Transposon-derived Brucella abortus rough mutants are attenuated and exhibit reduced intracellular survival
Transposon-derived Brucella abortus rough mutants are attenuated and exhibit reduced intracellular survival
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DOI：
10.1128/iai.66.3.1008-1016.1998
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发表时间：
1998-03-01
影响因子：
3.1
通讯作者：
Ficht, TA
中科院分区：
文献类型：
--
作者：
Allen, CA;Adams, LG;Ficht, TA
牛种布鲁氏菌O抗原是其主要的毒力决定因子，但由于缺乏对牛种布鲁氏菌和犬种布鲁氏菌的致病力的遗传学定义，因此对牛种布鲁氏菌O抗原的致病力和存活力的研究存在一定的困难。流产粗糙突变缺陷的镉抗原呈递。Tn 5插入位点侧翼DNA的序列分析可用于验证编码脂多糖（LPS）生物合成功能的基因中的插入。毫不奇怪，每种粗糙突变体在小鼠中的存活率都被减弱，但观察到突变体之间意想不到的差异。为了确定观察到的差异的基础，在体外研究了粗糙LPS的结构和这些突变体对个体杀伤机制的敏感性。所有的精灵都是B。与光滑亲本菌株相比，牛流产粗糙突变体在对补体介导的裂解的敏感性方面表现出4- 5个对数单位的增加。这些微生物对过氧化氢的敏感性变化不大，这与O抗原无法排除相对较小的分子一致。敏感性多粘菌素B，这是作为一个模型的阳离子，两亲性肽类似于在吞噬细胞中发现的防御素，揭示了粗糙的突变体之间的生存差异类似于在小鼠中观察到的。特别是一个突变体表现出对多粘菌素Hf的超敏反应，并降低了小鼠的存活率。该突变体的特征在于截短的粗糙LIPS。该突变体的DNA序列分析揭示了在编码磷酸甘露变位酶（pmm）的基因中的转座子中断，这表明除了O抗原之外，这种活性可能是合成全长核心多糖所必需的。B。流产O抗原似乎是小鼠细胞外和细胞内存活所必需的。
The O antigen of Brucella abortus has been described as a major virulence determinant based an the attenuated survival of fortuitously isolated rough variants, However, the lack of genetic definition of these mutants and the virulence of naturally occurring rough species, Brucella ovis and Brucella canis, has confused interpretation, To better characterize the role of O antigen in virulence and survival, transposon mutagenesis was used to generate B. abortus rough mutants defective in Cd-antigen presentation. Sequence analysis of DNA flanking the site of Tn5 insertion mas used to verify insertion in genes encoding lipopolysaccharide (LPS) biosynthetic functions. Not surprisingly, each of the rough mutants was attenuated for survival in mice, but unexpected differences among the mutants were observed. In an effort to define the basis for the observed differences, the structure of the rough LPS and the sensitivity of these mutants to individual killing mechanisms were examined in vitro. AII elf the B. abortus rough mutants exhibited a 4- to 5-log-unit increase, compared to the smooth parental strain, in sensitivity to complement-mediated lysis. Little change was evident in the sensitivity of these organisms to hydrogen peroxide, consistent with an inability of O antigen to exclude relatively small molecules. Sensitivity to polymyxin B, which was employed as a model cationic, amphipathic peptide similar to defensins found in phagocytic cells, revealed survival differences among the rough mutants similar to those observed in the mouse. One mutant in particular exhibited hypersensitivity to polymyxin Hf and reduced survival in mice. This mutant was characterized by a truncated rough LIPS. DNA sequence analysis of this mutant revealed a transposon interruption in the gene encoding phosphomannomutase (pmm), suggesting that this activity may be required for the synthesis of a full-length core polysaccharide in addition to O antigen. B. abortus O antigen appears to be essential for extra-and intracellular survival in mice.