Vibrio parahaemolyticus is a bacterial pathogen causative of food-borne gastroenteritis. Whole-genome sequencing of V. parahaemolyticus strain RIMD2210633, which exhibits Kanagawa phenomenon (KP), revealed the presence of two sets of the genes for the type III secretion system (T3SS) on chromosomes 1 and 2, T3SS1 and T3SS2, respectively. Although T3SS2 of the RIMD2210633 strain is thought to be involved in human pathogenicity, i.e., enterotoxicity, the genes for T3SS2 have not been found in trh-positive (KP-negative) V. parahaemolyticus strains, which are also pathogenic for humans. In the study described here, the DNA region of approximately 100 kb that surrounds the trh gene of a trh-positive V. parahaemolyticus strain, TH3996, was sequenced and its genetic organization determined. This revealed the presence of the genes for a novel T3SS in this region. Animal experiments using the deletion mutant strains of a gene (vscC2) for the novel T3SS apparatus indicated that the T3SS is essential for the enterotoxicity of the TH3996 strain. PCR analysis showed that all the trh-positive V. parahaemolyticus strains tested possess the novel T3SS-related genes. Phylogenetic analysis demonstrated that although the novel T3SS is closely related to T3SS2 of KP-positive V. parahaemolyticus, it belongs to a distinctly different lineage. Furthermore, the two types of T3SS2 lineage are also found among pathogenic Vibrio cholerae non-O1/non-O139 strains. Our findings demonstrate that these two distinct types are distributed not only within a species but also beyond the species level and provide a new insight into the pathogenicity and evolution of Vibrio species.Vibrio parahaemolyticus is a gram-negative halophilic marine and estuarine bacterium which is an important pathogen causative of food-borne gastroenteritis and traveler's diarrhea (1). Although most V. parahaemolyticus strains are nonpathogenic for humans, a limited population of these organisms causes human diseases. Almost all clinical V. parahaemolyticus isolates produce the thermostable direct hemolysin (TDH) and/or the TDH-related hemolysin (TRH), which are encoded by the tdh and trh genes, respectively (5, 21). The Kanagawa phenomenon (KP), a beta-type hemolysis on a special blood agar (Wagatsuma agar) (28), is known as a good marker of pathogenic strains (5, 21). V. parahaemolyticus strains which exhibit KP possess the two tdh genes tdhA (tdh2) and tdhS (tdh1) but not the trh gene (6,19,21). In contrast, KP-negative clinical V. parahaemolyticus strains possess the trh gene only or both the trh and tdh genes, while the majority of the nonpathogenic strains possess neither tdh nor trh.TDH and TRH, which have several biological activities in common (5,20,30,33), are considered to be the major virulence factors in clinical V. parahaemolyticus strains (5, 30).However, several studies have demonstrated that although the enterotoxicity was reduced in tdh-or trh-deleted mutant strains from that in the parent strains, the enterotoxic activity of these mutant strains partiall...

SummaryVibrio parahaemolyticus strain RIMD2210633 has two sets of genes encoding two separate type III secretion systems (T3SSs), called T3SS1 and T3SS2. T3SS2 has a role in enterotoxicity and is present only in Kanagawa phenomenon-positive strains, which are pathogenic to humans. Accordingly, T3SS2 is considered to be closely related to V. parahaemolyticus human pathogenicity. Despite this, the biological actions of T3SS2 and the identity of the effector protein(s) secreted by this system have not been well understood. Here we report that T3SS2 induces a cytotoxic effect in Caco-2 and HCT-8 cells. Moreover, it was revealed that VPA1327 (vopT), a gene encoded within the proximity of T3SS2, is partly responsible for this cytotoxic effect. The VopT shows approximately 45% and 44% identity with the ADPribosyltransferase (ADPRT) domain of ExoT and ExoS, respectively, which are two T3SS-secreted effectors of Pseudomonas aeruginosa. T3SS2 was found to be necessary not only for the secretion, but also for the translocation of the VopT into host cells. We also demonstrate that VopT ADP-ribosylates Ras, a member of the low-molecular-weight G (LMWG) proteins both in vivo and in vitro. These results indicate that VopT is a novel ADPRT effector secreted via V. parahaemolyticus T3SS.

This letter reports on the reduction in extended-defect densities in a-plane (112̄0) GaN films achieved via lateral epitaxial overgrowth (LEO) by hydride vapor phase-epitaxy. A variety of dielectric mask patterns was used to produce 8–125-μm-thick, fully coalesced nonpolar GaN films. The nanometer-scale pit densities in the overgrown regions were less than 3×106 cm−2 compared to ∼1010 cm−2 in the direct-growth a-plane GaN. Cathodoluminescence revealed a fourfold increase in luminous intensity in the overgrown material compared to the window material. X-ray rocking curves indicate the films were free of wing tilt within the sensitivity of the measurements. Whereas non-LEO a-plane GaN exhibits basal plane stacking fault and threading dislocation densities of 105 cm−1 and 109 cm−2, respectively, the overgrown LEO material was essentially free of extended defects. The basal plane stacking fault and threading dislocation densities in the wing regions were below the detection limits of ∼5×106 cm−2 and 3×103 cm−1, respectively.

