Similarity in the Structure of tetD-Carrying Mobile Genetic Elements in Bacterial Strains of Different Genera Isolated from Cultured Yellowtail.

Structure analysis was performed on the antibiotic-resistance-gene region of conjugative plasmids of four fish farm bacteria.The kanamycin resistance gene, IS26, and tetracycline resistance gene (tetA(D)) were flanked by two IS26s in opposite orientation in Citrobacter sp. TA3 and TA6, and Alteromonas sp. TA55 from fish farm A. IS26-Inner was disrupted with ISRSB101. The chloramphenicol resistance gene, IS26 and tetA (D) were flanked by two IS26s in direct orientation in Salmonella sp. TC67 from farm C. Structures of tetA (D) and IS26 were identical among the four bacteria, but there was no insertion within the IS26-Inner of Salmonella sp. TC67. Horizontal gene transfer between the strains of two different genera in fish farm A was suggested by the structure homologies of mobile genetic elements and antibiotic resistance genes.

Inhibitory Effects of Echinochrome A, Isolated from Shell of the Sea Urchin Anthocidaris crassispina, on Antigen-Stimulated Degranulation in Rat Basophilic Leukemia RBL-2H3 Cells through Suppression of Lyn Activation.

Echinochrome A (Echi-A) was isolated from the sea urchin Anthocidaris crassispina and its structure determined using ID and 2D-NMR. In the present study, we examined the inhibitory effect of Echi-A on antigen-stimulated degranulation in rat basophilic leukemia RBL-2H3 cells, which were suppressed in a dose dependent manner.. The antigens bind to the high affinity immunoglobulin E receptor, which is expressed on the surface of mast cells and basophils and activate intracellular signal transduction, resulting in the release of biologically active mediators such as histamine. In order to disclose the inhibitory mechanisms of degranulation by Echi-A, we examined the elevation in intracellular C²âº concentration ([Ca²âº]i), production levels of intracellular reactive oxygen species (ROS) and early intracellular signaling events. Both elevation of [Ca²âº] and intracellular ROS production were markedly suppressed in cells treated with Echi-A. Echi-A also suppressed the activation of Lyn, Syk, and PLCyl/2 in antigen-stimulated cells. These results indicated that inhibition of antigen-stimulated degranulation in RBL-2H3 cells by Echi-A is mainly due to the inactivation of Lyn/Syk/PLCy signaling pathways. Our findings suggest that Echi-A could be a beneficial agent for alleviating the symptoms of type I allergy.

Biochemical analyses of the antioxidative activity and chemical ingredients in eight different Allium alien monosomic addition lines.

We measured the antioxidant contents and antioxidative activities in eight Allium fistulosum-shallot monosomic addition lines (MAL; FF+1A-FF+8A). The high antioxidative activity lines (FF+2A and FF+6A) showed high polyphenol accumulation. These additional chromosomes (2A and 6A) would therefore have anonymous genes related to the upregulation of polyphenol production, the antioxidative activities consequently being increased in these MALs.

Supplementation with Japanese bunching onion (Allium fistulosum L.) expressing a single alien chromosome from shallot increases the antioxidant activity of Kamaboko fish jelly paste in vitro.

Kamaboko is a traditional type of processed seafood made from fish jelly paste that is unique to Japan. We supplemented Kamaboko with Japanese bunching onion (Allium fistulosum L.) with an alien monosome from shallot (Allium cepa L. Aggregatum group) and we measured in vitro the oxygen radical absorbance capacity (ORAC) value, an index of antioxidant activity. We also evaluated the results of sensory testing. The ORAC value of plain Kamaboko was 166±14 µmol trolox equivalent (TE)/100 g fresh weight (FW). The values of the edible Alliaceae powder, i.e., Japanese bunching onion (JBO, genome FF, 2n=2x=16) and the alien addition line of JBO carrying the 6A chromosome from shallot (FF+6A, 2n=2x+1=17), were 6,659±238 and 14,096±635 µmol TE/100 g dry weight (DW). We hypothesized that the 6A chromosome encoded the enhancement of polyphenol production. Subsequently, we created Kamaboko containing 4.8% JBO powder or 4.8% FF+6A powder. The ORAC value of each modified Kamaboko product was increased to 376±24 µmol TE/100 g FW for the JBO powder and to 460±16 µmol TE/100 g FW for the FF+6A powder, respectively. We next created Kamaboko containing 9.0% JBO powder or 9.0% FF+6A powder and the ORAC values of the respective modified Kamaboko products was increased to 671±16 and 740±21 µmol TE/100 g FW, i.e., 4.1- and 4.5-times the value of plain Kamaboko. Consequently, taking into consideration the sensory evaluation regarding taste and appearance as well, the use of Kamaboko supplemented with 4.8% FF+6A powder is recommended.

