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Brucella melitensis: The evaluation of a putative hemagglutinin gene's effect on virulence in the caprine model.

机译：布鲁氏菌：在鼠模型中评估推定的血凝素基因对毒力的影响。

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摘要

Brucella melitensis is a facultative intracellular bacterial pathogen that causes abortions in goats and sheep and Malta fever in humans. The zoonotic disease brucellosis causes severe economic losses in the Mediterranean region and parts of Africa, Asia, and Latin America.; With the completion of the genomic sequences of B. abortus 2308 and B. melitensis 16M, no classical virulence factors were found; and the chromosomes were virtually identical. However, in B. melitensis, a putative hemagglutinin gene was identified which is absent in B. abortus. The possibility of the hemagglutinin gene being a potential virulence factor was evaluated via gene replacement/deletion in B. melitensis and expression in trans in B. abortus.; The hemagglutinin gene was PCR-amplified, cloned into pBBRIMCS-4, and electroporated into B. abortus 2308 yielding B. abortus 2308-QAE. A kanamycin-Region E-kanamycin disrupted gene fragment (KAN-E-KAN) was also generated and electroporated into B. melitensis 16M. The resulting mutants were characterized biochemically to confirm its Brucella origin and screened by antibiotic selective pressure.; A colonization study of non-pregnant goats infected with B. abortus 2308, B. melitensis 16M, B. abortus 2308-QAE, or B. melitensis 16MDeltaE revealed no attenuation of the 16MDeltaE mutant when compared to 16M at 4, 7, and 21 days post inoculation. The study also showed that both the variant and the mutant were capable of infecting and disseminating throughout the host.; All four strains were introduced into the pregnant goat model and evaluated for pathogenicity. Pregnancy/delivery results revealed 27%, 78%, 67%, and 50% abortion rates in goats infected with 2308, 16M, 2308-QAE, and 16MDeltaE, respectively. Bacterial culture of tissues from 2308, 16M, 2308-QAE, 16MDeltaE-exposed goats revealed 45 %, 79%, 75%, and 100% colonization of dam/kid pairs, respectively. The expression of the B. melitensis 16M hemagglutinin gene in trans in 2308-QAE revealed a significant (p≤0.05) increase in colonization and abortion rates when compared to B. abortus 2308, mimicking the virulence of B. melitensis 16M in pregnant goats. The B. melitensis 16MDeltaE disruption mutant colonization and abortion rates demonstrated no attenuation in colonization but did show a 28% reduction in abortions when compared to B. melitensis 16M.
机译：布鲁氏菌是一种兼性的细胞内细菌病原体，会导致山羊和绵羊的流产和人类的马耳他热。人畜共患疾病布鲁氏菌病在地中海区域以及非洲，亚洲和拉丁美洲的部分地区造成严重的经济损失。随着流产双歧杆菌2308和melitensis 16M的基因组序列的完成，没有发现经典的毒力因子。染色体实际上是相同的。然而，在肉芽孢杆菌中，鉴定了推定的血凝素基因，该基因在流产芽孢杆菌中不存在。血凝素基因是潜在的致病因子的可能性通过在棉铃虫中的基因置换/缺失以及在流产柏油中的反式表达来评估。将血凝素基因PCR扩增，克隆到pBBRIMCS-4中，并电穿孔到流产双歧杆菌2308中，产生流产双歧杆菌2308-QAE。还产生了卡那霉素-区域E-卡那霉素破坏的基因片段（KAN-E-KAN），并将其电穿孔到meliensis B. 16M中。对所得突变体进行生化鉴定，以确认其布鲁氏菌起源，并通过抗生素选择压力进行筛选。一项针对未怀孕山羊感染了流产双歧杆菌2308，melitensis 16M，流产双歧杆菌2308-QAE或melitensis 16MDeltaE的山羊定殖研究表明，与4、7和21日的16M相比，16MDeltaE突变体没有减弱。接种后几天。研究还表明，变异体和突变体均能够感染并在整个宿主中传播。将所有四个菌株引入怀孕的山羊模型并评估致病性。怀孕/分娩结果显示，分别感染2308、16M，2308-QAE和16MDeltaE的山羊的流产率分别为27％，78％，67％和50％。暴露于2308、16M，2308-QAE和16MDeltaE的山羊的组织的细菌培养分别显示了dam / kids对的定殖率为45％，79％，75％和100％。相比于流产双歧杆菌2308，在2308-QAE中B. melitensis 16M血凝素基因的反式表达表明定殖和流产率显着提高（p≤0.05），从而模仿了怀孕山羊中B.melitensis 16M的毒力。 B. melitensis 16MDeltaE破坏突变体的定殖和流产率表明定殖没有减弱，但与B. melitensis 16M相比，流产减少了28％。

著录项

  • 作者

    Perry, Quinesha Laticia.;

  • 作者单位

    Louisiana State University and Agricultural & Mechanical College.;

  • 授予单位 Louisiana State University and Agricultural & Mechanical College.;
  • 学科 Biology Molecular.; Biology Microbiology.; Biology Animal Physiology.; Biology Veterinary Science.
  • 学位 Ph.D.
  • 年度 2007
  • 页码 100 p.
  • 总页数 100
  • 原文格式 PDF
  • 正文语种 eng
  • 中图分类 分子遗传学;微生物学;生理学;动物学;
  • 关键词

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