The Vibrio pathogenicity island (VPI) in epidemic Vibrio cholerae is an essential virulence gene cluster. Like many pathogenicity islands, the VPI has at its termini a phage-like integrase gene (int), a transposase-like gene (vpiT), and phage-like attachment (att) sites, and is inserted at a tRNA-like locus (ssrA). We report that the VPI precisely excises from the chromosome and that its left and right ends join to form an extrachromosomal circular excision product (pVPI). Two-stage nested PCR analysis and DNA sequencing confirmed the int-attvpiT junction and that the core attP of pVPI is identical to the chromosomal VPI attR site. Excision was independent of toxR and toxT. Excision was independent of recA, suggesting that it is mediated by site-specific recombination. Interestingly, while excision was detected in int and vpiT mutants, excision was abolished in a double (int vpiT) mutant and was restored by plasmids containing genes for either recombinase. Excision results in deletion of A361 in the ssrA locus, which flanks the right junction of the VPI. Since A361 encodes U70 in the critical G ⅐ U base pair in the acceptor stem of the ssrA RNA that is the determinant for aminoacylation with alanine, this deletion might have deleterious effects on ssrA function. Also, vpiT may have undergone interchromosomal translocation or may represent an independent integration event, as it was found downstream of hutA in some isolates. Our results provide new insight into the molecular biology of the VPI, and we propose that the process of excision and circularization is important in the emergence, pathogenesis, and persistence of epidemic V. cholerae.Cholera, attributed to the bacterium Vibrio cholerae, is a diarrheal disease of humans that results in significant morbidity and mortality (25). Available records show that there have been seven worldwide pandemics of cholera (40). The current seventh pandemic began on the island of Sulawesi in Indonesia and then rapidly spread throughout Asia (1, 24). Because of its high death-to-case ratio, transmissibility, and persistence in the environment and its ability to occur in explosive epidemic form, V. cholerae continues to be a public health concern.Only toxigenic strains of V. cholerae that possess two essential virulence gene clusters, i.e., the CTX element, which is a prophage encoding cholera toxin (39, 49), and the Vibrio pathogenicity island (VPI) (26, 30), can cause epidemic cholera. The chromosomal VPI is 41.2 kb in size in both sixthpandemic (classical biotype) and seventh-pandemic (El Tor biotype) strains and encodes 29 potential proteins (26, 27). The VPI encodes proteins with essential roles in virulence, such as toxin-coregulated pili (TCP) (17, 44), and proteins that regulate virulence, such as ToxT, TcpP, and TcpH (4,6,7,14). In addition, the VPI contains several open reading frames with no known or demonstrated function.The VPI has many features that are typical of pathogenicity islands (PAIs) (2, 12, 13). It has a low percent GϩC content (35%) compared to the re...

Aims:  To evaluate enumeration of Vibrio parahaemolyticus in oyster tissues following artificial contamination and depuration. Methods and Results:  After inoculating with V. parahaemolyticus (ATCC 17802) and incubating for 24 h, the contaminated oysters were depurated with artificial seawater for 14 days. At each step, the tissue homogenate supernatants of oysters were spread‐plated onto thiosulfate‐citrate‐bile salt‐sucrose agar, followed by colony confirmation by the polymerase chain reaction. The pathogen was detected in the gills, digestive glands (including stomach, digestive ducts and digestive diverticula), adductor muscle and mantle cilia. After a 48‐h depuration period at 17–19°C, the retention rate of V. parahaemolyticus in the gills (28·1%) and digestive glands (13·5%) was higher than that in adductor muscle and mantle cilia (1·4 and 2·4%, respectively). Conclusions:  The population of V. parahaemolyticus in the digestive glands was the highest among all tissues tested, followed by the gills. The data indicate that digestive glands and gills are good sample candidates for direct monitoring of V. parahaemolyticus contamination in oysters. Significance and Impact of the Study:  This is the first report on the dynamics of V. parahaemolyticus in various oyster tissues following artificial contamination and depuration. This study provides information to help in monitoring for V. parahaemolyticus in commercial oysters.

We have designed a new type of composite materials which not only has a Young’s modulus much larger than the Voigt limit, but also is always nearly isotropic. Moreover, its Poisson’s ratio can be designed at a desired value, e.g. positive, or negative, or zero. We have also demonstrated that structural hierarchy can help to enhance the stiffness of this type of composite materials. The results obtained in this paper provide a very useful insight into the development of new functional materials and structures.

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