Antioxidant activity of the giant jellyfish Nemopilema nomurai measured by the oxygen radical absorbance capacity and hydroxyl radical averting capacity methods.

The giant jellyfish Nemopilema nomurai (reaching sizes of up to 2 m diameter and 150 kg), which forms dense blooms, has caused extensive damage to fisheries by overloading trawl nets, while its toxic nematocysts cause dermatological symptoms. Giant jellyfish are currently discarded on the grounds of pest control. However, the giant jellyfish is considered to be edible and is part of Chinese cuisine. Therefore, we investigated whether any benefits for human health may be derived from consumption of the jellyfish in order to formulate medicated diets. Antioxidant activity of Nemopilema nomurai was measured using the oxygen radical absorbance capacity (ORAC) and hydroxyl radical averting capacity (HORAC) methods. Based on the results, the ORAC value of the giant jellyfish freeze-dried sample was 541 µmol trolox equivalent (TE)/100 g and the HORAC value was 3,687 µmol gallic acid equivalent (GAE)/100 g. On the other hand, the IC50 value of hydroxyl radical scavenging activity measured by using the electron spin resonance method was 3.3%. In conclusion, the results suggest that the freeze-dried powder of the giant jellyfish Nemopilema nomurai is a potentially beneficial food for humans.

Structural variation of Tn10 that carries tetB found in fish farm bacteria.

Three variants of the composite transposon Tn10 were extracted from transferable plasmids of fish farm bacteria. These variants were identical in insertions with IS10, but differed in another class I transposon insertion and a region of homologous recombination downstream of tetB.

Antioxidative activity of the buckwheat polyphenol rutin in combination with ovalbumin.

Buckwheat flour is well known for its highly antioxidative ingredient, rutin. We have undertaken to examine alterations in the characteristics of rutin treated with various proteins. In this study, the radical scavenging activities of a rutin-ovalbumin complex were examined. Dissolved rutin hydrate and ovalbumin were combined and boiled in water for 10 min. In the resulting rutin-ovalbumin complex, a new high molecular weight peak was detected using gel permeation chromatography analysis, and an existing high molecular weight area of ovalbumin was observed to be increased by the addition of rutin. This suggested that ovalbumin molecules produce a complex through their interaction with rutin. Alkaline luminol chemiluminescence and electron spin resonance analysis revealed the formation of a rutin-ovalbumin complex that markedly enhanced the peroxyl, but not the hydroxyl, radical scavenging activity of rutin. Rutin also demonstrated antioxidative activity against hydroxyl radicals in a DNA protection assay. We therefore conclude that, compared with ovalbumin or rutin alone, the rutin-ovalbumin complex has improved antioxidative activities in the form of enhanced peroxyl radical scavenging activity and DNA protection from apurinic/apyrimidinic site formation caused by hydroxyl radicals.

Antioxidant activity of fish sauces including puffer (Lagocephalus wheeleri) fish sauce measured by the oxygen radical absorbance capacity method.

