Alkyl hydroperoxide reductase subunit C (AhpC) is the catalytic subunit responsible for the detoxification of reactive oxygen species that form in bacterial cells or are derived from the host; thus, AhpC facilitates the survival of pathogenic bacteria under environmental stresses or during infection. This study investigates the role of AhpC in the induction and maintenance of a viable but nonculturable (VBNC) state in Vibrio parahaemolyticus. In this investigation, ahpC1 (VPA1683) and ahpC2 (VP0580) were identified in chromosomes II and I of this pathogen, respectively. Mutants with deletions of these two ahpC genes and their complementary strains were constructed from the parent strain KX-V231. The growth of these strains was monitored on tryptic soy agar-3% NaCl in the presence of the extrinsic peroxides H 2 O 2 and tert-butyl hydroperoxide (t-BOOH) at different incubation temperatures. The results revealed that both ahpC genes were protective against t-BOOH, while ahpC1 was protective against H 2 O 2 . The protective function of ahpC2 at 4°C was higher than that of ahpC1. The times required to induce the VBNC state (4.7 weeks) at 4°C in a modified Morita mineral salt solution with 0.5% NaCl and then to maintain the VBNC state (4.7 weeks) in an ahpC2 mutant and an ahpC1 ahpC2 double mutant were significantly shorter than those for the parent strain (for induction, 6.2 weeks; for maintenance, 7.8 weeks) and the ahpC1 mutant (for induction, 6.0 weeks; for maintenance, 8.0 weeks) (P < 0.03). Complementation with an ahpC2 gene reversed the effects of the ahpC2 mutation in shortening the times for induction and maintenance of the VBNC state. This investigation identified the different functions of the two ahpC genes and confirmed the particular role of ahpC2 in the VBNC state of V. parahaemolyticus.

The new PCR method has allowed us to monitor ginger for the presence of P. myriotylum as a way of disease diagnosis or healthy seed ginger examination.

Vibrio parahaemolyticus is a common marine food-borne enteropathogen. In this study, we examined the antioxidative activity, growth, biofilm formation, and cell mobility of an oxyR deletion mutant and its genetically complementary strain of V. parahaemolyticus. oxyR is the regulator of catalase and ahpC genes. Protection against extrinsic H 2 O 2 and against the organic peroxides cumene hydroperoxide and tert-butyl hydroperoxide was weaker in the deletion mutant than in its parent strain. Expression of the major functional antioxidative genes, ahpC1 and VPA1418, was markedly decreased in the oxyR mutant. Growth of this mutant on agar medium was significantly inhibited by autoclaved 0.25% glucose and by 0.25% dipotassium hydrogen phosphate, 0.5% monosaccharides (glucose, galactose, xylose, and arabinose), or 114.8 mM phosphates. The inhibition of the growth of this oxyR mutant by extrinsic peroxides, autoclaved sugars, and phosphates was eliminated by the complementary oxyR gene or by the addition of catalase to the autoclaved medium, while no inhibition of growth was observed when filter-sterilized sugars were used. The formation of biofilm and swimming mobility were significantly inhibited in the oxyR mutant relative to that in the wild-type strain. This investigation demonstrates the antioxidative function of oxyR in V. parahaemolyticus and its possible roles in biofilm formation, cell mobility, and the protection of growth in heated rich medium. Vibrio parahaemolyticus is a halophilic Gram-negative bacterium that is commonly associated with food-borne gastroenteritis (1), and it has exhibited global significance since the occurrence of pandemic O3:K6 strains in 1996 (2).The incomplete reduction of oxygen during aerobic metabolism or by exposure to metals, redox-active chemicals, or some environmental stresses produces various reactive oxygen species (ROS) in bacteria (3-5). ROS can damage all cellular components, including protein, DNA, and membrane lipids (6, 7). Therefore, antioxidative activity is required by pathogenic bacteria for their successful growth and survival under environmental stresses and is sometimes associated with their virulence (8), whereas characteristic antioxidative functions have been demonstrated in V. parahaemolyticus (9, 10).Several common antioxidative factors are used to scavenge ROS, including superoxide dismutases (SOD), catalases, and alkyl hydroperoxide reductase subunit C's (AhpC) (11). The expressions of catalase and ahpC genes are usually regulated by OxyR (12), which is a redox-sensitive transcriptional regulator of the LysR family in Escherichia coli, Salmonella spp., and other bacteria (13-17). OxyR also participates in pathogenesis by promoting biofilm formation, fimbrial expression, and mucosal colonization in pathogenic bacteria (18); nevertheless, the mechanism of oxidative stress defense in these phenomena is not clear (19).The function of oxyR has been examined in a few Vibrio species but not in V. parahaemolyticus. The effect of oxyR on the survival of bacteria a...

