Serum lipid level was investigated in mice by the intraperitoneal injection of . At a sublethal dose (0.3 mg wet cells) a considerable increase in triglyceride content was observed in mouse serum at 1 day postinjection. In total cholesterol levels no appreciable difference was shown between the control and injected groups. However, serum triglyceride level in mice injected with nonpathogenic strain also increased in serum at 4 hr postinjection similarly to that injected with pathogenic strain. Serum triglyceride level in mice changed effectively with increasing doses (0.2-0.8 mg) of the microorganism, and a marked increase of the level at 6 hr postinjection was seen in the injected animals with an increase in doses, but total cholesterol levels did not change in both the control and injected groups. The heatstable hemolysin from pathogenic strain could not cause an appreciable change in triglyceride level in mouse serum. When was administered into mice and after ten minutes the endotoxin antiserum was injected, the triglyceride content in serum was not different from that of the control mice. Judging from the hypertriglyceridemia of mouse by the endotoxin in the previous report and the results in the present study, it is presumed that the endotoxin is responsible for the increase in serum triglyceride level in mice injected with pathogenic strain.

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Vibrio

(V. parahaemolyticus), one of the most important foodborne pathogens in many maritime Asian countries, is frequently associated with the consumption of seafood. Thirty eight strains of V. parahaemolyticus were isolated from seafood in Hebei province of China. Resistance to 13 antibiotics was determined using broth microdilution methods. These strains were typed by pulsed-field gel electrophoresis (PFGE) technique with short pre-processing following SfiI digestion and a typing scheme was generated. The 38 strains were grouped into 5 types with 71% pattern similarity. All the type E were isolated from Shijiazhuang, Baoding and Langfang and simultaneously resistant to ampicillin, sulfisoxazole, streptonigrin and vancomycin, suggesting a clonal relationship between these strains. The data of antimicrobial susceptibility test and PFGE profiles in this study showed a good correlation among antimicrobial susceptibility test, PFGE profiles and geographic distribution.

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The relationship of acid adaptation to the resistance of other environmental stresses was examined in

Vibrio

. Acid-adapted cells were found to have increased resistance to various stresses, including heat, crystal violet, bile, and deoxy cholic acid. However, heat-adapted cells showed no increased resistance against acid stress. Adaptation required protein synthesis, since treatment with chloramphenicol during adaptation to pH 5.3 prevented the development of acid resistance. Acid-adapted cells showed an increased amount of outer membrane protein with an apparent molecular weight of 27,000. These results show that acid-induced cross-protection involved changes in outer membrane protein composition and the known enhancement of intracellular pH homeostasis.

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Hfq plays a fundamental role in bacterial cell physiology. It can stimulate or repress the expression of certain target genes, and there is a possibility that Hfq regulates the oxidative stress response. However, how Hfq functions that in

Vibrio

parahaemolyticus

remains speculative. In this paper, we explain the functions Hfq plays in V. parahaemolyticus in the gene expression of superoxide dismutase gene and catalase gene, comparing the hfq deletion mutant strain to the parental strain. The results show that the hfq deletion mutant V. parahaemolyticus has a stronger ability to resist H₂O₂. Superoxide dismutase (SOD) and catalase (CAT) activities in the hfq deletion mutant were remarkably higher than in the parental strain. Genetic experiments indicated that the gene expression of sod and kat was up-regulated in the mutant strain. These results indicate that Hfq down-regulates CAT and SOD activity, and Hfq is associated with the oxidative stress response.

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われわれが谷津干潟土壌より単離した

Vibrio

KN1699株は，キチン分解によりヘテロ二糖 [β-N-アセチル-D-グルコサミニル-(1,4)-D-グルコサミン(GlcNAc-GlcN)] を生成するユニークな分泌性のキチン分解酵素系を有している．GlcNAc-GlcNの生成メカニズムを調べたところ，本ヘテロ二糖は，GHファミリー18キチナーゼ(Pa-Chi)の作用によるキチンからジ-N-アセチルキトビオース [(GlcNAc)₂] の生成，続いてCEファミリー4キチンオリゴ糖デアセチラーゼ(Pa-COD)の作用による(GlcNAc)₂の還元性末端側の糖の脱アセチル化により生成することを明らかにした．二つの酵素を利用してキチンから(GlcNAc)₂とGlcNAc-GlcNを量産するために，大腸菌によるリコンビント酵素の大量調製について検討した．KN1699株のゲノムから分泌シグナル配列を含む各ORFのクローニングを行い，プラスミドを作製した後，大腸菌に導入した．これを用いて各リコンビナント酵素(Pa-rChi, Pa-rCOD)の発現を行った結果，各酵素を大量に発現させ，さらに培養液中に効率的に分泌させることに成功した．続いて，これら2種のリコンビナント酵素を用いたオリゴ糖生産について検討を行った．その結果，Pa-rChi分泌組換え大腸菌を2%キチン含有培地で培養することで，粉末キチンから(GlcNAc)₂を収率60%で生産することに成功した．さらに，Pa-rCOD分泌組換え大腸菌から調製した粗酵素溶液を用いて，(GlcNAc)₂からGlcNAc-GlcNを高収率で生産することができた．今後は，生産したGlcNAc-GlcNの機能性について解析する予定である．

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Some changes of serum lipid in mice were observed by the intraperitoneal injection of endotoxin extracted from . A remarkable increase in the serum triglyceride content was shown in the poisoned mice after 12-24 hr. FFA level in the control mice gradually increased in response to the fasting periods for 12-24 hr whereas the level of FFA in the endotoxin-posioned mice was lower than that in the control mice. Total cholesterol, however, did not show an appreciable difference in the level of the control animals. The hepatic triglyceride content in the poisoned mice increased at 14 hr postintoxication, and the lysosomal triglyceride lipase activity slightly increased 3 hr after intoxication, and declined after 14 hr. There was not very much change of the triglyceride level in the serum in cortisone plus endotoxininjected mice. An increase of serum triglyceride in CCl₄-poisoned mice was not observed at 14 hr postinjection of endotoxin in spite of its high level in the serum of endotoxin-injected mice. The mechanism responsible for the development of the hypertriglyceridemia in mice after injection of endotoxin is unkown at present.

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The effects of intraperitoneal administration of pathogenic strain on carbohydrate metabolism in mice at varying administration doses were examined. The content of liver glycogen in mice at 5 hr after injection decreased in inverse proportion to the increase of the dose of viable pathogen. On the other hand, the level of blood sugar in mice changed effectively with an increase in the dose, and at the dose of 0.8 mg (2.72×10⁵ cells) a distinct hypoglycemia appeared within 5 hr after injection. The activities of hepatic phosphorylase and G-6-Pase showed a tendency similar to the rise and the fall in the levels of blood sugar. The result of this experiment suggests a complexicity of metabolic response in host to the aggression of pathogen.

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Thermostable direct hemolysin (TDH) is a major virulence factor of

Vibrio

that causes pandemic food-borne enterocolitis mediated by seafood. TDH exists as a tetramer in solution, and it possesses extreme hemolytic activity. Here, we present the crystal structure of the TDH tetramer at 1.5 Å resolution. The TDH tetramer forms a central pore with dimensions of 23 Å in diameter and ∼50 Å in depth. π-cation interactions between protomers comprising the tetramer were indispensable for hemolytic activity of TDH. The N-terminal region was intrinsically disordered outside the pore. Molecular dynamics (MD) simulations suggested that water molecules permeate freely through the central and side channel pores. These findings imply a novel membrane attachment mechanism by a soluble tetrameric pore-forming toxin.

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