Fish sauces are fermented seasonings traditionally used throughout Asia, including Japan. Here, we report on the antioxidant activity of 30 fish sauces, among them a puffer fish sauce developed specifically for this study. To determine the antioxidant activity (i.e., the peroxyl radical elimination capacity) of the fish sauces, the oxygen radical absorbance capacity (ORAC) was measured. ORAC values ranged between 104 µmol (flatfish sauce 1) and 103 µmol (sandfish sauce) trolox equivalent (TE)/100 ml of fish sauce. Hydroxyl radical scavenging activity (IC50) was measured using electron spin resonance. IC50 values ranged between 0.081% (puffer fish sauce) and 0.653% (sardine fish sauce 7). Puffer fish sauce had a high ORAC value (8,365 µmol TE/100 ml) and the highest hydroxyl radical scavenging activity (0.081). The relationship between the ORAC and IC50 values of the 30 fish sauces was determined to be intermediate (r =-0.521, p=0.01).

Efficacy of puffer fish (Takifugu rubripes) sauce in reducing hydroxyl radical damage to DNA assessed using the apurinic/apyrimidinic site method.

Apurinic/apyrimidinic (AP) sites are frequently observed DNA lesions when cells are exposed to hydroxyl radicals. We developed a new method for measurement of the antioxidative activity of foods using the occurrence frequency of AP sites on DNA. Combined with the electron spin resonance (ESR) method as a standard method, we examined whether fish and soy sauces including puffer fish [Takifugu rubripes (Temminck et Schlegel)] sauce could protect DNA from damage caused by hydroxyl radicals. The results showed that the ratios of DNA protection by puffer fish sauce, salmon fish sauce, sandfish fish sauce (Shottsuru), colorless soy sauce, squid fish sauce (Ishiru), dark color soy sauce and light color soy sauce were 68.9, 67.0, 60.1, 49.7, 34.1, 28.2 and -4.4%, respectively. Puffer, salmon, and sandfish fish sauces showed high ratios of DNA protection against hydroxyl radicals. On the other hand, IC(50) values of hydroxyl radical scavenging of the puffer, salmon, sandfish, squid fish sauces and colorless, dark and light color soy sauces were 0.20, 0.09, 4.16, 0.26% and 0.28, 0.14 and 0.18%, respectively. Though the puffer fish sauce exhibited the highest level of DNA protection among the examined samples and a high hydroxyl radical scavenging capability, a correlation between the radical scavenging capability and DNA protection against hydroxyl radicals among the examined fish and soy sauces was not found.

Isolation of multidrug-resistant Stenotrophomonas maltophilia from cultured yellowtail (Seriola quinqueradiata) from a marine fish farm.

Six strains of multidrug-resistant Stenotrophomonas maltophilia were isolated from cultured yellowtail. The strains were divided into two clusters based on the 16S rRNA genes, and all of them contained L1 metallo-beta-lactamase and L2 beta-lactamase genes. Differences in the intercluster divergence between the lactamase genes suggest that horizontal transfer of the genes occurred.

Similarity of tetracycline resistance genes isolated from fish farm bacteria to those from clinical isolates.

Tetracycline-resistant (Tet(r)) bacteria were isolated from fishes collected at three different fish farms in the southern part of Japan in August and September 2000. Of the 66 Tet(r) gram-negative strains, 29 were identified as carrying tetB only. Four carried tetY, and another four carried tetD. Three strains carried tetC, two strains carried tetB and tetY, and one strain carried tetC and tetG. Sequence analyses indicated the identity in Tet(r) genes between the fish farm bacteria and clinical bacteria: 99.3 to 99.9% for tetB, 98.2 to 100% for tetC, 99.7 to 100% for tetD, 92.0 to 96.2% for tetG, and 97.1 to 100% for tetY. Eleven of the Tet(r) strains transferred Tet(r) genes by conjugation to Escherichia coli HB-101. All transconjugants were resistant to tetracycline, oxycycline, doxycycline, and minocycline. The donors included strains of Photobacterium, Vibrio, Pseudomonas, Alteromonas, Citrobacter, and Salmonella spp., and they transferred tetB, tetY, or tetD to the recipients. Because NaCl enhanced their growth, these Tet(r) strains, except for the Pseudomonas, Citrobacter, and Salmonella strains, were recognized as marine bacteria. Our results suggest that tet genes from fish farm bacteria have the same origins as those from clinical strains.