發(fā)布時(shí)間：2018-04-13 22:26

  本文選題：布魯菌 + 鐵轉(zhuǎn)錄調(diào)控因子��； 參考：《動(dòng)物醫(yī)學(xué)進(jìn)展》2017年10期

【摘要】：為了構(gòu)建牛布魯菌S2308(簡(jiǎn)稱(chēng)S2308)的鐵轉(zhuǎn)錄調(diào)控因子rirA基因突變株(S2308ΔrirA),探討該基因?qū)ψ陨砩L(zhǎng)的影響以及其對(duì)不同類(lèi)型細(xì)胞黏附侵襲和胞內(nèi)繁殖的作用。利用同源重組和抗性替換的方法,以卡那抗性基因替換rirA基因,獲得突變株S2308ΔrirA。將親本株S2308、疫苗株RB51和突變株S2308ΔrirA在相同營(yíng)養(yǎng)條件下培養(yǎng),觀察其振蕩培養(yǎng)時(shí)的生長(zhǎng)變化趨勢(shì)和靜置培養(yǎng)時(shí)的聚集狀態(tài)。將各菌株侵染人胚胎滋養(yǎng)層細(xì)胞(HPT-8)和小鼠巨噬細(xì)胞(RAW264.7),分別檢測(cè)其黏附侵襲能力和胞內(nèi)生存能力。結(jié)果顯示,成功獲得了布魯菌rirA基因突變株且10代內(nèi)未發(fā)生回復(fù)性突變;與親本株相比,S2308ΔrirA在相同體外培養(yǎng)條件下其生長(zhǎng)趨勢(shì)未發(fā)生明顯改變,且其凝集狀態(tài)與親本株類(lèi)似;黏附侵襲試驗(yàn)顯示,S2308ΔrirA對(duì)HPT-8細(xì)胞的黏附侵襲能力顯著強(qiáng)于親本株,而其對(duì)RAW264.7的黏附侵襲能力在一定程度上弱于親本株;布魯菌胞內(nèi)生存試驗(yàn)發(fā)現(xiàn),布魯菌侵染HPT-8細(xì)胞24h后,其胞內(nèi)細(xì)菌數(shù)量顯著升高,而侵染巨噬細(xì)胞24h后,S2308ΔrirA的繁殖能力明顯低于親本株。這些結(jié)果表明,鐵轉(zhuǎn)錄調(diào)控因子rirA基因參與了布魯菌胞內(nèi)寄生的過(guò)程,并與菌株的毒力強(qiáng)弱存在密切聯(lián)系。
[Abstract]:In order to construct the mutant of iron transcription regulator rirA gene of Brucella bovis S2308 螖 RIA, the effect of this gene on its own growth and its effects on the adhesion invasion and intracellular reproduction of different types of cells were investigated.By means of homologous recombination and resistance substitution, the rirA gene was replaced by kana-resistant gene, and the mutant S2308 螖 rirA was obtained.The parent strain S2308, vaccine strain RB51 and mutant strain S2308 螖 rirA were cultured under the same nutritional conditions.Human embryonic trophoblastic cells (HPT-8) and mouse macrophages (RAW264.7) were infected with each strain.The results showed that the mutant of Brucella rirA gene was obtained successfully and there was no reverse mutation within 10 generations, and the growth trend of S2308 螖 rirA was not changed obviously under the same culture condition, and its agglutination state was similar to that of parent strain.The adhesion and invasion test showed that S2308 螖 rirA had significantly stronger adhesion and invasion ability to HPT-8 cells than its parent strain, but its adhesion and invasion ability to RAW264.7 was weaker than that of parent strain to some extent, and Brucella spp. After 24 h invasion of HPT-8 cells, Brucella spp.The number of intracellular bacteria increased significantly, and the reproductive ability of S2308 螖 rirA was significantly lower than that of parent strain after 24 h infection of macrophages.These results suggest that the iron transcription regulator rirA gene is involved in the intracellular parasitism of Brucella and is closely related to the virulence of the strain.
【作者單位】： 石河子大學(xué)兵團(tuán)動(dòng)物疾病防控重點(diǎn)實(shí)驗(yàn)室;石河子大學(xué)新疆地方與民族高發(fā)病教育部重點(diǎn)實(shí)驗(yàn)室;新疆畜牧科學(xué)院獸醫(yī)研究所;
【基金】：新疆生產(chǎn)建設(shè)兵團(tuán)動(dòng)物疾病防控重點(diǎn)實(shí)驗(yàn)室開(kāi)放課題(BTDJ06) 國(guó)家自然科學(xué)基金項(xiàng)目(31572491,31660705)
【分類(lèi)號(hào)】：S852.614

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