Virulence, infection, and pathogenicity related genes of fish pathogens

鱼类病原体的毒力、感染性和致病性相关基因

• 批准号: 10460085
• 负责人: AOKI Takashi
• 金额: $ 7.74万
• 依托单位: Tokyo University of Fisheries
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 1998
• 资助国家: 日本
• 起止时间: 1998 至 1999
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-10460085/
• 关键词: Virulene gene; Fish pathogen; Pasteurella piscicida; Lactococcus garivieae; Edwardsiella tarda; Vibrio; genome analysis; Pasteurellapiscicida; LPS合成遺伝子群; マクロファージ耐性遺伝子

In this study, we analysed the virulence related genes of Pasteurella piscicida, Lactococcus garvieae, Edwardsiella tarda, Vibrio anguillarum and V. parahaemolyticus.Genome analysis was conducted to detect the virulence genes in P. piscicida KP9038. We have sequenced the both ends of approximately 900 clones. As a result of comparisons of these clones to the GeneBank database, it was determined that there are 13 toxin related genes, and 6 capsule and LPS synthesis genes. Characteristics of some of these virulence related genes have not detected in vitro. These results suggest that P. piscicida may express several virulence related genes only in vivo.Five different clones were isolated from the gene library of the L garvieae SA8201 (KG-) strain by immunological screening using rabbit serum against L. garvieae (KG-) phenotype cells. The amino acid sequences of the immunologically detected proteins were homologous to a processing protease, dihydropteroate synthase, trigger factor, N-acetylglucosamine-6-phosphate deacetylase, and anti-phagocytosis M protein. Five genes were specifically expressed in the virulence KG- phenotype strains.The eth hemolysin gene locus of E. tarda encodes the hemolysin EthA protein and its accessory protein EthB. We constructed mutants by site-directed marker insertion mutagenesis in ethA or ethB. Both the ethAィイD1-ィエD1 and ethBィイD1-ィエD1 mutants lacked extracellular, cell-associated, and intracellular hemolytic activities. The pathogenicity of these single gene mutants became weaker than that of original stain.Chitinase genes from V. anguillarum KV9001 and V. parahaemolyticus ATCC17802 were cloned. The deduced amino acid sequences of these genes have 71.6% identity. The vac gene was highly prevalent in V. anguillarum, and the DNA probe of the vac gene hybridized to V. alginolyticus and Beneckea proteolytica DNA. The DNA probe of the vpc gene hybridized to V. alginolyticus, V. harveyi, and V. ordalii DNA.

本研究对杀鱼巴斯德菌、加维耶乳球菌、迟缓爱德华菌、鳗弧菌和副溶血弧菌的毒力相关基因进行了分析，并对杀鱼巴斯德菌KP 9038进行了基因组分析。我们已经测序了大约900个克隆的两端。将这些克隆与GeneBank数据库比较的结果是，确定有13个毒素相关基因，以及6个荚膜和LPS合成基因。其中一些毒力相关基因的特性尚未在体外检测到。用兔抗L. garvieae SA 8201（KG-）的血清免疫筛选，从L. garvieae SA 8201（KG-）的基因文库中分离到5个不同的克隆。garvieae（KG-）表型细胞。免疫学检测的蛋白质的氨基酸序列同源的加工蛋白酶，二氢蝶酸合酶，触发因子，N-乙酰葡糖胺-6-磷酸脱乙酰酶，和抗吞噬M蛋白。KG-毒力表型菌株中特异性表达五个基因。tarda编码溶血素EthA蛋白及其辅助蛋白EthB。我们通过定点标记插入突变在ethA或ethB中构建突变体。ethAイD1-D1和ethBイD1-D1突变体都缺乏细胞外、细胞相关和细胞内溶血活性。克隆了鳗弧菌KV 9001和副溶血弧菌ATCC 17802的几丁质酶基因。推导的氨基酸序列同源性为71.6%。vac基因在鳗弧菌中高度流行，并且vac基因的DNA探针与溶藻弧菌和解蛋白贝氏菌DNA杂交。vpc基因的DNA探针与溶藻弧菌、哈维氏弧菌和奥氏弧菌DNA杂交。

项目成果

Hirono, I., S-J. Lee, and T. Aoki: "An accessory protein of the iron-regulated hemolysin of Edwardsiella tarda is necessary for hemolytic activity"Fisheries Science. 64(69. 924-928 (1998)

广野，I.，S-J。

Hirono,I.,H.Yamashita,C-L.Park,T.Yoshida,and T.Aoki: "Identification of genes in KG- phenotype of Lactococcus garvieae, a fish pathogenic bacterium, whose proteins react with anti-KG- rabbit serum."Microbial Pathogenesis. 27. 407-417 (1999)

Hirono,I.、H.Yamashita、C-L.Park、T.Yoshida 和 T.Aoki：“加氏乳球菌（一种鱼类致病菌）KG-表型中基因的鉴定，其蛋白质与抗 KG- 兔血清发生反应。

Hirono,I.: "Idetification of genes in KG phenotype of Lactococcus garvieae,a fish pathogenic bacterium, whose proteins react with anti-KG rabbit serum."Microbial Pathogenesis. 27. 407-417 (1999)

Hirono,I.：“加氏乳球菌（一种鱼类病原菌）KG 表型基因的鉴定，其蛋白质与抗 KG 兔血清发生反应。”微生物发病机制。

Hirono,I.: "An accessory protein of the iron-regulated hemolysin of Edwardsiella tarda is necessary for hemolytic activity" Fish.Sci.64(6). 924-928 (1998)

Hirono,I.：“迟缓爱德华氏菌铁调节溶血素的辅助蛋白对于溶血活性是必需的”Fish.Sci.64(6)。

Hirono, I.: "Molecular cloning of chitinase genes from Vibrio anguillarum and V. parahaemolyticus"Journal of Applied Microbiology. 84. 1175-1179 (1998)

Hirono, I.：“来自鳗弧菌和副溶血弧菌的几丁质酶基因的分子克隆”应用微生物学杂志。