The marine foodborne enteropathogen Vibrio parahaemolyticus has four putative catalase genes. The functions of two katEhomologous genes, katE1 (VPA1418) and katE2 (VPA0305), in the growth of this bacterium were examined using gene deletion mutants with or without complementary genes. The growth of the mutant strains in static or shaken cultures in a rich medium at 37°C or at low temperatures (12 and 4°C), with or without competition from Escherichia coli, did not differ from that of the parent strain. When 175 M extrinsic H 2 O 2 was added to the culture medium, bacterial growth of the ⌬katE1 strain was delayed and growth of the ⌬katE1 ⌬katE2 and ⌬katE1 ⌬ahpC1 double mutant strains was completely inhibited at 37°C for 8 h. The sensitivity of the ⌬katE1 strain to the inhibition of growth by H 2 O 2 was higher at low incubation temperatures (12 and 22°C) than at 37°C. The determined gene expression of these catalase and ahpC genes revealed that katE1 was highly expressed in the wild-type strain at 22°C under H 2 O 2 stress, while the katE2 and ahpC genes may play an alternate or compensatory role in the ⌬katE1 strain. This study demonstrated that katE1 encodes the chief functional catalase for detoxifying extrinsic H 2 O 2 during logarithmic growth and that the function of these genes was influenced by incubation temperature.

Alkyl hydroperoxide reductase subunit C gene (ahpC) functions were characterized in Vibrio parahaemolyticus, a commonly occurring marine food-borne enteropathogenic bacterium. Two ahpC genes, ahpC1 (VPA1683) and ahpC2 (VP0580), encoded putative two-cysteine peroxiredoxins, which are highly similar to the homologous proteins of Vibrio vulnificus. The responses of deletion mutants of ahpC genes to various peroxides were compared with and without gene complementation and at different incubation temperatures. The growth of the ahpC1 mutant and ahpC1 ahpC2 double mutant in liquid medium was significantly inhibited by organic peroxides, cumene hydroperoxide and tert-butyl hydroperoxide. However, inhibition was higher at 12°C and 22°C than at 37°C. Inhibiting effects were prevented by the complementary ahpC1 gene. Inconsistent detoxification of H 2 O 2 by ahpC genes was demonstrated in an agar medium but not in a liquid medium. Complementation with an ahpC2 gene partially restored the peroxidase effect in the double ahpC1 ahpC2 mutant at 22°C. This investigation reveals that ahpC1 is the chief peroxidase gene that acts against organic peroxides in V. parahaemolyticus and that the function of the ahpC genes is influenced by incubation temperature. Vibrio parahaemolyticus is a halophilic Gram-negative bacterium which frequently causes food-borne gastroenteritis in some Asian countries (1) as well as globally since the isolation of the first pandemic O3:K6 strains in 1996 (2). This bacterium inhabits seawater and is often isolated from seafood (1). Most clinical isolates of this pathogen are hemolytic on Wagatsuma agar (Kanagawa phenomenon-positive, KP ϩ ) and produce the major virulence factor, thermostable direct hemolysin (TDH).Pathogenic bacteria such as V. parahaemolyticus have evolved sophisticated mechanisms to survive oxidative stresses caused by their metabolic activities, host defense systems, or environmental factors. Various reactive oxygen species (ROS) such as superoxide anion (O 2 Ϫ ), hydrogen peroxide (H 2 O 2 ), and hydroxyl radical ( ⅐ OH) form in bacteria (3-5), and ROS are known to cause damage in all cellular components, including protein, DNA, and membrane lipids.Several common antioxidative factors are typically used to detoxify ROS. One is alkyl hydroperoxide reductase subunit C (AhpC), which is the catalytic subunit of a family of peroxidases collectively known as peroxyredoxins or thiol peroxidases (TPx family) (6). A genome search of V. parahaemolyticus RIMD2210633 (7) revealed several putative AphC factors, including the VPA1683 and VP0580 genes, which have been designated ahpC1 and ahpC2, respectively (8).The antioxidative activity of pathogenic bacteria is related to their survival, growth, and virulence under some environmental stresses. Our earlier studies revealed enhanced production of AhpC in V. parahaemolyticus cells subjected to concurrent cold stress and starvation (9). Our most recent study demonstrated that, at 4°C, ahpC2 has a stronger protective effect than ahpC1 and that ahpC2 i...