Bacteria that have sustained long-standing close associations with eukaryotic hosts have evolved specific adaptations to survive and replicate in this environment. Perhaps one of the most remarkable of those adaptations is the type III secretion system (T3SS)--a bacterial organelle that has specifically evolved to deliver bacterial proteins into eukaryotic cells. Although originally identified in a handful of pathogenic bacteria, T3SSs are encoded by a large number of bacterial species that are symbiotic or pathogenic for humans, other animals including insects or nematodes, and plants. The study of these systems is leading to unique insights into not only organelle assembly and protein secretion but also mechanisms of symbiosis and pathogenesis.

Pathogenic bacteria of the species Yersinia, including Yersinia pestis, block phagocytosis by macrophages. This process involves the YopE protein, which induces disruption of the host cell actin microfilament structure. Here, we show that the contact between the pathogen and the mammalian cell induces expression and then polarized transfer of YopE into the eukaryotic cell. While the bacteria remain at the surface of the target cell, the YopE cytotoxin is transferred through the host cell plasma membrane and YopE is only recovered within the cytosol of the target cell. The results suggest that the pathogen senses cell structures and focuses the transfer of YopE to occur solely at the interaction zone between the bacterium and the eukaryotic cell. The regulation of this process is shown to involve surface‐located YopN sensor protein of the bacterium.

A flagellin gene from the fish pathogen Vibrio anguillarum was cloned, sequenced, and mutagenized. The DNA sequence suggests that the flaA gene encodes a 40.1-kDa protein and is a single transcriptional unit. A polar mutation and four in-frame deletion mutations (180 bp deleted from the 5 end of the gene, 153 bp deleted from the 3 end of the gene, a double deletion of both the 180-and 153-bp deletions, and 942 bp deleted from the entire gene) were made. Compared with the wild type, all mutants were partially motile, and a shortening of the flagellum was seen by electron microscopy. Wild-type phenotypes were regained when the mutations were transcomplemented with the flaA gene. Protein analysis indicated that the flaA gene corresponds to a 40-kDa protein and that the flagellum consists of three additional flagellin proteins with molecular masses of 41, 42, and 45 kDa. N-terminal sequence analysis confirmed that the additional proteins were flagellins with N termini that are 82 to 88% identical to the N terminus of FlaA. Virulence studies showed that the N terminal deletion, the double deletion, and the 942-bp deletion increased the 50% lethal dose between 70-and 700-fold via immersion infection, whereas infection via intraperitoneal injection showed no loss in virulence. In contrast, the polar mutant and the carboxy-terminal deletion mutant showed approximately a 10 4 -fold increase in the 50% lethal dose by both immersion and intraperitoneal infection. In summary, FlaA is needed for crossing the fish integument and may play a role in virulence after invasion of the host.Vibrio anguillarum, which causes vibriosis in marine fish, is a highly pathogenic bacterium and has become a severe problem for the fish farming industry. An earlier study (3) has correlated the virulence of V. anguillarum with the possession of more than one flagellum. The importance of the flagellum as a potential virulence factor has been demonstrated for other bacteria. In Pseudomonas aeruginosa (5), three wild-type strains and their respective isogenic motility mutants were tested in the burned mouse model. The nonmotile mutants proliferated in the wound but did not cause the characteristic systemic infection, indicating that motility contributes to the invasive capabilities of this organism. For Campylobacter jejuni, the flagellum is the best-characterized virulence factor. In vitro studies using epithelial cell lines have shown that the flagellum aids the bacterium either in adherence to (30) or in internalization within (11) the cultured cells. A more recent study (47) shows that motility and not flagellin A is required for the invasion of intestinal cells by C. jejuni. However, flagellin A can serve as a secondary adhesin for the adherence to intestinal cells, whereas other adhesins are present to aid the motilitydependent internalization in an intestinal cell line. Similarly, for Vibrio cholerae, studies using spontaneous mutants or chemically derived motility mutants indicate that either motility (12) or an adhesin, proposed to be asso...

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It has previously been shown that the plasmid-encoded YopE protein of Yersinia pseudotuberculosis is a virulence determinant. In this study, HeLa cells, macrophages and mice were used as different model systems to determine the actual role of YopE in the virulence process. The YopE protein mediates a cytotoxic response on a confluent layer of HeLa cells. A prerequisite of this activity is that the pathogen binds to the cell surface. YopE also induces a cytotoxic response on mouse macrophages where it influences the ability of the pathogen to resist phagocytosis. Bacterial mutants defective in their ability to express YopE are avirulent after oral or intraperitoneal infection but virulent following intravenous injection. On the basis of these results, we propose a role for YopE in the virulence process of Yersinia.