Vibrio parahaemolyticus, a Gram-negative halophilic bacterium that causes acute gastroenteritis in humans, is characterized by two type III secretion systems (T3SS), namely T3SS1 and T3SS2. T3SS2 is indispensable for enterotoxicity but the effector(s) involved are unknown. Here, we identify VopV as a critical effector that is required to mediate V. parahaemolyticus T3SS2-dependent enterotoxicity. VopV was found to possess multiple F-actin-binding domains and the enterotoxicity caused by VopV correlated with its F-actin-binding activity. Furthermore, a T3SS2-related secretion system and a vopV homologous gene were also involved in the enterotoxicity of a non-O1/non-O139 V. cholerae strain. These results indicate that the F-actin-targeting effector VopV is involved in enterotoxic activity of T3SS2-possessing bacterial pathogens.

This letter reports on extended defect density reduction in m-plane (11¯00) GaN films achieved via lateral epitaxial overgrowth (LEO) by hydride vapor phase epitaxy. Several dielectric mask patterns were used to produce 10 to 100 μm-thick, partially and fully coalesced nonpolar GaN films. X-ray rocking curves indicated the films were free of wing tilt. Transmission electron microscopy showed that basal plane stacking fault (SF) and threading dislocation (TD) densities decreased from 105cm−1 and 109cm−2, respectively, less than 3×103cm−1 and ∼5×106cm−2, respectively, in the Ga-face (0001) wing of the LEO films. SFs persisted in ⟨0001⟩-oriented stripe LEO films, though TD reduction was observed in the windows and wings. Band-edge cathodoluminescence intensity increased 2 to 5 times in the wings compared to the windows depending on the stripe orientation. SFs in the low TD density wings of ⟨0001⟩-stripe films did not appear to act as nonradiative recombination centers.

Influence of the carrier gas composition on morphology, dislocations, and microscopic luminescence properties of selectively grown GaN by hydride vapor phase epitaxy

Clostridium perfringens is a causative agent of food-borne gastroenteritis for which C. perfringens enterotoxin (CPE) has been considered an essential factor. Recently, we experienced two outbreaks of food-borne gastroenteritis in which non-CPE producers of C. perfringens were strongly suspected to be the cause. Here, we report a novel enterotoxin produced by C. perfringens isolates, BEC (binary enterotoxin of C. perfringens). Culture supernatants of the C. perfringens strains showed fluid-accumulating activity in rabbit ileal loop and suckling mouse assays. Purification of the enterotoxic substance in the supernatants and high-throughput sequencing of genomic DNA of the strains revealed BEC, composed of BECa and BECb. BECa and BECb displayed limited amino acid sequence similarity to other binary toxin family members, such as the C. perfringens iota toxin. The becAB genes were located on 54.5-kb pCP13-like plasmids. Recombinant BECb (rBECb) alone had fluid-accumulating activity in the suckling mouse assay. Although rBECa alone did not show enterotoxic activity, rBECa enhanced the enterotoxicity of rBECb when simultaneously administered in suckling mice. The entertoxicity of the mutant in which the becB gene was disrupted was dramatically decreased compared to that of the parental strain. rBECa showed an ADP-ribosylating activity on purified actin. Although we have not directly evaluated whether BECb delivers BECa into cells, rounding of Vero cells occurred only when cells were treated with both rBECa and rBECb. These results suggest that BEC is a novel enterotoxin of C. perfringens distinct from CPE, and that BEC-producing C. perfringens strains can be causative agents of acute gastroenteritis in humans. Additionally, the presence of becAB on nearly identical plasmids in distinct lineages of C. perfringens isolates suggests the involvement of horizontal gene transfer in the acquisition of the toxin genes.C lostridium perfringens, a spore-forming anaerobic rod, is a member of the normal intestinal flora in humans and animals and a component of soil and sewage microbiota (1-4). C. perfringens is the causative agent of various human diseases, including gas gangrene and food-borne gastroenteritis (5-10). The pathogenicity of C. perfringens is attributed to various toxins produced by the organism, including alpha, beta, epsilon, and iota toxins that classify C. perfringens isolates into five toxin types (A to E), theta toxin, NetB, and C. perfringens enterotoxin (CPE) (10-13).CPE, which is mainly produced by type A C. perfringens, is associated with human gastrointestinal (GI) illnesses, such as food-borne gastroenteritis, antibiotic-associated diarrhea, and sporadic diarrhea (14-16). CPE is a 35-kDa protein, and the cpe gene is located in the chromosome or on a plasmid (17)(18)(19)(20). After orally ingested CPE-positive C. perfringens reaches the GI tract, sporulating C. perfringens produces CPE, and the toxin causes clinical symptoms, such as diarrhea and abdominal cramping. In the clinical diagnosis of gastroente...

Summary Vibrio parahaemolyticus is a Gram-negative marine bacterium that causes acute gastroenteritis in humans. The virulence of V. parahaemolyticus is dependent upon a type III secretion system (T3SS2). One effector for T3SS2, VopC, is a homolog of the catalytic domain of cytotoxic necrotizing factor (CNF), and was recently reported to be a Rho family GTPase activator and to be linked to internalization of V. parahaemolyticus by nonphagocytic cultured cells. Here, we provide direct evidence that VopC deamidates Rac1 and CDC42, but not RhoA, in vivo. Our results also suggest that VopC, through its activation of Rac1, contributes to formation of actin stress fibers in infected cells. Invasion of host cells, which occurs at a low frequency, does not seem linked to Rac1 activation, but instead appears to require CDC42. Finally, using an infant rabbit model of V. parahaemolyticus infection, we show that the virulence of V. parahaemolyticus is not dependent upon VopC-mediated invasion. Genetic inactivation of VopC did not impair intestinal colonization nor reduce signs of disease, including fluid accumulation, diarrhea, and tissue destruction. Thus, although VopC can promote host cell invasion, such internalization is not a critical step of the disease process, consistent with the traditional view of V. parahaemolyticus as an extracellular pathogen.