Bacteria have a tendency to attach to surfaces and grow as structured communities called biofilms. Chronic biofilm infections are a problem because they tend to resist antibiotic treatment and are difficult to eradicate. Bacterial biofilms have an extracellular matrix that is usually composed of a mixture of polysaccharides, proteins, and nucleic acids. This matrix has long been assumed to play a passive structural and protective role for resident biofilm cells. Here we show that this view is an oversimplification and that the biofilm matrix can play an active role in stimulating its own synthesis. Working with the model biofilm bacterium Pseudomonas aeruginosa , we found that Psl, a major biofilm matrix polysaccharide for this species, acts as a signal to stimulate two diguanylate cyclases, SiaD and SadC, to produce the intracellular secondary messenger molecule c-di-GMP. Elevated intracellular concentrations of c-di-GMP then lead to the increased production of Psl and other components of the biofilm. This mechanism represents a unique positive feedback regulatory circuit, where the expression of an extracellular polysaccharide promotes biofilm growth in a manner analogous to autocrine signaling in eukaryotes.

Therapeutic enzyme treatment disrupts Pseudomonas biofilms, potentiating antibiotics and ameliorating the innate immune system.

Mycobacterium tuberculosis is a facultative intracellular pathogen that parasitizes macrophages by modulating properties of the Mycobacterium-containing phagosome. Mycobacterial phagosomes do not fuse with late endosomal/lysosomal organelles but retain access to early endosomal contents by an unknown mechanism. We have previously reported that mycobacterial phosphatidylinositol analog lipoarabinomannan (LAM) blocks a trans-Golgi network-to-phagosome phosphatidylinositol 3-kinase-dependent pathway. In this work, we extend our investigations of the effects of mycobacterial phosphoinositides on host membrane trafficking. We present data demonstrating that phosphatidylinositol mannoside (PIM) specifically stimulated homotypic fusion of early endosomes in an ATP-, cytosol-, and N-ethylmaleimide sensitive factor-dependent manner. The fusion showed absolute requirement for small Rab GTPases, and the stimulatory effect of PIM increased upon partial depletion of membrane Rabs with RabGDI. We found that stimulation of early endosomal fusion by PIM was higher when phosphatidylinositol 3-kinase was inhibited by wortmannin. PIM also stimulated in vitro fusion between model phagosomes and early endosomes. Finally, PIM displayed in vivo effects in macrophages by increasing accumulation of plasma membrane-endosomal syntaxin 4 and transferrin receptor on PIM-coated latex bead phagosomes. In addition, inhibition of phagosomal acidification was detected with PIM-coated beads. The effects of PIM, along with the previously reported action of LAM, suggest that M. tuberculosis has evolved a two-prong strategy to modify its intracellular niche: its products block acquisition of late endosomal/lysosomal constituents, while facilitating fusion with early endosomal compartments. INTRODUCTIONMycobacterium tuberculosis is an intracellular parasite that survives in infected macrophages by modulating phagosomal biogenesis and maturation. After phagocytosis, mycobacteria reside in phagosomes that do not acquire late endosomal and lysosomal characteristics (Armstrong and Hart, 1971;Russell et al., 2002). Another important aspect of the mycobacterial phagosome is that the vacuole seems to retain access to markers derived from plasma membrane (Clemens and Horwitz, 1996;Russell et al., 1996; SturgillKoszycki et al., 1996), allowing acquisition of nutrients, such as iron (Schaible et al., 2002), by M. tuberculosis.The mycobacterial factors affecting phagosome maturation are only now beginning to be identified (Vergne et al., 2003a). In recent studies (Fratti et al., 2001(Fratti et al., , 2003b, lipoarabinomannan (LAM), a heavily glycosylated phosphatidylinositol exclusively produced by mycobacteria, has been shown to interfere with phagosomal acquisition of late endosomal constituents. LAM, and a number of other mycobacterial lipids, have been reported to traffic within the infected macrophages and intercalate into the host cell endomembranes (Beatty et al., 2000). One of the M. tuberculosis glycolipids shown to impregnate host cell endomembranes i...