. burn etii; and the secretion of factors that could favour the survival of M. avi um within chimeric vacuoles.